The Effect And Mechanisms Of Rhein On Renal Interstitial Fibrosis In IgA Nephropathy Rats

Objective:To investigate the effect of rhein(RH)on renal interstitial fibrosis in IgA nephropathy and its relationship with TGF-β1/Smad signaling pathway.Methods:The 5-week-old SPF female Sprague Dawley(SD)rats were randomly divided into four groups(n=7),namely the control group,IgA nephropathy group,RH-treated group and RH-prevented group.IgA nephropathy model was established through BSA+LPS+CCl4 protocol.The RH-prevented group and the RH-treatment group were performed by intragastric administration of RH at different time points.Animals were sacrificed at the end of the 10 th week.Twenty-four-hour-Urine was collected before the animals were sacrificed to count urinary sediment red blood cells and 24-hour urine protein.After the animals were sacrificed,blood and kidney tissues were collected.Concentration of serum IgA was measured by Enzyme-linked immunosorbent assay.Immunofluorescence staining was used to detect glomerular IgA deposition.The pathological changes of kidney tissues in each group were detected by hematoxylin-eosin staining.Expression of TGF-β1,Smad3,α-SMA and FN proteins in renal tissues were detected by the two-step immunohistochemistry.Stereological analysis was performed by image analyzer.The expression levels of TGF-β1,Smad3,α-SMA and FN mRNA in renal tissue of each group was assessed by real-time quantitative PCR.Result:1.In the control group,no microscopic hematuria was found in the urine.Compared with the control group,microscopic hematuria was found in the urine of the IgA nephropathy group.The numbers of red blood cells in the urine sediment increased(P<0.01).Compared with the IgA nephropathy group,the numbers of red blood cells in the RH-treated group and RH-prevented group were decreased,and the difference between the RH prevention group and IgA nephropathy group was statistically significant(P<0.01).2.Compared with the control group,the quantity of 24-hour urine protein increased(P<0.01).Compared with the IgA nephropathy group,the quantity of 24-hour urine protein significantly was decreased in the RH-treated group and RH-prevented group(respectively P<0.05,P<0.01).3.There was no IgA deposition in the glomeruli mesangial of the control group.Strong green fluorescence with IgA deposition was observed in the glomeruli mesangial of the IgA nephropathy group,which showed discontinuous granular deposition(P<0.01).Compared with the IgA nephropathy group,the density of IgA green fluorescence was decreased in the glomeruli mesangial of the RH-treated group and RH-prevented group(P<0.01),particularly in the RH-prevented group.4.Compared with the control group,the concentration of serum IgA was significantly increased(P<0.01).Compared with the IgA nephropathy group,the concentration of the serum IgA was decreased in the RH-treated group and RH-prevented group.There are statistical difference between the RH-prevention group and IgA nephropathy group(P<0.01).5.In the control group,there was no swelling of the renal capsule space,no proliferation of mesangial cells and matrix,and no significant changes in renal interstitial and renal tubule.Histological data from the IgA group showed renal capsule space was dilated,glomerulus became smaller,mesangial cells and matrix were increased with fibrosis,swollen tubules and limited lumen.Compared with the IgA nephropathy group,Histological examination from both RH-treated group and RH-prevented group showed renal capsule space dilation,mesangial cells and matrix proliferation and tubule swollen were alleviated.6.The results of immunohistochemistry showed that expression of TGF-β1,Smad3,α-SMA and FN was increased in the IgA nephropathy group compared with the control group(P<0.01).Compared with the IgA nephropathy group,expression of TGF-β1,Smad3,α-SMA and FN was decreased in the RH-treated group and RH-prevented group(P<0.01).7.Real-time quantitative PCR data showed that the expression levels of TGF-β1,Smad3,α-SMA and FN mRNA in the IgA nephropathy group were increased compared with the control group(P<0.01).Compared with the IgA nephropathy group,the mRNA expression of TGF-β1,Smad3,α-SMA and FN was decreased in the RH-treated group(P<0.05).The mRNA levels of TGF-β1,Smad3,α-SMA and FN were significantly decreased in the RH-prevented group(respectively P<0.01,P<0.01,P<0.05,P<0.05).Conclusion:RH can alleviate renal interstitial fibrosis with IgA nephropathy in rats,which might be mediated by TGF-β1/Smad signaling pathway.