| ObjectiveThe aim of this study was to evaluate the relation between expressions of nuclear factor-kappaB (NF-κB)p65 and monocyte chemoattractant protein-1(MCP-1) in renal tissues and renal structural and functional lesions in type 2 diabetic rat models induced by low-dose streptozotocin(STZ) after fore-administrated with the sucrose and fat enriched diets for 4 weeks, and to observe the effects of pyrrolidine dithiocarbamate(PDTC), pioglitazone(PIO),and PDTC combined with PIO on the renal expressions of NF-κB p65 and MCP-1,and to investigate their possible renoprotective mechanism.MethodsAfter adaptive feeding for 1 week,a total of 55 specific pathogen-free male SD rats were randomly divided into 2 groups:normal group for comparison purpose (NC group,n=10)who were administrated with normal diet and experimental model group(n=45) who were administrated with the sucrose and fat enriched diets.The rats of experimental model group were intraperitoneally given low-dose STZ (30mg·kg-1,dissolved in the 0.1mmol·L-1 citrate buffer solution) for once after the sucrose and fat enriched diets for 4 weeks.After 72 hours,measured randomly blood glucose twice in tail vena and selected 40 ones of 45 experimental model group rats whose blood glucose was equal or above 16.7 mmol·L-1 into experimental group.Then the modeling rats were further randomly divided into four groups: diabetic rats without treatment(DM group,n=10),diabetic rats treated with PDTC(PDTC group,n=10,100 mg·kg-1·d-1),diabetic rats treated with pioglitazone(PIO group,n=10, 10 mg·kg-1·d-1) and diabetic rats treated with combined PDTC with PIO(PDTC+PIO group,n=10, 100 mg·kg-1·d-1 and 10 mg·kg-1·d-1). The rats of treatmental groups were performed by intragastric administration for 8 weeks.The rats of NC group and DM group were performed by intragastric administration with equivalent distilled water for 8 weeks.Before the rats were sacrificed,we collected 24 hours urine for measurement of 24h urinary protein(24hUP),then killed the rats,gathered blood from inferior venous cave,and measured the fasting plasma glucose(FPG),total cholesterol(TC), triglyceride(TG), low density lipoprotein cholesterol(LDL-C),high density lipoprotein cholesterol(HDL-C),blood urea nitrogen (BUN) ,serum creatinine(Scr), superoxide dismutase(SOD),and malondialdehyde(MDA) were detected.Kidneys were removed and weighed to calculate the ratio of kidneys'weight vs body weight (KW/BW,KI) . Put the kidneys into 10% formalin to be fixed and made paraffinslices. Observed the pathological changes of the kidneys by PASM-HE,and studied the expressions of NF-κB p65 and MCP-1 in the kidneys with immunohistochemistry and immunoblotting. The data were analyzed by SPSS13.0 system.ResultsThe features of renal involvement and typical early morphological changes of diabetic nephropathy appeared in type 2 diabetic rat models induced by low-dose STZ combined with the sucrose and fat enriched diets at the end of experiment.Compared with NC group,the levels of FPG,TC,TG,LDL-C,BUN,Scr, 24hUP,KW,KI,MDA,extracellular matrix (ECM) accumulation index and the expressions of NF-κB p65 and MCP-1 were significantly increased in renal tissue (P<0.01),but body weights(BW), HDL-C,the enzyme activity of SOD were greatly decreased in DM group, PDTC group,PIO group and PDTC+PIO group(P<0.01).Compared with DM group,the levels of FPG,TC,TG,LDL-C, BUN,Scr,24hUP, KW,KI,MDA, ECM accumulation index and the expressions of NF-κB P65 and MCP-1 in renal tissue were significantly decreased in PDTC group,PIO group and PDTC+PIO group (P<0.01),but the levels of BW,HDL-C, the enzyme activity of SOD in PDTC group,PIO group and PDTC+PIO group were significantly increased (P<0.01).Compared with P group and PIO group,the obove indexes of PDTC+PIO group were further improved,and the expressions of NF-κB P65 and MCP-1 were more decreased with further improvement of renal pathological and functional changes (P<0.01,except for PDTC+PIO group's glucose vs PIO group: P>0.05).The expressions of NF-κB P65 and MCP-1 in renal tissues of type 2 diabetic rats were positively related with the levels of 24hUP,BUN,Scr,ECM accumulation index(P<0.01).At the same time,the expression of MCP-1 was positively related with the level of NF-κB p65 in renal tissues of type 2 diabetic rats.Conclusion(1)Overt persistent proteinuria and typical early morphological changes were observed in diabetic nephropathy in the type 2 diabetic rats.Increasing expressions of NF-κB p65 and MCP-1were positively related with functional and pathological changes in kidneys of type 2 diabetic rats.(2)The drugs of PDTC and PIO can obviously improve glucolipid metabolism, reduce 24hUP and the expressions of NF-κB p65 and MCP-1 in renal tissues of type 2 diabetic rats.So these drugs could prevent the courses of diabetic nephropathy by the improvement of renal pathology and function.The protective effects on kidneys of diabetic rats produced by combined use of PDTC with PIO were much more benefical than by application of either PDTC or PIO alone.(3)The mechanism of the two drugs appears to be correlated with decreasing oxidative stress,inhibiting overexpression of MCP-1 and macrophage infiltration in renal tissues of nephritic rats by down-regulating NF-κB p65 activation.In a word,PDTC and PIO can control the progression of DN by inhibiting renal inflammatory reaction. |
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