The Regulation of PPAR-γ Agonist on TGF-β1and c-Ski in Kidney Tissue of Diabetic Rats
|School||Tianjin Medical University|
|Course||Biochemistry and Molecular Biology|
|Keywords||Diabetic nephropathy transforming growth factor-β/Smads signalingpathway transforming growth factor-beta1 cellular Sloan-Kettering institute Peroxisome proliferator-activated receptor-gamma agonist pioglitazone|
Objective:Diabetic nephropathy is the most common cause of endstage kidney disease, and the leading cause of death in diabetic patients. The fundamental mechanism responsible for nephropathy in type2diabetes involves tubulointerstitial fibrosis and glomerulosclerosis, caused among other processes by oxidative stress, activation of the aldosterone-renin-angiotensin system, inflammation, release of profibrotic factors such as transforming growth factor β1(TGFβ1) and plasminogen activator inhibitor1(PAI-1). TGF-β1is also a critical meditor in glomerulosclerosis and renal tubulointerstitial fibrosis. The co-suppress factor of nuclear transcription c-Ski (cellular Sloan-Kettering institute) is the intra-cellular congener of v-Ski (virus Sloan-Kettering institutein) in human organization. Its main biological function is the negative regulation of TGF-β/Smad signaling pathway, to repress the formation of Smad complex and activate the transcription of target genes of TGF-β downstream. But weather they have the similar mechanism in the development of DN, and the relationship with Peroxisome proliferator-activated receptor y (PPAR-y) is still remain to be elucidated. PPAR-y is a class of ligand-activated nuclear receptors, TZDs (thiazolidine two ketones) is a new type of insulin sensitizer as the most important exogenous activator of PPAR-y. In this study, we built diabetic rat model with streptozotocin(STZ), to observe the expressions of c-Ski and TGF-β1in renal tissue of diabetic rats, to investigate the regulation of PPAR-y on TGF-β1and c-Ski, the repressor of TGF-(3/Smad pathway, and the relationship with diabetic nephropathy, to provide new evidence for the mechanism of DN.Method:Murine diabetic models were induced with streptozotocin (STZ,40mg/kg). Rats were randomized into normal control group (NC), diabetes group (DM) and Pioglitazone treatment group [PT, PIO15mg/(kg·d)]. All the rats were weighed every week, and detected blood glucose every two weeks. One day before they dead, we collected the24hour urine to detect24h urinary microalbumin (UMA). Other indicators related to renal function such as blood urea nitrogen (BUN)、 Triglyceride(TG) were detected with blood. The kidney hypertrophy index (LKW/BWT) was accounted with left kidney. We can use hematoxylin and eosin stain to observe pathological morphology of nephridial tissues. The liquid nitrogen frozen specimens from renal cortex were taken to use to analyze the level of TGF-β1 and c-Ski mRNA by the method of Real-time PCR. The protein expression of TGF-β1and c-Ski were determined by immunohistochemistry, then analyze the result by semi-quantitative method.Result:1. Compared with the normal control group, diabetic rats showed significant increased plasma glucose level and decrease in body weight (P＜0.01); The level of24h urinary volume、BUN、24h UMA、TG and LKW/BWT were significantly higher (P＜0.01). All the above indicated that diabetic rats had got nephropathy complication.2. TGF-β1mRNA expressed in renal cortex were increased obviously in diabetic group compared with normal rats, as well as its protein expression(P＜0.05); on the other hand, the protein level of c-Ski was significantly lower (P＜0.01), and the difference of c-Ski and SnoN mRNA level between two groups showed insignificance. These suggested that the effect of c-Ski as a repressor of TGF-β1 could act in protein level.3. After treatment with Pioglitazone, compared with diabetic rats, The level of serum glucose、24h urinary volume、BUN、24h UMA、TG、LKW/BWT was significantly lower (P＜0.05), and the body weight control was not obvious at first two weeks and began to increase from the fourth week (P＜0.01). After8weeks of treatment, the expression levels of TGF-β1mRNA and protein decreased significantly in the kidney of pioglitazon treated group compared with diabetic group(P＜0.05), and the protein level of c-Ski increased(P＜0.05). however, the differences of c-Ski and SnoN mRNA level between pioglitazon treated group and diabetic group were not significant (P>0.05).Conclution:1. TGF-β1is a critical mediator of glomerulosclerosis and renal tubulointerstitial fibrosis. c-Ski could inhibited its gene transdction of downstream from prevent forming of Smad protein complex. TGF-β1was significantly increased in the early stage of DN, while c-Ski was significantly decreased. Maybe it is the sharply less expression of c-Ski in DN decreased the blocking of TGF-β/Smad signaling pathway, and created such an ideal environment that TGF-β1could transduce signal to promote fibrosis without controll. Therefore, it is meaningful for down-regulating the profibrotic effect of TGF-β1, preventing renal fibrosis and delaying DN progression to have the expression of c-Ski under control.2. According to the results of real-time PCR, we suggested that the effect of c-Ski as a repressor of TGF-β1could act in protein level.3. PPAR-gamma agonist pioglitazone may up-regulate the expression and function of c-Ski, to inhibite the overexpression of TGF-β1and make control of its unlimited profibrotic effect. Thus, we speculated that pioglitazone could protect renal function, reduce the renal damage caused by diabetic mellitus, so as to achieve the limited control of DN.