The Experimental Study Of Antioxidation Effect Of Hepatocyte Growth Factor Gene On Rat Exposed To High Glucose Conditions And The Role Of Promoting Angiogenesis

Objective1. To study the effect of HGF on oxidative stress in Sprague-Dawley rats cultured in high glucose conditions.2. To explore the underlying mechanism of angiogenesis promoting with HGF in Sprague-Dawley rats cultured in high glucose conditions.Methods and Material1. The model of occlusive peripheral arterial disease was constructed in Sprague-Dawley rats cultured in high glucose conditions.2. The rats were divided into three groups randomly. After24hours of modeling, rats in the high dosage group were injected with pUDKH at200μg for each, those in the low dosage group were injected with pUDKH at100μg for each, and those in the control groups which were treated with aqua pro injectione for injection at the same volume. The period of treatment is15days.3. Detection methods:general observation; HE staining for examining morphologic changes after ischemic hind limb damage in rats; ELISA was used to detect the total ROS, HGF and beta-Gal expression in local ischemic hind limb tissue by the reagent kit and analyse quantitatively; the generation of endothelial progenitor cells number was detected by Flow Cytometry.4. All results are expressed as mean±SEM. One-way analysis of variance ANOVA and Post hoc Bonferroni t test comparisons were used to compare each paramerer. P<0.05was considered significant.Results1. Comparion of data, the blood glucose showed no statistics difference among each experimental group (P>0.05), and modeling can measured up to the standard. About24hour later, all modeling groups of the hind limbs couldn’t walk normally, otherwise one rat died in the placebo group within24hours after the operation. The wound was arescent without exudation in treatment group and exudate was reduced the placebo group within72hours. All rats were survived, without any swelling, pyorrhea or fissuration in treatment group. The time for onset and whole decrustation one week later and one rat was died in the placebo group. The effect of gross observation in treatment group by pUDKH was better than in control group.2. Histological observation by analysis of hematoxylin and eosin staining: Sarcoplasmic atrophy and degeneration, fibrotic hyperplasia and hyalinization, invasive degeneration of plasma cells, widened muscle space, and focal abscess could be seen in the control group. Plentiful capillaries were formed after the treatment by pUDKH, and stripes of muscle fibers were well kept in the high dosage group compared with the low dosage group. It is indicated that focal necrosis seen in as well as inflammatory cell infiltration.3. The results showed that with Elisa in the tissue homogenate of muscles around the femoral arteries, the formation amount of reactive oxygen species in high dosage injected group were much than that in the control group which were treated with aqua pro injectione for injection (P<0.05). The expression of HGF were much higher in high dosage group (P<0.05). Detection of endothelial progenitor cells showed high expression of quantitative fluorescence with CD34and CD133by Flow Cytometry.Conclusions1. It is indicated that pUDK carrying hepatocyte growth factors has the therapeutic effects on Limb ischemia of diabetic rats.2. HGF can protect body tissues against HG-mediated oxidative stress. HGF may balance the ROS production and ROS scavenging in body tissues through promoting the protective effects.3. HGF has the Effects on promotes angiogenesis and increaseing mobilization of endothelial progenitor cells recruitment.