1.Influence on the expression of type I and type II transforming growth factor-? receptors in cultured rat mesangial cells transfected by decorin gene
Xiuyan FENG ; Zhigang ZHANG ; Zhonghua ZHAO ; Qi CHEN ; Muyi GUO
Chinese Journal of Nephrology 1997;0(05):-
Objective To explore whether the antagonistic effect of decorin (DCN)on progression of glomeruloselerosis is associated with the inhibition of the expression of type I and type II transforming growth factor-? receptors (TGF-?R I and TGF-?R II )in mesangial cells (MsC). Methods RT-RCR and Western blot analysis were used to detect the expression of TGF-?R I and TGF-?R II mRNA and their proteins on cultured rat MsC stimulated by exogenous TGF-?1. Lipofectin-mediated method was used to transfect DCN vector into MsC. After screening and identifying of transfected MsC, RT-PCR and Western blot analysis were adapted to detect the changes of TGF-?R I and TGF-?R II expression respectively. Results The expression of TGF-?R I and TGF-?R II mRNA and their proteins on normal MsC stimulated by exogenous TGF-?1 increased in time-dependent manner and reached the peak at 24th hour. Compared with normal and untransfected MsC (1P-1), the mRNA and protein expression of TGF-?R I and TGF-?R II on MsC (3D-5, 7D-1) transfecled by DCN gene decreased significantly, and DCN gene transfection could antagonize the increase of mRNA and protein expression of both receptors caused by exogenous TGF-?1. Conclusions The expression of both TGF-?R I and TGF-?R II decreases obviously in MsC overexpressing DCN gene, which may be one of the importan antagonistic mechanisms of decorin involved in the development of glomeruloselerosis mediated by TGF-?.
2.Study on the inhibitory effect of decorin on rat mesangial cell growth and its signal transduction pathway
Xiuyan FENG ; Zhigang ZHANG ; Zhonghua ZHAO ; Qi CHEN ; Muyi GUO
Chinese Journal of Nephrology 2005;0(09):-
Objective To explore the inhibitory effect of decorin(DCN) on rat mesangial cell (MsC) growth and on the expression of mitogen-activated protein kinases (MAPKs) and p21 protein. Methods Lipofectin-mediated method was used to transfect DCN vector into MsC. After the screen and identification of transfected MsC, DCN-containing supernatant was collected and added into the culture medium of normal MsC, then flow cytometer was used to detect the cell cycle. Western blot analysis was used to explore the changes of MAPK and p21 protein. Immunofluorescence was adapted to detect the expression of p21 on cultured MsC. Results Compared with normal MsC, the number of G2-M cells treated with DCN-containing supernatant decreased to 35%(P
3.Effects of tripterine on local expressions of collagen type I and type IV in BW F1 mice kidney
Chen XU ; Zhaolong WU ; Zhigang ZHANG ; Muyi GUO
Chinese Journal of Rheumatology 2001;0(04):-
Objective To study the protective effects of tripterine on experimental lupus nephritis glomerulosclerosis.Methods Different doses of tripterine were injected peritoneally to BW F1 mice at different stages.The levels of 24 hour urine protein excretion and serum anti dsDNA antibodies,and the expressions of renal collagen type Ⅰ,type Ⅳ,MMP 2,TIMP 2,and transforming growth factor (TGF) ? 1 mRNA were analyzed.Results ①Tripterine suppressed the development of proteinuria,decreased the level of serum anti dsDNA antibodies,reduced the local expressions of TGF ? 1,collagen type Ⅰ,type Ⅳ,TIMP 2 and improved the expression of MMP 2 in murine kidney.②The use of tripterine before occurence of proteinuria got more obvious protective effects than it did after the occurence of proteinuria.③No significance was found between both 3 mg/kg (a week) tripterine treated and 6 mg/kg (a week) groups.Conclusion Tripterine has a definite protective effect on glomerulosclerosis of the lupus murine model.The decrease of renal collagen type Ⅰ and type Ⅳ is probably due to its suppressive effects on the expression of local TGF ? 1,TIMP 2 and its improvement effect on the local expression of MMP 2.
4.A Study of Apoptosis in Renal Tissue of Diffuse Proliferative Lupus Nephritis
Wen SHEN ; Fuming LU ; Xiurong ZHANG ; Muyi GUO ; Shanyan LIN
Fudan University Journal of Medical Sciences 2001;28(1):35-38
Purpose To evaluate apoptosis in renal tissue of diffuse proliferative lupus nephritis and therelationship between the existence of apoptosis cells in renal tissue and histopathological or clinical changes.Methods Apoptosis was detected by in situ nick-end labeling techniques (TUNEL) in renal biopsies from 25patients with type Ⅳ LN, 12 patients with IgAN, 4 patients with MsPGN, and 3 patients with APSGN. Normalrenal tissue obtained at nephrectorny for hypemephroma in 4 adults was used as control. In addition, proliferatingcells were identified by proliferating cell nuclear antigen(PCNA) in these patients. Results Compared to otherproliferative glomerulonephritis and control,the patients with lupus nephritis had less apoptosis cells, higher ratio ofPCNA+ cells/TdT+ cells/(P/T) in renal tissues;Ratio of P/T in glomeruli and tubulointerstitium correlated withthe chronicity index, r=0. 498 3(P = 0. 013 2), r = 0. 839 9(P< 0.001 ), r = 0. 661 4(P = 0. 003 3),respectively. Ratio of P/T in glomerulus and tubule had positive correlation with 24 hour urinary protein, r =0.855 4(P<0.001),r=0.713 4(P=0. 001); negative correlation with Ccr, r = - 0. 488 0(P =0. 013 3)and r = - 0. 722 9(P = 0. 001), which in tubules positively correlated with Scr, r = 0. 410 7 (P = 0.041 4 ).Conclusions Apoptosis is insufficient in proliferative lupus nephritis. Intense proliferation without followingincrease in apoptosis may be related to chronic progressive renal histopatholcgical changes.
5.Significance of u-PA and PAI-1 Expression on Diseased Glomeruli
Yinfeng HONG ; Zhigang ZHANG ; Xiurong ZHANG ; Muyi GUO
Fudan University Journal of Medical Sciences 2001;28(1):17-20
Purpose To investigate the significance of u-PA and PAI-1 expression in various types ofglomerulonephritis. Methods The expression level of u-PA, PAI-1, type-Ⅳ collagen and PCNA positivenuclei in 120 cases of renal biopsies from patients with various types of glomerulonephritis were detected byimmunohistochemical method. Both the staining intensity of u-PA and PAI-1 in glomeruli were quantitativelyanalyzed by image analysis. Results The expression intensity of u-PA and PAI-1 was different in varoustypes of glomerulonephritis, which were significantly higher than that of the minor lesion group. Meanwhile,the intensity of PAI-1 stain was significantly higher than that of u-PA in various types of glomerulonephritis( P < 0.05). The increase of u-PA expression was closely related to increase of type- Ⅳ collagen synthesis andhypercellularity in glomeruli(r = 0. 761 and 0. 811, P< 0.05), while the expression of PAI-1 was closelyrelated to the increase of Col-Ⅳ synthesis other than the cell proliferation in the glomeruli. ConclusionsThe over expression of u-PA and PAI-1 may play an important role in contributing to the pathogenesis anddevelopment of glomerulonephritis.
6.Constitution of Rat Glomerular Mesangial Cell Stably Expressing Decorin Gene
Huijun WANG ; Zhigang ZHANG ; Wensheng LIN ; Qi CHEN ; Muyi GUO ; Ping GUANG
Fudan University Journal of Medical Sciences 2001;28(2):97-99,102
PurposeTo study the possibility of transferring decorin gene to rat glomerular mesangial cell. Methods Amplification of the rat decorin (DCN) eDNA by RT-PCR for constructing the plasmid pcDNA3.1A-DCN and lipofectin method for transfecting DCN gene into MsC; G418 scanning, Western blot and RT-PCR analysis for detecting DCN protein and mRNA in D-A6 cell clone.Results The recombinant eukaryotic expression plasmid, pcDNA3.1A-DCN was successfully constructed and 2 cell clones positively expressing DCN were selected. ConclusionsThese cell clones positively expressing DCN is valuable for providing favorable experimental bases of gene therapy to model with glomerular diseases.
7.Effect of Heparin on the Cell Proliferation and u-PA/PAI-1 Expression on Glomeruli in Rat Anti-thy1 Glomerulonephritis
Rong QIN ; Nong ZHANG ; Chen LIU ; Guangping CHEN ; Xiurong ZHANG ; Muyi GUO
Fudan University Journal of Medical Sciences 2000;27(6):423-426
Purpose To investigate the significance of u-PA and PAI-1 expression on the glomeruli,and the effect of heparin on their expressions in rat anti-thy1 glomerulonephritis. Methods We analyzed the cell proliferation and the expression of u-PA/PAI-1 on the glomeruli by immunohistochemistry and quantitative analysis of immunostaining. Results The cell proliferation of the glomeruli decreased significantly at 7 th,14 th,21 st day after heparin treatment in comparison to the glomerulonephritic group(P<0.05 or 0.01).The expression of u-PA and PAI-1 on the glomeruli in glomerulonephritic and heparin-treated groups was higher than that in the control group.At 3 rd,7 th,14 th,21 st day,the glomerular hypercellularity in the glomerulonephritic group was closely related to the increased expression of u-PA and PAI-1(P<0.05 or 0.01).At 3 rd,7 th day,the decreased cell proliferation of the glomeruli in heparin-treated group had close relationship with the decreased expression of PAI-1(P<0.05). Conclusions In rat anti-thy 1 glomerulonephritis model,the expression of u-PA and PAI-1 increased with glomerular hypercellularity;heparin treatment can decrease the extent of glomerular hypercellularity in rat anti-thy 1 glomerulonephritis.The treatment function of heparin might be related with the inhibitory effect of PAI-1 expression on the glomeruli.
8.Pathogenesis of renal lesion in Fechtner syndrome
Haiyan YANG ; Zhaoyue WANG ; Guoyuan LU ; Xiaojuan ZHAO ; Zhigang ZHANG ; Muyi GUO ; Xia BAI ; Changgeng RUAN
Chinese Journal of Nephrology 2008;24(5):328-331
Objective To investigate the pathogenesis of renal lesion in Fechtner syndrome . Methods Pathological characteristics of kidney tissues from Fechtner syndrome patients were explored by HE staining, immunochemistry, immunofluorescence and electron microscopy . Results Immunochemistry analysis showed that non-muscle myosin heavy chain IIA (NMMHC-IIA)was expressed in podocytes of giomeruli and distal convoluted tube, and was faintly expressed in the brush border of proximal tube . Histological examination demonstrated glomerulosclerosis and decreased expression of NMMHC-IIA in abnormal podocytes . Through standard immunofluorecence, the expression of NMMHC-IIA in patient's podocyte was higher than that in normal pedocytes . The fusion of foot process and microvillus were detected by electron microscopy . Conclusion Abnormal NMMHC-IIA aggregates in the glomeruli podocyts and foot process fusion accompanied with appearance of microvillus leads to renal lesion in Fechtuer syndrome .
9.The expressions of TGF-beta1 and Smad 2 mRNA on diseased glomeruli and their significance in the development of glomerulosclerosis.
Yonghua RUAN ; Zhigang ZHANG ; Xiurong ZHANG ; Chen LIU ; Muyi GUO
Chinese Journal of Pathology 2002;31(4):314-317
OBJECTIVETo explore the expressions of transforming growth factor-beta1 (TGF-beta1) and its signaling transduction molecule Smad 2 and their significance in the development of glomerulosclerosis.
METHODSUsing in situ hybridization and immunohistochemistry to detect Smad 2 mRNA expression and TGF-beta1, collagen IV, fibronectin expression in renal biopsies from 61 cases with a spectrum of glomerulonephritis including IgA nephropathy (40 cases), membranous glomerulonephritis (10 cases) and sclerosing glomerulonephritis (11 cases), compared with 11 cases of glomerular mild lesion with image analysis system.
RESULTSWith the exception of Smad 2 mRNA expression in mild type IgA nephropathy, all other types of diseased glomeruli showed increased expression of both TGF-beta1 and Smad 2 mRNA when compared with the 11 cases of mild glomerular lesions. The expressions of glomerular TGF-beta1 and Smad 2 mRNA positively correlated with collagen IV and fibronectin deposition in the glomeruli.
CONCLUSIONSTGF-beta1 and Smad 2 may be involved in the excessive deposition of glomerular extracellular matrix and play an important role in the development of glomerulosclerosis.
Collagen Type IV ; analysis ; DNA-Binding Proteins ; genetics ; Fibronectins ; analysis ; Glomerulonephritis ; metabolism ; Humans ; Immunohistochemistry ; Kidney Glomerulus ; chemistry ; RNA, Messenger ; analysis ; Smad2 Protein ; Trans-Activators ; genetics ; Transforming Growth Factor beta ; analysis ; Transforming Growth Factor beta1
10.The localization of adrenomedullin in rat kidney tissue and its inhibitory effect on the growth of cultured rat mesangial cells.
Xueguang LIU ; Zhigang ZHANG ; Xiurong ZHANG ; Hongguang ZHU ; Qi CHEN ; Muyi GUO
Chinese Medical Sciences Journal 2002;17(3):129-133
OBJECTIVETo observe the localization of adrenomedullin (AM) in rat kidney tissue and its inhibitory effect on the growth of cultured rat mesangial cells (MsC).
METHODSA monoclonal antibody against AM developed by our laboratory was used to detect the localization of AM protein in rat kidney tissue by avidin-biotin complex immunohistochemistry. The expressions of AM and its receptor CRLR mRNA on cultured glomerular epithelial cells (GEC) and MsC were investigated by Northern blot assay, and the possible effect of AM secreted by GEC on MsC proliferation was observed using [3H]thymidine incorporation as an index.
RESULTSA specific monoclonal antibody against AM was succesfully developed. AM was immunohistochemically localized mainly in glomeruli (GEC and endothelial cells), some cortical proximal tubules, medullary collecting duct cells, interstitial cells, vascular smooth muscle cells and endothelial cells. Northern blot assay showed that AM mRNA was expressed only on cultured GEC, but not on MsC, however, AM receptor CRLR mRNA was only expressed on MsC. GEC conditioned medium containing AM can inhibit MsC growth and AM receptor blocker CGRP8-37 may partially decreased this inhibitory effect.
CONCLUSIONAM produced by GEC inhibits the proliferation of MsC, which suggests that AM as an important regulator is involved in glomerular normal physiological functions and pathologic processes.
Adrenomedullin ; Animals ; Antibodies, Monoclonal ; analysis ; Calcitonin Receptor-Like Protein ; Cell Division ; Cells, Cultured ; Epithelial Cells ; cytology ; metabolism ; Female ; Glomerular Mesangium ; cytology ; metabolism ; Kidney Glomerulus ; cytology ; Mice ; Mice, Inbred BALB C ; Peptides ; immunology ; metabolism ; physiology ; RNA, Messenger ; biosynthesis ; Receptors, Calcitonin ; biosynthesis ; genetics