Effects Of Low Frequency Pulsing Electromagnetic Fields On Endothelial Progenitor Cells Derived From Rat Bone Marrow

Background and ObjectiveIschemic heart disease is detrimental to human health, and the main measure to treat it is to encourage endothelial repair and angiogenesis. Previous studies have found that Endothelial Progenitor Cells (EPCs) participate not only in the process of vasculogenesis, but also in the process of postnal neovascularization and repair of injured endothelium, and bone marrow derived EPCs (BM-EPCs) account for 5%-25% of neovascularization. Laboratary evidence indicated that the bionomics of EPCs such as cell number and function have an inverse correlation with risk factors of cardiovascular disease incluing hyperlipemia,diabetes and C-rective protein(CRP), the potential of angiogenesis descend in this situation. Megnetic Field as a foreign interventional instrument have already found that it could enhance endothelium repair and promote Fibroblast growth factor (FGF) secretion, and form a 3D blood vessel structure in a fibrin-coated condition. This indicated that Magnetic Field could promote vasculogenesis. In the present study, we examined the effect of low frequency electromagnetic field at different intensity and exposure time on proliferation, apoptosis and NO secrtion of EPCs derived from rat Bone Marrow in order to offer a new rationale foundation for therapeutic angiogenesis. MethodsPart one: Isolation, purification and identification of rat BM-derived EPCs in vitro. Hollow femurs and tibias of rat were prepared by standard surgical procedures and whole bone marrow was harvested by flushing marrow. Total mononuelear cells (MNCs) were isolated from rat bone Marrow by density gradient centrifugation , then the cells were plated on culture dishes supplemented with vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF).After 4 days, the adherent cells were collected. EPCs were characterized by FITC-UEA-I and Dil-acLDL, then these cells were detected by fluorescent staining under a laser scanning confocal microscope. Part two: Effect of low frequency electromagnetic field on the proliferation,NO secretion and apoptosis of BM-EPCs EPCs cultured for 4 days were collected, and were exposed to different intensity and time point of low frequency pulsing electromagnetic field every day. After 3 days exposure, proliferation capacity was measured by MTT assay, cell cycle and apoptosis was detected by FCM, NO secretion of EPC was measured by the Griess assay.Results1. Rat BM-derived EPCs were cultured and identified in vitro successfully.2. Compared with control group, 4h and 8h exposure group at 0.6 mT and 2h exposure group at 1.0mT enhance the prolifrative proficiency of EPCs(P<0.01), 4h and 8h exposure group at 1.0mT reduced EPCs proliferation (P<0.01), The other exposure group showed no statistical difference (P > 0.05),3. Compared with control group, 8h exposure group at 0.6 mT and 2h exposure group at 1.0mT promote NO secretion of EPC(sP<0.01). There was no statistical difference between 2h exposure group at 1.0mT and 8h exposure group at 0.6 mT (P > 0.05). 8h exposure group at 1.0mT reduced NO secretion (P<0.01), The other exposure group showed no statistical difference (P > 0.05),4. Compared with control group, 8h exposure group at 1.0mT induced EPCs apoptosis (P<0.01), while the other groups showed no statistical difference (P > 0.05),ConclusionsLow frequency pulsing electromagnetic field had an effect on biology property of EPCs in a dose-depedent and time-dependent manner. At some intensity and exposure time it could enhance EPCs proliferation, NO secretion, and had no effect on cell apoptosis; with the longer exposure time and stronger intensity, low frequency pulsing electromagnetic field could inhibite cell proliferation, reduce NO secretion and induce cell apoptosis.