Study On Mechanism Of Yiqi Yangyin Qingre Tongluo Formula On Diabetic Foot Mice

Diabetic foot is a systemic disease.The longer foot ulcer lasts,the more likely foot ulcer complicates infection and greater risks patients have of amputation.Therefore,it is of vital importance to promote diabetic ulcer healing efficiently.This study suggests that according to Luobing theory,diabetic foot is caused by dysfunction of spleen in transport,impairment of both qi and yin and obstruction of collaterals by blood stasis and heat.Deficieny and blood stasis in collaterals coexist.Pathological changes locate in collaterals and muscles.Promoting collateral blood circulation is considered as a general treatment principle.Yiqi Yangyin Qingre Tongluo Formula is created in accordance with the rule.Through theoretical analysis and experimental study,curative effect of the formula on diabetic foot is evaluated and its potential mechanism is discussed to improve its clinical application value.Objective:To observe the effect of Yiqi Yangyin Qingre Tongluo Formula on foot ulcer of type 2 diabetes mice,discuss potential pathogenesis of diabetic foot and mechanism of Yiqi Yangyin Qingre Tongluo Formula on diabetic foot ulcer,and provide new ideas for prevention and treatment of diabetic foot and scientific basis for clinical promotion.Methods:10 mice,randomly selected from 70 SPF male C57BL/6J mice,were named normal group.Other mice,fed by high fat and sugar fodder,given 120 mg/kg streptozotocin(STZ)intraperitoneal injection once and lower extremity vessel ligature,were established as mouse models of type 2 diabetic mellitus.Successful mouse models were divided into 6 groups according to random number table:model group,Metformin and Clostazol group,Tongsaimai group,and high,middle and low dose of Yiqi Yangyin Qingre Tongluo Formula groups.General state,body weight,fasting blood glucose(FBG)and ulcer square of all the mice were observed dynamically.The experiments also oberserved the effects of Yiqi Yangyin Qingre Tongluo Formula on inflammatory factors,vascular endothelial function,angiogenesis indexes,oxidative stress indexes,fasting insulin(FINS),fasting C-Peptide(FC-P),insulin-like growth factor 1(IGF-1),islet cell morphology and function by such means as biochemistry,optical microscope,immunohistochemistry and three-dimensional quantitative stereology.Results:(1)Compared with model group,body weight in high dose of Yiqi Yangyin Qingre Tongluo Formula group and Metformin and Clostazol group increased(P<0.05),with increase tendency close to that of normol group.(2)Compared with model group,F3G in Metformin and Clostazol group,Tongsaimai group,high,middle and low dose of Yiqi Yangyin Qingre Tongluo Formula groups decreased steadily(P<0.001).There was a downward trend in FBG bwteen high,middle and low dose groups.(3)Compared with model group,ulcer square reduced and scabbing time shortened in Metformin and Clostazol group,high,middle and low dose of Yiqi Yangyin Qingre Tongluo Formula groups(P<0.001,P<0.01 or P<0.05).Swelling of lower limb ulcers gradually subsided.Claudication and hyperaphia were improved.(4)High sensitive C reaction protein(hs-CRP)in high,middle and low dose of Yiqi Yangyin Qingre Tongluo Formula groups lowered.Interleukin-6(IL-6),tumor necrosis factor-a(TNF-a),thromboxane B2(TXB2)and endothelin-1(ET-1)in high dose group fell(P<0.001,or P<0.01).6-Keto-PGF-l?(6-K-PGF-la)in high dose group rose(P<0.01).(5)Compared with model group,vascular endothelial growth factor(VEGF)in high dose of Yiqi Yangyin Qingre Tongluo Formula group also rose(P<0.05).(6)Compared with model group,superoxide dismutase(SOD)and glutathione(GSH)increased(P<0.05,P<0.01,or P<0.001)and malondialdehyde(MDA)decreased(P<0.01)in Metformin and Clostazol group,Tongsaimai group,high and middle dose of Yiqi Yangyin Qingre Tongluo Formula groups.(7)Compared with model group,except low dose of Yiqi Yangyin Qingre Tongluo Formula group,FC-P increased in other groups(P<0.05 or P<0.001).Compared with low dose group,FC-P increased in Tongsaimai group and high dose group(P<0.01,or P<0.001).(8)Compared with model group,IGF-1 increased in Metformin and Clostazol group and high dose of Yiqi Yangyin Qingre Tongluo Formula group(P<0.05).Compared with model group,FINS in Metformin and Clostazol group rosed(P<0.05).Compared with low dose group,FINS in Metformin and Clostazol group,high and middle dose groups rose(P<0.05,or P<0.001).(9)Compared with model group,homeostasis model assessment of insulin resistance index(HOMA-IR)in treatment groups fell(P<0.001)·(10)Hematoxylin-eosin(HE)staining,immunohistochemical staining and three-dimensional quantitative stereology showed:Yiqi Yangyin Qingre Tongluo Formula can delay mouse pancreatic atrophy,promote islet struture and islet ? cell repaire,reduce pathological damage of islet cells,regulate systemic status of mice,and improve diabetic foot symptoms,so as to accelerate foot ulcers healing.Conclusion:(1)Yiqi Yangyin Qingre Tongluo Formula,especially at high dose,can ameliorate tissue ischemic and hypoxic status,and promote granulation tissue grow and foot ulcer healing through lowering blood glucose,inibiting weight continuous decline,beneficially regulating vascular endothelial function and depressing inflammatory factors release.(2)High dose Yiqi Yangyin Qingre Tongluo Formula can monitor serum VEGF concentration,activate cell proliferation and granulation tissue growth,promote revascularization and accelerate foot ulcer healing.Meanwhile,the formula can also enhance pancreatic antioxidative ability,and thus prevent or delay pancreatic function lesion.(3)High dose Yiqi Yangyin Qingre Tongluo Formula can decrease insulin resistance and enhance insulin sensitivity to promote islet structure recovery and islet ? cell repair,and better insulin resistance,? cell apoptosis and dysfunction.These indexes above in high dose group improve more than those in middle and low dose groups.(4)High dose Yiqi Yangyin Qingre Tongluo Formula has better effects in lowering glucose,reducing ulcer square and reducing insulin resistance than Tongsaimai Tablet.The effects of comprehensively improving endothelial function,reducing inflammatory cytokines,reducing islet atrophy,and improving the overall state of mice are superior to the effect of Metformin and Clostazol and that of Tongsaimai Tablet.Yiqi Yangyin Qingre Tongluo Formula can not only enhance antioxidative stress of diabetic foot mice and decrease insulin resistance as a whole,but also alleviate ulcerous oxidative inflammatory injury and improve wound microenvironment.These aspects give play to good therapeutic effect,especially in high dose Yiqi Yangyin Qingre Tongluo Formula group.