| Objectives:1.To investigate the changes of mitogen-activated protein kinase MAPK(P38MAPK)expression in the renal tissue of diabetic rats induced by STZ.2.To detection the the influence of pioglitazone on the expression of P38 MAPK and TLR4/P38 MAPK signaling pathway of diabetic rat renal tissue.Methods:After adaptivie feed for 1 week,60 male Sprague-dawlry(SD)rats were randomly divided into normal group(group A,n=10) and model group(n=50).Normal control group was fed with the normal diet, drinking freely,model group was fed with high sucrose-fat diets(HSFDs),also drinking freely. After HSFDs fed for 4 weeks, weight from rats were tested,blood samples from rats were collected and the fast blood glucose(FBG), fasting insulin(FINS) level, total cholesterol(TC), triglyceride(TG), low density lipoprotein cholesterol(LDL-C),high density lipoprotein cholesterol(HDL-C)were assayed to determined whether the insulin resistance was induced. Model group was fed with high sucrose-fat diets for 4 weeks,the subjects in model group were injected intraperitoneally with streptozotocin(STZ,35mg/kg) after fasting for 12 hours. These diabetic rats were determined by randomly blood was higher than VIII16.7mmol/L of three consecutive times after 72 hours of STZ injection(four cases was excluded because of failing to achieve the diagnosis standard of diabetes).These diabetic rats were randomly divided into five groups: small-dose pioglitazone intervention group(2.5mg/kg daily [group B, n=9]), large-dose pioglitazone intervention group(10mg/ kg daily [group C, n=9]), small-dose pioglitazone plus intraperitoneal injection P38 MAPK inhibitor SB203580 1mg/kg.d intervention group(group D, n=9) and large-dose pioglitazone plus intraperitoneal injection P38 MAPK inhibitor SB203580 1mg/kg.d intervention group(group E, n=9), no intervention group(group F, n=10). Pioglitazone was solubled in physiological saline and filled into stomach at 9:00 AM daily for eight weeks. These rats of group A were given physiological saline. The group D and group E were began given intraperitoneal injection P38 MAPK inhibitor SB203580 1mg/kg daily in the tenth weeks.At the end of the 12 th week, weight from rats were tested. At the same time, 24 h urine samples were collected to examine 24 h microalbuminuria. Blood samples of kidney from rats were assayed FBG, FINS, SCr, BUN, CRP, TG, TC, LDL-C, HDL-C. Then the rats were killed to take out the kidney. Blood sample were The morphological changes of kidney were observed through light microscopy after hematoxylin-eosin(HE) staining.Immunohistochemistry staining was used to examine the expression of P38 MAPK,TLR4, TGF-β1 protein in the kidney. The expression of P38 MAPK m RNA was analyzed by RT-PCR.Results:1.After four weeks, compared with group A, BW, FBG, FINS, HOMA-IR, TG, TC, LDL-C of the rats in model group were much higher(P<0.05).2.After 12 weeks, compared with group A, BW of the rats in group B, C, D, E, F were significantly lower(P<0.05).Compared with group A,FBG, TG, TC, LDL-C, SCR, CRP, BUN, 24 h urine protein in group B, C, D, E, F were significantly higher(P<0.05).Compared with group F, FBG, TG, TC, LDL-C, SCR, CRP, BUN,24 h urine protein in group B, C, D, E were significantly lower(P<0.05).3.The histopathological examination of rat renal suggests, from the naked eye and light microscopy: the normal control group A glomerular morphology neat, glomerular cells arranged in rules, basement membrane was clear and tidy, no thickening, glomerular capillary lumens stenosis, renal tubular-interstitial inflammatory cell infiltration. Group F glomerular volume increase, glomerular capillary loops collapse, occlusion of the lumen, thickening of basement membrane increased, a large number of inflammatory cell infiltration in renal interstitial. And the lesions in kidney of the rats of group B,C,D and group E with drug intervention was significantly lighter than group F.4.Immunohistochemical results showed that,Compared with group A, the renal levels of TLR4,P38 MAPK,TGF-β1 by group B, C, D, E were increased(P<0.05). Compared with group C and group B, group D and group B, group E and group C, the renal levels of TLR4,P38 MAPK,TGF-β1 by group B, C, D, E were decreased(P<0.05).5.Rat kidney P38 MAPKm RNA expression level: compared with control group, group B and C, D, E, F group in the rat kidney P38 MAPK m RNA expression were significantly higher(P < 0.05);Compared with group F, P38 MAPK m RNA expression of the rats in group B, C, D, E were significantly lower(P<0.05).Compared with group B, Group C P38 MAPKm RNA expression had decreased(P < 0.05);While using the P38 MAPK inhibitor SB203580 D, E group compared with group B, C, P38 MAPK m RNA expression significantly lower(P < 0.05).Conclusions:1.The expression of TLR4,P38 MAPK,TGF-β1 in the renal tissue of diabetic rats is increased induced by STZ and high sucrose-fat diets.2.Pioglitazone can relieve renal inflammation in diabetic rats, the mechanism may be through regulating the expression of TLR4,then down-regulation P38 MAPK expression in renal inflammation, thus inhibiting TLR4/P38 MAPK signaling pathway. |
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