Effects Of Pulsed Electromagnetic Fields On Acute Hindlimb Ischemia Diabetic Rats In Microcirculation Angiogenesis

BackgroundIschemic diseases, particularly in diabetics, result in significant morbidity and mortality and have a profound economic impact. To ischemic disease, therapeutic angiogenesis is the focus of current research.Stem and progenitor cell therapy, protein of promoting angiogenesis factor therapy and gene therapy were the main methods to improve neovascularization and function of ischemic tissue. These methods in ischemic diseases have certain effect, but they were expensive, technically, especially to our country of lack of medical resources,it is still very difficult for the general clinical promotion. Therefore, we need an effective treatment..It is well documented that electromagnetic fields play a role in the repair of human issues.Electrical stimulation by means of capacitive coupling, combined magnetic fields and pulsed electromagnetic fields has been used for over 30 years to augment healing. and subsequently demonstrated electromagnetic fields increase angiogenesis in animal experiments . Based on the above findings, we hypothesized that pulsed electromagnetic fields might be beneficial in diabetic limb ischemia . In this study, we discuss pulsed electromagnetic fields on lower limb ischemia in diabetic rats the impact of angiogenesis, and explore its mechanism.AIMTo observe the effects of pulsed electromagnetic fields (PEMF) on acute hindlimb ischemia diabetic rats in microcirculation angiogenesis. Establishing diabetic rats acute hindlimb ischemia model, the experimental group was given pulsed electromagnetic field treatment, analysis effects of pulsed electromagnetic fields on hindlimb ischemia diabetic rat microcirculation angiogenesis, and explore its mechanism.MethodsDiabetes were induced by intraperitoneal injection of streptozotocin(60μg/g body with in 50 mmol/L citric acid buffer, pH 4.5) once to Sprague-Dawley rats. Then, 1 week later, blood was obtained periorbitally after 8 hours of fasting. Only rats with blood glucose values >300 mg/dl were kept in the protocol and randomized for experiment.Diabetic rats were anesthetized intraperitoneally with 3% pentobarbital. An incision was made along the inner right hind limb along the line of the femoral artery and vein. The proximal end of the femoral artery was tied with 4-0 silk suture .The femoral artery was dissected free from the limb and its peripheral branches. The distal end was severed, the artery was removed, and the skin was sutured closed. After surgery, rats were randomly divided into experimental group and control group(n=60).After operation, rats in experiment group were exposed to pulsed electromagnetic fields for 2 hours every day while those in control group were not given any treatment. Laser-Doppler perfusion measurements was used to determine the blood flow of ischemia hindlimbs respectively on the day 0, 7, 14 and 28 after operation, and immunohistochemistry analysis of CD31 andα-SMA were used to evaluate the changes in angiogenesis,immunofluorescence to detect expression of RECA-1. ELISA and Western blot were analyzed for the protein levels of VEGF,VEGFR2,FGF-2,FGFR1, P-ERK1/2,ERK1/2, P-P38 and P38.ResultsThe perfusion ratios were significantly higher in pulsed electromagnetic field-treated diabetic rats at day 14 and 28 compared with those in controls (0.64±0.02,0.85±0.021vs0.48±0.023,0.61±0.021,P<0.05)。CD31 density in tissues measured by immunohistochemistry significantly increased in pulsed electromagnetic field-treated groups on 14d and 28d(677.4±15.63/mm~2,837.2±25.60/mm~2vs495.2±25.31/mm~2,619.4±19.24/mm~2 , P<0.05 ) . Immunohistochemical analysis revealed significantly higher numbers ofα-SMA in animals exposed to PEMF at day 14 and 28(35.8±2.08/mm~2,50.6±3.08/mm~2vs 25.6±1.94/mm~2,32.4±1.72/mm~2,P<0.05), . Immunofluorescence analysis of RECA-1 density in tissues were significantly increased in pulsed electromagnetic field–treated groups at day 14 and 28 ( 179.8±6.95vs150.4±5.35/mm~2,253.0±8.67vs184.6±6.09/ mm~2,P<0.05).The levels of FGF-2 and FGFR1 in ischemic hindlimbs significantly increased in PEMF group at all time points and the same were p-ERK1/2 /total ERK1/2. No significant differences were observed in VEGF, VEGFR2 and p-p38/total p38 between the two groups.ConclusionPulsed electromagnetic fields can promote angiogenesis on acute hindlimb ischemia diabetic rat by up-regulation of FGF-2. As an indispensable signal pathway for FGF2-induced angiogenesis, MAPK/ERK1/2 is responsible for coupling FGFR1 to multiple downstream pathways to mediate blood vessel formation.