多囊蛋白2对低频脉冲电磁场促进成骨细胞分化成熟的影响北大核心CSCD

已知低频脉冲电磁场(pulsed electromagnetic fields,PEMFs)可以促进体外培养大鼠颅骨成骨细胞(rat calvarial osteoblasts,ROBs)的分化成熟,但ROBs感知PEMFs物理信号并启动成骨性分化的机制至今不明。本研究探究了0.6 mT 50 Hz PEMFs促进ROBs成骨性分化与位于ROBs表面的初级纤毛上的多囊蛋白2(polycystin2,PC2)的关系。首先用免疫荧光染色法研究了PC2是否位于ROBs初级纤毛内,然后通过Western blotting检测了经PEMFs处理不同时间后,ROBs内PC2蛋白表达的变化情况。接着用PC2阻断剂盐酸阿米洛利(amiloride HCl,AMI)预处理ROBs,检测碱性磷酸酶(alkline phosphatase,ALP)活性和PC2蛋白表达受PEMFs处理的影响情况,以及与骨形成相关的Runx-2、Bmp-2、Col-1、Osx的蛋白及基因表达的变化情况。再用RNA干扰法抑制ROBs内PC2的表达后,检测与骨形成相关基因表达情况。结果发现,PC2被定位于ROBs初级纤毛上,PEMFs处理增加了PC2蛋白表达量。PC2被AMI阻断后,PEMFs不再能提高PC2蛋白表达水平及ALP活性,PEMFs对成骨相关蛋白和基因表达的促进作用也被抵消。使用RNA干扰法抑制PC2的表达后,PEMFs也不再能提高骨形成相关基因的表达。结果表明:存在于成骨细胞初级纤毛表面的PC2在感知并传递PEMFs发出的物理信号中扮演着不可或缺的角色,PEMFs促进ROBs成骨性分化依赖于PC2的存在。本研究为阐明低频脉冲电磁场促进骨形成及治疗骨质疏松的机制研究奠定了基础。     

IN VITRO EFFECTS OF PEMFs ON BONE CELL CULTURES OF NORMAL AND OSTEOPENIC RAT

The aim of this study was to examine the effects of low-frequency, low-energy pulsed electromagnetic fields (PEMFs) on cell proliferation and differentiation in rat osteoblast primary cultures. Cells were obtained from normal and osteopenic rat bone and were named NB and OB, respectively. The osteoblastic phenotype was assessed by stimulation with 1,25(OH₂) vitamin D₃. NB and OB cells were seeded in multiwell plates and exposed to PEMFs for two different periods. Control cultures of both groups were incubated under the same conditions, with the pulse generator off. Assessment of PEMF effects was performed for the following parameters for each culture: alkaline phosphatase (ALP) activity, osteocalcin level, and MTT test. Results showed that OB and NB cell proliferation was significantly improved (p < 0.03, p = 0.04 respectively) after 48 h of PEMF exposure. Osteocalcin production of OB after 5 days of PEMF exposure was significantly higher than normal (p = 0.007) and osteopenic (p = 0.033) bone-derived controls. These results show that PEMFs act on osteopenic bone-derived osteoblasts, stimulating proliferation of cells and then, after a longer exposure, activating them.     

Pulsed electromagnetic fields on postmenopausal osteoporosis in southwest China: A randomized,active‐controlled clinical trial

A randomized, active‐controlled clinical trial was conducted to examine the effect of pulsed electromagnetic fields (PEMFs) on women with postmenopausal osteoporosis (PMO) in southwest China. Forty‐four participants were randomly assigned to receive alendronate or one course of PEMFs treatment. The primary endpoint was the mean percentage change in bone mineral density of the lumbar spine (BMDL), and secondary endpoints were the mean percentage changes in left proximal femur bone mineral density (BMDF), serum 25OH vitamin D3 (25(OH)D) concentrations, total lower‐extremity manual muscle test (LE MMT) score, and Berg Balance Scale (BBS) score. The BMDL, BMDF, total LE MMT score and BBS score were recorded at baseline, 5, 12, and 24 weeks. Serum concentrations of 25(OH)D were measured at baseline and 5 weeks. Using a mixed linear model, there was no significant treatment difference between the two groups in the BMDL, BMDF, total LE MMT score, and BBS score (P ≥ 0.05). For 25(OH)D concentrations, the effects were also comparable between the two groups (P ≥ 0.05) with the Mann–Whitney's U‐test. These results suggested that a course of PEMFs treatment with specific parameters was as effective as alendronate in treating PMO within 24 weeks. Bioelectromagnetics 34:323–332, 2013. © 2012 Wiley Periodicals, Inc.     

Influence of Electromagnetic Signal of Antibiotics Excited by Low-Frequency Pulsed Electromagnetic Fields on Growth of Escherichia coli

Energy medicine (EM) provides a new medical choice for patients, and its advantages are the noninvasive detection and nondrug treatment. An electromagnetic signal, a kind of EM, induced from antibiotic coupling with weak, extremely low-frequency pulsed electromagnetic fields (PEMFs) is utilized for investigating the growth speed of Escherichia coli (E. coli). PEMFs are produced by solenoidal coils for coupling the electromagnetic signal of antibiotics (penicillin). The growth retardation rate (GRR) of E. coli is used to investigate the efficacy of the electromagnetic signal of antibiotics. The E. coli is cultivated in the exposure of PEMFs coupling with the electromagnetic signal of antibiotics. The maximum GRR of PEMFs with and without the electromagnetic signal of antibiotics on the growth of E. coli cells in the logarithmic is 17.4 and 9.08 %, respectively. The electromagnetic signal of antibiotics is successfully coupled by the electromagnetic signal coupling instrument to affect the growth of E. coli. In addition, the retardation effect on E. coli growth can be improved of by changing the carrier frequency of PEMFs coupling with the electromagnetic signal of antibiotics. GRR caused by the electromagnetic signal of antibiotics can be fixed by a different carrier frequency in a different phase of E. coli growth.     

Effect of Low-Frequency Low-Energy Pulsing Electromagnetic Fields on the Response to Lectin Stimulation of Human Normal and Chronic Lymphocytic Leukemia Lymphocytes

Many literature reports suggest that at least one of the mechanisms of action of low-frequency pulsing electromagnetic fields (PEMFs) is to favor Ca++ movement into the cell. Ca¹ influx Is a fundamental step In the activation process of lymphocytes by lectins. We report here the results of the exposure of human normal and chronic lymphocytic leukemia (CLL) lymphocytes to lectins and PEMFs. Simultaneous exposure to PEMFs and mitogens significantly increased the number of normal responsive lymphocytes compared to those exposed to the mitogens alone. Almost all the normal lymphocytes entered into the mitotic cycle. The number of CLL lymphocytes stimulated by simultaneous exposure to PEMFs and lectins was doubled compared with lectin exposure alone. Ca^?1 influx was increased when the cultures were exposed to PEMFs. The stimulatory effect of PEMFs was mediated by an increased release of some B-cell growth factor by the T-Zymphocytes.     

Pulsed Electromagnetic Fields Improved Peripheral Nerve Regeneration After Delayed Repair of One Month

The goal of this study was to determine if postoperative pulsed electromagnetic fields (PEMFs) could improve the neuromuscular rehabilitation after delayed repair of peripheral nerve injuries. Thirty-six Sprague–Dawley rats were randomly divided into sham group, control group, and PEMFs group. The sciatic nerves were transected except for the control group. One month later, the nerve ends of the former two groups were reconnected. PEMFs group of rats was subjected to PEMFs thereafter. Control group and sham group received no treatment. Four and 8 weeks later, morphological and functional changes were measured. Four and eight weeks postoperatively, compared to sham group, the sciatic functional indices (SFIs) of PEMFs group were higher. More axons regenerated distally in PEMFs group. The fiber diameters of PEMFs group were larger. However, the axon diameters and myelin thicknesses were not different between these two groups. The brain-derived neurotrophic factor and vascular endothelial growth factor expressions were higher in PEMFs group after 8 weeks. Semi-quantitative IOD analysis for the intensity of positive staining indicated that there were more BDNF, VEGF, and NF200 in PEMFs group. It's concluded that PEMFs have effect on the axonal regeneration after delayed nerve repair of one month. The upregulated expressions of BDNF and VEGF may play roles in this process. © 2023 Bioelectromagnetics Society.     

Pulsed electromagnetic field potentiates etoposide-induced MCF-7 cell death

Etoposide is a chemotherapeutic medication used to treat various types of cancer, including breast cancer. It is established that pulsed electromagnetic field (PEMF) therapy can enhance the effects of anti-cancer chemotherapeutic agents. In this study, we investigated whether PEMFs influence the anti-cancer effects of etoposide in MCF-7 cells and determined the signal pathways affected by PEMFs. We observed that co-treatment with etoposide and PEMFs led to a decrease in viable cells compared with cells solely treated with etoposide. PEMFs elevated the etoposide-induced PARP cleavage and caspase-7/9 activation and enhanced the etoposide-induced down-regulation of survivin and up-regulation of Bax. PEMF also increased the etoposide-induced activation of DNA damage-related molecules. In addition, the reactive oxygen species (ROS) level was slightly elevated during etoposide treatment and significantly increased during co-treatment with etoposide and PEMF. Moreover, treatment with ROS scavenger restored the PEMF-induced decrease in cell viability in etoposide-treated MCF-7 cells. These results combined indicate that PEMFs enhance etoposide-induced cell death by increasing ROS induction–DNA damage–caspase-dependent apoptosis.     

脉冲电磁场促进大鼠成骨细胞成熟与矿化依赖于NO/cGMP信号途径

脉冲电磁场(pulse electromagnetic fields, PEMFs)能够促进大鼠成骨细胞（rat osteoblast cells, ROB）成熟矿化,但是其作用机制并不明确。本实验主要研究PEMFs促进大鼠成骨细胞成熟矿化与NO/cGMP信号途径的关系,进而阐明PEMFs促进成骨细胞成熟矿化的机理。首先将成骨细胞经50Hz、0.6mT脉冲电磁场作用不同时间后,检测细胞培养液中一氧化氮(Nitric Oxide, NO)和细胞内3?-5?-环鸟苷一磷酸(3?-5?-cyclic-GMP, cGMP)的含量,以探明电磁场是否影响NO和cGMP的合成;其次,提取总蛋白,应用蛋白质印迹检测细胞内eNOS、iNOS和PKG-1的蛋白表达量;最后,利用NOS的阻断剂L-NAME抑制NO信号通路后,检测成骨性相关指标,包括碱性磷酸酶（ALP）活性、钙化结节数量、成骨性基因Bmp-2、Collagen-1、Osterix及破骨细胞调节因子Rankl基因的表达量。结果发现经PEMFs处理后,NO含量及cGMP含量均有明显升高;细胞内eNOS、iNOS和PKG-1蛋白表达量较空白对照组均有显著升高,说明PEMFs能够激活NO/cGMP信号途径。且经PEMFs处理的成骨细胞,ALP活性升高,BMP-2、Collagen-1和Osterix基因表达量显著增加,Rankl基因表达量下降,成骨细胞形成钙化结节的能力增强,当加入L-NAME,PEMFs引起的ALP活性增加、成骨性基因表达升高和钙化结节形成能力增强的趋势均被显著抑制。上述结果表明,经PEMFs处理成骨细胞成熟矿化过程中 NO/cGMP信号通路被激活;如该通路被抑制,则电磁场促成骨作用被抵消。说明脉冲电磁场促进成骨细胞成熟矿化依赖于NO/cGMP信号通路。     

Pulsed electromagnetic fields accelerate proliferation and osteogenic gene expression in human bone marrow mesenchymal stem cells during osteogenic differentiation

Osteogenesis is a complex series of events involving the differentiation of mesenchymal stem cells to generate new bone. In this study, we examined the effect of pulsed electromagnetic fields (PEMFs) on cell proliferation, alkaline phosphatase (ALP) activity, mineralization of the extracellular matrix, and gene expression in bone marrow mesenchymal stem cells (BMMSCs) during osteogenic differentiation. Exposure of BMMSCs to PEMFs increased cell proliferation by 29.6% compared to untreated cells at day 1 of differentiation. Semi‐quantitative RT‐PCR indicated that PEMFs significantly altered temporal expression of osteogenesis‐related genes, including a 2.7‐fold increase in expression of the key osteogenesis regulatory gene cbfa1, compared to untreated controls. In addition, exposure to PEMFs significantly increased ALP expression during the early stages of osteogenesis and substantially enhanced mineralization near the midpoint of osteogenesis. These results suggest that PEMFs enhance early cell proliferation in BMMSC‐mediated osteogenesis, and accelerate the osteogenesis. Bioelectromagnetics 31:209–219, 2010. © 2009 Wiley‐Liss, Inc.     

脉冲电磁场对家猪淋巴细胞的细胞遗传学效应

以家猪外周血淋巴细胞为材料,研究了脉冲电磁场（pulsing electromagnetic fields,简称PEMFS)树细胞的遗传学效应,实验发现,100和200kHz的PEMFs对家猪的淋巴细胞照射培养12,24,48h后,染色体畸变（包括非整倍体,染色体断裂等）频率明显高于对照组（P＜0．05）,其中,56％的染色体或染色单体断裂和42％的间隙发生在家猪常见染色体脆性位点部位,同时, 经100kHz和200kHz的PEMFs照射48h后,淋巴细胞姐妹染色单体交换（SCE）频率也明显高于对照组（P＜0．05）,实验结果表明,PEMFS能诱导DNA损伤和染色体畸变。     

Protective effect of low frequency low energy pulsing electromagnetic fields on acute experimental myocardial infarcts in rats.

This series of experiments assesses the effect of exposure to low-frequency pulsing electromagnetic fields (PEMFs) in 340 rats with acute experimental myocardial infarcts. The left anterior descending artery was ligated with suture thread, and the rats underwent total body exposure to PEMFs until they were killed. Twenty-four hours after surgery, the necrotic area was evaluated by staining with triphenyltetrazolium chloride. A significant reduction of the necrotic area was observed in the animals exposed to PEMFs compared with the nonexposed controls. Exposure for up to 6 days does not appear to affect the area of necrosis, although in exposed animals an increase of vascular invasion of the necrotic area is observed: 24.3 % as against 11.3 % in controls. No effect on the necrotic area size from exposure was found when the left anterior descending artery was occluded for 60 min, followed by reperfusion. The results reported show that exposure to PEMFs is able to limit the area of necrosis after an acute ischemic injury caused by permanent ligation of the left anterior descending artery. These data are in agreement with the protective effect of PEMFs observed on acute ischemia in skin free flaps in rats and in cerebral infarcts in rabbits.     

Pulsed electromagnetic fields promote bone formation by activating the sAC–cAMP–PKA–CREB signaling pathway

The application of pulsed electromagnetic fields (PEMFs) in the prevention and treatment of osteoporosis has long been an area of interest. However, the clinical application of PEMFs remains limited because of the poor understanding of the PEMF action mechanism. Here, we report that PEMFs promote bone formation by activating soluble adenylyl cyclase (sAC), cyclic adenosine monophosphate (cAMP), protein kinase A (PKA), and cAMP response element-binding protein (CREB) signaling pathways. First, it was found that 50 Hz 0.6 millitesla (mT) PEMFs promoted osteogenic differentiation of rat calvarial osteoblasts (ROBs), and that PEMFs activated cAMP–PKA–CREB signaling by increasing intracellular cAMP levels, facilitating phosphorylation of PKA and CREB, and inducing nuclear translocation of phosphorylated (p)-CREB. Blocking the signaling by adenylate cyclase (AC) and PKA inhibitors both abolished the osteogenic effect of PEMFs. Second, expression of sAC isoform was found to be increased significantly by PEMF treatment. Blocking sAC using sAC-specific inhibitor KH7 dramatically inhibited the osteogenic differentiation of ROBs. Finally, the peak bone mass of growing rats was significantly increased after 2 months of PEMF treatment with 90 min/day. The serum cAMP content, p-PKA, and p-CREB as well as the sAC protein expression levels were all increased significantly in femurs of treated rats. The current study indicated that PEMFs promote bone formation in vitro and in vivo by activating sAC–cAMP–PKA–CREB signaling pathway of osteoblasts directly or indirectly.     

The effects of pulsed electromagnetic fields on the cellular activity of SaOS-2 cells

Although pulsed electromagnetic fields (PEMFs) have been used for treatments of nonunion bone fracture healing for more than three decades, the underlying cellular mechanism of bone formation promoted by PEMFs is still unclear. It has been observed that a series of parameters such as pulse shape and frequency should be carefully controlled to achieve effective treatments. In this article, the effects of PEMFs with repetitive pulse burst waveform on the cellular activity of SaOS-2 osteoblast-like cells were investigated. In particular, cell proliferation and mineralization due to the imposed PEMFs were assessed through direct cell counts, the MTT assay, tissue nonspecific alkaline phosphatase (ALP) and Alizarin Red S (ARS) staining. PEMF stimulation with repetitive pulse burst waveform did not affect metabolic activity and cell number. However, the ALP activity of SaOS-2 cells and mineral nodule formation increased significantly after PEMF stimulation. These observations suggest that repetitive pulse burst PEMF does not affect cellular metabolism; however, it may play a role in the enhancement of SaOS-2 cell mineralization. We are currently investigating cellular responses under different PEMF waveforms and Western blots for protein expression of bone mineralization specific proteins.     

In vitro exposure of human chondrocytes to pulsed electromagnetic fields

The effect of pulsed electromagnetic fields (PEMFs) on the proliferation and survival of matrix-induced autologous chondrocyte implantation (MACI)-derived cells was studied to ascertain the healing potential of PEMFs. MACI-derived cells were taken from cartilage biopsies 6 months after surgery and cultured. No dedifferentiation towards the fibro- blastic phenotype occurred, indicating the success of the surgical implantation. The MACI-derived cultured chondrocytes were exposed to 12 h/day (short term) or 4 h/day (long term) PEMFs exposure (magnetic field intensity, 2 mT; frequency, 75 Hz) and proliferation rate determined by flow cytometric analysis. The PEMFs exposure elicited a significant increase of cell number in the SG2M cell cycle phase. Moreover, cells isolated from MACI scaffolds showed the presence of collagen type II, a typical marker of chondrocyte functionality. The results show that MACI membranes represent an optimal bioengineering device to support chondrocyte growth and proliferation in surgical implants. The surgical implant of MACI combined with physiotherapy is suggested as a promising approach for a faster and safer treatment of cartilage traumatic lesions.     

Effects of electromagnetic stimulation on the functional responsiveness of isolated rat osteoclasts

We report the effects of pulsed electromagnetic fields (PEMFs) on the responsiveness of osteoclasts to cellular, hormonal, and ionic signals. Osteoclasts isolated from neonatal rat long bones were dispersed onto either slices of devitalised cortical bone (for the measurement of resorptive activity) or glass coverslips (for the determination of the cytosolic free Ca²⁺ concentration, [Ca²⁺]). Osteoclasts were also cocultured on bone with osteoblastlike, UMR-106 cells. Bone resorption was quantitated by scanning electron microscopy and computer-assisted morphometry. PEMF application to osteoblast–osteoclast cocultures for 18 hr resulted in a twofold stimulation of bone resorption. In contrast, resorption by isolated osteoclasts remained unchanged in the presence of PEMFs, suggesting that osteoblasts were necessary for the PEMF-induced resorption simulation seen in osteoblast–osteoclast cocultures. Furthermore, the potent inhibitory action of the hormone calcitonin on bone resorption was unaffected by PEMF application. However, PEMFs completely reversed another quite distinct action of calcitonin on the osteoclast: its potent inhibitory effect on the activation of the divalent cation-sensing (or Ca²⁺) receptor. For these experiments, we made fura 2-based measurements of cytosolic [Ca²⁺] in single osteoclasts in response to the application of a known Ca²⁺ receptor agonist, Ni²⁺. We first confirmed that activation of the osteoclast Ca²⁺ receptor by Ni²⁺ (5 mM) resulted in a characteristic monophasic elevation of cytosolic [Ca²⁺]. As shown previously, this response was attenuated strongly by calcitonin at concentrations between 0.03 and 3 nM but remained intact in response to PEMFs. PEMF application, however, prevented the inhibitory effect of calcitonin on Ni²⁺-induced cytosolic Ca²⁺ elevation. This suggested that the fields disrupted the interaction between the calcitonin and Ca²⁺ receptor systems. In conclusion, we have shown that electromagnetic fields stimulate bone resorption through an action on the osteoblast and, by abolishing the inhibitory effects of calcitonin, also restore the responsiveness of osteoclasts to divalent cations. J. Cell. Physiol. 176:537–544, 1998. © 1998 Wiley-Liss, Inc.     

Effect of localized pulsed electromagnetic fields on tail-suspension osteopenia in growing mice

Pulsed magnetic fields (PEMFs) have been used effectively to treat bone fractures and sciatic-nerve-section-induced osteopenias. Properly applied PEMFs are presumed to stimulate osteogenesis. Mouse-tail suspension has been implemented as a means of inducing an osteopenic response in the long bones of the hind limbs. To evaluate localized PEMF effects, the mouse-suspension model was modified to accommodate the use of miniature wire coils affixed directly to the rear legs. Laterally and axially orientated PEMF effects were compared. Three test groups of mice included (C) control mice, (S) tail-suspended mice with treatment apparatus attached, and (SF) tail-suspended mice with apparatus attached and PEMFs delivered. The SF group was divided into mice receiving axial or lateral PEMFs. Significant bone changes occurred in suspended as compared with control mice after a 2-week test period. The PEMF mice showed significantly fewer osteopenic effects than did untreated, suspended mice. These findings are based on biomechanical measures of stiffness, strength, ductility, and energy as well as whole-bone mass and porosity. The effects of PEMFs on these properties differ for axial and lateral exposures. The results are discussed in terms of mechanisms underlying PEMF effects.     

不同强度50 Hz脉冲电磁场促进大鼠颅骨成骨细胞矿化成熟最佳参数的筛选

为研究不同强度脉冲电磁场(pulse electromagnetic fields,PEMFs)对大鼠颅骨成骨细胞（rat skull osteoblasts,OB）增殖及成熟矿化的影响,将大鼠颅骨成骨细胞随机分为 7 组. 检测大鼠颅骨成骨细胞的增殖,细胞内碱性磷酸酶（ALP）活性变化,细胞沉积钙盐的情况,组织化学染色以及成骨细胞内标志性分子表达量的改变.结果显示,0.6 mT组促细胞增殖作用最强（P <0.01）;0.6 mT、1.8 mT、3.0 mT和3.6 mT均能提高ALP活性,其中0.6 mT ALP活性最高（P＜0.01）;在磁场处理4 ～12 d时细胞沉积钙盐逐渐增加,6种强度的脉冲电磁场均能促进钙盐沉积,尤以0.6 mT水平最高; ALP 染色、茜素红染色0.6 mT 组均显著高于对照组（P<0.01）;0.6 mT组 Bmp-2和Collagen-1 mRNA 的表达明显（P<0.01）高于对照组,磁场处理组Rankl mRNA 的表达均比对照组低. 0.6 mT 50 Hz 脉冲电磁场是促进成骨细胞增殖和矿化成熟的最佳参数,这为采用脉冲电磁场治疗骨质疏松症提供了治疗参数的基础支持.