Objective: To explore the effect of L158, 809 (angiatensin Ⅱ receptorMockers, ARBs) and Cilazapril (Angiotensin converting enzyme inhibitor, ACEI) on the expression oftransforming growth factor-β_1 (TGF-β_1) and se...Objective: To explore the effect of L158, 809 (angiatensin Ⅱ receptorMockers, ARBs) and Cilazapril (Angiotensin converting enzyme inhibitor, ACEI) on the expression oftransforming growth factor-β_1 (TGF-β_1) and secretion of fibronectin, laminin and type Ⅳcollagen from the cultured human mesangial cells . Methods: Human mesangial cells were cultured indifferent glucose (5.6 mmol/L and 30 mmol/L) and agents (1, 10, 100 and 500 μmol/L) concentrations. The proliferation of mesangial cells were detected at 24, 48 and 72 h . Then the mesangial cellsare divided into four groups, low glucose (5.6 mmol/L) control group, high glucose (30 mmol/L)control group , L158, 809 (10 μmol/L) group and cilazapril (10 μmol/L) group. Forty- eight hourslater, the expression of TGF-β_1 were detected by RT-PCR. Concentrations of TGF-β_1 ,fibronection, laminin and type Ⅳ collagen in the su-pematants of the, mesangial cells weredetermined by EUSA and radioimmunoassay methods. Results: Compared with low glucose control group,the mesangial cells under high glucose medium show excessive proliferation and more TGF-β_1,fibronectin, laminin and type Ⅳ collagen in the supernatant. The expression of TGF-β_1 mRNA wasalso significantly increased under high glucose. The levels of TGF-β_1 and ECM (extracellularmatrix) proteins in the L158, 809 group and cilazapril group are obviously lower than that of thehigh glucose control group. The expression of TGF-β_1 mRNA was markedly decreased in the L158, 809group and cilazapril group compared with that of high glucose control group . Conclusion: Highglucose stimulated the cultured human mesangial cells to excessively proliferate, express TGF-β_1and secrete ECM proteins, and the high glucose-indeced changes were suppressed by either L158, 809and cilazapril.展开更多
Objective:To investigate the effect of shikonin on the proliferation, apoptosis and extracellular matrix (ECM) of human mesangial cells (MC). Methods: MC was cultured in vitro with different concentrations of glucose ...Objective:To investigate the effect of shikonin on the proliferation, apoptosis and extracellular matrix (ECM) of human mesangial cells (MC). Methods: MC was cultured in vitro with different concentrations of glucose (30, 50, 80 mmol/L). The cell growth was observed by using MTT method and apoptosis by using an aunexin-V-Fluos. Immunohistochemical studies for Laminin (LN), Fibronectin (FN) and type Ⅳ Collagens (Col Ⅳ) were measured. Results: Shikonin inhibited their growth (P<0.05) and apoptosis in the glycated cultured cells. Shikonin 0.05 mmol/L significantly reduced the secretion of LN, FN and Col Ⅳ from MC (P<0.05) cultured in 30, 50 and 80 mmol/L glucose. Conclusion: Shikonin could prevent or treat diabetic nephropathy (DN) and glomerulosclerosis (GS).展开更多
Objective:To examine whether lipoxin A 4(LXA 4) has an inhibitory effect on tumor necrosis factor-α(TNF-α)-induced proliferation of glomerular mesangial cells of rat, and explore the molecular mechanisms of signal...Objective:To examine whether lipoxin A 4(LXA 4) has an inhibitory effect on tumor necrosis factor-α(TNF-α)-induced proliferation of glomerular mesangial cells of rat, and explore the molecular mechanisms of signal pathway in LXA 4 actions. Methods: Glomerular mesangial cells of rat were cultured and treated with TNF-α(10 ng/ml), with or without preincubation with LXA 4 at different concentrations. Cell proliferation was evaluated by monotetrazolium (MTT) colorimetric assay. The expression of cyclin E mRNA was measured by RT-PCR. Phosphorylated Akt1(Thr308) and p27 kip1 were analyzed by Western blotting. Results: TNF-α-stimulated proliferation of mesangial cells was inhibited by LXA 4 in a dose-dependent manner. The marked increments in cyclin E mRNA expression induced by TNF-α during proliferation of mesangial cells were down-regulated by LXA 4. Threonine phosphorylated Akt1 proteins at 308 site stimulated by TNF-α was reduced by LXA 4. TNF-α-induced decrements in expression of p27 kip1 proteins was ameliorated by LXA 4 in a dose-dependent manner. Conclusion: TNF-α-induced proliferation of rat mesangial cells can be inhibited by TXA 4 through the mechanism of Akt 1/p27 kip1 pathway-dependent signal transduction.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 usin...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： 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. ...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-u）, 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
<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
BACKGROUND： This study aimed to explore the effects of TNF-α on the expression of IP3R1 mRNA and protein in human mesangial cells （HMCs）, and to elucidate the mechanism of TNF-α relating to IP3R1 expression in th...BACKGROUND： This study aimed to explore the effects of TNF-α on the expression of IP3R1 mRNA and protein in human mesangial cells （HMCs）, and to elucidate the mechanism of TNF-α relating to IP3R1 expression in the occurrence of hepatorenal syndrome （HRS）.METHODS： HMCs were stimulated by tumor （TNF-α） with 100 ng/mL for different hours （2, 4, 8, and 24 hours）. The expression changes of IP3R1 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 of TNF-α-regulated IP3R1 in HMCs.RESULTS： The levels of IP3R1 mRNA at 2 hours after TNF-α 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 IP3R1 protein at 4 hours after TNF-α exposure were obviously increased and peaked at 24 hours after TNF-α exposure （P〈0.01）. Compared to the control group, safingol （PKCa inhibitor） and D609 （phosphatidylcholine-specific phospholipase C inhibitor） significantly blocked the TNF-α- induced expression of IP3R1 mRNA （3.30±0.81 vs. 1.95±0.13, P〈0.05; 2.10±0.49, P〈0.01） and IP3R1 protein （3.09±0.13 vs. 1.86±0.39, P〈0.01; 1.98±0.02, P〈0.01）. TNF-α 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-α increased the expression of IP3R1 and this was mediated, at least in part, through the PC-PLC/PKCa signaling pathways 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. lntracellula...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. lntracellular 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. Tripteriglum 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 supematants 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-roB （NF-kB） 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-kB. P-p42/44 MAPK blockade inhibited the CTgF-induced expression of P-p42/44 MAPK and partially decreased the level of RANTES in supematants. P- PI3-K blockade downregulated the CTGF-stimulated expression of P-PI3-K, P-PKB and NF-kB, and partially decreased the release of RANTES. NF-kB blockade abrogated the CTGF-activated NF-kB 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 CTGFinduced release of RANTES and expression of the P-p42/44 MAPK. In conclusion, LXA4 inhibits CTGFinduced production of RANTES via PI3-K/PKB/NF-kB and p42/44 MAPK-dependent signal pathway, which is mediated by LRHG in rat mesangial cells.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 investigate whether hPTH1-34 regulate the synthesis of fibronectin (FN) from cultured rat mesangial cells and its possible mechanism. Methods: (1) MCs seeded at a density of 1 × 104 per well in 24-w...Objective: To investigate whether hPTH1-34 regulate the synthesis of fibronectin (FN) from cultured rat mesangial cells and its possible mechanism. Methods: (1) MCs seeded at a density of 1 × 104 per well in 24-well plates were treated with medium containing various concentrations of hPTH1-34(10-12 mol/l -10-8 mol/l) for 6 h, 12 h, 24 h and 48 h, control cells were treated with vehicle only. The FN levels (in the supernatant) were measured by ELISA assay. (2) MCs were co-cultured with 10 ng/l of anti-TGF-β antibody and various concentrations of hPTH1-34(10-12 mol/l - 10-8 mol/l ). Forty-eight hours later, FN were tested by ELISA. (4) MCs were co-cultured with 10 ng/l of anti-TGF-β antibody and 10-8 mol/l hPTH1-34 for 6 h, 12 h, 24 h and 48 h 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 stimulation 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 important role in deteriorating the residual renal function at the early stage of chronic renal disease.展开更多
Objective: To explore whether there is phenotypic modulation of mesangial cells in streptozotocin (STZ) induced diabetic rats and study the effect of Tujian Mixture (TJM) on it. Methods: SD rats were divided into the ...Objective: To explore whether there is phenotypic modulation of mesangial cells in streptozotocin (STZ) induced diabetic rats and study the effect of Tujian Mixture (TJM) on it. Methods: SD rats were divided into the normal control group , the unilateral nephrectomized control group , the STZ induced diabetes mellitus with unilateral nephrectomy model group , the Valsartan treated group (VT group, n=8) and the TJM treated group , rats in the latter two groups were modeled as in the DM group and treated with Valsartan (20 mg/kg·d) and TJM (20g/kg·d) respectively for 12 weeks. The expression of α-smooth muscle actin (α-SMA) and transforming growth factor-β 1 (TGF-β 1) in rats’ glomeruli were observed by immunohistochemistry assay, and the ratio of α-SMA and TGF-β 1 positive area/total glomerule tuft area (SMA/GT and TGF/GT) were analyzed using computer-assisted image analysis software. Results: In the NC and the QC groups, only trace of α-SMA positive staining was found. But there was prominant α-SMA positive staining in glomeruli of the DM group, with SMA/GT and TGF/GT increased significantly , and marked increase of 24 hrs proteinuria excretion ( P<0 01). As compared with the DM group, the three indexes were all significantly lower in the VT and ZY groups , and the lowering of proteinuria was more significant in the ZY group than that in the VT group (P<0 01). Conclusion: The expression of α-SMA in glomeruli in STZ induced diabetic rats with unilateral nephrectomy is pronounced, indicating that phenotypic modulation of mesangial cells involvement in the pathogenesis of diabetic nephropathy. TJM and Valsartan can reduce 24 hrs proteinuria excretion, inhibit the phenotypic modulation of mesangial cells and the expression of TGF-β 1 in glomeruli of diabetic rats, and the effect of TJM is more potent than that of Valsartan in lowering urinary protein excretion..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 observe the localization of adrenomedullin (AM) in rat kidney tissue and its inhibitory effect on the growth of cultured rat mesangial cells (MsC). METHODS: A monoclonal antibody against AM developed by ...OBJECTIVE: To observe the localization of adrenomedullin (AM) in rat kidney tissue and its inhibitory effect on the growth of cultured rat mesangial cells (MsC). METHODS: A 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. RESULTS: A 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. CONCLUSION: AM 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.展开更多
Objective: To examine whether lipoxin A 4 (LXA 4) 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 LXA 4 ...Objective: To examine whether lipoxin A 4 (LXA 4) 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 LXA 4 actions. Methods: The glomerular mesangial cells of rat were cultured and treated with IL-1β, with or without preincubation with LXA 4 at different concentrations. The amount of IL-6 in the supernatant of cells was analyzed by enzyme-linked immunosorbent assay(ELISA). The expressions of mRNA of IL-6 were determined by RT-PCR. The expressions of Src homology 2(SH 2) 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-stimulated secretion of protein and expression of mRNA of IL-6 in mesangial cells were inhibited by LXA 4 in a dose-dependent manner. LXA 4 antagonizes the phosphorylation of Shp-2 and activities of NF-κB induced by IL-1β. Conclusion: LXA 4 antagonists IL-1β-induced synthesis of IL-6 in glomerular mesangial cells through the mechanism of Shp-2/NF-κB pathway-dependent signal transduction.展开更多
The effects of mycophenolic acid （MPA） on high glucose-induced expression of transforming growth factor-β （TGF-β） and connective tissue growth factor （CTGF） in mesangial cells （MC） were investigated. Rat MC ...The effects of mycophenolic acid （MPA） on high glucose-induced expression of transforming growth factor-β （TGF-β） and connective tissue growth factor （CTGF） in mesangial cells （MC） were investigated. Rat MC were cultured in the presence of different concentrations of MPA （1.0 and 10.0 μmol/L） or MPA plus high glucose for 72 h. The expression of TGF-β and CTGF was detected by Western blot. The results showed that high glucose could induce the expression of TGF-β and CTGF in MC, but MPA could inhibit this effects. MPA did not influence the expression of TGF-β and CTGF in normal glucose. It was concluded that MPA might prevent the progression of diabetic nephropathy by inhibiting the expression of TGF-β and CTGF in MC.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 in-ducible nitric oxide sy...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 in-ducible 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. Results: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-regulated 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, upregulafion 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
Diabetic nephropathy(DN)is a common type of end-stage renal disease and glomerular mesangial cells(GMCs)are widely used as a cell model for DN.This study firstly investigated the inhibitory effects of the Apostichopus...Diabetic nephropathy(DN)is a common type of end-stage renal disease and glomerular mesangial cells(GMCs)are widely used as a cell model for DN.This study firstly investigated the inhibitory effects of the Apostichopus japonicus and Acaudina leucoprocta hydrolysates on cellular growth under high-glucose treatment,better inhibitory effect of A.japonicus hydrolysate was observed compared to that of A.leucoprocta hydrolysate.Subsequently,the global transcription profiles obtained via microarray analysis showed that 6070 and 7015 genes were identified in the A.japonicus and A.leucoprocta groups compared with the model group,respectively.Among them,transcriptions of the slc30a4,slc35dl,tppp3,tp53inpl,bcl-2,apafl,alox12b and adrala genes were restored from the levels of the model group to those of the control group,contributed to cell mitosis and proliferation in both treatment groups.In addition,other apoptosis-related genes,such as bcl-6,clu,foxo3 and akt,showed opposite trends between two groups,which might cause the difference in inhibitory effect.We preliminarily proposed that the regulation effects of A.japonicus and A.leucoprocta on the genes involved in cellular mitosis,proliferation and apoptosis,might contribute to their inhibitory activity on GMCs under high-glucose environment.展开更多
Objective To elucidate the renoprotective effect of resveratrol(RSV)on sphingosine kinase 1(SphK1)signaling pathway and expression of its downstream molecules including activator protein 1(AP-1)and transformation grow...Objective To elucidate the renoprotective effect of resveratrol(RSV)on sphingosine kinase 1(SphK1)signaling pathway and expression of its downstream molecules including activator protein 1(AP-1)and transformation growth factor-β1(TGF-β1)in lipopolysaccharide(LPS)-induced glomerular mesangial cells(GMCs).Methods The rat GMCs line(HBZY-1)were cultured and randomly divided into 5 groups,including control,LPS(100 ng/mL),and 5,10,20µmol/L RSV-treated groups.In addition,SphK1 inhibitor(SK-II)was used as positive control.GMCs were pretreated with RSV for 2 h and treated with LPS for another 24 h.GMCs proliferation was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide(MTT)assay.The proteins expression of SphK1,p-c-Jun and TGF-β1 in GMCs were detected by Western blot,and DNA-binding activity of AP-1 was performed by electrophoretic mobility shift assay(EMSA).The binding activity between RSV and SphK1 protein was detected by AutoDock Vina and visualized by Discovery Studio 2016.Results LPS could obviously stimulate GMCs proliferation,elevate SphK1,p-c-Jun and TGF-β1 expression levels and increase the DNA-binding activity of AP-1(P<0.05 or P<0.01),whereas these effects were significantly blocked by RSV pretreatment.It was also suggested that the effect of RSV was similar to SK-II(P>0.05).Moreover,RSV exhibited good binding affinity towards SphK1,with docking scores of−8.1 kcal/moL and formed hydrogen bonds with ASP-178 and LEU-268 in SphK1.Conclusion RSV inhibited LPS-induced GMCs proliferation and TGF-β1 expression,which may be independent of its hypoglycemic effect on preventing the development of mesangial cell fibrosis and closely related to the direct inhibition of SphK1 pathway.展开更多
基金National Science and Technology Ninth 5-year Project of Medicine(96-906-05-0)
文摘Objective: To explore the effect of L158, 809 (angiatensin Ⅱ receptorMockers, ARBs) and Cilazapril (Angiotensin converting enzyme inhibitor, ACEI) on the expression oftransforming growth factor-β_1 (TGF-β_1) and secretion of fibronectin, laminin and type Ⅳcollagen from the cultured human mesangial cells . Methods: Human mesangial cells were cultured indifferent glucose (5.6 mmol/L and 30 mmol/L) and agents (1, 10, 100 and 500 μmol/L) concentrations. The proliferation of mesangial cells were detected at 24, 48 and 72 h . Then the mesangial cellsare divided into four groups, low glucose (5.6 mmol/L) control group, high glucose (30 mmol/L)control group , L158, 809 (10 μmol/L) group and cilazapril (10 μmol/L) group. Forty- eight hourslater, the expression of TGF-β_1 were detected by RT-PCR. Concentrations of TGF-β_1 ,fibronection, laminin and type Ⅳ collagen in the su-pematants of the, mesangial cells weredetermined by EUSA and radioimmunoassay methods. Results: Compared with low glucose control group,the mesangial cells under high glucose medium show excessive proliferation and more TGF-β_1,fibronectin, laminin and type Ⅳ collagen in the supernatant. The expression of TGF-β_1 mRNA wasalso significantly increased under high glucose. The levels of TGF-β_1 and ECM (extracellularmatrix) proteins in the L158, 809 group and cilazapril group are obviously lower than that of thehigh glucose control group. The expression of TGF-β_1 mRNA was markedly decreased in the L158, 809group and cilazapril group compared with that of high glucose control group . Conclusion: Highglucose stimulated the cultured human mesangial cells to excessively proliferate, express TGF-β_1and secrete ECM proteins, and the high glucose-indeced changes were suppressed by either L158, 809and cilazapril.
文摘Objective:To investigate the effect of shikonin on the proliferation, apoptosis and extracellular matrix (ECM) of human mesangial cells (MC). Methods: MC was cultured in vitro with different concentrations of glucose (30, 50, 80 mmol/L). The cell growth was observed by using MTT method and apoptosis by using an aunexin-V-Fluos. Immunohistochemical studies for Laminin (LN), Fibronectin (FN) and type Ⅳ Collagens (Col Ⅳ) were measured. Results: Shikonin inhibited their growth (P<0.05) and apoptosis in the glycated cultured cells. Shikonin 0.05 mmol/L significantly reduced the secretion of LN, FN and Col Ⅳ from MC (P<0.05) cultured in 30, 50 and 80 mmol/L glucose. Conclusion: Shikonin could prevent or treat diabetic nephropathy (DN) and glomerulosclerosis (GS).
文摘Objective:To examine whether lipoxin A 4(LXA 4) has an inhibitory effect on tumor necrosis factor-α(TNF-α)-induced proliferation of glomerular mesangial cells of rat, and explore the molecular mechanisms of signal pathway in LXA 4 actions. Methods: Glomerular mesangial cells of rat were cultured and treated with TNF-α(10 ng/ml), with or without preincubation with LXA 4 at different concentrations. Cell proliferation was evaluated by monotetrazolium (MTT) colorimetric assay. The expression of cyclin E mRNA was measured by RT-PCR. Phosphorylated Akt1(Thr308) and p27 kip1 were analyzed by Western blotting. Results: TNF-α-stimulated proliferation of mesangial cells was inhibited by LXA 4 in a dose-dependent manner. The marked increments in cyclin E mRNA expression induced by TNF-α during proliferation of mesangial cells were down-regulated by LXA 4. Threonine phosphorylated Akt1 proteins at 308 site stimulated by TNF-α was reduced by LXA 4. TNF-α-induced decrements in expression of p27 kip1 proteins was ameliorated by LXA 4 in a dose-dependent manner. Conclusion: TNF-α-induced proliferation of rat mesangial cells can be inhibited by TXA 4 through the mechanism of Akt 1/p27 kip1 pathway-dependent signal transduction.
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.
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-u）, 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.
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supported by a grant from Health Bureauof Jiangxi Province
文摘BACKGROUND： This study aimed to explore the effects of TNF-α on the expression of IP3R1 mRNA and protein in human mesangial cells （HMCs）, and to elucidate the mechanism of TNF-α relating to IP3R1 expression in the occurrence of hepatorenal syndrome （HRS）.METHODS： HMCs were stimulated by tumor （TNF-α） with 100 ng/mL for different hours （2, 4, 8, and 24 hours）. The expression changes of IP3R1 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 of TNF-α-regulated IP3R1 in HMCs.RESULTS： The levels of IP3R1 mRNA at 2 hours after TNF-α 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 IP3R1 protein at 4 hours after TNF-α exposure were obviously increased and peaked at 24 hours after TNF-α exposure （P〈0.01）. Compared to the control group, safingol （PKCa inhibitor） and D609 （phosphatidylcholine-specific phospholipase C inhibitor） significantly blocked the TNF-α- induced expression of IP3R1 mRNA （3.30±0.81 vs. 1.95±0.13, P〈0.05; 2.10±0.49, P〈0.01） and IP3R1 protein （3.09±0.13 vs. 1.86±0.39, P〈0.01; 1.98±0.02, P〈0.01）. TNF-α 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-α increased the expression of IP3R1 and this was mediated, at least in part, through the PC-PLC/PKCa signaling pathways 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. lntracellular 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. Tripteriglum 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 supematants 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-roB （NF-kB） 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-kB. P-p42/44 MAPK blockade inhibited the CTgF-induced expression of P-p42/44 MAPK and partially decreased the level of RANTES in supematants. P- PI3-K blockade downregulated the CTGF-stimulated expression of P-PI3-K, P-PKB and NF-kB, and partially decreased the release of RANTES. NF-kB blockade abrogated the CTGF-activated NF-kB 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 CTGFinduced release of RANTES and expression of the P-p42/44 MAPK. In conclusion, LXA4 inhibits CTGFinduced production of RANTES via PI3-K/PKB/NF-kB and p42/44 MAPK-dependent signal pathway, which is mediated by LRHG in rat mesangial cells.
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 investigate whether hPTH1-34 regulate the synthesis of fibronectin (FN) from cultured rat mesangial cells and its possible mechanism. Methods: (1) MCs seeded at a density of 1 × 104 per well in 24-well plates were treated with medium containing various concentrations of hPTH1-34(10-12 mol/l -10-8 mol/l) for 6 h, 12 h, 24 h and 48 h, control cells were treated with vehicle only. The FN levels (in the supernatant) were measured by ELISA assay. (2) MCs were co-cultured with 10 ng/l of anti-TGF-β antibody and various concentrations of hPTH1-34(10-12 mol/l - 10-8 mol/l ). Forty-eight hours later, FN were tested by ELISA. (4) MCs were co-cultured with 10 ng/l of anti-TGF-β antibody and 10-8 mol/l hPTH1-34 for 6 h, 12 h, 24 h and 48 h 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 stimulation 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 important role in deteriorating the residual renal function at the early stage of chronic renal disease.
文摘Objective: To explore whether there is phenotypic modulation of mesangial cells in streptozotocin (STZ) induced diabetic rats and study the effect of Tujian Mixture (TJM) on it. Methods: SD rats were divided into the normal control group , the unilateral nephrectomized control group , the STZ induced diabetes mellitus with unilateral nephrectomy model group , the Valsartan treated group (VT group, n=8) and the TJM treated group , rats in the latter two groups were modeled as in the DM group and treated with Valsartan (20 mg/kg·d) and TJM (20g/kg·d) respectively for 12 weeks. The expression of α-smooth muscle actin (α-SMA) and transforming growth factor-β 1 (TGF-β 1) in rats’ glomeruli were observed by immunohistochemistry assay, and the ratio of α-SMA and TGF-β 1 positive area/total glomerule tuft area (SMA/GT and TGF/GT) were analyzed using computer-assisted image analysis software. Results: In the NC and the QC groups, only trace of α-SMA positive staining was found. But there was prominant α-SMA positive staining in glomeruli of the DM group, with SMA/GT and TGF/GT increased significantly , and marked increase of 24 hrs proteinuria excretion ( P<0 01). As compared with the DM group, the three indexes were all significantly lower in the VT and ZY groups , and the lowering of proteinuria was more significant in the ZY group than that in the VT group (P<0 01). Conclusion: The expression of α-SMA in glomeruli in STZ induced diabetic rats with unilateral nephrectomy is pronounced, indicating that phenotypic modulation of mesangial cells involvement in the pathogenesis of diabetic nephropathy. TJM and Valsartan can reduce 24 hrs proteinuria excretion, inhibit the phenotypic modulation of mesangial cells and the expression of TGF-β 1 in glomeruli of diabetic rats, and the effect of TJM is more potent than that of Valsartan in lowering urinary protein excretion..
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 observe the localization of adrenomedullin (AM) in rat kidney tissue and its inhibitory effect on the growth of cultured rat mesangial cells (MsC). METHODS: A 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. RESULTS: A 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. CONCLUSION: AM 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.
文摘Objective: To examine whether lipoxin A 4 (LXA 4) 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 LXA 4 actions. Methods: The glomerular mesangial cells of rat were cultured and treated with IL-1β, with or without preincubation with LXA 4 at different concentrations. The amount of IL-6 in the supernatant of cells was analyzed by enzyme-linked immunosorbent assay(ELISA). The expressions of mRNA of IL-6 were determined by RT-PCR. The expressions of Src homology 2(SH 2) 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-stimulated secretion of protein and expression of mRNA of IL-6 in mesangial cells were inhibited by LXA 4 in a dose-dependent manner. LXA 4 antagonizes the phosphorylation of Shp-2 and activities of NF-κB induced by IL-1β. Conclusion: LXA 4 antagonists IL-1β-induced synthesis of IL-6 in glomerular mesangial cells through the mechanism of Shp-2/NF-κB pathway-dependent signal transduction.
文摘The effects of mycophenolic acid （MPA） on high glucose-induced expression of transforming growth factor-β （TGF-β） and connective tissue growth factor （CTGF） in mesangial cells （MC） were investigated. Rat MC were cultured in the presence of different concentrations of MPA （1.0 and 10.0 μmol/L） or MPA plus high glucose for 72 h. The expression of TGF-β and CTGF was detected by Western blot. The results showed that high glucose could induce the expression of TGF-β and CTGF in MC, but MPA could inhibit this effects. MPA did not influence the expression of TGF-β and CTGF in normal glucose. It was concluded that MPA might prevent the progression of diabetic nephropathy by inhibiting the expression of TGF-β and CTGF in MC.
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 in-ducible 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. Results: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-regulated 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, upregulafion 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.
基金sponsored by Regional Demonstration Project of Marine Economic Innovation and Development in 2014 and 2016,the National Key Research and Development Program of China(2018YFD0901102)K.C.Wong Magna Fund of Ningbo University.
文摘Diabetic nephropathy(DN)is a common type of end-stage renal disease and glomerular mesangial cells(GMCs)are widely used as a cell model for DN.This study firstly investigated the inhibitory effects of the Apostichopus japonicus and Acaudina leucoprocta hydrolysates on cellular growth under high-glucose treatment,better inhibitory effect of A.japonicus hydrolysate was observed compared to that of A.leucoprocta hydrolysate.Subsequently,the global transcription profiles obtained via microarray analysis showed that 6070 and 7015 genes were identified in the A.japonicus and A.leucoprocta groups compared with the model group,respectively.Among them,transcriptions of the slc30a4,slc35dl,tppp3,tp53inpl,bcl-2,apafl,alox12b and adrala genes were restored from the levels of the model group to those of the control group,contributed to cell mitosis and proliferation in both treatment groups.In addition,other apoptosis-related genes,such as bcl-6,clu,foxo3 and akt,showed opposite trends between two groups,which might cause the difference in inhibitory effect.We preliminarily proposed that the regulation effects of A.japonicus and A.leucoprocta on the genes involved in cellular mitosis,proliferation and apoptosis,might contribute to their inhibitory activity on GMCs under high-glucose environment.
基金Supported by the National Natural Science Foundation of China(No.81603355,81900745)。
文摘Objective To elucidate the renoprotective effect of resveratrol(RSV)on sphingosine kinase 1(SphK1)signaling pathway and expression of its downstream molecules including activator protein 1(AP-1)and transformation growth factor-β1(TGF-β1)in lipopolysaccharide(LPS)-induced glomerular mesangial cells(GMCs).Methods The rat GMCs line(HBZY-1)were cultured and randomly divided into 5 groups,including control,LPS(100 ng/mL),and 5,10,20µmol/L RSV-treated groups.In addition,SphK1 inhibitor(SK-II)was used as positive control.GMCs were pretreated with RSV for 2 h and treated with LPS for another 24 h.GMCs proliferation was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide(MTT)assay.The proteins expression of SphK1,p-c-Jun and TGF-β1 in GMCs were detected by Western blot,and DNA-binding activity of AP-1 was performed by electrophoretic mobility shift assay(EMSA).The binding activity between RSV and SphK1 protein was detected by AutoDock Vina and visualized by Discovery Studio 2016.Results LPS could obviously stimulate GMCs proliferation,elevate SphK1,p-c-Jun and TGF-β1 expression levels and increase the DNA-binding activity of AP-1(P<0.05 or P<0.01),whereas these effects were significantly blocked by RSV pretreatment.It was also suggested that the effect of RSV was similar to SK-II(P>0.05).Moreover,RSV exhibited good binding affinity towards SphK1,with docking scores of−8.1 kcal/moL and formed hydrogen bonds with ASP-178 and LEU-268 in SphK1.Conclusion RSV inhibited LPS-induced GMCs proliferation and TGF-β1 expression,which may be independent of its hypoglycemic effect on preventing the development of mesangial cell fibrosis and closely related to the direct inhibition of SphK1 pathway.
