Low-dose Rapamycin Alters The Expression Of Nephrin And Podocin In Rats With Diabetic Nephropathy

Objective This experiment evaluated the effect of low dose rapamycin on rats with diabetic nephropathy, including blood glucose, weight-adjusted endogenous creatinine clearance rate (Ccr), kidney to body weight ratio, renal pathology and the expression of two podocyte-specific protein nephrin and podocin. Based on the experiment results, we tried to illustrate our hypothesis that low dose Rapamycin could delay the progress of kidney disease in diabetic rats by relieving damage to podocytes. Methods 24 SD male rats, weighted 200-250g, were randomly divided into normal control group (Group C, n=8) and diabetes group (n=16). After 12 hours'fasting, rats in diabetes group received 1% streptozotocin (STZ) by an intraperitoneal injection with a dose of 75mg/kg. 72 hours after injection, blood was f drawn from caudal vein and the level of blood glucose was measured. Those whose blood glucose level showed higher or equal to 16.7mmol/L were thought to be successful in model-building. Levels of blood glucose were measured on day 7, 14, 28 after model building. Only qualified rats were preserved, otherwise removed from the experiment. Qualified rats in diabetes group were further divided into untreated group (Group DN, n=8) and Rapamycin-treated group (Group DN+RAPA, n=8). From the fifth week, rats in Group DN+RAPA were lavaged with 0.3% Rapamycin solution at 1mg/kg; Group C and Group DN were lavaged with 0.5% carboxymethyl cellulose (CMC) at the same volume. Treatment lasted for 8 weeks. At the end of 12-week experiment, changes in blood glucose, 24-hour urine protein, weight-adjusted endogenous creatinine clearance rate (Ccr), kidney to body weight ratio were measured. Pathological changes of glomerulus were tested by HE and PASM staining, and ultramicrostructure of glomerulus by electron microscope. The expressions of nephrin and podocin tested by using immunohistochemistry and RT-PCR assay. Results Compared with Group C at the end of 12-week experiment, blood glucose, weight-adjusted creatinine (Ccr), 24-hour urine protein and kidney to body weight ratio were significantly elevated in Group DN(P<0.01); kidneys were enlarged, podocyte foot process widened and partially in fusion; GBM segmentally thickened, the expression of nephrin, podocin and their mRNA were significantly reduced. Group RAPA's weight-adjusted creatinine (Ccr), 24-hour urine protein were significantly lower than Group DN(P<0.01); the results of HE and PASM staining in Group RAPA showed glomerulus hypertrophy, matrix deposition in mesangium were obviously alleviated, as compared to Group DN; Podocyte foot process widening, fusion, and GBM thickening seen under electron microscope were mitigated; Nephrin and podocin's expression increased(P<0.01). Conculsion Nephrin and podocin are two important proteins to make up slit diaphragm(SD),which are critical to maintain SD structure and function. Reduction in expression of nephrin and podocin can cause integrity damage of filter membrane and proteinuria. Reduced expressions of nephrin and podocin in diabetic nephropathy may be one of the mechanisms accounting for proteinuria in the disease. Treatment of DN rats with low dose Rapamycin can significantly alleviate proteinuria, maintain the integrity of glomerulus filtration barrier and podocyte structure by increasing nephrin and podocin expression, and by relieving matrix deposition in mesangium.