| Objective: To explore the relationgship between hepatic fibrosis indiabetes rats and stromal ce11-derived factor-1(SDF-1) and its receptor CXCchemokine receptor4(CXCR4) through detecting the changes of SDF-1andCXCR4in hepatic tissue with diabetic rats. Simultaneously, observe theeffects of rosiglitazone on hepatic fibrosis and its influence to expression ofSDF-1and CXCR4in the hepatic tissue of diabetic rats, providing experimentproof to treat for diabitic hepatic fibrosis.Methods: The model of type2diabetes in rats was established by highfat diet combined with one-time intraperitoneal injection of low dosestreptozotocin (STZ,30mg/kg). Meanwhile, the control group was established.At week10, insulin resistance in the diabetic rats was confirmed byhyperinsulinemic–euglycemic clamp. The fasting blood glucose (FBG),fasting insulin (FINS), triglycerides (TG), cholesterol (TC), alanineaminotransferase (ALT) and aspartate aminotransferase (AST) were measured.Then, diabetic rats were randomly divided into untreated diabetic rats (DMgroup) and treated diabetic rats (RSG group) that received rosiglitazone(1.42mg/(kg·d),gavage). At week24, FBG, FINS, TG, TC, ALT, AST andglycosylated hemoglobin (HbA1c) were measured. The morphology of hepatictissue was observed by hematoxylin and eosin (HE) staining. The volume ofcollagen was evaluated by masson staining and the hydroxyproline (HYP)determination. The site and level of expression of SDF-1and CXCR4proteinwere examined by immunohistochemistry staining. The expression of SDF-1and CXCR4mRNA was examined by RT-PCR.Results:1Symptoms and Signs Eating, drinking, movement and color of fur of the rats in control groupwere normal. The diabetic rats appeared polyphagia, polydipsia and polyuria.The movement was significantly reduced and the color of fur was dark indiabeticrats. There was no obvious improvement of symptoms mentionedabove in the rosiglitazone group.2Glucose clamp experimentThe level of GIR in control group was11.34±0.76mg/kg/min, the level ofGIR in the DM group was5.45±0.84mg/kg/min, the level of GIR in diabeticrats was significantly lower than control group (P<0.01), indicating that thediabetic rats existed insulin resistance. The modle of experimental type2diabetic rats were established successfully.3The change of biochemistry markersThe level of fasting plasma glucose, fast insulin, triglyceride, cholesterol(P<0.05) and alanine aminotransferase in the diabetic group at the end of10week were higher than those in control group (P<0.01). The level of fastingplasma glucose, fast insulin, triglyceride, cholesterol, alanine aminotransferaseand glycosylated hemoglobin were significantly increased in diabetic group atthe end of24week, compared with control rats in the corresponding timeperiod (P<0.01). The level of fasting insulin, triglyceride, cholesterol andalanine aminotransferase in the rosiglitazone group decreased significantlycompared with diabetic rats at the end of24week (P<0.01), but fasting plasmaglucose, glycosylated hemoglobin and aspartate aminotransferase decreasedslightly (P>0.05).4Determination of hydroxyproline in hepatic tissueConcentrations of hydroxyproline in the diabetic group at the end of24week(229.99±2.96μg/(g·prot)) were higher than that of the controlgroup(138.07±6.05μg/(g·prot)) in the corresponding time period (P<0.01).Concentrations of hydroxyproline were increased in the rosiglitazonegroup(189.64±6.00μg/(g·prot)), compared with control group at the end of24week (P<0.01). The level of hydroxyproline decreased markedly in therosiglitazone group compared with that in diabetic group at24w (P<0.01). 5Masson stainingThere was less collgen fibers between portal area and central venous inthe control group, facio-density of collagen was0.0052±0.0011. Collagen wasincreased and structure was deranged between portal area and central venousin the diabetic rats at the end of24w, and the fiber widened and extended.facio-density of collagen was0.1215±0.0018, and had statistical difference(P<0.01). Facio-density of collagen of hepatic in the rosiglitazone group was0.0865±0.0023, accumulation of collagen in hepatic tissues decreasedsignificantly compared with diabetic group, and had statistical difference(P<0.01).6Hematoxylin and eosin stainingLiver cell cord arranged radiatedly around the central veins in the controlgroup The liver cells lined and the disse space was normal. However, in thediabetic rats at the end of24w, liver cells arranged disorderly, disse spacebecame broaden, necrosis of liver cells, fat-vacuolization formed,inflammatory cells increased, fibre hyperplasia. The alterations mentionedabove were all obviously ameliorated in rosiglitazone group.7ImmunohistochemistryIn control group, there were faint positive expression of SDF-1in hepaticcells, mainly in the cytoplasm. The positive staining optical density value was0.1231±0.0089. In the diabetic group at the end of24w, the positive stainingoptical density value was0.2768±0.0089, which was higher than that incontrol group in the corresponding time period, and had statistical difference(P<0.01). The positive expression of SDF-1in tissue of hepatic in therosiglitazone group was0.3669±0.0110, which was more than that in controlgroup and diabetic group in the corresponding time period, and had statisticaldifference (P<0.01). In control group, there were faint positive expression ofCXCR4in hepatic cells, mainly in the cytoplasm and plasmalemma. Thepositive staining optical density value was0.1089±0.0078. In the diabetic ratsat the end of24w, the positive staining optical density value was0.2954±0.0079, which was higher than that in control group in the corresponding time period, and had statistical difference (P<0.01). Thepositive expression of CXCR4in tissue of hepatic in rosiglitazone group was0.3994±0.0097, which was more than that in control group and diabetic groupin the corresponding time period, and had statistical difference (P<0.01).8The expression of SDF-1mRNA and CXCR4mRNAThe expression of SDF-1mRNA in diabetic group (0.82±0.04) wassignificantly higher than that in control group (0.64±0.05)(P<0.01). Theexpression of SDF-1in rosiglitazone group (1.02±0.06) was significantlyhigher than that in control group and diabetic group (P<0.01). The expressionof CXCR4mRNA in diabetic group (0.15±0.01) was significantly higher thanthat in control group (0.11±0.02)(P<0.01). The expression of CXCR4mRNAin rosiglitazone group (0.19±0.02) was significantly higher than that in controlgroup and diabetic group (P<0.01).Conclusions:1Hyperglycemia in rats could be caused by feeding high fat food andinjecting low dose streptozotocin (STZ), and then the GIR in glucose clampexperiment was conspicuously declined, which indicated a successful modelof type2diabetes rats. The morphology changes in hepatic tissue could befound by light microscope. Collagen accumulation in hepatic tissue could betested by Masson staining and the determination of hydroxyproline in hepatictissue, which indicated that the hepatic is also a target organ of diabetes, whilehepatic fibrosis is one of the main morphologic changes.2The pathological alteration of hepatic tissue in rosiglitazone group mitigateobviously, accumulation of collagen abatement, the expression of SDF-1andCXCR4increases, which indicating the intervention of rosiglitazone canmarkly reduce the progression of hepatic fibrosis in diabetic rats. Themechanism is, at least in part, through the induction of SDF-1and CXCR4. |
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