BACKGROUND:This study aimed to explore the effects of TNF-a on the expression of IP_3R1mRNA and protein in human mesangial cells(HMCs),and to elucidate the mechanism of TNF-a relating to IP_3R1 expression in the occur...BACKGROUND:This study aimed to explore the effects of TNF-a on the expression of IP_3R1mRNA and protein in human mesangial cells(HMCs),and to elucidate the mechanism of TNF-a relating to IP_3R1 expression in the occurrence of hepatorenal syndrome(HRS).METHODS:HMCs were stimulated by tumor(TNF-a) with 100 ng/mL for different hours(2,4,8,and 24 hours).The expression changes of IP_3R1 mRNA and protein were detected by quantitative real-time polymerase chain reaction and immunoblotting.Several inhibitors including D609,U73122,PP1,safingol,rottlerin and non-radioactive protein kinase C(PKC) were used to examine the mechanism of signal transduction ofTNF-a-regulated IP_3R1 in HMCs.RESULTS:The levels of IP_3R1 mRNA at 2 hours after TNF-a exposure were significantly enhanced and peaked at 8 hours in HMCs(P<0.01),then descended at 24 hours(P<0.01).The levels of IP_3R1 protein at 4 hours after TNF-a exposure were obviously increased and peaked at24 hours after TNF-a exposure(P<0.01).Compared to the control group,safingol(PKCa inhibitor)and D609(phosphatidylcholine-specific phospholipase C inhibitor) significantly blocked the TNF-ainduced expression of IP_3R1 mRNA(3.30±0.81 vs.1.95±0.13,P<0.05;2.10±0.49,P<0.01) and IP_3R1protein(3.09±0.13 vs.1.86+0.39,P<0.01;1.98±0.02,P<0.01).TNF-a promoted PKCa activation with maximal PKCa phosphorylation that occurred 8 hours after stimulation measured by non-radioactive PKC assay,and the effect was markedly attenuated by pretreatment with D609 or safingol.CONCLUSION:TNF-a increased the expression of IP_3R1 and this was mediated,at least in part,through the PC-PLC/PKCa signaling pathways in HMCs.展开更多
<Abstrat>The proliferation of mesangial cells on cyclosporin (CsA) test mediumwas studied by MTT assay and TNF-Q in cultured supernatant was examined byusing ELISA. The results showed that cyclosporin A signific...<Abstrat>The proliferation of mesangial cells on cyclosporin (CsA) test mediumwas studied by MTT assay and TNF-Q in cultured supernatant was examined byusing ELISA. The results showed that cyclosporin A significantly inhibited theproliferation of mesangial cells at the concentration between 0. 25 - 15 μg/ml(IC50 1μg/ml). This action appeared to be dose-dependent. Release of TNF-αfrom mesangial cells stimulated by LPS was also dose-dependently suppressed. Itis suggested that cyclosporin A play an important role in antiproliferation mecha-nism of mesangial cells in vitro.展开更多
Diabetic kidney disease (DKD)is a microvascular complication of type 2 diabetes.The study of DKD mechanisms is the most important target for the prevention of DKD.Renal senescence is one of the important pathogeneses ...Diabetic kidney disease (DKD)is a microvascular complication of type 2 diabetes.The study of DKD mechanisms is the most important target for the prevention of DKD.Renal senescence is one of the important pathogeneses for DKD,but the mechanism of renal and cellular senescence is unclear.Decreased expression of circulating miR-126 is associated with the development of DKD and may be a promising blood-based biomarker for DKD.This study is to probe the effect and mechanism of miR-126 on the aging of human glomerular mesangial cells (HGMCs)induced by high glucose.HGMCs were cultured with Roswell Park Memorial Institute (RPMI-1640)in vitro.The effect of high glucose on morphology of HGMCs was observed 72h after intervention.The cell cycle was examined by flow cytometry.The telomere length was measured by Southern blotting.The expression levels of p53,p21 and Rb proteins in p53-p21-Rb signaling pathway and p-statl,p-stat3 in JAK/STAT signaling pathway were detected by Western blotting respectively.The expression of miR-126 was examined by qRT-PCR.MiR-126 mimics was transfected into HGMCs.The effects of miR-126 mimics transfection on cell morphology,cell cycle,telomere length,p53,p21,Rb,p-stat1 and p-stat3 were observed. The results showed that high glucose not only arrested the cell cycle in G1phase but also shortened the telomere length.High glucose led to high expression of p53,p21,Rb,p-statl and p-stat3 and premature senescence of HGMCs by activating the telomere-p53-p21-Rb and JAK/STAT signaling pathways.Moreover,the miR-126 was decreased in HGMCs induced by high glucose.It was suggested that the transfection of miR-126 mimics could inhibit the telomere-p53-p21-Rb and JAK/STAT signaling pathway activity in vitro and delay the senescence of HGMCs.The results may serve as a new strategy for the treatment of DKD.展开更多
Background The pathogenesis of diabetic nephropathy (DN) is a complex pathophysiological process.Its precise mechanism is not fully known. In recent years it has been recognized that synthesis of various extracelluar ...Background The pathogenesis of diabetic nephropathy (DN) is a complex pathophysiological process.Its precise mechanism is not fully known. In recent years it has been recognized that synthesis of various extracelluar matrix (ECM) components may increase, and that degradation of ECM may decrease in DN. It was reported heparin could inhibit mesangial cells proliferation in vitro. The main aim of this study is to explore whether heparin inhibits proliferation of mesangial cells grown in high glucose concentration and to measure the effect of heparin on matrix metalloproteinases (MMPs) expression in mesangial cells. Methods The medium contained either low glucose (5 mmol/L) or high glucose (25 mmol/L). The concentrations of heparin in the culture medium were 0, 25, 50,100, 200 or 400 μg/mL. A metabolic (WST-1) assay was used to measure mesangial cell proliferation and Western blot analysis was used to measure MMPs expression of mesangial cells. Results Normal human mesangial cell (NHMC) proliferation was higher in high glucose (HG) medium than in low glucose (LG) medium. They showed a 1.93 fold expansion after 72 h in high glucose in contrast to a 1.63 fold expansion in low glucose. In the presence of heparin, mesangial cells proliferation was inhibited, which was more obvious at high glucose concentrations than at low glucose concentrations. In high glucose, with heparin concentration of 50, 100, 200 and 400 μg/mL, the mesangial cells showed a 0. 61 fold, 0.52 fold, 0.52 fold and 0.41 fold reductions in cell number compared to cells grown without heparin. In low glucose, only concentrations of 200 μg/mL and 400 μg/mL showed reduction in cell number, namely 0.54 fold and 0.45 fold, when compared to cells grown without heparin. In Western blot analysis,MMP1, MMP2, MMP3 and MMP9 was expressed by mesangial cells expressed in both high and low glucose concentrations, which was more prominent in high glucose medium. Incubation of heparin further increased expression of MMP1, MMP2, MMP3 and MMP9. Conclusions This study suggests that glucose can accelerate mesangial cell proliferation while heparin can reduce proliferation, being more obvious at high glucose concentrations. Higher glucose concentrations led to increased MMP expression, which may take part in the regulation of mesangial matrix synthesis and degradation. Addition of heparin resulted in a corresponding increase in MMP expression, most notably at high glucose concentrations, indicating a potentially renoprotective role in DN.展开更多
Objective:Monocytes/macrophages,proinflammatory cytokines and chemokines are important in the pathogenesis of glomerulonephritis.Interleukin(IL)-13 has been shown to exert potent anti-inflammatory properties. This stu...Objective:Monocytes/macrophages,proinflammatory cytokines and chemokines are important in the pathogenesis of glomerulonephritis.Interleukin(IL)-13 has been shown to exert potent anti-inflammatory properties. This study was designed to investigate the effect of IL-13 on the expression of proinflammatory cytokines,chemokines and profibrogenic cytokines and the involved molecular mechanism in cultured human mesangial cells (HMCs).Methods:The expressions of proinflammatory cytokines,chemokines and profibrogenic cytokines were determined by ribonuclease protection assay(RPA).Activity of nuclear factor-kappa B(NF-κB)and activa- tor protein-1(AP-1)was examined by electrophoretic mobility shift assay(EMSA).NF-κB subunit p65 nuclear transportation and c-Jun N-terminal kinase(JNK)activity were assayed by immunoblot.Results:Recombinant IL-13 inhibited tumor necrosis factor-α(TNF-α),IL-1α,IL-1β,monocyte chemoattractant protein-1(MCP-1), IL-8,and transforming growth factor-β1(TGF-β1)mRNA expressions in a dose-dependent manner.Lipopoly-sacchorides(LPS)dramatically increased NF-κB DNA binding activity of HMCs,which was inhibited by IL-13 in a dose-dependent manner.LPS-activated NF-κB contained p50 and p65 dimers,but not c-Rel subunit.IL-13 blocked LPS-induced NF-κB subunit p65.LPS stimulated JNK/AP-1 activation,which was inhibited by IL-13 in a dose-dependent manner.Conclusion:IL-13 inhibits proinflammatory cytokines,chemokines,and profibrogenic cytokines synthesis by blocking NF-κB and JNK/AP-1 activation.These observations point to the importance of IL-13 in the modulation of inflammatory processes in the renal glomerulus.展开更多
The X gene of HBV encodes a 17-kD protein,termed HBx,which has been shown to function as a transcriptional trans-activator of a variety of viral and cellular promoter/enhancer elements.The aim of this study was to inv...The X gene of HBV encodes a 17-kD protein,termed HBx,which has been shown to function as a transcriptional trans-activator of a variety of viral and cellular promoter/enhancer elements.The aim of this study was to investigate the effect of HBx on gene expression of interleukin(IL)-1β and IL-6,and proliferation of rat mesangial cells in vitro.The X gene of HBV was amplified by PCR assay,and inserted into the eukaryotic expression vector pCI-neo.The structure of recombinant pCI-neo-X plasmid was proved by restrict endonuclease digestion and sequencing analysis.pCI-neo-X was transfected into cultured rat mesangial cell line in vitro via liposome.HBx expression in transfected mesangial cells was detected by Western blot.The IL-1β and IL-6 mRNA expression in those cells was assayed by semiquantitative RT-PCR.Mesangial cell proliferation was tested by MTT.The results showed that HBx was obviously expressed in cultured mesangial cell line at 36th and 48th h after transfection.The expression of IL-1β and IL-6 mRNA was simultaneously increased.The cell proliferation was also obvious at the same time.It was concluded that HBx gene transfection could induce IL-1β and IL-6 gene expression and mesangial cell proliferation.HBx may play a critical role in mesangial cell proliferation through upregulation of the IL-1β and IL-6 gene expression.展开更多
The crude polysaccharide was obtained by means of the decolorization of porphyrized Cordyceps minlitaris stroma with organic solvent, extraction with hot water, precipitation in 80% ethanol, and protein removal with t...The crude polysaccharide was obtained by means of the decolorization of porphyrized Cordyceps minlitaris stroma with organic solvent, extraction with hot water, precipitation in 80% ethanol, and protein removal with the Sevag method. After purification with Sephadex G-75, two of its components, CMP-1 and CMP-2, were obtained. Through the assay of gel chromatography and polarimetry, CMP-1 was identified as pure polysaccharide. The results demonstrated that CMP-1 had favorable oxidation resistance activity, which could scavenge not only oxygen-free radicals in the self-oxidation system of pyrogallic acid, but also the hydroxide-free radicals in the Fenton system. The study focused on the effects of low, medium, and high dosages of CMP-1 in rat blood serum on the proliferation of glomerular mesangial cells in vitro. Through MTT Colorimetric analysis, the activities were compared among the blank control group and the Niaoduqing positive control group CMP-1 and CMP-2. The results shows that CMP-1 was able to inhibit the proliferation of rat glomerular mesangial cells effectively. Therefore, CMP-1, one component of polysaccharides of Cordyceps minlitaris, was certainly a potential remedy for hyperplastic glomerular nephritis, whose antioxidant activity could slow down the process of chronic renal failure(CRF) to some extent.展开更多
Objective: To examine whether lipoxin A4 (LXA4) has an antagonistic effect on IL-1β-induced synthesis of IL-6 in glomerular mesangial cells, and to explore the molecular mechanisms of signal pathway in LXA4 actions. ...Objective: To examine whether lipoxin A4 (LXA4) has an antagonistic effect on IL-1β-induced synthesis of IL-6 in glomerular mesangial cells, and to explore the molecular mechanisms of signal pathway in LXA4 actions. Methods: The glomerular mesangial cells of rat were cultured and treated with IL-1β with or without preincubation with LXA4 at different concentrations. The amount of IL-6 in the supernatant of cells was analyzed by enzymelinked immunosorbent assay(ELISA). The expressions of mRNA of IL-6 were determined by RT-PCR. The expressions of Src homology 2( SH2 ) containing protein-tyrosine phosphatase 2(Shp-2) were assessed by immunoprecipitation and immunoblotting. Activities of DNA-binding of nuclear factor-kappa B(NF-κB) were measured by electrophoretic mobility shift assay(EMSA). Results:IL-1β- snulated secretion of protein and expression of mRNA of IL-6 in mesangial cells were inhibited by LXA4 in a dose-dependent manner. LXA4 antagonizes the phosphorylation of Shp-2 and activities of NF-κB induced by IL-1β Conclusion: LXA4 antagonists IL-1β-induced synthesis of IL-6 in glomerular mesangial cellsthrough the mechanism of Shp-2/NF-κB pathway-dependent signal transduction.展开更多
Moutan Cortex (MC) has been demonstrated to have an inhibitive effect on inflammation and oxidative stress responses in mesangial cells in our previous study. However, little is known about the components of MC contri...Moutan Cortex (MC) has been demonstrated to have an inhibitive effect on inflammation and oxidative stress responses in mesangial cells in our previous study. However, little is known about the components of MC contributing to this benefit. In the present study, cell membrane immobilized chromatography (CMC), a fast and useful method, was presented for screening potential active components of MC. HBZY-1 cells were incubated with MC (200 μg/mL) at the optimal incubation time (90 min). HPLC-DAD analysis and LC/ESI/MS/MS were performed to distinguish the active components and identify its structural ion fragments. The results showed that eight components binding to HBZY-1 cells were mudanoside B, paeoniflorin sulfonate, paeoniflorin, tetragalloyl glucose (isomeride), hexagalloyl glucose, mudanopiside A, and paeonol. In conclusion, our established CMC might be a useful method for screening potential active components in complicated traditional Chinese medicines. These components might be associated with the efficacy of MC on prevention and treatment of diabetic nephropathy.展开更多
The proliferation of glomerular mesangial cells (GMC) and secretion of the extracellular matrix (ECM) in rat with Thy-1 nephritis (Thy-1N) resembling human mesangioproliferative glomerulonephritis have been explored f...The proliferation of glomerular mesangial cells (GMC) and secretion of the extracellular matrix (ECM) in rat with Thy-1 nephritis (Thy-1N) resembling human mesangioproliferative glomerulonephritis have been explored for many years; however, the molecular mechanisms of GMC proliferation and ECM production remain unclear. Our previous studies have demonstrated that the thrombospondin-1 (TSP-1) gene was involved in mediating rat GMC proliferation and ECM synthesis induced by sublytic C5b-9 in vitro. In the present study, the roles of the TSP-1 gene in GMC proliferation, ECM production, and urinary protein secretion in Thy-1N rats were determined by using TSP-1 small hairpin RNA, and the results revealed that silencing of the TSP-1 gene in rat renal tissues could diminish GMC proliferation (P < 0.01) and ECM secretion (P < 0.01) as well as urinary protein secretion (P < 0.05) in Thy-1N rats. Together, the current findings suggested that TSP-1 gene expression was required for GMC proliferation and ECM production in Thy-1N rats.展开更多
The role of serum and glucocorticoid-induced kinase 1 (SGK1) pathway in the connective tissue growth factor (CTGF) expression was investigated in cultured human mesangial cells (HMCs) under high glucose. By using RT-P...The role of serum and glucocorticoid-induced kinase 1 (SGK1) pathway in the connective tissue growth factor (CTGF) expression was investigated in cultured human mesangial cells (HMCs) under high glucose. By using RT-PCR and Western blot, the effect of SGK1 on the CTGF expression in HMCs under high glucose was examined. Overexpression of active SGK1 in HMCs transfected with PIRES2-EGFP- S422D hSGK1 (SD) could increase the expression of phosphorylated SGK1 and CTGF as compared with HMCs groups transfected with PIRES2-EGFP (FP) under high glucose or normal glucose. Overexpression of inactive SGK1 in HMCs transfected with PIRES2-EGFP- K127N hSGK1 (KN) could decrease phosphorylated SGK1 and CTGF expression as compared with HMCs groups transfected with FP under high glucose. In conclusion, these results suggest that high glucose-induced CTGF expression is mediated through the active SGK1 in HMCs.展开更多
Objective To investigate the relationship between the alteration of intracellular calcium concentration and proliferation in cultured glomerular mesangial cells. Methods Rat mesangial cells were cultured. Intracellula...Objective To investigate the relationship between the alteration of intracellular calcium concentration and proliferation in cultured glomerular mesangial cells. Methods Rat mesangial cells were cultured. Intracellular calcium concentrations were measured by confocal Laser Scanning Microscopy and Fura-3 fluorescence dyeing techniques. Cell growth was measured by MTT assay. Results PDGF-BB increased intracellular calcium concentrations in a dose-dependent manner, and at the same time promote the proliferation of mesangial cells. After preincubation with calcium channel blocker nifedipine or angiotensin converting enzyme inhibitor captopril, both the increase of intracellular calcium concentrations and cell proliferations induced by PDGF-BB were inhibited. Tripterigium Wilfordii Glycosides (TMG) significantly inhibited the mesangial cell proliferations, but it had no significant effect on intracellular calcium concentrations. Conclusion There was a positive relationship between the elevation of intracellular calcium concentration and cell proliferation in glomerular mesangial cells, but the increase of intracellular calcium concentrations wasn’t the only way for proliferation.展开更多
In order to investigate the regulatory role of connective tissue growth factor （CTGF） on production of RANTES （regulated on activation, normal T cell expressed and secreted） in rat glomerular mesangial cells, and ...In order to investigate the regulatory role of connective tissue growth factor （CTGF） on production of RANTES （regulated on activation, normal T cell expressed and secreted） in rat glomerular mesangial cells, and the modulatory effect of lipoxin A4（LXA4） on action of CTGF, and to explore the mechanisms of action of CTGF and LXA4, cultured rat mesangial cells were treated with CTGF, with or without preincubation with LXA4. Expression of mRNA was analyzed by RT-PCR. Protein of RANTES in the supernatants was determined by ELISA. Monocyte transmigration was assessed by in vitro chemotaxis assay. Expression of p42/44 mitogen-activated protein kinase （MAPK）, phosphoinositide 3-kinase （PI3-K） and protein kinase B （PKB） was assessed by Western blotting. DNA-binding activity of nuclear factor-κB （NF-κB） was determined by electrophoretic mobility shift assay （EMSA）. To observe whether transfection of LXA4 receptor homologue gene （LRHG） into mesangial cells intensified these modulatory effects of LXA4, mesangial cells were transfected with pcDNA3.1/LRHG vector. The results showed that CTGF enhanced the mRNA expression and protein release of RANTES, and the expression of phospho （P）-p42/44 MAPK, P-PI3-K, P-PKB and NF-κB. P-p42/44 MAPK blockade inhibited the CTGF-induced expression of P-p42/44 MAPK and partially decreased the level of RANTES in supernatants. P-PI3-K blockade downregulated the CTGF-stimulated expression of P-PI3-K, P-PKB and NF-κB, and partially decreased the release of RANTES. NF-κB blockade abrogated the CTGF-activated NF-κB and partially decreased the secretion of RANTES. LXA4 dose-dependently inhibited the CTGF-stimulated above action. Transfection of LRHG into mesangial cells intensified these inhibitory effects of LXA4 on CTGF-induced release of RANTES and expression of the P-p42/44 MAPK. In conclusion, LXA4 inhibits CTGF-induced production of RANTES via PI3-K/PKB/NF-κB and p42/44 MAPK-dependent signal pathway, which is mediated by LRHG in rat mesangial cells.展开更多
Abstract Objective:To investigate whether hPTH1-34 regulate the synthesis of fibronectin(FN) from cul-trued rat mesangial cells and its possible mechanism.Methods:(1) MCs seeded at a density of 1×10^4 per well in...Abstract Objective:To investigate whether hPTH1-34 regulate the synthesis of fibronectin(FN) from cul-trued rat mesangial cells and its possible mechanism.Methods:(1) MCs seeded at a density of 1×10^4 per well in 24-well plates were treated with medium containing various concentrations of hPTH1-34(10^-12mol/l-10^-8mol/l)for 6h,12h,24h and 48h,control cells were treated with vehicle only.The FN levels (in the supernatant)were measured by ELISA assay.(2) MCs were co-cultured with 10ng/l of anti-TGF-βanti-body and various concentrations of hPTH1-34(10^-12mol/l-10^-6mol/l).Forty-eight hours later, FN were tested by ELISA.(4)MCs were co-cultured with 10ng/l of anti-TGF-β antibody and 10^-8 mol/l hPTH1-34 for 6h,12h,24h and 48h and then FN were tested.Results(1)hPTH1-34 stimulated FN synthesis in a dose-and time-dependent way with a peak at 10^-8 mol/l(P<0.01).(2)Anti-TGF-β antibody inhibited the stimu-lation effect of hPTH1-34 on synthesis of FN in cultured rat mesangial cells(P<0.05).Conclusion:hPTH1-34 up-regulates FN synthesis in cultured rat mesangial cells via TGF-β ,suggesting that PTH may play an im-portant role in deteriorating the residual renal function at the early stage of chroic renal disease.展开更多
To evaluate the role of glucose transporter- l (GLUT1) in the glucose uptake of glomerular mesangial cells. Methods. Cultured C57/SJL mouse mesangial cells were used in the study. The expression of GLUT1 mRNA was dete...To evaluate the role of glucose transporter- l (GLUT1) in the glucose uptake of glomerular mesangial cells. Methods. Cultured C57/SJL mouse mesangial cells were used in the study. The expression of GLUT1 mRNA was detected by RT- PCR. The expression of GLUT1 protein was detected by immunofluorescence and flow cytometry. The uptake of glucose and its kinetics were determined by 2- deoxy- [3H]- D- glucose uptake. Results. Both GLUT1 mRNA and protein were found in mouse glomerular mesangial cells. 2- deoxy- D- glucose uptake and kinetics assay showed that this glucose transporter had high affinity for glucose and the glucose uptake specificity was further confirmed by phloretin. Conclusion. Functional GLUT1 did present in mouse mesangial cells cultured in vitro and it might be the predominant transporter mediated the uptake of glucose into mesangial cells.展开更多
Objective:To observe the expression of Long non-coding RNA antisense mitochondrial non-coding RNA-2 (ASncmtRNA-2) in high glucose (HG) treated human renal mesangial cells (HRMCs) and the role of ASncmtRNA-2 in oxidati...Objective:To observe the expression of Long non-coding RNA antisense mitochondrial non-coding RNA-2 (ASncmtRNA-2) in high glucose (HG) treated human renal mesangial cells (HRMCs) and the role of ASncmtRNA-2 in oxidative stress mediated diabetic nephropathy (DN) fibrosis.Methods: The expression levels of ASncmtRNA-2、transforming growth factorβ1 (TGF-β1) and fibronectin (FN) mRNA in cultured HRMCs were measured by qRT-PCR. In addition, relative reactive oxygen species (ROS) levels in HRMCs were detected with the non-fluorescent probe DCFH-DA assays.Results: Compared with 0h, the expression of ASncmtRNA-2 remained unchanged in all groups at 8 h post treatment. However, the level of ASncmtRNA-2 mRNA was increased significantly in HG and HG+NG-nitro-L-Arginine methylester (L-NAME) treated cells compared with low glucose (LG) treated cells from 16h onwards, while the level of ASncmtRNA-2 mRNA in the HG+L-NAME group was decreased compared with the HG group. Moreover, ROS fluorescence was significantly up-regulated in HG-treated cells compared with LG-treated cells, while the ROS fluorescence in HG+L-NAME group was suppressed compared with HG-treated cells. In addition, Levels of ASncmtRNA-2, TGF-β1 and FN mRNA were significantly up-regulated in HG treated cells compared with LG treated cells while Levels of ASncmtRNA-2, TGF-β1 and FN mRNA in HG+L-NAME group were down-regulated compared with HG group. Finally, the expression of ASncmtRNA-2, TGF-β1 and FN mRNA were significantly decreased in HG+ASncmtRNA-2 siRNA group compared with HG group.Conclusion: ASncmtRNA-2 was up-regulated in HG treated cells and may promote glomerular fibrosis in DN via positively regulating the expression of pro-fibrotic factors. These findings may provide novel potential therapeutic treatments for DN.展开更多
Objective: To explore the effect of the Nephritis No. 3 (N-3) recipe on nitric oxide (NO),nitric oxide synthase (NOS) secreted by cultured mesangial cells (MC) and its gene expression of the inducible nitric oxide syn...Objective: To explore the effect of the Nephritis No. 3 (N-3) recipe on nitric oxide (NO),nitric oxide synthase (NOS) secreted by cultured mesangial cells (MC) and its gene expression of the inducible nitric oxide synthase (iNOS). Methods. The drug (nephritis No. 3)-containing serum was prepared with serum pharmacological technique, and then was applied to react on mesangial cells cultured in fetal calf serum (FCS) and cells cultured in FCS plus lipopolysaccharide. To observe the secretion of NO and NOS and the gene expression of iNOS by means of RT-PCR. Renltsz Under the two kinds of culture conditions, the content of NO and NOS in the groups with drug-containing serum were higher than those without drug-containing serum (P<0. 05, P<0. 01), and the expression of iNOS mRNA was up-regula-ted too. Conclusion. The N-3 could significantly promote the secretion of NO and NOS and the mRNA expression of iNOS in rats.展开更多
Objective: To examine whether lipoxin A4 （LXA4） has an inhibitory effect on tumor necrosis factor-α（TNF-α）-induced DNA synthesis of glomerular mesangial cells of rat, and explore the molecular mechanisms of LXA4...Objective: To examine whether lipoxin A4 （LXA4） has an inhibitory effect on tumor necrosis factor-α（TNF-α）-induced DNA synthesis of glomerular mesangial cells of rat, and explore the molecular mechanisms of LXA4 action. Methods: Glomerular mesangial cells of rat were cultured and preincubated with LXA4 at different concentrations, and then treated with TNF-α（10 ng/ml）. DNA synthesis was assessed by the incorporation of [ 3H]-thymidine in mesangial cells. Expression of cyclin E protein was determined by Western blotting analysis. Activities of signal transducers and activators of transcription-3 （STAT3） were analyzed by electrophoretic mobility shift assay （EMSA）. Results: TNF-α-stimulated DNA synthesis of mesangial cells, upregulation of cyclin E protein and STAT3 activities were inhibited by LXA4 in a dose-dependent manner. Conclusion: TNF-α-induced DNA synthesis of mesangial cells can be inhibited by TXA4 probably through the mechanism of Jak1/STAT3 pathway-dependent signal transduction.展开更多
基金supported by a grant from Health Bureauof Jiangxi Province
文摘BACKGROUND:This study aimed to explore the effects of TNF-a on the expression of IP_3R1mRNA and protein in human mesangial cells(HMCs),and to elucidate the mechanism of TNF-a relating to IP_3R1 expression in the occurrence of hepatorenal syndrome(HRS).METHODS:HMCs were stimulated by tumor(TNF-a) with 100 ng/mL for different hours(2,4,8,and 24 hours).The expression changes of IP_3R1 mRNA and protein were detected by quantitative real-time polymerase chain reaction and immunoblotting.Several inhibitors including D609,U73122,PP1,safingol,rottlerin and non-radioactive protein kinase C(PKC) were used to examine the mechanism of signal transduction ofTNF-a-regulated IP_3R1 in HMCs.RESULTS:The levels of IP_3R1 mRNA at 2 hours after TNF-a exposure were significantly enhanced and peaked at 8 hours in HMCs(P<0.01),then descended at 24 hours(P<0.01).The levels of IP_3R1 protein at 4 hours after TNF-a exposure were obviously increased and peaked at24 hours after TNF-a exposure(P<0.01).Compared to the control group,safingol(PKCa inhibitor)and D609(phosphatidylcholine-specific phospholipase C inhibitor) significantly blocked the TNF-ainduced expression of IP_3R1 mRNA(3.30±0.81 vs.1.95±0.13,P<0.05;2.10±0.49,P<0.01) and IP_3R1protein(3.09±0.13 vs.1.86+0.39,P<0.01;1.98±0.02,P<0.01).TNF-a promoted PKCa activation with maximal PKCa phosphorylation that occurred 8 hours after stimulation measured by non-radioactive PKC assay,and the effect was markedly attenuated by pretreatment with D609 or safingol.CONCLUSION:TNF-a increased the expression of IP_3R1 and this was mediated,at least in part,through the PC-PLC/PKCa signaling pathways in HMCs.
文摘<Abstrat>The proliferation of mesangial cells on cyclosporin (CsA) test mediumwas studied by MTT assay and TNF-Q in cultured supernatant was examined byusing ELISA. The results showed that cyclosporin A significantly inhibited theproliferation of mesangial cells at the concentration between 0. 25 - 15 μg/ml(IC50 1μg/ml). This action appeared to be dose-dependent. Release of TNF-αfrom mesangial cells stimulated by LPS was also dose-dependently suppressed. Itis suggested that cyclosporin A play an important role in antiproliferation mecha-nism of mesangial cells in vitro.
基金This project was supported by grants from the Key Science and Technology Development Program of Nanjing City of the People's Republic of China (No. YKK15057 and No.YKK16097)and the National Natural Science Foundation of China (No.81473684).
文摘Diabetic kidney disease (DKD)is a microvascular complication of type 2 diabetes.The study of DKD mechanisms is the most important target for the prevention of DKD.Renal senescence is one of the important pathogeneses for DKD,but the mechanism of renal and cellular senescence is unclear.Decreased expression of circulating miR-126 is associated with the development of DKD and may be a promising blood-based biomarker for DKD.This study is to probe the effect and mechanism of miR-126 on the aging of human glomerular mesangial cells (HGMCs)induced by high glucose.HGMCs were cultured with Roswell Park Memorial Institute (RPMI-1640)in vitro.The effect of high glucose on morphology of HGMCs was observed 72h after intervention.The cell cycle was examined by flow cytometry.The telomere length was measured by Southern blotting.The expression levels of p53,p21 and Rb proteins in p53-p21-Rb signaling pathway and p-statl,p-stat3 in JAK/STAT signaling pathway were detected by Western blotting respectively.The expression of miR-126 was examined by qRT-PCR.MiR-126 mimics was transfected into HGMCs.The effects of miR-126 mimics transfection on cell morphology,cell cycle,telomere length,p53,p21,Rb,p-stat1 and p-stat3 were observed. The results showed that high glucose not only arrested the cell cycle in G1phase but also shortened the telomere length.High glucose led to high expression of p53,p21,Rb,p-statl and p-stat3 and premature senescence of HGMCs by activating the telomere-p53-p21-Rb and JAK/STAT signaling pathways.Moreover,the miR-126 was decreased in HGMCs induced by high glucose.It was suggested that the transfection of miR-126 mimics could inhibit the telomere-p53-p21-Rb and JAK/STAT signaling pathway activity in vitro and delay the senescence of HGMCs.The results may serve as a new strategy for the treatment of DKD.
文摘Background The pathogenesis of diabetic nephropathy (DN) is a complex pathophysiological process.Its precise mechanism is not fully known. In recent years it has been recognized that synthesis of various extracelluar matrix (ECM) components may increase, and that degradation of ECM may decrease in DN. It was reported heparin could inhibit mesangial cells proliferation in vitro. The main aim of this study is to explore whether heparin inhibits proliferation of mesangial cells grown in high glucose concentration and to measure the effect of heparin on matrix metalloproteinases (MMPs) expression in mesangial cells. Methods The medium contained either low glucose (5 mmol/L) or high glucose (25 mmol/L). The concentrations of heparin in the culture medium were 0, 25, 50,100, 200 or 400 μg/mL. A metabolic (WST-1) assay was used to measure mesangial cell proliferation and Western blot analysis was used to measure MMPs expression of mesangial cells. Results Normal human mesangial cell (NHMC) proliferation was higher in high glucose (HG) medium than in low glucose (LG) medium. They showed a 1.93 fold expansion after 72 h in high glucose in contrast to a 1.63 fold expansion in low glucose. In the presence of heparin, mesangial cells proliferation was inhibited, which was more obvious at high glucose concentrations than at low glucose concentrations. In high glucose, with heparin concentration of 50, 100, 200 and 400 μg/mL, the mesangial cells showed a 0. 61 fold, 0.52 fold, 0.52 fold and 0.41 fold reductions in cell number compared to cells grown without heparin. In low glucose, only concentrations of 200 μg/mL and 400 μg/mL showed reduction in cell number, namely 0.54 fold and 0.45 fold, when compared to cells grown without heparin. In Western blot analysis,MMP1, MMP2, MMP3 and MMP9 was expressed by mesangial cells expressed in both high and low glucose concentrations, which was more prominent in high glucose medium. Incubation of heparin further increased expression of MMP1, MMP2, MMP3 and MMP9. Conclusions This study suggests that glucose can accelerate mesangial cell proliferation while heparin can reduce proliferation, being more obvious at high glucose concentrations. Higher glucose concentrations led to increased MMP expression, which may take part in the regulation of mesangial matrix synthesis and degradation. Addition of heparin resulted in a corresponding increase in MMP expression, most notably at high glucose concentrations, indicating a potentially renoprotective role in DN.
基金supported by grants from the National Natural Science Foundation of China(No.30872803 to Aihua Zhang,No.30772365 to Songming Huang)the Jiangsu Key Medical Talent Foundation(No.RC2007015 to Aihua Zhangthe Scientific Research Foundation for the Returned Overseas Chinese Scholars,State Education Ministry of China
文摘Objective:Monocytes/macrophages,proinflammatory cytokines and chemokines are important in the pathogenesis of glomerulonephritis.Interleukin(IL)-13 has been shown to exert potent anti-inflammatory properties. This study was designed to investigate the effect of IL-13 on the expression of proinflammatory cytokines,chemokines and profibrogenic cytokines and the involved molecular mechanism in cultured human mesangial cells (HMCs).Methods:The expressions of proinflammatory cytokines,chemokines and profibrogenic cytokines were determined by ribonuclease protection assay(RPA).Activity of nuclear factor-kappa B(NF-κB)and activa- tor protein-1(AP-1)was examined by electrophoretic mobility shift assay(EMSA).NF-κB subunit p65 nuclear transportation and c-Jun N-terminal kinase(JNK)activity were assayed by immunoblot.Results:Recombinant IL-13 inhibited tumor necrosis factor-α(TNF-α),IL-1α,IL-1β,monocyte chemoattractant protein-1(MCP-1), IL-8,and transforming growth factor-β1(TGF-β1)mRNA expressions in a dose-dependent manner.Lipopoly-sacchorides(LPS)dramatically increased NF-κB DNA binding activity of HMCs,which was inhibited by IL-13 in a dose-dependent manner.LPS-activated NF-κB contained p50 and p65 dimers,but not c-Rel subunit.IL-13 blocked LPS-induced NF-κB subunit p65.LPS stimulated JNK/AP-1 activation,which was inhibited by IL-13 in a dose-dependent manner.Conclusion:IL-13 inhibits proinflammatory cytokines,chemokines,and profibrogenic cytokines synthesis by blocking NF-κB and JNK/AP-1 activation.These observations point to the importance of IL-13 in the modulation of inflammatory processes in the renal glomerulus.
基金a grant from National Natural Science Foundation of China (No. 30772360)
文摘The X gene of HBV encodes a 17-kD protein,termed HBx,which has been shown to function as a transcriptional trans-activator of a variety of viral and cellular promoter/enhancer elements.The aim of this study was to investigate the effect of HBx on gene expression of interleukin(IL)-1β and IL-6,and proliferation of rat mesangial cells in vitro.The X gene of HBV was amplified by PCR assay,and inserted into the eukaryotic expression vector pCI-neo.The structure of recombinant pCI-neo-X plasmid was proved by restrict endonuclease digestion and sequencing analysis.pCI-neo-X was transfected into cultured rat mesangial cell line in vitro via liposome.HBx expression in transfected mesangial cells was detected by Western blot.The IL-1β and IL-6 mRNA expression in those cells was assayed by semiquantitative RT-PCR.Mesangial cell proliferation was tested by MTT.The results showed that HBx was obviously expressed in cultured mesangial cell line at 36th and 48th h after transfection.The expression of IL-1β and IL-6 mRNA was simultaneously increased.The cell proliferation was also obvious at the same time.It was concluded that HBx gene transfection could induce IL-1β and IL-6 gene expression and mesangial cell proliferation.HBx may play a critical role in mesangial cell proliferation through upregulation of the IL-1β and IL-6 gene expression.
基金the Science Technology Development Project of Jilin Province, China(No.20020503-2).
文摘The crude polysaccharide was obtained by means of the decolorization of porphyrized Cordyceps minlitaris stroma with organic solvent, extraction with hot water, precipitation in 80% ethanol, and protein removal with the Sevag method. After purification with Sephadex G-75, two of its components, CMP-1 and CMP-2, were obtained. Through the assay of gel chromatography and polarimetry, CMP-1 was identified as pure polysaccharide. The results demonstrated that CMP-1 had favorable oxidation resistance activity, which could scavenge not only oxygen-free radicals in the self-oxidation system of pyrogallic acid, but also the hydroxide-free radicals in the Fenton system. The study focused on the effects of low, medium, and high dosages of CMP-1 in rat blood serum on the proliferation of glomerular mesangial cells in vitro. Through MTT Colorimetric analysis, the activities were compared among the blank control group and the Niaoduqing positive control group CMP-1 and CMP-2. The results shows that CMP-1 was able to inhibit the proliferation of rat glomerular mesangial cells effectively. Therefore, CMP-1, one component of polysaccharides of Cordyceps minlitaris, was certainly a potential remedy for hyperplastic glomerular nephritis, whose antioxidant activity could slow down the process of chronic renal failure(CRF) to some extent.
文摘Objective: To examine whether lipoxin A4 (LXA4) has an antagonistic effect on IL-1β-induced synthesis of IL-6 in glomerular mesangial cells, and to explore the molecular mechanisms of signal pathway in LXA4 actions. Methods: The glomerular mesangial cells of rat were cultured and treated with IL-1β with or without preincubation with LXA4 at different concentrations. The amount of IL-6 in the supernatant of cells was analyzed by enzymelinked immunosorbent assay(ELISA). The expressions of mRNA of IL-6 were determined by RT-PCR. The expressions of Src homology 2( SH2 ) containing protein-tyrosine phosphatase 2(Shp-2) were assessed by immunoprecipitation and immunoblotting. Activities of DNA-binding of nuclear factor-kappa B(NF-κB) were measured by electrophoretic mobility shift assay(EMSA). Results:IL-1β- snulated secretion of protein and expression of mRNA of IL-6 in mesangial cells were inhibited by LXA4 in a dose-dependent manner. LXA4 antagonizes the phosphorylation of Shp-2 and activities of NF-κB induced by IL-1β Conclusion: LXA4 antagonists IL-1β-induced synthesis of IL-6 in glomerular mesangial cellsthrough the mechanism of Shp-2/NF-κB pathway-dependent signal transduction.
文摘Moutan Cortex (MC) has been demonstrated to have an inhibitive effect on inflammation and oxidative stress responses in mesangial cells in our previous study. However, little is known about the components of MC contributing to this benefit. In the present study, cell membrane immobilized chromatography (CMC), a fast and useful method, was presented for screening potential active components of MC. HBZY-1 cells were incubated with MC (200 μg/mL) at the optimal incubation time (90 min). HPLC-DAD analysis and LC/ESI/MS/MS were performed to distinguish the active components and identify its structural ion fragments. The results showed that eight components binding to HBZY-1 cells were mudanoside B, paeoniflorin sulfonate, paeoniflorin, tetragalloyl glucose (isomeride), hexagalloyl glucose, mudanopiside A, and paeonol. In conclusion, our established CMC might be a useful method for screening potential active components in complicated traditional Chinese medicines. These components might be associated with the efficacy of MC on prevention and treatment of diabetic nephropathy.
基金supported by grants from National Natural Science Foundations of China (No. 31000396, and No.81072402)grants from Natural Science Foundations of Jiangsu Province in China (No. BK2009417, No. 10KJB310006, and No. 09hx43)
文摘The proliferation of glomerular mesangial cells (GMC) and secretion of the extracellular matrix (ECM) in rat with Thy-1 nephritis (Thy-1N) resembling human mesangioproliferative glomerulonephritis have been explored for many years; however, the molecular mechanisms of GMC proliferation and ECM production remain unclear. Our previous studies have demonstrated that the thrombospondin-1 (TSP-1) gene was involved in mediating rat GMC proliferation and ECM synthesis induced by sublytic C5b-9 in vitro. In the present study, the roles of the TSP-1 gene in GMC proliferation, ECM production, and urinary protein secretion in Thy-1N rats were determined by using TSP-1 small hairpin RNA, and the results revealed that silencing of the TSP-1 gene in rat renal tissues could diminish GMC proliferation (P < 0.01) and ECM secretion (P < 0.01) as well as urinary protein secretion (P < 0.05) in Thy-1N rats. Together, the current findings suggested that TSP-1 gene expression was required for GMC proliferation and ECM production in Thy-1N rats.
基金a grant from the National Natural Sciences Foundation of China (No. 30600810)
文摘The role of serum and glucocorticoid-induced kinase 1 (SGK1) pathway in the connective tissue growth factor (CTGF) expression was investigated in cultured human mesangial cells (HMCs) under high glucose. By using RT-PCR and Western blot, the effect of SGK1 on the CTGF expression in HMCs under high glucose was examined. Overexpression of active SGK1 in HMCs transfected with PIRES2-EGFP- S422D hSGK1 (SD) could increase the expression of phosphorylated SGK1 and CTGF as compared with HMCs groups transfected with PIRES2-EGFP (FP) under high glucose or normal glucose. Overexpression of inactive SGK1 in HMCs transfected with PIRES2-EGFP- K127N hSGK1 (KN) could decrease phosphorylated SGK1 and CTGF expression as compared with HMCs groups transfected with FP under high glucose. In conclusion, these results suggest that high glucose-induced CTGF expression is mediated through the active SGK1 in HMCs.
文摘Objective To investigate the relationship between the alteration of intracellular calcium concentration and proliferation in cultured glomerular mesangial cells. Methods Rat mesangial cells were cultured. Intracellular calcium concentrations were measured by confocal Laser Scanning Microscopy and Fura-3 fluorescence dyeing techniques. Cell growth was measured by MTT assay. Results PDGF-BB increased intracellular calcium concentrations in a dose-dependent manner, and at the same time promote the proliferation of mesangial cells. After preincubation with calcium channel blocker nifedipine or angiotensin converting enzyme inhibitor captopril, both the increase of intracellular calcium concentrations and cell proliferations induced by PDGF-BB were inhibited. Tripterigium Wilfordii Glycosides (TMG) significantly inhibited the mesangial cell proliferations, but it had no significant effect on intracellular calcium concentrations. Conclusion There was a positive relationship between the elevation of intracellular calcium concentration and cell proliferation in glomerular mesangial cells, but the increase of intracellular calcium concentrations wasn’t the only way for proliferation.
文摘In order to investigate the regulatory role of connective tissue growth factor （CTGF） on production of RANTES （regulated on activation, normal T cell expressed and secreted） in rat glomerular mesangial cells, and the modulatory effect of lipoxin A4（LXA4） on action of CTGF, and to explore the mechanisms of action of CTGF and LXA4, cultured rat mesangial cells were treated with CTGF, with or without preincubation with LXA4. Expression of mRNA was analyzed by RT-PCR. Protein of RANTES in the supernatants was determined by ELISA. Monocyte transmigration was assessed by in vitro chemotaxis assay. Expression of p42/44 mitogen-activated protein kinase （MAPK）, phosphoinositide 3-kinase （PI3-K） and protein kinase B （PKB） was assessed by Western blotting. DNA-binding activity of nuclear factor-κB （NF-κB） was determined by electrophoretic mobility shift assay （EMSA）. To observe whether transfection of LXA4 receptor homologue gene （LRHG） into mesangial cells intensified these modulatory effects of LXA4, mesangial cells were transfected with pcDNA3.1/LRHG vector. The results showed that CTGF enhanced the mRNA expression and protein release of RANTES, and the expression of phospho （P）-p42/44 MAPK, P-PI3-K, P-PKB and NF-κB. P-p42/44 MAPK blockade inhibited the CTGF-induced expression of P-p42/44 MAPK and partially decreased the level of RANTES in supernatants. P-PI3-K blockade downregulated the CTGF-stimulated expression of P-PI3-K, P-PKB and NF-κB, and partially decreased the release of RANTES. NF-κB blockade abrogated the CTGF-activated NF-κB and partially decreased the secretion of RANTES. LXA4 dose-dependently inhibited the CTGF-stimulated above action. Transfection of LRHG into mesangial cells intensified these inhibitory effects of LXA4 on CTGF-induced release of RANTES and expression of the P-p42/44 MAPK. In conclusion, LXA4 inhibits CTGF-induced production of RANTES via PI3-K/PKB/NF-κB and p42/44 MAPK-dependent signal pathway, which is mediated by LRHG in rat mesangial cells.
文摘Abstract Objective:To investigate whether hPTH1-34 regulate the synthesis of fibronectin(FN) from cul-trued rat mesangial cells and its possible mechanism.Methods:(1) MCs seeded at a density of 1×10^4 per well in 24-well plates were treated with medium containing various concentrations of hPTH1-34(10^-12mol/l-10^-8mol/l)for 6h,12h,24h and 48h,control cells were treated with vehicle only.The FN levels (in the supernatant)were measured by ELISA assay.(2) MCs were co-cultured with 10ng/l of anti-TGF-βanti-body and various concentrations of hPTH1-34(10^-12mol/l-10^-6mol/l).Forty-eight hours later, FN were tested by ELISA.(4)MCs were co-cultured with 10ng/l of anti-TGF-β antibody and 10^-8 mol/l hPTH1-34 for 6h,12h,24h and 48h and then FN were tested.Results(1)hPTH1-34 stimulated FN synthesis in a dose-and time-dependent way with a peak at 10^-8 mol/l(P<0.01).(2)Anti-TGF-β antibody inhibited the stimu-lation effect of hPTH1-34 on synthesis of FN in cultured rat mesangial cells(P<0.05).Conclusion:hPTH1-34 up-regulates FN synthesis in cultured rat mesangial cells via TGF-β ,suggesting that PTH may play an im-portant role in deteriorating the residual renal function at the early stage of chroic renal disease.
基金This work was supported by the National Natural Science Foundation of China (No.39870288)
文摘To evaluate the role of glucose transporter- l (GLUT1) in the glucose uptake of glomerular mesangial cells. Methods. Cultured C57/SJL mouse mesangial cells were used in the study. The expression of GLUT1 mRNA was detected by RT- PCR. The expression of GLUT1 protein was detected by immunofluorescence and flow cytometry. The uptake of glucose and its kinetics were determined by 2- deoxy- [3H]- D- glucose uptake. Results. Both GLUT1 mRNA and protein were found in mouse glomerular mesangial cells. 2- deoxy- D- glucose uptake and kinetics assay showed that this glucose transporter had high affinity for glucose and the glucose uptake specificity was further confirmed by phloretin. Conclusion. Functional GLUT1 did present in mouse mesangial cells cultured in vitro and it might be the predominant transporter mediated the uptake of glucose into mesangial cells.
文摘Objective:To observe the expression of Long non-coding RNA antisense mitochondrial non-coding RNA-2 (ASncmtRNA-2) in high glucose (HG) treated human renal mesangial cells (HRMCs) and the role of ASncmtRNA-2 in oxidative stress mediated diabetic nephropathy (DN) fibrosis.Methods: The expression levels of ASncmtRNA-2、transforming growth factorβ1 (TGF-β1) and fibronectin (FN) mRNA in cultured HRMCs were measured by qRT-PCR. In addition, relative reactive oxygen species (ROS) levels in HRMCs were detected with the non-fluorescent probe DCFH-DA assays.Results: Compared with 0h, the expression of ASncmtRNA-2 remained unchanged in all groups at 8 h post treatment. However, the level of ASncmtRNA-2 mRNA was increased significantly in HG and HG+NG-nitro-L-Arginine methylester (L-NAME) treated cells compared with low glucose (LG) treated cells from 16h onwards, while the level of ASncmtRNA-2 mRNA in the HG+L-NAME group was decreased compared with the HG group. Moreover, ROS fluorescence was significantly up-regulated in HG-treated cells compared with LG-treated cells, while the ROS fluorescence in HG+L-NAME group was suppressed compared with HG-treated cells. In addition, Levels of ASncmtRNA-2, TGF-β1 and FN mRNA were significantly up-regulated in HG treated cells compared with LG treated cells while Levels of ASncmtRNA-2, TGF-β1 and FN mRNA in HG+L-NAME group were down-regulated compared with HG group. Finally, the expression of ASncmtRNA-2, TGF-β1 and FN mRNA were significantly decreased in HG+ASncmtRNA-2 siRNA group compared with HG group.Conclusion: ASncmtRNA-2 was up-regulated in HG treated cells and may promote glomerular fibrosis in DN via positively regulating the expression of pro-fibrotic factors. These findings may provide novel potential therapeutic treatments for DN.
基金The item was supported by Tianjin Scientific Committee (No. 953101311)won the second grade Science and Technology Progress award of Tianjin in 2001
文摘Objective: To explore the effect of the Nephritis No. 3 (N-3) recipe on nitric oxide (NO),nitric oxide synthase (NOS) secreted by cultured mesangial cells (MC) and its gene expression of the inducible nitric oxide synthase (iNOS). Methods. The drug (nephritis No. 3)-containing serum was prepared with serum pharmacological technique, and then was applied to react on mesangial cells cultured in fetal calf serum (FCS) and cells cultured in FCS plus lipopolysaccharide. To observe the secretion of NO and NOS and the gene expression of iNOS by means of RT-PCR. Renltsz Under the two kinds of culture conditions, the content of NO and NOS in the groups with drug-containing serum were higher than those without drug-containing serum (P<0. 05, P<0. 01), and the expression of iNOS mRNA was up-regula-ted too. Conclusion. The N-3 could significantly promote the secretion of NO and NOS and the mRNA expression of iNOS in rats.
文摘Objective: To examine whether lipoxin A4 （LXA4） has an inhibitory effect on tumor necrosis factor-α（TNF-α）-induced DNA synthesis of glomerular mesangial cells of rat, and explore the molecular mechanisms of LXA4 action. Methods: Glomerular mesangial cells of rat were cultured and preincubated with LXA4 at different concentrations, and then treated with TNF-α（10 ng/ml）. DNA synthesis was assessed by the incorporation of [ 3H]-thymidine in mesangial cells. Expression of cyclin E protein was determined by Western blotting analysis. Activities of signal transducers and activators of transcription-3 （STAT3） were analyzed by electrophoretic mobility shift assay （EMSA）. Results: TNF-α-stimulated DNA synthesis of mesangial cells, upregulation of cyclin E protein and STAT3 activities were inhibited by LXA4 in a dose-dependent manner. Conclusion: TNF-α-induced DNA synthesis of mesangial cells can be inhibited by TXA4 probably through the mechanism of Jak1/STAT3 pathway-dependent signal transduction.
