Sci Rep. 2016; 6: 30783. Published online 2016 Jul 29. doi:  10.1038/srep30783 PMCID: PMC4965791 PMID: 27470078

Noninvasive low-frequency electromagnetic stimulation of the left stellate ganglion reduces myocardial infarction-induced ventricular arrhythmia

Songyun Wang,^1,* Xiaoya Zhou,^1,* Bing Huang,¹ Zhuo Wang,¹ Liping Zhou,¹ Menglong Wang,¹ Lilei Yu,^a,1 andHong Jiang^b,11Department of Cardiology, Renmin Hospital of Wuhan University, Cardiovascular Research Institute of Wuhan University, Wuhan, 430060, Hubei, China ^aEmail: moc.361@ieliluyuhw^bEmail: moc.361@gnohgnaijuhw^*These authors contributed equally to this work. Author information ? Article notes ? Copyright and License information ? Disclaimer Received 2016 Jan 29; Accepted 2016 Jul 11. Copyright © 2016, Macmillan Publishers Limited This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/

Abstract

Noninvasive magnetic stimulation has been widely used in autonomic disorders in the past few decades, but few studies has been done in cardiac diseases. Recently, studies showed that low-frequency electromagnetic field (LF-EMF) might suppress atrial fibrillation by mediating the cardiac autonomic nervous system. In the present study, the effect of LF-EMF stimulation of left stellate ganglion (LSG) on LSG neural activity and ventricular arrhythmia has been studied in an acute myocardium infarction canine model. It is shown that LF-EMF stimulation leads to a reduction both in the neural activity of LSG and in the incidence of ventricular arrhythmia. The obtained results suggested that inhibition of the LSG neural activity might be the causal of the reduction of ventricular arrhythmia since previous studies have shown that LSG hyperactivity may facilitate the incidence of ventricular arrhythmia. LF-EMF stimulation might be a novel noninvasive substitute for the existing implant device-based electrical stimulation or sympathectomy in the treatment of cardiac disorders.

Previous studies have demonstrated that the activation and remodeling of left stellate ganglion (LSG) induced by myocardial infarction^1,2 might be the immediate triggering mechanisms of ventricular arrhythmia (VA) and sudden cardiac death^3,4, and suppressing LSG neural activity might be a feasible antiarrhythmic therapy⁵. In the past decades, LSG denervation and blocking have been shown to be benefit for reducing VA⁶. However, undesirable side effects, such as cervical injury and Horner’s syndrome, have limited the clinic use of LSG denervation or blocking. Therefore, exploring a novel noninvasive approach is necessary.

Transcranial magnetic stimulation (TMS), a neurostimulation and neuromodulation technique based on the principle of electromagnetic induction of an electric field in the brain, has been proposed for treatment of a variety of neurological disorders. Previous studies has shown that TMS might mediate the cardiac rhythm by modulating the autonomic nervous system⁷. Scherlag et al.⁸ showed that exposure the vagal trunks or the chest to the low-frequency magnetic field (LF-EMF) might suppress atrial fibrillation, whereas exposure to the high-frequency field might induce atrial fibrillation by autonomic modulating. Recently, Yu et al.⁹ further demonstrated that LF-EMF stimulation of the vagal trunks or chest might suppress atrial fibrillation by inhibiting the neural activity of atrial ganglionated plexus. In this study, we hypothesized that exposure LSG to the LF-EMF might inhibit the LSG neural activity, thereby reducing VAs after acute myocardial infarction⁶.

Results

LSG was exposed to intermittent LF-EMF stimulation before left anterior descending artery occlusion in LF-EMF group (Fig. 1A–C). Both the blood pressure and heart rate were kept at a stable level during the LF-EMF stimulation. No visible damage was shown in LSG or cardiac tissue after 90?min LF-EMF treatment. All dogs developed ECG ST-segment and/or T-wave changes acutely after ligating the left anterior descending artery.

Open in a separate windowFigure 1

Schematic representation of the position of the LF-EMF (A), stimulus pattern (B) and the experimental design flow chart (C). LSG, left stellate ganglion; LF-EMF, low-frequency electromagnetic field; LAD, left anterior descending artery; MAP, monophasic action potential; HRV, heart rate variability; VA, ventricular arrhythmia.

Effect of LF-EMF stimulation on myocardial infarction-induced VAs

Figure 2A shows the representative examples of VAs in the Control group and LF-EMF group. As compared to the Control group, both the number of ventricular premature beat (VPB) and the number of non-sustained ventricular tachycardia (VT) were significantly decreased (Fig. 2B,C). Furthermore, the incidence of sustained VT/VF was significantly suppressed (75.0% vs 12.5%, P?<?0.05, Fig. 2D) in the LF-EMF group.

Figure 2

Representative examples (A) and the incidence (B–E) of AMI-induced VAs in the Control group (n?=?8) and EMF group (n?=?8). *P?<?0.05 and **P?<?0.05 as compared to the Control group. AMI, acute myocardial infarction; VPB, ventricular premature beats; VT, ventricular tachycardia; VF, ventricular fibrillation; other abbreviations as in Fig. 1.

Effect of LF-EMF stimulation on MAP

Figure 3A–F demonstrates the effect of LF-EMF on action potential duration at 90% repolarization (APD₉₀, Fig. 3A–C), pacing cycle length of action potential duration alternans (PCL, Fig. 3D–F) and the maximal slope of the restitute curve (S_max, Fig. 3G–I). As compared to group baseline, no significant change was shown in APD₉₀, PCL or S_max obtained from different sites of left ventricle in the Control group, whereas a significant change was shown in APD₉₀, PCL and Smax of those sites both at 30?min and 90?min after LF-EMF stimulation in the LF-EMF group (Fig. 3A–F).

Figure 3

Effect of LF-EMF stimulation on APD₉₀ (A,B), PCL (C,D) and Smax (E,F) in the Control group (n?=?8) and EMF group (n?=?8). *P?<?0.05 and **P?<?0.05 as compared to the group baseline; ^#P?<?0.05 and ^##P?<?0.05 as compared to the Control group. LVA, left ventricular apex; LVM, the median of left ventricle; LVB, left ventricular base; MAP, monophasic action potential; APD, action potential duration; APD₉₀, monophasic action potential duration determined at 90% of repolarization; PCL, pacing cycle length of APD alternans; BH, baseline; Smax, the maximal slope of the restitution curve, other abbreviations are identical to Fig. 1.

Effect of LF-EMF stimulation on heart rate variability

Figure 4 demonstrates that both low frequency component (LF) and the ratio between LF the high component (LF/HF) were significantly decreased by LF-EMF stimulation both at 30?min and 90?min later but not by sham LF-EMF stimulation as compared to group baseline. In comparison with group baseline, acute myocardial infarction resulted in a significant change in LF (2.54?±?0.23?ms² vs 1.72?±?0.12?ms², P?<?0.01, Fig. 4A), high frequency component (HF, 1.01?±?0.08?ms² vs 1.43?±?0.18?ms², P?<?0.01, Fig. 4B) and LF/HF (2.51?±?0.34 vs 1.20?±?0.20, P?<?0.01, Fig. 4C) in the Control group, whereas those were kept at a normal level in the LF-EMF group (LF, 1.52?±?0.1?1?ms² vs 1.68?±?0.10?ms²; HF, 1.43?±?0.12?ms² vs 1.48?±?0.13?ms²; LF/HF, 1.06?±?0.10 vs 1.14?±?0.19, all P?>?0.05, Fig. 4A–C).

Figure 4

Effect of LF-EMF stimulation on LF (A), HF (B) and LF/HF (C) in the Control group (n?=?8) and EMF group (n?=?8). *P?<?0.05 and **P?<?0.01 vs group baseline; ^#P?<?0.05 and ^##P?<?0.05 as compared to the Control group. LF, low frequency; HF, high frequency; LF/HF, the ratio between LF and HF; BH, baseline. Other abbreviations are identical to those in Fig. 1.

Effect of LF-EMF stimulation on serum norepinephrine and LSG function

In comparison with group baseline, serum norepinephrine was decreased from 180.3?±?6.8?pg/ml to 162.5?±?5.8?pg/ml at 30?min later and to 160.3?±?5.2?pg/ml at 90?min later in the LF-EMF group, whereas kept a stable level in the Control group (Fig. 5A). Furthermore, the systolic blood pressure increase in response to LSG stimulation was kept a baseline level in the Control group (Fig. 5B), whereas significantly attenuated by LF-EMF in the LF-EMF group at a voltage of 20–30?V as compared to group baseline (Fig. 5C). Take 25?V for example, the maximal systolic blood pressure increase induced by LSG stimulation was decreased from 88.3?±?15.4% to 43.1?±?6.2% (P?<?0.01) at 90?min later, whereas kept at about 90% in the Control group (Fig. 5B,C).

Figure 5

Effect of LF-EMF stimulation on serum NE (A) and LSG function (B,C) in the Control group (n?=?8) and EMF group (n?=?8). NS, P?>?0.05, *P?<?0.05 and **P?<?0.01 as compared to the Control group at the same time point. NE, norepinephrine. Other abbreviations are alike to those in Fig. 1.

Effect of LF-EMF stimulation on the neural activity of LSG

Figure 6A shows the representative examples of LSG neural activity at baseline, 30?min after LF-EMF stimulation, 90?min after LF-EMF stimulation and 15?min after acute myocardial infarction. Figure 6B,Cdemonstrates that no significant difference was shown both in the frequency and the amplitude of LSG neural activity between the Control group and the LF-EMF group. As compared to group baseline, LF-EMF stimulation resulted in a significant decrease in LSG neural activity at 30?min and 90?min later, whereas no significant change was caused by sham LF-EMF stimulation (Fig. 6B,C). Furthermore, as compared to baseline, the neural activity was significantly increased after acute myocardial infarction in the Control group (Frequency: 62.5?±?5.2impulse/min vs 112.2?±?8.1impulse/min, P?<?0.01; Amplitude: 0.18?±?0.03?mV vs 0.33?±?0.05?mV, P?<?0.01) but kept at a comparable level in the LF-EMF group (Frequency: 60.8?±?4.8impulse/min vs 65.6?±?4.8impulse/min, P?>?0.05; Amplitude: 0.19?±?0.02?mV vs 0.18?±?0.02?mV, P?>?0.05).

Open in a separate windowFigure 6

Representative examples (A) and quantitative analysis (B,C) of LSG neural activity in the Control group (n?=?8) and EMF group (n?=?8). **P?<?0.01 as compared to group baseline; ^#P?<?0.05 and ^##P?<?0.05 as compared to the Control group. All abbreviations are identical to Figs 1 and ?and22.

Discussion

In the present study, we applied LF-EMF at the body surface of LSG. Both the ventricular electrophysiological parameters (APD₉₀, PCL, Smax) and autonomic neural activity (serum norepinephrine, LSG function and LSG neural activity) were significantly affected by LF-EMF stimulation. Furthermore, the acute myocardial infarction-induced increased neural activity of LSG was significantly attenuated and the VAs was significantly reduced by LF-EMF. These findings suggested that exposure the LSG to LF-EMF might significantly reduce the neural activity of LSG, therefore reducing the incidence of VAs.

Previous studies have shown that activation of LSG facilitates, whereas inhibition of LSG protects against VAs^4,10. In the past two decades, TMS has been widely used in clinical neurology^11,12. Amounts of studies have shown that high-frequency stimulation increases cortical excitability, whereas low-frequency stimulation decreases neuronal excitability^11,12. Recently, studies also demonstrated that TMS might affect the cardiac rhythm by modulating the autonomic nervous system⁷. Scherlag et al.⁸ showed that high-frequency magnetic stimulation of the vagal nerves might induce atrial tachycardia and atrial fibrillation, which was eliminated after propranolol and atropine injection. Low-frequency stimulation of the vagal nerves, however, reduced the heart rate and decreased the voltage required to induce atrioventricular conduction block⁸. Furthermore, recent study demonstrated that exposure the heart to the LF-EMF might significantly suppress atrial fibrillation and the mechanism might be by modulating the neural activity of atrial ganglionated plexus⁹. In the present study, we found that exposure the LSG to the LF-EMF significantly reduced the serum norepinephrine, neural activity of LSG and VAs. All these indicate that noninvasive LF-EMF might reduce VAs by facilitating the autonomic rebalance, but what underlie the beneficial effects of LF-EMF on LSG was poorly defined.

In the present study, we suggested some possible mechanisms underlying the suppressing of LSG neural activity. Firstly, TMS, as an effective treatment for patients with neural disorders, has been implicated long-lasting therapeutic effects after the cessation of TMS treatment¹³. Most researchers have contributed these effects to be long-term depression (LTD) and long-term potentiation (LTP) cause the duration of the effects seemed to implicate changes in synaptic plasticity¹³. LTD is caused by low-frequency stimulation or the stimulation of a postsynaptic neuron, whereas LTP is caused by high-frequency stimulation or the stimulation of a presynaptic neuron¹³. Ca++ signal, which is known to regulate membrane excitability and modulate second messengers related to multiple receptors and signal transduction pathways, has been shown to be the major determinant whether LTD or LTD arises^14,15. Recently, Scherlag et al.⁸ also suggested that LTP or LTD was existed cause exposure the chest to the low-frequency electromagnetic field for 35?mins might result in the suppression of atrial fibrillation for 3 to 4?hours after the application of LF-EMF. In the present study, we also found that pretreatment with LF-EMF might significantly attenuated the acute myocardial infarction-induced activation of LSG neural activity and VAs, suggesting that LTP or LTD might be a potential explain for the salutary effects of LF-EMF stimulation. Secondly, previous studies have shown that TMS might also affect the expression levels of various receptors and other neuromediators, such as ?-adrenoreceptors, dopamine^11,16,17. In the present study, serum norepinephrine was significantly decreased after exposure to the LF-EMF, indicating that modulating the neurotransmitters might be one of the underlying mechanisms underlying the salutary effects of LF-EMF stimulation. Thirdly, previous studies also showed that TMS might also modulate dentritic sprouting (axon growth) and the density of synaptic contacts, and the authors suggested that these results are associated with the Brain-derived neurotrophic factor (BDNF)-tyrosine kinase B (TrkB) signaling system^18,19. BDNF, as the most abundant neurotrophin in the brain, was reported to be a major contributor to the N-methyl-D-aspartate receptor-dependent LTP and LTD processes²⁰. Wang et al.²¹ demonstrated that low-frequency TMS might reduce BDNF levels. High-frequency stimulation, however, might increase serum BDNF levels and the affinity of BDNF for TrkB receptors. Furthermore, previous studies also showed that trancranial stimulation might result in the changes in neural-related proteins, such as c-fos and tyrosine hydroxylase, which are closely related with the neural remodeling processes^6,13,20. Autonomic neural remodeling, however, plays a key role in the initiation and maintenance of VAs^4,10. All these implicate that modulating autonomic neural remodeling might be another mechanism of the antiarrhythmic effect of LF-EMF stimulation. Fourthly, the above mainly shows the underlying mechanisms of LF-EMF stimulation, but how can the LSG perceive the LF-EMF remains unknown. During the past few decades, many mechanisms, which might provide the basis for how the animals detect magnetic fields, have been proposed²². However, the magnetoreceptors have not been identified with certainty in any animal, and the mode of transduction for the magnetic sense remains unknown²³. Recently, Xie et al. hypothesized that the putative magnetoreceptor, the iron-sulphur cluster protein, might combine with the magnetoreception-related photoreceptor cryptochromes to form the basis of magnetoreception in animals and this was corroborated in pigeon retina²⁴. Furthermore, Zhang et al. further showed that the cells which had been transfected iron-sulphur cluster protein might response to the remote magnetic stimulation²⁵. All these indicate that the iron-sulphur cluster protein might be the potential magnetoreceptor for the animals to detect the magnetic fields.

Though the present study showed wonderful results, but there are some limitations in this study. First, anesthesia with pentobarbital might affect the autonomic nervous system. However, this could be counteracted cause anesthesia was maintained continuously during the whole surgery and conducted in a same fashion in both groups. Second, the coil used in this study is too large to achieve LSG-targeted stimulation without affecting the surrounding tissues. It would be a great step forward if the coils could be technically improved. Third, we only observed the effect of LF-EMF in acute canine model. Fourth, we mainly focused on the autonomic nervous system imbalance, one of the major contributors of post-infarction VAs, cause we intervened the LSG with LF-EMF in this study. It’s a great limitation that some other major factors, like area at risk, infarct size, degree of collateral flow and the possibility of any preconditioning pathway were not involved in this study. However, previous studies have shown that LSG activation might facilitate the incidence of VAs, whereas pre-emptive or post-ischemic/infarction LSG inhibition by blockage or denervation might decrease the incidence of VAs and improve the infarct size, collateral flow, contractile force both in animals^26,27,28,29 and patients^30,31. Furthermore, studies have shown that LSG stimulation might increase the likelihood of early or delayed afterdepolarization development and the initiation of reentry, thereby resulting in the incidence of VAs^32,33,34. In this study, LF-EMF stimulation of the LSG might significantly inhibit the neural activity of LSG, thereby reducing the incidence of VAs. Therefore, it’s reasonable to refer that improving the above factors might also be the potential mechanisms underlying the beneficial effects of LF-EMF stimulation, but further studies with optimized parameters and all-round considerations are required in the future.

In conclusion, the present study showed that LF-EMF stimulation might significantly reduce the neural function and neural activity of LSG. Exposure the LSG to the LF-EMF might be a feasible method for preventing the acute myocardial infarction-induced VAs. However, larger studies with optimized parameters should be done in the chronic models to verify the beneficial effect of LF-EMF stimulation.

Methods

Animal preparation

Sixteen canines weighing between 20 and 25?kg were included in this study. The experiments were approved by the Animal Ethics Committee of Wuhan University under approval number 2015–0445 and followed the guidelines outlined by the Care and Use of Laboratory Animals of the National Institutes of Health. All surgeries were performed under anesthesia with sodium pentobarbital at an initial dose of 30?mg/kg and a maintenance dose of 60?mg/h. The depth of anesthesia was evaluated by monitoring corneal reflexes, jaw tone, and alterations in cardiovascular indices. The body surface electrocardiogram was recorded throughout the experiment with a computer-based Lab System (Lead 2000B, Jingjiang Inc., Wuhan, China). The core body temperature of the dogs was kept at 36.5?±?1.5?°C. Left thoracotomy was conducted at the fourth intercostal space. At the end of the experiment, canines were a lethal dose of pentobarbital (100?mg/kg, iv).

LF-EMF

Repeated LF-EMF was supplied by the magnetic stimulation machine (YRD CCY-I, YiRuiDe Inc., Wuhan, China) with the curve 8 coil located at the body surface of the LSG (Fig. 1A). The LSG was stimulated by intermittent (8?s ON, 10?s OFF) LF-EMF stimulation with the frequency set at 1?HZ and intensity at approximately 90% of motor threshold (Fig. 1B). Motor threshold was defined as the lowest electromagnetic intensity that induced muscle contractions in the proximal forepaw and shoulder.

Monophasic action potential recording

Monophasic action potentials from the left ventricle were recorded with a custom-made Ag–AgCl catheter. A dynamic steady state pacing protocol (S1S1) was performed to determine action potential duration alternans³⁵. The pulse train was delivered at an initial cycle length slightly shorter than the sinus cycle length and the drive train of stimuli was maintained for 30?s to ensure a steady state, and then a 2-min interruption was taken to minimize the pacing memory effects. After that, another pulse train with the PCL decreased by 10?ms was delivered until action potential duration alternans appeared. Action potential duration alternans was defined as ?APD₉₀?10?ms for ?5 consecutive beats³⁶. The monophasic action potential recordings were analyzed by the LEAD 2000B work station system (Lead 2000B, Jingjiang Inc. China). The APD₉₀ was defined as the 90% repolarization duration and the diastolic interval was the time interval from the previous APD₉₀ point to the activation time of the following beat. As described in previous studies, the dynamic action potential duration restitution curves were constructed from (Diastolic interval, APD₉₀) pairs using Origin 8.0 (OriginLab, Co., Northampton, MA, USA)^35,37. Slope of the shortest diastolic interval was defined as S_max.

Measurements of heart rate variability

Spectral power for heart rate variability was analyzed on 5-minute electrocardiogram recording segments and an autoregressive algorithm was used to analyze digitized signals from the electrocardiographic recordings. The following power spectral variables were determined: HF, LF and LF/HF³⁸.

Neural recording from the LSG

To record the neural activity of the LSG, one tungsten-coated microelectrode was inserted into the fascia of the LSG and one ground lead was connected to the chest wall. The signal of the LSG was recorded with a PowerLab data acquisition system (8/35, AD Instruments, Australia) and amplified by an amplifier (DP-304, Warner Instruments, Hamden, CT, USA). The band-pass filters were set at 300?Hz to 1?kHz and the amplification ranges from 30 to 50 times³⁹. The neural activity, deflections with a signal-to-noise ratio greater than 3:1, was manually determined as described in our previous studies^39,40,41.

LSG function

LSG function was measured as the LSG stimulation-induced maximal change in systolic blood pressure as described in our previous study³⁸. High frequency stimulation (20?Hz, 0.1?ms pulse duration) was applied to the LSG using a stimulator (Grass-S88; Astro-Med, West Warwick, RI, USA). The voltage ranged from 20?V to 30?V and increased by 5?V. To eliminate the residual effect of the LSG stimulation, each stimulation should be less than 30?s and the next stimulation should be not be taken until the blood pressure returned to a normal level.

Blood sampling

Venous blood samples were collected. Serum was separated by centrifuging at 3000?rpm for 15?min at 4?°C, and stored at ?80?°C until assayed. The serum norepinephrine level was measured with a canine-specific high sensitivity ELISA kit (Nanjing Jiancheng Bioengineering Institute, Nanjing City, China)³⁸.

Measurement of the acute myocardial infarction-induced VAs

The left anterior descending coronary artery was ligated at approximately 2.5?centimeters away from its origin to induce acute myocardial infarction. The incidence and duration of the VAs induced by acute myocardial infarction during the first hour was analyzed. The VAs recorded on the ECG were defined as following⁴²: VPBs, identifiable premature QRS complexes; VT, three or more consecutive VPBs; non-sustained VT, VT terminating spontaneously within 30?s; sustained VT, VT sustained for more than 30?s; and VF, a tachycardia with random ECG morphology and an associated loss of arterial blood pressure that degenerates into ventricular asystole.

Experimental protocol

Sixteen dogs were randomly divided into LF-EMF group (n?=?8, with LF-EMF) and Control group (n?=?8, with sham LF-EMF). LF-EMF (1?HZ; stimulation time 8?s; interstimulus interval, 5?s) was delivered to the surface area of LSG for 90?minutes. As shown in Fig. 1C, monophasic action potential, heart rate variability, serum norepinephrine, LSG function and LSG neural activity were measured at baseline, 30?min and 90?min after LF-EMF treatment. Measurements of heart rate variability and LSG neural activity were repeated at 15?min after acute myocardial infarction. Furthermore, the incidence of VAs was recorded during the first hour after acute myocardial infarction.

Statistical analysis

Continuous variables are presented as the mean?±?SEM and were analyzed by t test, one-way ANOVA, or two-way repeated-measures ANOVA with a Bonferroni posthoc test. To compare the incidence of VF between groups, Fisher’s exact test was used. All data was analyzed by GraphPad Prism version 5.0 software (GraphPad Software, Inc., San Diego, CA), and two-tailed P???0.05 was considered significant.

Additional Information

How to cite this article: Wang, S. et al. Noninvasive low-frequency electromagnetic stimulation of the left stellate ganglion reduces myocardial infarction-induced ventricular arrhythmia. Sci. Rep. 6, 30783; doi: 10.1038/srep30783 (2016).

Acknowledgments

This work was supported by the grants from National Natural Science Foundation of China No. 81270339, No. 81300182, No. 81530011, No. 81570463, grant from the Natural Science Foundation of Hubei Province No. 2013CFB302, and grants from the Fundamental Research Funds for the Central Universities No. 2042014kf0110 and No. 2042015kf0187.

Footnotes

Author Contributions S.W. and X.Z. wrote the main manuscript text and prepared figures; B.H., Z.W., L.Z. and M.W. performed experiments and anlalyzed data; L.Y. and H.J. designed the project and revised the paper. All authors reviewed and approved the final version.

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Articles from Scientific Reports are provided here courtesy of Nature Publishing Group

Biosci Rep. 2016 Dec; 36(6): e00420. Published online 2016 Dec 5. Prepublished online 2016 Oct 25. doi:  10.1042/BSR20160082 PMCID: PMC5137536

Novel protective effects of pulsed electromagnetic field ischemia/reperfusion injury rats

Fenfen Ma,^*,1 Wenwen Li,^†‡,1 Xinghui Li,^† Ba Hieu Tran,^§ Rinkiko Suguro,^§ Ruijuan Guan,^† Cuilan Hou,^† Huijuan Wang,^? Aijie Zhang,^† Yichun Zhu,^† and YiZhun Zhu^?¶,2*Department of Pharmacy, Shanghai Pudong Hospital, Fudan University, Shanghai 201399, China ^†Shanghai Institute of Immunology & Department of Immunobiology and Microbiology, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China ^‡Shanghai Key Laboratory of Bioactive Small Molecules and Research Center on Aging and Medicine, Department of Physiology and Pathophysiology, Shanghai Medical College, Fudan University, Shanghai 200032, China ^§Department of Pharmacology, School of Pharmacy, Fudan University, Shanghai 201203, China ^?Longhua Hospital, Shanghai University of Tradition Chinese Medicine, Shanghai 201203,China ^¶Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore 119228, Singapore ¹These authors contributed equally to the article. ²To whom correspondence should be addressed (email nc.ude.naduf@zyuhz). Author information ? Article notes ? Copyright and License information ? Received 2016 Mar 17; Revised 2016 Oct 11; Accepted 2016 Oct 17. Copyright © 2016 The Author(s) This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution Licence 4.0 (CC BY).

Abstract

Extracorporeal pulsed electromagnetic field (PEMF) has shown the ability to regenerate tissue by promoting cell proliferation. In the present study, we investigated for the first time whether PEMF treatment could improve the myocardial ischaemia/reperfusion (I/R) injury and uncovered its underlying mechanisms.

In our study, we demonstrated for the first time that extracorporeal PEMF has a novel effect on myocardial I/R injury. The number and function of circulating endothelial progenitor cells (EPCs) were increased in PEMF treating rats. The in vivo results showed that per-treatment of PEMF could significantly improve the cardiac function in I/R injury group. In addition, PEMF treatment also reduced the apoptosis of myocardial cells by up-regulating the expression of anti-apoptosis protein B-cell lymphoma 2 (Bcl-2) and down-regulating the expression of pro-apoptosis protein (Bax). In vitro, the results showed that PEMF treatment could significantly reduce the apoptosis and reactive oxygen species (ROS) levels in primary neonatal rat cardiac ventricular myocytes (NRCMs) induced by hypoxia/reoxygenation (H/R). In particular, PEMF increased the phosphorylation of protein kinase B (Akt) and endothelial nitric oxide synthase (eNOS), which might be closely related to attenuated cell apoptosis by increasing the releasing of nitric oxide (NO). Therefore, our data indicated that PEMF could be a potential candidate for I/R injury.Keywords: apoptosis, Bax, B-cell lymphoma 2 (Bcl-2), ischaemia/reperfusion (I/R) injury, pulsed electromagnetic field (PEMF)

INTRODUCTION

Hypertension, arrhythmia, myocardial infarction (MI) and myocardial ischaemia/reperfusion (I/R) injury are all the most common cardiac diseases, which are the major causes of mortality in the world [1]. Among them, myocardial I/R injury is the most important cause of cardiac damage. Its pathological process is closely related to postoperative complications [2,3] caused by coronary artery vascular formation, coronary revascularization and heart transplantation. After myocardium suffered severe ischaemia, restoration of the blood flow is a prerequisite for myocardial salvage [2]. However, reperfusion may induce oxidative stress [4], inflammatory cell infiltration and calcium dysregulation [5]. All these players contribute to the heart damage such as contraction and arrhythmias [6], generally named myocardial I/R injury. Recently, more and more evolving therapies have been put into use for I/R injury.

Pulsed electromagnetic field (PEMF) is the most widely tested and investigated technique in the various forms of electromagnetic stimulations for wound healing [7], alleviating traumatic pain and neuronal regeneration [8,9]. The rats were randomly divided into PEMF-treated (5 mT, 25 Hz, 1 h daily) and control groups. They hypothesized the possible mechanism that PEMF would increase the myofibroblast population, contributing to wound closure during diabetic wound healing. It is a non-invasive and non-pharmacological intervention therapy. Recent studies indicated that PEMF also stimulated angiogenesis in patients with diabetes [10], and could improve arrhythmia, hypertension and MI [1]. The MI rats were exposed to active PEMF for 4 cycles per day (8 min/cycle, 30±3 Hz, 6 mT) after MI induction. In vitro, PEMF induced the degree of human umbilical venous endothelial cells tubulization and increased soluble pro-angiogenic factor secretion [VEGF and nitric oxide (NO)] [7]. However, the role of PEMF in ischaemia and reperfusion diseases remains largely unknown. Our study aimed to investigate the effects of PEMF preconditioning on myocardial I/R injury and to investigate the involved mechanisms.

In our study, we verified the cardioprotective effects of PEMF in myocardial I/R rats and the anti-apoptotic effects of PEMF in neonatal rat cardiac ventricular myocytes (NRCMs) subjected to hypoxia/reoxygenation (H/R). We hypothesized that PEMF treatment could alleviate myocardial I/R injury through elevating the protein expression of B-cell lymphoma 2 (Bcl-2), phosphorylation of protein kinase B (Akt). Meanwhile, it could decrease Bax. We emphatically made an effort to investigate the MI/R model and tried to uncover the underlying mechanisms.

MATERIALS AND METHODS

Animals

Male, 12-week-old Sprague Dawley (SD) rats (250–300 g) were purchased from Shanghai SLAC Laboratory Animal. Animals were housed in an environmentally controlled breeding room and given free access to food and water supplies. All animals were handled according to the “Guide for the Care and Use of Laboratory Animals” published by the US National Institutes of Health (NIH). Experimental procedures were managed according to the Institutional Aminal Care and Use Committee (IACUC), School of Pharmacy, Fudan University.

The measurement of blood pressure in SHR rats

At the end of 1 week treatment with PEMF, the rats were anesthetized with chloral hydrate (350 mg/kg, i.p.), the right common carotid artery (CCA) was cannulated with polyethylene tubing for recording of the left ventricle pressures (MFlab 200, AMP 20130830, Image analysis system of physiology and pathology of Fudan University, Shanghai, China).

Myocardial I/R injury rat model and measurement of infarct size

All the rats were divided into three groups: (1) Sham: The silk was put under the left anterior descending (LAD) without ligation; (2) I/R: Hearts were subjected to ischaemia for 45 min and then reperfusion for 4 h; (3) I/R + PEMF: PEMF device was provided by Biomobie Regenerative Medicine Technology. The I/R rats were pre-exposed to active PEMF for 2 cycles per day (8 min per cycle), whereas other two groups were housed with inactive PEMF generator. I/R was performed by temporary ligation of the LAD coronary artery for 45 min through an incision in the fourth intercostal space under anaesthesia [11]. Then, the ligature was removed after 45 min of ischaemia, and the myocardium was reperfused for 4 h. Ischaemia and reperfusion were confirmed and monitored by electrocardiogram (ECG) observation. The suture was then tightened again, and rats were intravenously injected with 2% Evans Blue (Sigma–Aldrich). After explantation of the hearts, the left ventricles were isolated, divided into 1 mm slices, and subsequently incubated in 2% 2,3,5-triphenyltetrazolium chloride (TTC; Sigma–Aldrich) in 0.9% saline at 37°C for 25 min, to distinguish infarcted tissue from viable myocardium. These slices were flushed with saline and then fixed in 10% paraformaldehyde in PBS (pH 7.4) for 2 h. Next, the slices were placed on a glass slice and photographed by digital camera, the ImageJ software (NIH) was used in a blind fashion for analysis. Infarct size was expressed as a ratio of the infarct area and the area at risk [12].

Pulsed electromagnetic field treatment

PEMF were generated by a commercially available healing device (length × width × height: 7 cm × 5cm × 3cm) purchased from Biomoble Regenerative Medicine Technology. The adapter input voltage parameter is approximately 100–240 V and output parameter is 5 V. Fields were asymmetric and consisted of 4.5 ms pulses at 30±3 Hz, with an adjustable magnetic field strength range (X-axis 0.22±0.05 mT, Y-axis 0.20±0.05 mT, Z-axis 0.06±0.02 mT). The I/R rats were housed in custom designed cages and exposed to active PEMF for 2 cycles per time (8 min for 1 cycle), whereas the I/R rats were housed in identical cages with inactive PEMF generator. For in vitro study, culture dishes were directly exposed to PEMF for 1–2 cycles as indicated (8 min for 1 cycle, 30 Hz, X-axis 0.22 mT, Y-axis 0.20 mT, Z-axis 0.06 mT) [1]. The background magnetic field in the room area of exposure animals/samples and controls is 0 mT.

Detection of myocardium apoptosis

Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) assay was applied to analyse cardiomyocyte apoptosis. Heart samples were first fixed in 10% formalin and then paraffin embedded at day 14. Then, the hearts were cut into 5 ?m sections. TUNEL staining was carried out as described previously [12]. When apoptosis occurred, cells would look green.

Determination of myocardial enzymes in plasma

Blood samples were collected after haemodynamic measurement and centrifuged at 3000 g for 15 min to get the plasma. Creatine kinase (CK), lactate dehydrogenase (LDH), creatine kinase isoenzyme-MB (CKMB) and ?-hydroxybutyrate dehydrogenase (HBDH) were quantified by automatic biochemical analyzer (Cobas 6000, Roche). All procedures were performed according to the manufacturer’s protocols.

Myocardium cells morphology via TEM

At the end of the experiment, sections from myocardial samples of left ventricular were immediately fixed overnight in glutaraldehyde solution at 4°C and then incubated while protected from light in 1% osmium tetroxide for 2 h. After washing with distilled water for three times (5 min each), specimens were incubated in 2% uranyl acetate for 2 h at room temperature and then dehydrated in graded ethanol concentrations. Finally, sections were embedded in molds with fresh resin. The changes in morphology and ultrastructure of the myocardial tissues were observed and photographed under a TEM [13].

Scal-1⁺/flk-1⁺ cells counting of endothelial progenitor cells

We applied antibodies to the stem cell antigen-1 (Sca-1) and fetal liver kinase-1 (flk-1) to sign endothelial progenitor cells (EPCs) as described before, and used the isotype specific conjugated anti-IgG as a negative control. The amount of Scal-1⁺/flk-1⁺ cells would be counted by flow cytometry technique [14].

Measurement of nitric oxide concentration and Western blotting

Plasma concentrations of NO were measured with Griess assay kit (Beyotime Institute of Biotechnology) according to the manufacturer’s protocol. The expressions of Bax, Bcl-2, p-Akt, Akt, p-endothelial nitric oxide synthase (eNOS), eNOS and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were assessed using Western blot as described recently [15]. Proteins were measured with Pierce BCA Protein Assay Kit (Thermo). Hippocampal protein lysates (50 mg/well) were separated using (SDS/PAGE) under reducing conditions. Following electrophoresis, the separated proteins were transferred to a PVDF membrane (Millipore). Subsequently, non-specific proteins were blocked using blocking buffer (5% skim milk or 5% BSA in T-TBS containing 0.05% Tween 20), followed by overnight incubation with primary rabbit anti-rat antibodies specific for target proteins as mentioned before (Cell Signaling Technology) at 4°C. Blots were rinsed three times (5 min each) with T-TBS and incubated with horseradish peroxidase (HRP)-conjugated secondary antibody (1:10000, Proteintech) for 2 h at room temperature. The blots were visualized by using enhanced chemiluminescence (ECL) method (Thermo). GAPDH was applied to be the internal control protein. Intensity of the tested protein bands was quantified by densitometry.

Cell culture

Primary neonatal rat cardiac ventricular myocytes (NRCMs) were collected as previously described [15]. Briefly, the ventricles of new born SD rats (1–3 days old) were minced and digested with 0.125% trypsin. Isolated cardiomyocytes were cultured in Dulbecco’s modified Eagle’s medium/F-12 (DMEM/F12, Life Technologies) supplemented with 10% (v/v) FBS (Life Technologies), 100 units/ml penicillin and 100 mg/ml streptomycin. The following experiments used spontaneously beating cardiomyocytes 48–72 hours after plating. (37°C with 5% CO₂).

Cell treatment (hypoxia/reoxygenation)

NRCMs were prepared according to the methods recently described [15]. To establish the H/R model, the cells were cultured in DMEM/F-12 without glucose and serum. The cells were exposed to hypoxia (99% N₂+5% CO₂) for 8 h, followed by reoxygenation for 16 h. The cells were pretreated with PEMF for 30 min before the H/R procedure. The control group was cultured in DMEM/F-12 with low glucose (1000 mg/l) and 2% serum under normoxic air conditions for the corresponding times.

Cell viability assays

The viability of NRCMs cultured in 96-well plates was measured by using the Cell Counting Kit-8 (CCK-8) (Dojindo Molecular Technologies) according to the manufacturer’s instructions. The absorbance of CCK-8 was obtained with a microplate reader at 450 nm.

Measurement of intracellular reactive oxygen species levels

Reactive oxygen species (ROS) levels in NRVMs were determined by dihydroethidium (DHE, Sigma–Aldrich) fluorescence using confocal microscopy (Zeiss, LSM 710). After different treatments, cells were washed with D-PBS and incubated with DHE (10 ?mol/l) at 37°C for 30 min in the dark. Then, residual DHE was removed by PBS-washing. Fluorescent signals were observed (excitation, 488 nm; emission, 610 nm) under a laser confocal microscope (Zeiss).

Data analysis

All the data were presented as means ± S.E.M. Differences were compared by one-way ANOVA analysis by using SPSS software version 19.0 (SPSS) and P value <0.05 was taken as statistically significant.

RESULTS

PEMF could lower blood pressure under treatment of certain PEMF intensity in SHR rat model (double-blind)

To determine whether PEMF has any effects on blood pressure of SHR rats, we treated SHR rats with different PEMF intensity 1–4 cycles per day for 7 days and measured the blood pressure changes via CCA. We observed that PEMF treatment could significantly lower the blood pressure in the Bioboosti WIN235 and WI215-stimulating groups than that in non-treated ones (Figures 1A and ?and1B).1B). But Bioboosti WIN221 and WC65 treating groups did not have any effects on the blood pressure in SHR rats, compared with the non-treated ones (Figures 1C and ?and1D).1D). Fields were asymmetric and consisted of 4.5 ms pulses at 30±3 Hz, with an adjustable magnetic field strength range (X-axis 0.22±0.05 mT, Y-axis 0.20±0.05 mT, Z-axis 0.06±0.02 mT). The I/R rats were housed in custom designed cages and exposed to active PEMF for 2 cycles per time (8 min for 1 cycle), whereas the I/R rats were housed in identical cages with inactive PEMF generator.

Figure 1The effect of PEMF on SHR rats in vivo. PEMF could lower the blood pressure in SHR rats. At day 7 treatment with different intensity PEMF, blood pressure was recorded via CCA [1(A), 1(B), 1(C) and 1(D)]. Data were represented as the mean ±…

According to this result, we chose Bioboosti WIN235 as our needed PEMF to carry out the following experiments.

PEMF treatment could observably improve the abundance of EPCs

Amplifying EPCs abundance and function is an active focus of research on EPCs-mediated neovascularization after I/R. Thus, the number of circulating EPCs was identified by Sca-1/flk-1 dual positive cells as described. We determined that PEMF treatment could remarkably increase the number of Scal-1⁺/flk-1⁺ cells in peripheral blood at postoperative days 7 and 14 (Figure 2).

Figure 2The effect of PEMF on the number of Scal-1⁺/flk-1⁺ cells after treating EPSc for 7 and 14 days. PEMF treatment notably increased the number of Scal-1⁺/flk-1⁺ cells after treating EPSc for 7 and 14 days. Data were represented as the mean…

Preliminary assessment of PEMF showed great protective effect against myocardial infarction/reperfusion injury (MI/RI) rat model

To examine the effect of PEMF on myocardial I/R, male SD rats were divided into three groups: Sham, I/R and I/R+ PEMF (2 cycles per day, 8 min per cycle) per day until 28 days. We observed that PEMF stimulation could significantly decrease four plasma myocardial enzymes (LDH, CK, CKMB and HBDH) in I/R rats (Figure 3A). Additionally, we found that pre-stimulating PEMF could improve the cardiac morphology via TEM, compared with I/R+ PEMF group. TEM revealed the rupture of muscular fibres, together with mitochondrial swelling, and intracellular oedema in Group I/R. The shape of nucleus was irregular, with evidence of mitochondrial overflow after cell death. Compared with Group I/R+ PEMF, less muscular fibres were ruptured, with mild swelling of mitochondria, mild intercellular oedema and less cell death. In Group Sham, the ruptured muscular fibres, mitochondrial or intracellular oedema and dead cells were not observed (Figure 3B). To further confirm protective effect of PEMF, we measured the MI size by applying TTC and Evans Blue staining in all three groups. The MI area in I/R+ PEMF group could be reduced, compared with the model rats in I/R group (Figure 3C).

Figure 3Protective effect of PEMF on I/R rats in vivo. Plasma myocardial enzymes (LDH, CK, HBDH and CKMB) content was quantified by automatic biochemical analyzer (A) (n=18 in each group). Changes on cardiac cell morphology via TEM (B) (n=6 in…

In vivo, PEMF dramatically reduced cell apoptosis induced by I/R injury

As H/R of cardiomyocytes contributed to cell death, we also detected the effect on myocardial apoptosis by using TUNEL kit, as shown in Figure 4(A). We uncovered that PEMF pretreating could dramatically decrease apoptosis of myocardial cells in I/R + PEMF group, compared with I/R group. In addition, we also found that PEMF treatment could significantly increase the expression of anti-apoptosis protein Bcl-2, p-eNOS and p-Akt and down-regulated the expression of pro-apoptosis protein Bax in the heart tissue, as shown in Figure 4(B).

Figure 4Apoptotic cardiomyocyte was identified by TUNEL analysis, apoptotic cardiomyocyte appears green whereas TUNEL-negative appears blue (A), photomicrographs were taken at ×200 magnification. Apoptosis-related protein Bcl-2, Bax, p-Akt level of different…

The effect of PEMF on cell viability in neonatal rat cardiac ventricular myocytes

To further investigate whether PEMF has the same effect in vitro, we simulated the I/R injury model in vitro. We applied NRCMs and hypoxia incubator to mimic myocardial I/R injury via H/R as described in the section ‘Materials and Methods’. We found that PEMF treatment (2 cycles) could remarkably improve cell viability, compared with the H/R group (Figure 5). For in vitro study, culture dishes were directly exposed to PEMF for 1–2 cycles as indicated (8 min for 1 cycle, 30±3 Hz, X-axis 0.22±0.05 mT, Y-axis 0.20±0.05 mT, Z-axis 0.06±0.02 mT).

Figure 5NRCMs viability measured by CCK-8 assay at the end of the treatment for 72 h. PEMF treatment enhanced the cell viability of hypoxia NRCMs. Data were represented as the mean ± S.E.M.

Specific-density PEMF could decrease intracellular ROS levels of primary cardiomyocytes subjected to hypoxia/reperfusion

As shown in Figure 6(A), NRCMs that were subjected to H/R increased significantly the ROS level, whereas the ROS level had been decreased in PEMF group (2 cycles), in contrast with the H/R group. Representative images of the ROS level were displayed in Figure 6(B). At the same time, we identified the effect on NRCMs apoptosis after suffering H/R by using TUNEL kit. As shown in Figure 6(C), cell apoptosis in the H/R group was aggravated, whereas PEMF treatment could reduce the cell death. Representative images of TUNEL staining were shown in Figure 6(D).

Figure 6PEMF protected Neonatal rat cardiac ventricular myocytes (NRCMs) from hypoxia/reoxygenation (H/R)-induced apoptosis via decreasing ROS levelat the end of the treatment for 72 h in vitro.

Effect of PEMF on NO releasing via Akt/eNOS pathway

Cultured NRCMs were treated with PEMF stimulation for 1 to 2 cycles and the supernatant and cell lysate were collected. When cells suffered H/R, intracellular levels of p-Akt, p-eNOS and Bcl-2 were decreased, whereas PEMF treatment could increase the phosphorylation of Akt, p-eNOS and Bcl-2 (Figures 7A–7C). The expression of Bax was increased when cells subjected to H/R whereas PEMF treatment reversed such increase (Figure 7C). Western blot analysis was shown in Figure 7(D) for p-Akt/Akt, Figure 7(E) for p-eNOS/eNOS, Figure 7(F) for Bcl-2 and Figure 7(G) for Bax.

Figure 7The related protein expression about the effect of PEMF on apoptosis induced by hypoxia/reoxygenationat the end of the treatment for 72 h in vitro. PEMF increased the phosphorylation of Akt, endothelial nitric oxide synthase (eNOS), and the expression…Go to:

DISCUSSION

Our present study provides the first evidence that PEMF has novel functions as follows: (1) We treated SHR rats with different PEMF intensity (8 min for 1 cycle, 30±3 Hz, X-axis 0.22±0.05 mT, Y-axis 0.20±0.05 mT, Z-axis 0.06±0.02 mT) 1–4 cycles per day for 7 days. PEMF can lower blood pressure under treatment of certain PEMF intensity in SHR rat model (double-blind). (2) PEMF has a profound effect on improving cardiac function in I/R rat model. (3) PEMF plays a vital role in inhibiting cardiac apoptosis via Bcl-2 up-regulation and Bax down-regulation. (4) In vitro, PEMF treatment also has a good effect on reducing ROS levels by Akt/eNOS pathway to release NO and improving cell apoptosis in NRCMs subjected to hypoxia.

Many previous studies showed that extracorporeal PEMF-treated(5 mT, 25 Hz, 1 h daily) could enhance osteanagenesis, skin rapture healing and neuronal regeneration, suggesting its regenerative potency [8,16,17]. And some researchers had found that PEMF therapy (8 min/cycle, 30±3 Hz, 6 mT) could improve the myocardial infarct by activating VEGF–Enos [18] system and promoting EPCs mobilized to the ischaemic myocardium [1,19]. Consistent with the previous work, our present study demonstrated that PEMF therapy could significantly alleviate cardiac dysfunction in I/R rat model.

Recent evidence suggest that circulating EPCs can be mobilized endogenously in response to tissue ischaemia or exogenously by cytokine stimulation and the recruitment of EPCs contributes to the adult blood vessels formation [19,20,21]. We hypothesized that PEMF could recruit more EPCs to the vessels. To confirm our hypothesis, we applied antibodies to the Sca-1 and flk-1 to sign EPC. The results indicated that PEMF could remarkably increase the number of EPCs in the PEMF group, compared with the I/R group.

Previous evidence indicated that when heart suffered I/R, cardiac apoptosis would be dramatically aggravated [22–24]. Myocardial apoptosis plays a significant role in the pathogenesis of myocardial I/R injury. We assumed that PEMF might play its role in improving cardiac function through inhibiting cell apoptosis. The Bcl-2 family is a group of important apoptosis-regulating proteins that is expressed on the mitochondrial outer membrane, endoplasmic reticulum membrane and nuclear membrane. Overexpression of Bcl-2 proteins blocks the pro-apoptosis signal transduction pathway, thereby preventing apoptosis caused by the caspase cascade [25]. The role Bax plays in autophagy is a debatable. Recently, new genetic and biochemical evidence suggest that Bcl-2/Bcl-xL may affect apoptosis through its inhibition of Bax [26]. Overexpression of Bax protein promotes the apoptosis signal pathway. In the present study, we applied TUNEL staining to find that PEMF has a perfect effect on cardiac cell apoptosis by regulating apoptosis-related proteins Bcl-2 and Bax [25,26,27,28].

To verify our findings in the rat model, we mimicked I/R condition in vitro by hypoxia exposure in NRCMs. Results showed that not only in vivo, hypoxia could induce cell apoptosis in vitro. And we also found that PEMF treatment could significantly alleviate cell apoptosis induced by hypoxia. At the basal level, ROS play an important role in mediating multiple cellular signalling cascades including cell growth and stress adaptation. Conversely, excess ROS can damage tissues by oxidizing important cellular components such as proteins, lipids and DNA, as well as activating proteolytic enzymes such as matrix metalloproteinases [29]. Previous studies showed that when cells were subjected to hypoxia, the intracellular ROS level would be sharply increased, and the overproduction of ROS would result in cell damage [19,30,31]. In the present study, PEMF treatment could prominently down-regulate ROS levels. We also investigated how PEMF reduced the intracellular ROS level.

NO appears to mediate distinct pathways in response to oxidative stress via AKt–eNOS pathway [32,33]. NO is identified as gaseous transmitters. In vascular tissue, NO is synthesized from L-arginine by nitric oxide synthase (NOS) and it is considered to be the endothelium-derived relaxing factor. Evidence show that the NO generation in endothelium cells was damaged in hypertensive patients [34]. NO could also prevent platelet activation and promote vascular smooth muscle cells proliferation [35]. NO generation from eNOS is considered to be endothelium-derived relaxing and ROS-related factor [36,37]. Some researchers found that bradykinin limited MI induced by I/R injury via Akt/eNOS signalling pathway in mouse heart [38]. And bradykinin inhibited oxidative stress-induced cardiomyocytes senescence by acting through BK B2 receptor induced NO release [39]. Such evidence indicated that Akt phosphorylation could activate eNOS, which lead to NO releasing, and resulted in ROS reducing. In the present study, we found that PEMF decreased ROS via Akt/eNOS pathway.

In conclusion, this is the first study suggesting that PEMF treatment could improve cardiac dysfunction through inhibiting cell apoptosis. Furthermore, in vitro, we first clarified PEMF still plays a profound effect on improving cell death and removing excess ROS via regulating apoptosis-related proteins and Akt/eNOS pathway. All these findings highlight that PEMF would be applied as a potentially powerful therapy for I/R injury cure.

Acknowledgments

We thank all of the members of the Laboratory of Pharmacology of Chen Y., Ding Y.J. for their technical assistance.

Abbreviations

Akt	protein kinase B
Bax	Bcl-2 associated X protein
Bcl-2	B-cell lymphoma 2
CCA	common carotid artery
CCK-8	Cell Counting Kit-8
CK	creatine kinase
CKMB	creatine kinase isoenzyme-MB
DAPI	4,6?-diamidino-2?-phenylindole
DHE	dihydroethidium
DMEM/F12	Dulbecco’s modified Eagle’s medium/F-12
dUTP	deoxyuridine triphosphate
eNOS	endothelial nitric oxide synthase
EPCs	endothelial progenitor cells
flk-1	fetal liver kinase-1
GAPDH	glyceraldehyde-3-phosphate dehydrogenase
HBDH	?-hydroxybutyrate dehydrogenase
H/R	hypoxia/reoxygenation
HRP	horseradish peroxidase
I/R	ischaemia/reperfusion
LAD	left anterior descending
LDH	lactate dehydrogenase
MI	myocardial infarction
MI/R	myocardial infarction/reperfusion
MI/RI	myocardial infarction/reperfusion injury
NRCMs	neonatal rat cardiac ventricular myocytes
PEMF	pulsed electromagnetic field
ROS	reactive oxygen species
Sca-1	stem cell antigen-1
SD	Sprague Dawley
SHR	spontaneously hypertensive rats
TTC	2,3,5-triphenyltetrazolium chloride
TUNEL	terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling
VEGF	vascular endothelial growth factor

AUTHOR CONTRIBUTION

Fenfen Ma designed and performed experiments on MI/RI rat model, histological stain and Western blot. Wenwen Li assisted the in vivo experiments, validated the effect in vitro experiments, analysed data and wrote the manuscript. Xinghui Li interpreted data and formatted manuscript. Rinkiko Suguro, Ruijuan Guan, Cuilan Hou, Huijuan Wang and Aijie Zhang interpreted data and edited manuscript. Yichun Zhu and YiZhun Zhu proposed the idea and supervised the project.

FUNDING

This work was supported by the key laboratory program of the Education Commission of Shanghai Municipality [grant number ZDSYS14005].

References

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Pulsed electromagnetic field improves cardiac function in response to myocardial infarctionChang-Ning Hao,^1,3,*Jing-Juan Huang,^1,2,*Yi-Qin Shi,^1,3Xian-Wu Cheng,³Hao-Yun Li,¹Lin Zhou,¹Xin-Gui Guo,²Rui-Lin Li,^1,2Wei Lu,^5,*Yi-Zhun Zhu,^4,* and Jun-Li Duan^1,2,*1Department of Gerontology, Xin Hua Hospital, Shanghai Jiaotong University, Kongjiang Road 1665, Shanghai 200092, China ²Department of Cardiology, Hua Dong Hospital, Fudan University, West Yan’an Road 221, Shanghai 200040, China ³Department of Cardiology, Nagoya University Graduate School of Medicine, 65 Tsuruma-cho, Showa-ku, Nagoya 466-8550, Japan ⁴Department of Pharmacology, School of Pharmacy, Fudan University, Zhang-Heng Road 826, Shanghai 201203, China ⁵National Lab for Infrared Physics, Shanghai Institute of Technical Physics, Chinese Academy of Sciences, Yu-Tian Road 500, Shanghai 200083, China Address correspondence to: Jun-Li Duan, Department of Cardiology, Hua Dong Hospital, Fudan University, West Yan’an Road 221, Shanghai 200040, China. E-mail: moc.361@hxilnujnaud; Dr. Wei Lu, National Lab for Infrared Physics, Shanghai Institute of Technical Physics, Chinese Academy of Sciences, Yu-Tian Road 500, Shanghai 200083, China. E-mail: nc.ca.ptis.liam@iewul; Dr. Yi-Zhun Zhu, Department of Pharmacology, School of Pharmacy, Fudan University, Zhang-Heng Road 826, Shanghai 201203, China. E-mail: moc.liamg@uhznuhziy^*Equal contributors. Author information  Article notes Copyright and License information Received February 3, 2014; Accepted April 18, 2014. AJTR Copyright © 2014

Introduction

Coronary artery disease is a leading cause of morbidity and mortality in modern society. Massive loss of cardiac muscle after several ischemic episodes lead to compromised cardiac function, remodeling and low quality life of patients. A growing body of evidence in experimental models of cardiac injury suggests that early re-establishment of blood perfusion to the injured myocardium would restrict infarct expansion, prevent cardiac remodeling and maintain cardiac function [1–3]. Although several strategies for therapeutic angiogenesis including the delivery of growth factors, gene therapy and stem cell implantation have been investigated, unsolvable theoretical limitations are still remaining [4–8]. For instance, the limited survival of implanted stem cell, uncontrolled angiogenesis and others [9–11]. Therefore, a safe, effective and non-invasive treatment for myocardial ischemia may be an ideal approach.

The therapeutic efficacy of various forms of electromagnetic stimulations, including capacitative coupling, direct current, combined magnetic fields, and pulsed electromagnetic field (PEMF), have been intensely investigated [12]. Among them, extracorporeal PEMF is the most widely tested techniques in the topic of osteanagenesis [13], skin rapture healing [14] and neuronal regeneration [15,16]. Recently, several study also indicated that PEMF exhibited the capability to stimulate angiogenesis and endothelial proliferation [17–19], however the detailed mechanism remains modest understood.

In the present study, we investigated whether extracorporeal PEMF therapy was able to rescue ischemic myocardium through inhibiting cardiac apoptosis as well as promoting postnatal neovascularization in a rat model of myocardial infarction (MI).

Material and methods

Animals

Male Sprague-Dawley (SD) rats weighing 250-300 g were provided by Sino-British SIPPR/BK Laboratory Animal (Shanghai, China). Animals were housed with controlled temperature (22-25°C) and lighting (08:00-20:00 light, 20:00-08:00 dark), and free access to tap water and standard rat chow. All the animals in this work received humane care in compliance with institutional guidelines for health and care of experimental animals of Shanghai Jiao Tong University.

MI model

All rats (n=36) were subjected to permanent left anterior descending artery ligation to establish MI model. Briefly, left thoracotomy and pericardiectomy were performed, and the hearts were gently exteriorized. Left anterior descending artery was ligated 4 mm below the left atrium with a 5-0 silk suture. The chest wall was then closed and the animals were returned to home cages. MI rats were then randomly divided into PEMF treated and untreated groups.

PEMF treatment

PEMF were generated by a commercially available healing device purchased from Biomobie Regenerative Medicine Technology (Shanghai, China). Fields were asymmetric and consisted of 4.5 ms pulses at 30 ± 3 Hz, with a magnetic flux density increasing from 0 to 5 mT in 400 ?s. The MI rats were housed in custom-designed cages and exposed to active PEMF for 4 cycles per day (8 minutes for 1 cycle), while the control rats were housed in identical cages with inactive PEMF generator. For in vitro study, culture dishes were directly exposed to PEMF for 1-4 cycles as indicated (8 minutes for 1 cycle, 30 ± 3 Hz, 5 mT).

Echocardiography

Trans-thoracic echocardiographic analysis was performed using an animal specific instrument (VisualSonics, Vevo770; VisualSonicsInc, Toronto, Canada), at postoperative day 7, 14 and 28. Rats were anesthetized with 10% chloral hydrate solution. After shaving the chest, pre-warmed ultrasound transmission gel was applied to the chest and two dimensional-directed M-mode and Doppler echocardiographic studies were carried out. The ejection fraction (EF) and fractional shortening (FS) were used to assess left ventricular systolic function. All measurements were averaged for consecutive cardiac cycles and triplicated.

Capillary density

Capillary density in peri-infarcted zone (PIZ) was determined by anti-CD31 staining (R&D Systems, San Diego, CA, USA). Briefly, 14 days after MI, rats were euthanized and hearts were perfused with a 0.9% NaCl solution followed by 4% solution of paraformaldehyde in 0.1 mol/L phosphate buffer (pH 7.4), and then dissected and fixed in this solution for 24 h. Next, samples were washed, dehydrated in a graded ethanol series and embedded in paraffin. 5 ?m-sections were cut transversely at 200 ?m intervals from into 5 slices from the ligation site to the apex. Endothelial capillaries were identified by goat anti-rat antibody of CD31 (5 ?g/ml, Becton-Dickinson Biosciences, Franklin Lakes, NJ, USA), and followed by a secondary antibody (Invitrogen, Carlsbad, CA, USA). Capillary density was determined by counting of 10 randomly selected fields and is expressed as numbers of capillary/field (×400 magnification) [20,21].

Enzyme-linked immunosorbent assay (ELISA)

The concentration of vascular endothelial growth factor (VEGF) and nitric oxide (NO) contained in conditional media of cultured HUVECs was measured using ELISA kit purchased from R&D Systems (San Diego, CA, USA). The concentrations of VEGF contained in PIZ was determined by ELISA kits purchased from Raybiotech (Norcross, GA, USA) [22].

Western blotting

PIZ tissue and HUVECs were homogenized with ice-cold homogenizing buffer (20 ?l/gram tissue, 50 mmol/l Tris-HCl, 150 mmol/l NaCl, 1 mmol/l EDTA, and 0.5 mmol/l Triton X-100, pH 7.4) and protease inhibitor cocktail (5 mM, Roche, Berlin, Germany). Proteins were measured with Pierce BCA Protein Assay Kit (Thermo, Asheville, North Carolina, USA). Hippocampal protein lysates (50 mg/well) were separated using SDS-PAGE under reducing conditions. Following electrophoresis, the separated proteins were transferred to a polyvinylidene difluoride membrane (Millipore, Billerica, Massachusetts, USA). Subsequently, nonspecific proteins were blocked using blocking buffer (5% nonfat dried milk in T-TBS containing 0.05% Tween 20), followed by incubation with primary rabbit anti-rat antibodies specific for phospho-Akt (p-Akt), total Akt, hypoxic-inducible factor (HIF)-1? (Santa Cruz, California, USA), phospho-endothelial nitric oxide synthase (p-eNOS), total eNOS and ?-actin (Cell Signaling Technology, Beverly, MA, USA) overnight at 4°C. Blots were washed four times with 0.1% Tween 20 in PBS and incubated with HRP-conjugated secondary antibody (1/5000; Biochain, Newark, California, USA) for 1 h at room temperature. The bands were visualized using enhanced chemiluminescence method (Bioimaging System; Syngene, Cambridge, UK). Intensity of the tested protein bands was quantified by densitometry.

Detection of apoptosis

Heart samples were fixed in 10% formalin and then paraffin embedded at day 14. Then, the hearts were cut into 5-?m sections. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining was carried out using a commercially available kit according to the manufacturer’s instructions (Promega, Madison, Wisconsin, USA). Nuclei were stained by DAPI (Roche) [23]. Three mid-ventricular sections of each heart (from the apex to the base) were analyzed. Ten fields in the PIZ were randomly selected from each section for the calculation of the percentage of apoptotic nuclei (apoptotic nuclei/total nuclei) and the obtained ratios were averaged for statistical analysis.

Isolation of circulating endothelial progenitor cells (EPCs)

Circulating EPCs were obtained by cardiac puncture after animals were anesthetized. Peripheral blood-derived mononuclear cells (PB-MNCs) were then purified by Histopaque-1083 (Sigma-Aldrich, St. Louis, MO, USA) density gradient centrifugation at 400 g for 30 min. The mononuclear layer was then collected and re-suspended in endothelial growth medium-2 (EGM-2, Clonetics, San Diego, CA, USA). Antibodies to the stem cell antigen-1 (Sca-1) and Flk-1 were used to mark EPC as described before), and the isotype specific conjugated anti-IgG was used as a negative control. Sca-1⁺ and Flk-1⁺ cells were gated in the mononuclear cell fraction.

EPC migration assay

Migratory activity of PB-EPCs from PEMF-treated and untreated rats was evaluated by a 24-well modified Boyden chamber assay (Transwell, Corning, NY, USA) [24]. After cultured with EGM-2 for 4 days, PB-EPCs were trypsinized and 5×10⁵ cells in 100 ?l of EBM-2 with 0.1% BSA in placed in the upper compartments. 50 ng/mL recombinant vascular endothelial growth factor (VEGF, Clonetics) in 600 ?L of chemotaxis buffer (serum-free EBM-2, 0.1% BSA) was added to the lower compartment. The chamber was incubated at 37°C for 6 hrs. The cells were then fixed and stained with hematoxylin and eosin (H&E). Non-migrated cells on the filter’s upper surface were removed using a cotton swab. The numbers of migrated cells were counted in 4 random high-power fields (HPF, ×400 magnification) and averaged for each sample.

Tube formation assay

Matrigel-Matrix (BD Biosciences, Franklin Lakes, New Jersey, USA) was inserted in the well of a 48-well cell culture plate and a number of 5×10⁴ EPCs or HUVECs were seeded [25].

After incubation in EGM-2, images of tube morphology was taken and tube number was counted at random under four low power fields (magnifications ×40) per sample. Capillary tube branch points were counted in six randomly selected fields per well, and used as an index for tube formation.

Cell culture

Human umbilical vein endothelial cells (HUVECs, passage 3) were purchased from Clonetics (San Diego, CA, USA) and EGM-2 in a humidified atmosphere of 5% CO₂ and 95% air. HUVECs were reseeded into plates coated with Matrix gel and stimulated for 1-4 cycles of PEMF stimulation (5.5 mT, 8 minutes per cycle). Supernatant and cell lysates were collected at 24 hrs after reseeding. Additionally, HUVECs-formed vasculature was quantified by calculating its length under microscopic photography 24 hrs after reseeding [26].

Statistical analysis

Data are expressed as means ± standard deviation (SD). Student’s t-test was used for statistical analyses. SPSS software version 17.0 (SPSS Inc., Chicago, IL, USA) was used. A value of p<0.05 was considered significant.Go to:

Results

PEMF promotes cardiac function after MI

To determine whether PEMF could increase myocardial function in MI rats, echocardiographic studies were carried out at postoperative day 7, 14 and 28. We observed that PEMF had no effects on body weight and heart rates when compared with control group (Table 1). Meanwhile, higher EF and FS values were detected in PEMF-treated rats than control (Figure 1), indicating that PEMF preserves left ventricular contractility after MI damage.

Figure 1 Echocardiography after PEMF therapy. All rats were subjected to MI and randomly separated to control and PEMF group. The data of (A) ejection fractions and (B) fractional shorting in both groups collected in day 7, 14 and 28. Values are mean ±…

Table 1 Effect of PEMF on cardiac functions of MI rats

PEMF enhances angiogenesis in PIZ

To examine whether the changes in the cardiac function are associated with changes in capillary EC formation, we measured capillary densities of PEMF and control rats in PIZ through anti-CD31 immunofluorescence staining at postoperative day 14. Representative photomicrographs are shown in Figure 2A. Quantitative analyses by counting the CD31⁺ capillary ECs revealed that PEMF treatment significantly increases capillary densities in PIZ than control rats (Figure 2B). PEMF treatment also increased the protein levels of VEGF and HIF-1? in damaged hearts (Figure 2C and ?and2D),2D), as well as enhancing the phosphorylation of Akt signal pathway in ischemic myocardium at postoperative day 14 (Figure 2E).

Figure 2 Pro-angiogenic effect of PEMF in ischemic myocardium. A: Immunofluorescence staining of CD31-positive cells in the infarct border zone at postoperative day 14 in PEMF-treated and control rats. B: Quantitative analyses of capillary density between 2 groups…

Protective effect of PEMF to MI-induced cardiac apoptosis

We evaluated the effect of PEMF on the survival of myocardium in response to hypoxia in vivo at postoperative day 7. The number of TUNEL positive nucleus in PIZ significantly increased in PEMF-treated rats compared with the non-treated ones (Figure 3), indicating that PEMF treatment decreases the susceptibility of cardiomyocytes to hypoxic damage.

Figure 3 Anti-apoptotic benefit of PEMF in damaged myocardium. A: TUNEL staining for cardiac cell apoptosis (green) and DAPI (blue) for nuclear staining in the border zone 14 days after AMI (×400 magnification). B: Quantitative analysis of the TUNEL-positive…

PEMF augments EPC-mediated neovascularization

EPC-mediated neovascularization after myocardial infarction supported their therapeutic potential [27]. Thus, the strategy to amplify EPC abundance and function is an active focus of research. The number of circulating EPCs was identified by stem cell antigen-1 (Sca-1)/fetal liver kinase-1 (flk-1) dual positive cells as described. We found that PEMF treatment increased the number of Sca-1⁺/flk-1⁺ cells in peripheral blood at postoperative day 7 and 14 (Figure 4A). Additionally, EPCs isolated from PEMF-treated rats exhibited enhanced tube formative capacity and migratory ability when compared with control ones in vitro (Figure 4B and ?and4C),4C), which suggesting that PEMF increases the abundance and regenerative capacity of EPCs.

Figure 4 PEMF enhanced circulating endothelial progenitor cells (EPCs) function in MI Rats. 7 and 14 days after AMI induction, peripheral blood was collected from rats in both groups. A: Quantitative analysis of Sca-1/flk-1 dual positive PB-EPCs isolated from…

Pro-angiogenic beneficial of PEMF in vitro

Cultured HUVECs were treated with PEMF stimulation for 1 to 4 cycles and the supernatant and cell lysate were collected. PEMF promoted VEGF and NO releasing from cultured HUVECs in a dose-dependent manner (Figure 5A and ?and5B).5B). Additionally, the phosphorylation of eNOS in HUVECs was also enhanced in response to PEMF following a dose dependent manner (Figure 5C). Finally, the HUVEC-formed tubes were lengthened by PEMF in a dose dependent manner (Figure 6).

Figure 5 Enhancement of the expression of VEGF and nitric oxide in PEMF-treated HUVECs. PEMF stimulated vascular endothelial growth factors secretion concentration dependently. Bar graph of the concentrations of (A) VEGF and (B) nitric oxide released from HUVECs…

Figure 6 Effects of PEMF on tube formation of cultured HUVECs. Representative images of tube formation in HUVECs by stimulated PEMF for 1-4 cycles and quantitative analysis of tube length formed by PEMF-treated HUVECs. Values are mean ± SEM; n=4. *means…Go to:

Discussion

Major findings of our study are: (1) PEMF prevents cardiomyocytes against hypoxia-induced apoptosis and preserves cardiac systolic function in a rat MI model; (2) PEMF induces angiogenesis and vasculogenesis through activating VEGF-eNOS system and promoting EPCs mobilized to the ischemic myocardium.

We demonstrated that PEMF treatment preserved the cardiac systolic function after MI and prevented cardiac apoptosis. Previous report demonstrated that PEMF treatment activated voltage-gated calcium channels (VGCC) [28], which is crucial for maintaining cardiac contractility and cell survival [29,30]. Increased intracellular Ca²⁺produced by PEMF-mediated VGCC activation may lead to increase of NO through the action of eNOS, which is dominant modulator to prevent cardiomyocytes from apoptosis and enhance revascularization in PIZ after MI [31]. Consistent with the previous work, we demonstrated that the HIF-1?/Akt axis was activated in PIZ in PEMF rats. In addition, PEMF induced eNOS phosphorylation in vitro, which is a key molecular served in the survival pathway in both myocardium and endothelial cell lineage [32].

Another possible mechanism in cardiac protecting effect of PEMF is to stimulate neovascularization. Increasing evidence suggests that neovascularization limits infarct expansion and extension, improves cardiac remodeling [1,2]. Recent data demonstrated that PEMF stimulation induced angiogenesis and amplified endothelial cells function [17,20]. Some researchers believe that PEMF induces cellular proliferation, as evidenced by cAMP activation and uptake of tritiated thymidine [33]. In present study, we demonstrated that the capillary density in PIZ was increased after PEMF treatment. Moreover, PEMF therapy triggered the Akt/HIF-1?/VEGF cascade was activated in ischemic myocardium. In in vitro study, we confirmed PEMF-treated HUVECs released more VEGF and NO, which are the key factors response to endothelial proliferation and survival, suggesting that PEMF activates both autocrine and paracrine function of mature endothelial cells. Furthermore, Tepper and colleagues also reported that PEMF stimulated fibroblastic growth factor-2 (FGF-2) releasing and augment angiogenesis [14].

Recent evidence indicates that adult blood vessels may result from not only expansion of existing endothelial cells (angiogenesis), but also the recruitment of endothelial progenitor cells or EPCs (vasculogenesis) [24]. We hypothesized that besides mature endothelial cells, PEMF might also act as a stimulator of progenitor (EPC). To confirm the hypothesis, we examined the effect of PEMF on ex vivo angiogenesis. Our data demonstrated the number of Sca-1/flk-1 dual positive EPCs in peripheral blood increased in response to PEMF. Using the well-established Matrigel assay, we demonstrated that PEMF was able to dramatically enhance the tube formative capacity of either EPCs or mature endothelial cells in vitro. PEMF also accelerated the migratory ability of EPCs. Moreover, Goto et al reported that PEMF stimulation up-regulated the expression of angiopoietin-2 and FGF-2 in bone marrow, suggesting PEMF could promote the regenerative capacity of myeloid-derived cells (such as EPCs) in damaged tissue when recruited. From all these findings, we conclude that PEMF sufficiently re-establishes blood supply to the ischemic and hypoxic cardiomyocytes via enhancing both angiogenesis and vasculogenesis.

In conclusion, our findings indicate that extracorporeal PEMF treatment increases cardiac systolic function through inhibiting cardiac apoptosis and stimulating neovascularization in PIZ. These findings suggest that PEMF deserves further consideration of investigation in its regulation on the signaling pathway and new clinical strategies for ischemic vascular diseases.Go to:

Acknowledgements

This work was supported by the Shanghai Science and Technology Committee (11 nm 0503600), the China National Natural Science Foundation (11374213) and Foundation of National Lab for Infrared Physics (200901).Go to:

Disclosure of conflict of interest

The authors have nothing to disclose.Go to:

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Bioelectromagnetics. 2010 May;31(4):296-301.

Effects of weak static magnetic fields on endothelial cells.

Martino CF, Perea H, Hopfner U, Ferguson VL, Wintermantel E.

Department of Mechanical Engineering, University of Colorado at Boulder, Boulder, Colorado, USA. martino@colorado.edu

Abstract

Pulsed electromagnetic fields (PEMFs) have been used extensively in bone fracture repairs and wound healing. It is accepted that the induced electric field is the dose metric. The mechanisms of interaction between weak magnetic fields and biological systems present more ambiguity than that of PEMFs since weak electric currents induced by PEMFs are believed to mediate the healing process, which are absent in magnetic fields. The present study examines the response of human umbilical vein endothelial cells to weak static magnetic fields. We investigated proliferation, viability, and the expression of functional parameters such as eNOS, NO, and also gene expression of VEGF under the influence of different doses of weak magnetic fields. Applications of weak magnetic fields in tissue engineering are also discussed. Static magnetic fields may open new venues of research in the field of vascular therapies by promoting endothelial cell growth and by enhancing the healing response of the endothelium.

Eur J Appl Physiol. 2007 Nov;101(4):495-502. Epub 2007 Aug 3.

Short-term effects of pulsed electromagnetic fields after physical exercise are dependent on autonomic tone before exposure.

Grote V, Lackner H, Kelz C, Trapp M, Aichinger F, Puff H, Moser M.

Institute of Noninvasive Diagnosis, JOANNEUM RESEARCH, Weiz, Austria.

Abstract

The therapeutic application of pulsed electromagnetic fields (PEMFs) can accelerate healing after bone fractures and also alleviate pain according to several studies. However, no objective criteria have been available to ensure appropriate magnetic field strength or type of electromagnetic field. Moreover, few studies so far have investigated the physical principles responsible for the impact of electromagnetic fields on the human body. Existing studies have shown that PEMFs influence cell activity, the autonomic nervous system and the blood flow. The aim of this study is to examine the instantaneous and short-term effects of a PEMF therapy and to measure the impact of different electromagnetic field strengths on a range of physiological parameters, especially the autonomic nervous systems, determined by heart rate variability (HRV) as well as their influence on subjects’ general feeling of well-being. The study comprised experimental, double-blind laboratory tests during which 32 healthy male adults (age: 38.4+/-6.5 years) underwent four physical stress tests at standardised times followed by exposure to pulsed magnetic fields of varying intensity [HPM, High Performance magnetic field; Leotec; pulsed signal; mean intensity increase: zero (placebo), 0.005, 0.03 and 0.09 T/s]. Exposure to electromagnetic fields after standardised physical effort significantly affected the very low frequency power spectral components of HRV (VLF; an indicator for sympathetically controlled blood flow rhythms). Compared to placebo treatment, exposure to 0.005 T/s resulted in accelerated recovery after physical strain. Subjects with lower baseline VLF power recovered more quickly than subjects with higher VLF when exposed to higher magnetic field strengths. The application of electromagnetic fields had no effect on subjects’ general feeling of well-being. Once the magnetic field exposure was stopped, the described effects quickly subsided. PEMF exposure has a short-term dosage-dependent impact on healthy subjects. Exposure to PEMF for 20 min resulted in more rapid recovery of heart rate variability, especially in the very low frequency range after physical strain. The study also showed the moderating influence of the subjects’ constitutional VLF power on their response to PEMF treatment. These findings have since been replicated in a clinical study and should be taken into consideration when PEMF treatment is chosen.

Vopr Kurortol Fizioter Lech Fiz Kult. 2009 Sep-Oct;(5):9-11.

Rehabilitative medical technology for the correction of microcirculatory disorders in patients with arterial hypertension.

[Article in Russian]

Kul’chitskaia DB.

Abstract

The study with the use of laser Doppler flowmetry has revealed pathological changes in the microcirculatory system of patients with arterial hypertension. Their treatment with a low-frequency magnetic field showed that its effect on microcirculation depends on the regime and site of application of magnetotherapy as well as its combination with other physical factors. Frontal application of the magnetic field had the most pronounced beneficial effect on dynamic characteristics of microcirculation. Pulsed regime of magnetotherapy was more efficacious than conventional one. Amplipulse magnetotherapy produced better results than monotherapy.

Bioelectromagnetics. 2007 Jan;28(1):64-8.

A pilot investigation of the effect of extremely low frequency pulsed electromagnetic fields on humans’ heart rate variability.

Baldi E, Baldi C, Lithgow BJ.

Diagnostic and Neurosignal Processing Research Group, Electrical & Computer System Engineering, Monash University, Victoria, Australia. Emilio.Baldi@eng.monash.edu.au

Abstract

The question whether pulsed electromagnetic field (PEMF) can affect the heart rhythm is still controversial. This study investigates the effects on the cardiocirculatory system of ELF-PEMFs. It is a follow-up to an investigation made of the possible therapeutic effect ELF-PEMFs, using a commercially available magneto therapeutic unit, had on soft tissue injury repair in humans. Modulation of heart rate (HR) or heart rate variability (HRV) can be detected from changes in periodicity of the R-R interval and/or from changes in the numbers of heart-beat/min (bpm), however, R-R interval analysis gives only a quantitative insight into HRV. A qualitative understanding of HRV can be obtained considering the power spectral density (PSD) of the R-R intervals Fourier transform. In this study PSD is the investigative tool used, more specifically the low frequency (LF) PSD and high frequency (HF) PSD ratio (LF/HF) which is an indicator of sympatho-vagal balance. To obtain the PSD value, variations of the R-R time intervals were evaluated from a continuously recorded ECG. The results show a HR variation in all the subjects when they are exposed to the same ELF-PEMF. This variation can be detected by observing the change in the sympatho-vagal equilibrium, which is an indicator of modulation of heart activity. Variation of the LF/HF PSD ratio mainly occurs at transition times from exposure to nonexposure, or vice versa. Also of interest are the results obtained during the exposure of one subject to a range of different ELF-PEMFs. This pilot study suggests that a full investigation into the effect of ELF-PEMFs on the cardiovascular system is justified.

Georgian Med News. 2006 Jun;(135):109-13.

Influence of treatment with variable magnetic field of low frequency in low mountain environment on cardiohemodynamic index of patients with arterial hypertension.

[Article in Russian]

Tarkhan-Mouravi ID, Purtseladze NA.

Abstract

Pathological changes in function and action of cardiovascular system is the significant link in formation and progression of arterial hypertension. 68 patients were investigated. From these patients in 32 first stage of mentioned pathology, while in 36 – the II degree was found. It is established that treatment of arterial hypertension by variable magnetic field of low frequency in low mountain environment causes decrease of systolic, diastolic and heart dynamic blood pressure, normalizes heart index and pulse rate; decreases peripheral vascular specific resistance, increases amount of upset index accelerated of blood flow on the region “lung-ear”, improves electrocardiological data. Mentioned pathological displacements were more expressed at the first stage of arterial hypertension.

Bioelectromagnetics. 2005 Apr;26(3):161-72.

Decreased plasma levels of nitric oxide metabolites, angiotensin II, and aldosterone in spontaneously hypertensive rats exposed to 5 mT static magnetic field.

Okano H, Masuda H, Ohkubo C.

Department of Environmental Health, National Institute of Public Health, Tokyo 108-8638, Japan. okano@niph.go.jp

Previously, we found that whole body exposure to static magnetic fields (SMF) at 10 mT (B(max)) and 25 mT (B(max)) for 2-9 weeks suppressed and delayed blood pressure (BP) elevation in young, stroke resistant, spontaneously hypertensive rats (SHR). In this study, we investigated the interrelated antipressor effects of lower field strengths and nitric oxide (NO) metabolites (NO(x) = NO(2)(-) + NO(3)(-)) in SHR. Seven-week-old male rats were exposed to two different ranges of SMF intensity, 0.3-1.0 mT or 1.5-5.0 mT, for 12 weeks. Three experimental groups of 20 animals each were examined: (1) no exposure with intraperitoneal (ip) saline injection (sham-exposed control); (2) 1 mT SMF exposure with ip saline injection (1 mT); (3) 5 mT SMF exposure with ip saline injection (5 mT). Arterial BP, heart rate (HR), skin blood flow (SBF), plasma NO metabolites (NO(x)), and plasma catecholamine levels were monitored. SMF at 5 mT, but not 1 mT, significantly suppressed and retarded the early stage development of hypertension for several weeks, compared with the age matched, unexposed (sham exposed) control. Exposure to 5 mT resulted in reduced plasma NO(x) concentrations together with lower levels of angiotensin II and aldosterone in SHR. These results suggest that SMF may suppress and delay BP elevation via the NO pathways and hormonal regulatory systems.

Auton Neurosci. 2003 Apr 30;105(1):53-61.

Can extremely low frequency alternating magnetic fields modulate heart rate or its variability in humans?

Kurokawa Y, Nitta H, Imai H, Kabuto M.

Environmental Health Science Region, National Institute for Environmental Studies, 16-2 Onogawa, Ibaraki Tsukuba 305-0053, Japan.kurokawa@nies.go.jp

Abstract

This study is a reexamination of the possibility that exposure to extremely low frequency alternating magnetic field (ELF-MF) may influence heart rate (HR) or its variability (HRV) in humans. In a wooden room (cube with 2.7-m sides) surrounded with wire, three series of experiments were performed on 50 healthy volunteers, who were exposed to MFs at frequencies ranging from 50 to 1000 Hz and with flux densities ranging from 20 to 100 microT for periods ranging from 2 min to 12 h. In each experiment, six indices of HR/HRV were calculated from the RR intervals (RRIs): average RRI, standard deviation of RRIs, power spectral components in three frequency ranges (pVLF, pLF and pHF), and the ratio of pLF to pHF. Statistical analyses of results revealed no significant effect of ELF-MFs in any of the experiments, and suggested that the ELF-MF to which humans are exposed in their daily lives has no acute influence on the activity of the cardiovascular autonomic nervous system (ANS) that modulates the heart rate.

Klin Med (Mosk). 2003;81(1):24-7.

Clinico-functional efficacy of medicinal and photon stabilization in patients with angina pectoris.

[Article in Russian]

Vasil’ev AP, Senatorov IuN, Strel’tsova NN, Gorbunova TIu.

Modification of erythrocytic membrane and the trend in clinicofunctional indices were studied in 90 patients with angina of effort (FC I-IV) in the course of treatment with a combination of membranoprotective drugs (group 1), magneto-laser radiation (group 2) and imitation of laser radiation (group 3). In patients of groups 1 and 2 the treatment resulted in stabilization of cell membrane accompanied with a hypotensive effect and increased exercise tolerance due to more effective cardiac performance.

Saudi Med J. 2002 May;23(5):517-20.

The effect of magneto-treated blood autotransfusion on central hemodynamic values and cerebral circulation in patients with essential hypertension.

Alizade IG, Karayeva NT.

Department of Cardiology, Hospital of Ministry of Internal Affairs, Baku, Azerbaijan.

OBJECTIVE: The work was carried out to study the effect of magneto-treated blood autotransfusion on the values of central and cerebral hemodynamics in patients with essential hypertension.

METHODS: Sixty-six patients with stage II essential hypertension aged 31-60 years who underwent magneto-treated blood autotransfusion were evaluated and treated, at the Cardiology Department, Hospital of Ministry of Internal Affairs of the Azerbaijan Republic, over a period of 8 years. The diagnosis was based on clinical examination and generally accepted criteria of essential hypertension stages proposed in 1978 by the World Health Organization.

RESULTS: Sixty-six patients with stage II essential hypertension with stable drop in blood pressure, simultaneously showed a positive clinical effect. Central hemodynamic changes in the process of magneto-treated blood autotransfusion were different and depended on the initial state of circulation. High clinical effect showed the patients with hyperkinetic type of hemodynamics. Their blood pressure were significantly lower than the patients with hypokinetic type of circulation.

CONCLUSION: Rheoencephalographic study demonstrated that magneto-treated blood autotransfusion possessed insignificant effect on cerebral hemodynamics, mainly expressed by the reduction of arterial blood flow tension in the patients with hypokinetic type of hemodynamics.

Ter Arkh. 2001;73(10):70-3.

Changes in blood rheological properties in patients with hypertension.

[Article in Russian]

Shabanov VA, Terekhina EV, Kostrov VA.

AIM: To study hemorheology in patients with essential hypertension (EH), to improve EH treatment in terms of blood rheology.

MATERIAL AND METHODS: Blood rheology, microcirculation, lipid plasm spectrum, central hemodynamics were studied in 90 patients with mild and 83 patients with moderate or severe EH as well as 30 healthy controls before and after treatment (hypotensive drugs, essential phospholipids, intravenous laser blood radiation, plasmapheresis).

RESULTS: Hemorrheological disorders (subnormal deformability of the red cells and elastoviscosity of their membranes, disk-spherical transformation and hyperaggregation of blood cells, high dynamic viscosity) correlated with the disease severity, arterial pressure and total peripheral vascular resistance. Long-term (1-1.5 years) hypotensive therapy, especially with combination of beta-blockers with diuretics, has a negative effect on blood rheology. Optimisation of EH treatment in terms of blood rheology consists in using essential phospholipids in stable hypertension, intravenous laser radiation in complicated hypertension, plasmapheresis in drug-resistant hypertension. Such an approach not only significantly improves hemorheology but also provides good clinical and hypotensive effects in 75-80% patients.

CONCLUSION: Blood viscodynamics should be taken into consideration in individual treatment of hypertensive patients.

Med Tr Prom Ekol. 2001;(6):20-3.

Influence of low-frequency magnetotherapy and HF-puncture on the heart rhythm in hypertensive workers exposed to vibration.

[Article in Russian]

Drobyshev VA, Loseva MI, Sukharevskaia TM, Michurin AI.

Abstract

The authors present results concerning use of low-frequency magnetic fields and HF-therapy for correction of vegetative homeostasis in workers with variable length of service, exposed to vibration, having early forms of arterial hypertension. The most positive changes of vegetative status and central hemodynamics are seen in workers with low length of service.

Vopr Kurortol Fizioter Lech Fiz Kult. 2001 Mar-Apr;(2):11-5.

Therapeutic complexes of physical factors in mild arterial hypertension.

[Article in Russian]

Kniazeva TA, Nikiforova TI.

Three therapeutic complexes were compared clinically in patients with mild arterial hypertension. Complex 1 consisted of dry air–radon baths, bicycle exercise and exposure of the renal projection area to decimetric electromagnetic field. Its efficacy was 90%, mechanism of the hypotensive action is reduction of enhanced activity of the sympathico-adrenal and renin-angiotensin-aldosterone systems, improvement of water-mineral metabolism and lipid peroxidation. Complex 2 consisted of dry effervescent baths, anaprilin electrophoresis with sinusoidal modulated currents and exposure of the renal projection area to low-frequency alternating magnetic field. Its efficacy was 80%. It affects renin-angiotensin-aldosterone system, water-mineral metabolism and lipid peroxidation. Complex 3 consisted of electric sleep, laser therapy and general sodium chloride baths. Its efficacy was 63%. The effect was due to inhibition of high sympathico-adrenal system.

Klin Med (Mosk). 2000;78(3):23-5.

Characteristics of microcirculation and vascular responsiveness in elderly patients with hypertension and ischemic heart disease.

[Article in Russian]

Abramovich SG.

Microcirculation and vascular responsiveness were studied in 52 patients with arterial hypertension and ischemic heart disease versus 48 healthy elderly persons. The patients were found to have defects of the end blood flow in all links of microcirculation, longer and more severe vasoconstriction of conjunctival and skin vessels in response to norepinephrine and cold stimulation tests.

Vopr Kurortol Fizioter Lech Fiz Kult. 2000 May-Jun;(3):9-11.

The use of low-frequency magnetotherapy and EHF puncture in the combined treatment of arterial hypertension in vibration-induced disease.

[Article in Russian]

Drobyshev VA, Filippova GN, Loseva MI, Shpagina LA, Shelepova NV, Zhelezniak MS.

Combination of EHF therapy + magnetotherapy + drugs results in faster and persistent hypotensive and analgetic effect compared to standard drug therapy, potentiates action of vascular drugs on cerebral and peripheral circulation, reduces dose of hypotensive drugs in patients with arterial hypertension and vibration disease.

Crit Rev Biomed Eng. 2000;28(1-2):339-47.

The use of millimeter wavelength electromagnetic waves in cardiology.

Lebedeva AYu.

2nd Department of urgent cardiology at State Clinical Hospital, Russian State Medical University, Moscow.

Abstract

This paper concerns the problems of the use of millimeter wavelength electromagnetic waves for the treatment of cardiovascular disease. The prospects for this use are considered.

Vopr Kurortol Fizioter Lech Fiz Kult. 1999 Sep-Oct;(5):7-9.

The characteristics of the geroprotective action of magnetotherapy in elderly patients with combined cardiovascular pathology.

[Article in Russian]

Abramovich SG, Fedotchenko AA, Koriakina AV, Pogodin KV, Smirnov SN.

Central hemodynamics, diastolic and pumping functions of the heart, myocardial reactivity, microcirculation and biological age of cardiovascular system were studied in 66 elderly patients suffering from hypertension and ischemic heart disease. The patients received systemic magnetotherapy which produced a geroprotective effect as shown by improved microcirculation, myocardial reactivity, central hemodynamics reducing biological age of cardiovascular system and inhibiting its ageing.

Neuropsychobiology. 1998 Nov;38(4):251-6.

No effects of pulsed high-frequency electromagnetic fields on heart rate variability during human sleep.

Mann K, Roschke J, Connemann B, Beta H.

Department of Psychiatry, University of Mainz, Germany.

The influence of pulsed high-frequency electromagnetic fields emitted by digital mobile radio telephones on heart rate during sleep in healthy humans was investigated. Beside mean RR interval and total variability of RR intervals based on calculation of the standard deviation, heart rate variability was assessed in the frequency domain by spectral power analysis providing information about the balance between the two branches of the autonomic nervous system. For most parameters, significant differences between different sleep stages were found. In particular, slow-wave sleep was characterized by a low ratio of low- and high-frequency components, indicating a predominance of the parasympathetic over the sympathetic tone. In contrast, during REM sleep the autonomic balance was shifted in favor of the sympathetic activity. For all heart rate parameters, no significant effects were detected under exposure to the field compared to placebo condition. Thus, under the given experimental conditions, autonomic control of heart rate was not affected by weak-pulsed high-frequency electromagnetic fields.

Vopr Kurortol Fizioter Lech Fiz Kult. 1998 Jul-Aug;(4):31-6.

The combined action of infrared radiation and permanent and alternating magnetic fields in experimental atherosclerosis.

[Article in Russian]

Zubkova SM, Varakina NI, Mikhailik LV, Bobkova AS, Maksimov EB.

Paravertebral exposure to infrared radiation (0.87 micron, 5 mW) and permanent magnetic field in combination with one- and two-semiperiodic alternative magnetic fields (50 Hz, 15-30 mT) was studied in respect to the action on adaptive reactions in animals with experimental atherosclerosis. Complex consisting of infrared radiation, permanent magnetic field and one-semiperiodic pulse alternative magnetic field was most effective in restoration of vasomotor-metabolic and immune disturbances accompanying development of atherosclerosis.

Bioelectromagnetics. 1998;19(2):98-106.

Nocturnal exposure to intermittent 60 Hz magnetic fields alters human cardiac rhythm.

Sastre A, Cook MR, Graham C.

Midwest Research Institute, Kansas City, Missouri 64110, USA. Asastre@mriresearch.org

Abstract

Heart rate variability (HRV) results from the action of neuronal and cardiovascular reflexes, including those involved in the control of temperature, blood pressure and respiration. Quantitative spectral analyses of alterations in HRV using the digital Fourier transform technique provide useful in vivo indicators of beat-to-beat variations in sympathetic and parasympathetic nerve activity. Recently, decreases in HRV have been shown to have clinical value in the prediction of cardiovascular morbidity and mortality. While previous studies have shown that exposure to power-frequency electric and magnetic fields alters mean heart rate, the studies reported here are the first to examine effects of exposure on HRV. This report describes three double-blind studies involving a total of 77 human volunteers. In the first two studies, nocturnal exposure to an intermittent, circularly polarized magnetic field at 200 mG significantly reduced HRV in the spectral band associated with temperature and blood pressure control mechanisms (P = 0.035 and P = 0.02), and increased variability in the spectral band associated with respiration (P = 0.06 and P = 0.008). In the third study the field was presented continuously rather than intermittently, and no significant effects on HRV were found. The changes seen as a function of intermittent magnetic field exposure are similar, but not identical, to those reported as predictive of cardiovascular morbidity and mortality. Furthermore, the changes resemble those reported during stage II sleep. Further research will be required to determine whether exposure to magnetic fields alters stage II sleep and to define further the anatomical structures where field-related interactions between magnetic fields and human physiology should be sought.

Vopr Kurortol Fizioter Lech Fiz Kult. 1998 Jan-Feb;(1):16-8.

A comparative evaluation of the effect of an extremely high-frequency electromagnetic field on cerebral hemodynamics in hypertension patients exposed in different reflexogenic areas.

[Article in Russian]

Sokolov BA, Bezruchenko SV, Kunitsyna LA.

A single session and multiple sinocarotid and temporal exposures to EHF electromagnetic field in patients with stage I and II hypertension had different effects on cerebral circulation Variants of the above treatment are proposed.

Vopr Kurortol Fizioter Lech Fiz Kult. 1997 Jan-Feb;(1):8-11.

Prognostic criteria of the efficacy of magnetic and magnetic-laser therapy in patients with the initial stages of hypertension.

[Article in Russian]

Zadionchenko VS, Sviridov AA, Adasheva TV, Demicheva OIu, Bagatyrova KM, Beketova IL.

Study of the efficacy of a course of exposures to travelling pulsed magnetic field and magnetic laser sessions in 97 patients with stages I-II essential hypertension showed a high efficacy of travelling pulsed magnetic field in patients with hyperkinetic hemodynamics and initially just slightly shifted blood rheology and platelet hemostasis. Magnetic laser therapy is more effective in patients with eukinetic and hypokinetic hemodynamics and initially sharply expressed disorders of blood rheology and platelet hemostasis.

Biofizika. 1996 Jul-Aug;41(4):944-8.

Effect of a “running” pulse magnetic field on certain humoral indicators and physical ability to work in patients with neurocirculatory hypo- and hypertension.

[Article in Russian]

Orlov LL, Pochechueva GA, Makoeva LD.

The influence of “running” impulse magnetic field in patients with neurocirculatory hypo- and hypertension was studied. It has been determined that magnetotherapy in all patients increased physical load tolerability and at the same time produced different effects on hemodynamics (lowering blood pressure in hypertension and increasing it in hypotension). In patients with neurocirculatory hypotension the slightly expressed positive clinical effect was obtained, that makes “running” impulse magnetic field therapy useless in this pathology. At the same time in patients with neurocirculatory hypertension “running” impulse magnetic field therapy resulted in significant improvement of physical tolerability, improvement of patients general condition, blood pressure decrease, lowering of pressor power generation concentration, correcting effect on aldosterone blood content. These data witness for the usefulness of this method in treatment of patients with neurocirculatory hypertension.

Ter Arkh. 1996;68(5):63-7.

The therapeutic correction of disorders in thrombocyte-vascular hemostasis and of changes in the rheological properties of the blood in patients with arterial hypertension.

[Article in Russian]

Zadionchenko VS, Bagatyrova KM, Adasheva TV, Timofeeva NIu, Zaporozhets TP.

158 patients with essential hypertension received beta-adrenoblockers and were exposed to travelling impulse magnetic field, magnetolaser radiation. The study of platelet-vessel hemostasis and blood rheology revealed a relation of good clinical response and increased exercise tolerance with initial platelet dysfunction and rheological disorders which underwent positive changes in the course of treatment

Vopr Kurortol Fizioter Lech Fiz Kult. 1996 Mar-Apr;(2):5-8.

The effect of exposure to magnetics and lasers on the clinical status and the electrophysiological indices of the heart in patients with cardiac arrhythmias.

[Article in Russian]

Budnar’ LN, Antiuf’ev VF, Oranskii IE, Bekhter TV.

Magnetolaser radiation has a considerable influence on electrophysiological condition of the sinus node and sinoatrial zone. There are cases when patients with sick sinus syndrome get rid of arrhythmia. The treatment is safe and promising for further studies.

Vestn Khir Im I I Grek. 1996;155(5):37-9.

The potentials of laser and electromagnetic-laser therapy in the treatment of patients with arteriosclerosis obliterans of the vessels of the lower extremities.

[Article in Russian]

Galimzianov FV.

A comparative analysis of the laser and electromagnetic laser therapy was performed in the complex treatment of patients with obliterating atherosclerosis of the lower extremity vessels. Laser treatment exerts a therapeutic effect related with its influence upon microcirculation. The effectiveness of complex treatment becomes higher when using a combination of laser therapy with the impulse electromagnetic therapy of complex modulation at the expense of improvement of the regional blood circulation in all links of the vasculature.

Vopr Kurortol Fizioter Lech Fiz Kult. 1996 Mar-Apr;(2):8-10.

The effect of a low-frequency alternating magnetic field on the autonomic system in children with primary arterial hypertension.

[Article in Russian]

Konova OM, Khan MA.

The paper provides cardiointervalographic data assessing autonomic nervous system (ANS) function in children with primary arterial hypertension exposed to low-frequency alternating magnetic field. Favourable effects of such magnetotherapy manifest in attenuation of sympathetic and vagotonic symptoms.

Lik Sprava. 1996 Jan-Feb;(1-2):58-62.

The clinico-biochemical, functional, immunological and cellular characteristics of the body reactions in patients with the initial stages of hypertension to the effect of a magnetic field.

[Article in Ukrainian]

Myloslavs’kyi DK, Koval’ SM, Sheremet MS.

The article presents a comprehensive evaluation of major clinical, laboratory and functional indices in the time course of magnetotherapy as well as during administration of such treatments. The most promising alternative appears to be that involving the use of immunologic and cellular parameters as markers of efficacy of therapeutic action of magnetic fields in early stages of hypertensive disease. Causes for effectiveness and ineffectiveness of the above treatment option are analyzed, approaches to eliminating those are outlined, the main indications and contraindications are determined, merits and demerits of magnetotherapy are drawn attention to.

Vopr Kurortol Fizioter Lech Fiz Kult. 1994 May-Jun;(3):10-2.

The effect of the joint use of plasmapheresis and magnetic treatment of the blood on the indices of blood rheology and hemodynamics in hypertension patients.

[Article in Russian]

Alizade IG, Karaeva NT.

The results are presented obtained on combined application of plasmapheresis and magnetic blood treatment as regards hemorheology and hemodynamics in 41 patients with essential hypertension stage II. The course introduction of the above combined treatment led to positive shifts in arterial pressure irrespective of the patients’ hemodynamic type, in blood density, elasticity and dynamic properties.

Vopr Kurortol Fizioter Lech Fiz Kult. 1994 Jan-Feb;(1):8-9.

The efficacy of low-intensity exposures in hypertension.

[Article in Russian]

Kniazeva TA, Otto MP, Markarov GS, Donova OM, Markarova IS.

One hundred hypertensive subjects with labile and stable disease were exposed to low-intensity low-frequency electrostatic field generated by the unit “Infita-A”. In labile hypertension, the field produces a hypotensive effect, improves myocardial contractility, increases myocardial and coronary reserves due to reduced peripheral resistance and stimulation of myocardial propulsion. Therapeutic response to the treatment is attributed to normalization of deep brain structure functioning.

Vopr Kurortol Fizioter Lech Fiz Kult. 1994 Mar-Apr;(2):18-20.

The effect of a low-frequency magnetic field on erythrocyte membrane function and on the prostanoid level in the blood plasma of children with parasystolic arrhythmia.

[Article in Russian]

Vasil’eva EM, Danilova NV, Smirnov IE, Kupriianova OO, Gordeeva GF.

As shown by clinical and biochemical evidence on 23 parasystolic children, the treatment with low-frequency magnetic field improves humoral and cellular processes participating in cardiac rhythm regulation. There is activation of Ca, Mg-ATPase in the red cells, a reduction of plasma thromboxane levels. Red cell phospholipid composition insignificantly change. Further courses of magnetotherapy may lower the risk of recurrent arrhythmia.

Vopr Kurortol Fizioter Lech Fiz Kult. 1993 Sep-Oct;(5):4-9.

Changes in intracellular regeneration and the indices of endocrine function and cardiac microcirculation in exposure to decimeter waves.

[Article in Russian]

Korolev IuN, Geniatulina MS, Popov VI.

Abstract

An electron-microscopic study of rabbit heart with experimental myocardial infarction revealed that extracardiac exposure to decimetric waves (DW) activated intracellular regeneration in the myocardium. This was associated with enhanced circulation and endocrine activity in the heart. Most pronounced regeneration was registered in adrenal exposure, the effect of the parietal exposure being somewhat less.

Vopr Kurortol Fizioter Lech Fiz Kult. 1993 Sep-Oct;(5):22-5.

The use of magnetics and laser therapy in treating obliterating vascular disease of the extremities.

[Article in Russian]

Kirillov IuB, Shval’b PG, Lastushkin AV, Sigaev AA, Kachinskii AE, Shashkova SN.

The paper presents the results of treatment received by 60 patients suffering from lower limb vascular obliteration stage IIA-III. The treatment involved combined use of magnetic field and laser irradiation. Peripheral circulation and central hemodynamics were evaluated rheographically and using ultrasound Doppler sphygmomanometry. Combined application of the above two modalities produced a greater effect on central hemodynamics compared to them introduced alone.

Ter Arkh. 1993;65(1):44-9.

The comparative efficacy of nondrug and drug methods of treating hypertension.

[Article in Russian]

Ivanov SG.

Effectiveness of some physical therapeutic factors (constant magnetic field, impulse currents) and new hypotensive drugs (tobanum, prinorm, ormidol, minipress, arifon, arilix) was compared in the treatment of essential hypertension stage II. It is suggested that nonpharmaceutical therapy can regulate functions, correct hemodynamic and microcirculatory disorders, produce therapeutic effect without side effects typical for drugs.

Lik Sprava. 1992 May;(5):40-3.

The effect of combined treatment with the use of magnetotherapy on the systemic hemodynamics of patients with ischemic heart disease and spinal osteochondrosis.

[Article in Russian]

Dudchenko MA, Vesel’skii ISh, Shtompel’ VIu.

The authors examined 66 patients with ischemic heart disease and concomitant cervico-thoracic osteochondrosis and 22 patients without osteochondrosis. Differences were revealed in values of the systemic hemodynamics with prevalence of the hypokinetic type in patients with combined pathology. Inclusion of magnetotherapy in the treatment complex of patients with ischemic heart disease and osteochondrosis favours clinical improvement, normalization of indices of central and regional blood circulation.

Lik Sprava. 1992 Oct;(10):32-5.

A comparative evaluation of the efficacy of quantum methods for treating hypertension patients.

[Article in Ukrainian]

Nykul TD, Karpenko VV, Voitovych NS, Karmazyna OM.

A study is presented of the effect of laser and microwave resonance therapy on the hemodynamics and hemorheology in 56 patients with hypertensive disease. The hypotensive effect of intravascular laser therapy is related to the positive changes, reduction of blood viscosity and general peripheral vascular resistance. The effect of low molecular electromagnetic radiation on acupuncture points favoured clear reduction of peripheral vessel resistance. Combination of laser and microwave resonance therapy produces a positive effect due to potentiation of these methods and, thus, influencing the systems of hemodynamics, hemostasis and hemorheology.

Vopr Kurortol Fizioter Lech Fiz Kult. 1992 Sep-Dec;(5-6):13-8.

The effect of decimeter waves on the metabolism of the myocardium and its hormonal regulation in rabbits with experimental ischemia.

[Article in Russian]

Frenkel’ ID, Zubkova SM, Liubimova NN, Popov VI.

Abstract

Biochemical and morphometric methods were employed to study the effect of decimetric waves (460 MHz, 10 and 120 mW/cm2) in cardiac and thyroid exposure on oxygen metabolism, myocardial microcirculation and contractility, thyroid and adrenal hormonal activity, kallikrein-kinin system activity in rabbits with experimental myocardial ischemia. Hypoxia discontinued in all the treatment regimens, but the exposure of the heart (field density 10 mW/sm2) had the additional effect on lipid peroxidation which reduced in the serum and normalized in the myocardium, on myocardial contractility, kallikrein-kinin system and on the adrenal and thyroid hormones.

Vopr Kurortol Fizioter Lech Fiz Kult. 1992 May-Jun;(3):14-7.

Magnetotherapy in obliterating vascular diseases of the lower extremities.

[Article in Russian]

Kirillov IuB, Shval’b PG, Lastushkin AV, Baranov VM, Sigaev AA, Zueva GV, Karpov EI.

The investigators have developed a polymagnetic system “Avrora-MK-01” employing running impulse magnetic field to treat diseases of the leg vessels by the action on peripheral capillary bed. At a pregangrene stage a positive effect on peripheral capillaries was achieved in 75-82% of the patients treated.

Kardiologiia. 1991 Feb;31(2):67-70.

Optimization of the treatment of patients with hypertensive disease from the rheological viewpoint.

[Article in Russian]

Shabanov VA, Kitaeva ND, Levin GIa, Karsakov VV, Kostrov VA.

The efficacy of various modes of correcting rheological disorders (membrane-protective agents, laser irradiation, plasmapheresis was compared in hypertensive patients. In 30% of the patients, the conventional antihypertensive therapy was demonstrated to deteriorate hemorheological parameters, which was due to its atherogenic impact on the blood lipid spectrum. Essential phospholipids, laser irradiation, and plasmapheresis, which are supplemented to the multimodality therapy promote a significant improvement of hemorheological parameters, which makes it possible to recommend them for management of hypertensive patients with a stable (essential phospholipids), complicated (laser irradiation), and refractory (plasmapheresis) course.

Khirurgiia (Mosk). 1990 Nov;(11):41-3.

Outpatient electromagnetic therapy combined with hyperbaric oxygenation in arterial occlusive diseases.

[Article in Russian]

Reut NI, Kononova TI.

The authors first applied hyperbaric oxygenation (HBO) in the outpatient clinic in 1968. Barotherapy was conducted in 107 outpatients whose ages ranged from 27 to 80 years; they had various stages of the disease of 5- to 20-year history. In 70 patients treated for obliterating diseases of the vessels by HBO in a complex with magnetotherapy by means of magnetophors, the remission lasted 1-2 years; patients treated by HBO alone had a 3-8 month remission. A prolonged positive effect was produced in 64 patients. The suggested effective and safe method is an additional one to the existing means of treating this serious and progressive disease, which can be applied successfully in outpatient clinics.

Ter Arkh. 1990;62(9):71-4.

The magnetotherapy of hypertension patients.

[Article in Russian]

Ivanov SG, Smirnov VV, Solov’eva FV, Liashevskaia SP, Selezneva LIu.

A study was made of the influence of the constant MKM2-1 magnets on patients suffering from essential hypertension. Continuous action of the magnetic field, created by such magnets, on the patients with stage II essential hypertension was noted to result in a decrease of arterial pressure without the occurrence of any side effects and in a simultaneous reduction of the scope of drug administration. Apart from that fact, magnetotherapy was discovered to produce a beneficial effect on the central hemodynamics and microcirculation. The use of the MKM2-1 magnets may be regarded as a feasible method of the treatment of essential hypertension patients at any medical institution.

Patol Fiziol Eksp Ter. 1989 May-Jun;(3):59-61.

Changes of central hemodynamics in rats with spontaneous hypertension under the effect of a low-frequency magnetic field.

[Article in Russian]

Buiavykh AG, Stukanov AF.

It was established that a course of exposures of the renal region of rats with spontaneous hypertension to the effect of low-frequency magnetic field influenced the central hemodynamic parameters significantly, which was displayed by reduction of total peripheral vascular resistance and normalization of the cardiac output.