Objective:To explore the regulatory mechanism of transient receptor potential melastatin-7(TRPM7)in high glucose-induced renal tubular epithelial cell injury.Methods:The expression of TRPM7 in the serum of diabetic ne...Objective:To explore the regulatory mechanism of transient receptor potential melastatin-7(TRPM7)in high glucose-induced renal tubular epithelial cell injury.Methods:The expression of TRPM7 in the serum of diabetic nephropathy patients and high glucose-induced HK-2 cells was detected by RT-qPCR.Then,the TRPM7 interference vector was constructed,and the downstream high mobility group box 1(HMGB1)/Toll-like receptor 4(TLR4)signaling pathway proteins were detected.Next,in addition to interference with TRPM7 expression,overexpression of HMGB1 in high glucose-induced HK-2 cells was performed.Cell activity,apoptosis,oxidative stress levels,and inflammation levels were determined by CCK8,TUNEL,Western blotting,immunofluorescence and related kits.Results:TRPM7 expression was upregulated in the serum of diabetic nephropathy patients and high glucose-induced HK-2 cells.Interference with TRPM7 reduced cell damage,epithelial-mesenchymal transition,oxidative stress,and inflammatory response in high glucose-induced HK-2 cells via inhibiting the HMGB1/TLR4 signaling pathway.However,the effects induced by TRPM7 silencing were abrogated by HMGB1 overexpression.Conclusions:Decreased TRPM7 alleviates high glucose-induced renal tubular epithelial cell injury by inhibiting the HMGB1/TLR4 signaling pathway.Further animal experiments and clinical trials are warranted to verify its effect.展开更多
In order to investigate the effects of connective tissue growth factor （CTGF） antisense oligodeoxynucleotide （ODN） on plasminogen activator inhibitor-1 （PAI-1） expression in renal tubular cells induced by transf...In order to investigate the effects of connective tissue growth factor （CTGF） antisense oligodeoxynucleotide （ODN） on plasminogen activator inhibitor-1 （PAI-1） expression in renal tubular cells induced by transforming growth factor β1 （TGF-β1） and to explore the role of CTGF in the degradation of renal extracellular matrix （ECM）, a human proximal tubular epithelial cell line （HKC） was cultured in vitro. Cationic lipid-mediated CTGF antisense ODN was transfected into HKC. After HKC were stimulated with TGF-β1 （5 μg/L）, the mRNA level of PAI-1 was detected by RT-PCR. Intracellular PAI-1 protein synthesis was assessed by flow cytometry. The secreted PAI-1 in the media was determined by Western blot. The results showed that TGF-β1 could induce tubular CTGF and PAI-1 mRNA expression. The PAI-1 mRNA expression induced by TGF-β1 was significantly inhibited by CTGF antisense ODN. CTGF antisense ODN also inhibited intracellular PAI-1 protein synthesis and lowered the levels of PAI-1 protein secreted into the media. It was concluded that CTGF might play a crucial role in the degradation of excessive ECM during tubulointerstitial fibrosis, and blocking the biological effect of CTGF may he a novel way in preventing renal fibrosis.展开更多
To elucidate the pathogenesis of acute renal failure (ARF) with nephropathia epidemic (NE), provide experimental evidence for the new therapy to NE and observe the effects of Arg-Gly-Asp (RGD) peptides on adhesion of ...To elucidate the pathogenesis of acute renal failure (ARF) with nephropathia epidemic (NE), provide experimental evidence for the new therapy to NE and observe the effects of Arg-Gly-Asp (RGD) peptides on adhesion of re-nal tubular epithelial cell (RTEC), urine specimens of patients were collected un-der sterile conditions. Detached RTECs were separated, cultured and identified.Hantan Virus antigen was determined by using indirect immunofluorescence method and effects of RGD on adhesion of RTECs was observed by subgroup counting as well as by flow cytometry. This study showed that: (1) sublethal RTECs existed in the urine of NE-ARF patients, which could be cultured in monolayer form; (2 ) there was NE antigen in RTECs; and (3) adhesion of RTECs could be inhibited by RGD.展开更多
Objective Ubiquitin-specific protease 4(USP4)facilitates the development of transforming growth factor-beta 1(TGF-β1)-induced epithelial-mesenchymal transition(EMT)in various cancer cells.Moreover,EMT of renal tubula...Objective Ubiquitin-specific protease 4(USP4)facilitates the development of transforming growth factor-beta 1(TGF-β1)-induced epithelial-mesenchymal transition(EMT)in various cancer cells.Moreover,EMT of renal tubular epithelial cells(RTECs)is required for the progression of renal interstitial fibrosis.However,the role of USP4 in EMT of RTECs remains unknown.The present study aimed to explore the effect of USP4 on the EMT of RTECs as well as the involved mechanism.Methods In established unilateral ureteral obstruction(UUO)rats and NRK-52E cells,immunohistochemistry and Western blot assays were performed.Results USP4 expression was increased significantly with obstruction time.In NRK-52E cells stimulated by TGF-β1,USP4 expression was increased in a time-dependent manner.In addition,USP4 silencing with specific siRNA indicated that USP4 protein was suppressed effectively.Meanwhile,USP4 siRNA treatment restored E-cadherin and weakened alpha smooth muscle actin(α-SMA)expression,indicating that USP4 may promote EMT.After treatment with USP4 siRNA and TGF-β1 for 24 h,the expression of TGF-β1 receptor type I(TβRI)was decreased.Conclusion USP4 promotes the EMT of RTECs through upregulating TβRI,thereby facilitating renal interstitial fibrosis.These findings may provide a potential target of USP4 in the treatment of renal fibrosis.展开更多
To investigate the role of connective tissue growth factor (CTGF) in transdifferentiation of human renal tubular epithelial cell (HKC), in vitro cultured HKC cells were divided into 3 groups: negtive control, low dose...To investigate the role of connective tissue growth factor (CTGF) in transdifferentiation of human renal tubular epithelial cell (HKC), in vitro cultured HKC cells were divided into 3 groups: negtive control, low dose CTGF-treated group (rh CTGF, 2.5 ng/ml) and high dose CTGF-treated (rhCTGF, 5.0 ng/ml). Then the expression of α-smooth muscle actin (α-SMA) were assessed by indirect immuno-fluorescence, and the percentage of α-SMA positive cells were assessed by flow cytometry. RT-PCR were also performed to examine the mRNA level of α-SMA. Upon the stimulation of different concentrations of rhCTGF, the expression of α-SMA were markedly stronger than that in negative controls. The percentages of α-SMA positive cells were significantly higher in the stimulated groups than that of negative controls (38.9 %, 65.5 % vs 2.4 %, P<0.01) .α-SMA mRNA levels were also up-regulated by the stimulation of rhCTGF (P<0.01). These results suggest that CTGF can promote the transdifferentiation of human renal tubular epithelial cells towards myofibroblast (Myo-F).展开更多
Objective To analyze and identify the differentially expressed proteins in human renal tubular epithelial ceils ( HK-2) after injury caused by oxalic acid and calcium oxalate monohydrate ( COM ) crystal,and to explore...Objective To analyze and identify the differentially expressed proteins in human renal tubular epithelial ceils ( HK-2) after injury caused by oxalic acid and calcium oxalate monohydrate ( COM ) crystal,and to explore the potential role of renal tubular cell injury in kidney stone formation. Methods Normal HK-2 cells展开更多
Objective To investigate the effect and mechanism of adrenomedullin ( AM ) on apoptosis of renal tubular epithelial cell in rats induced by renal ischemia reperfusion injury. Methods Thirty-two Wistar rats were random...Objective To investigate the effect and mechanism of adrenomedullin ( AM ) on apoptosis of renal tubular epithelial cell in rats induced by renal ischemia reperfusion injury. Methods Thirty-two Wistar rats were randomly divided into 4 groups: control group,IRI group, empty plasmid group and AM group. One week after re-展开更多
Objective Renal fibrosis is the ultimate pathway of various forms of acute and chronic kidney damage.Notably,the knockout of transient receptor potential channel 6(TRPC6)has shown promise in alleviating renal fibrosis...Objective Renal fibrosis is the ultimate pathway of various forms of acute and chronic kidney damage.Notably,the knockout of transient receptor potential channel 6(TRPC6)has shown promise in alleviating renal fibrosis.However,the regulatory impact of TRPC6 on renal fibrosis remains unclear.Methods In vivo,TRPC6 knockout(TRPC6−/−)mice and age-matched 129 SvEv(WT)mice underwent unilateral renal ischemia-reperfusion(uIR)injury surgery on the left renal pedicle or sham operation.Kidneys and serum were collected on days 7,14,21,and 28 after euthanasia.In vitro,primary tubular epithelial cells(PTECs)were isolated from TRPC6−/−and WT mice,followed by treatment with transforming growth factorβ1(TGFβ1)for 72 h.The anti-fibrotic effect of TRPC6−/−and the underlying mechanisms were assessed through hematoxylin-eosin staining,Masson staining,immunostaining,qRT-PCR,and Western blotting.Results Increased TRPC6 expression was observed in uIR mice and PTECs treated with TGFβ1.TRPC6−/−alleviated renal fibrosis by reducing the expression of fibrotic markers(Col-1,α-SMA,and vimentin),as well as decreasing the apoptosis and inflammation of PTECs during fibrotic progression both in vivo and in vitro.Additionally,we found that the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(AKT)/glycogen synthase kinase 3 beta(GSK3β)signaling pathway,a pivotal player in renal fibrosis,was down-regulated following TRPC6 deletion.Conclusion These results suggest that the ablation of TRPC6 may mitigate renal fibrosis by inhibiting the apoptosis and inflammation of PTECs through down-regulation of the PI3K/AKT/GSK3βpathway.Targeting TRPC6 could be a novel therapeutic strategy for preventing chronic kidney disease.展开更多
文摘Objective:To explore the regulatory mechanism of transient receptor potential melastatin-7(TRPM7)in high glucose-induced renal tubular epithelial cell injury.Methods:The expression of TRPM7 in the serum of diabetic nephropathy patients and high glucose-induced HK-2 cells was detected by RT-qPCR.Then,the TRPM7 interference vector was constructed,and the downstream high mobility group box 1(HMGB1)/Toll-like receptor 4(TLR4)signaling pathway proteins were detected.Next,in addition to interference with TRPM7 expression,overexpression of HMGB1 in high glucose-induced HK-2 cells was performed.Cell activity,apoptosis,oxidative stress levels,and inflammation levels were determined by CCK8,TUNEL,Western blotting,immunofluorescence and related kits.Results:TRPM7 expression was upregulated in the serum of diabetic nephropathy patients and high glucose-induced HK-2 cells.Interference with TRPM7 reduced cell damage,epithelial-mesenchymal transition,oxidative stress,and inflammatory response in high glucose-induced HK-2 cells via inhibiting the HMGB1/TLR4 signaling pathway.However,the effects induced by TRPM7 silencing were abrogated by HMGB1 overexpression.Conclusions:Decreased TRPM7 alleviates high glucose-induced renal tubular epithelial cell injury by inhibiting the HMGB1/TLR4 signaling pathway.Further animal experiments and clinical trials are warranted to verify its effect.
基金This project was supported by a grant from the Science & Technology Foundation of Hubei Province (No. 2003 AA 301C14).
文摘In order to investigate the effects of connective tissue growth factor （CTGF） antisense oligodeoxynucleotide （ODN） on plasminogen activator inhibitor-1 （PAI-1） expression in renal tubular cells induced by transforming growth factor β1 （TGF-β1） and to explore the role of CTGF in the degradation of renal extracellular matrix （ECM）, a human proximal tubular epithelial cell line （HKC） was cultured in vitro. Cationic lipid-mediated CTGF antisense ODN was transfected into HKC. After HKC were stimulated with TGF-β1 （5 μg/L）, the mRNA level of PAI-1 was detected by RT-PCR. Intracellular PAI-1 protein synthesis was assessed by flow cytometry. The secreted PAI-1 in the media was determined by Western blot. The results showed that TGF-β1 could induce tubular CTGF and PAI-1 mRNA expression. The PAI-1 mRNA expression induced by TGF-β1 was significantly inhibited by CTGF antisense ODN. CTGF antisense ODN also inhibited intracellular PAI-1 protein synthesis and lowered the levels of PAI-1 protein secreted into the media. It was concluded that CTGF might play a crucial role in the degradation of excessive ECM during tubulointerstitial fibrosis, and blocking the biological effect of CTGF may he a novel way in preventing renal fibrosis.
文摘To elucidate the pathogenesis of acute renal failure (ARF) with nephropathia epidemic (NE), provide experimental evidence for the new therapy to NE and observe the effects of Arg-Gly-Asp (RGD) peptides on adhesion of re-nal tubular epithelial cell (RTEC), urine specimens of patients were collected un-der sterile conditions. Detached RTECs were separated, cultured and identified.Hantan Virus antigen was determined by using indirect immunofluorescence method and effects of RGD on adhesion of RTECs was observed by subgroup counting as well as by flow cytometry. This study showed that: (1) sublethal RTECs existed in the urine of NE-ARF patients, which could be cultured in monolayer form; (2 ) there was NE antigen in RTECs; and (3) adhesion of RTECs could be inhibited by RGD.
文摘Objective Ubiquitin-specific protease 4(USP4)facilitates the development of transforming growth factor-beta 1(TGF-β1)-induced epithelial-mesenchymal transition(EMT)in various cancer cells.Moreover,EMT of renal tubular epithelial cells(RTECs)is required for the progression of renal interstitial fibrosis.However,the role of USP4 in EMT of RTECs remains unknown.The present study aimed to explore the effect of USP4 on the EMT of RTECs as well as the involved mechanism.Methods In established unilateral ureteral obstruction(UUO)rats and NRK-52E cells,immunohistochemistry and Western blot assays were performed.Results USP4 expression was increased significantly with obstruction time.In NRK-52E cells stimulated by TGF-β1,USP4 expression was increased in a time-dependent manner.In addition,USP4 silencing with specific siRNA indicated that USP4 protein was suppressed effectively.Meanwhile,USP4 siRNA treatment restored E-cadherin and weakened alpha smooth muscle actin(α-SMA)expression,indicating that USP4 may promote EMT.After treatment with USP4 siRNA and TGF-β1 for 24 h,the expression of TGF-β1 receptor type I(TβRI)was decreased.Conclusion USP4 promotes the EMT of RTECs through upregulating TβRI,thereby facilitating renal interstitial fibrosis.These findings may provide a potential target of USP4 in the treatment of renal fibrosis.
基金ThisworkwassupportedbyagrantfromtheScience&TechnologyFoundationofHubeiProvince (No .2 0 0 3AA30 1C14 )
文摘To investigate the role of connective tissue growth factor (CTGF) in transdifferentiation of human renal tubular epithelial cell (HKC), in vitro cultured HKC cells were divided into 3 groups: negtive control, low dose CTGF-treated group (rh CTGF, 2.5 ng/ml) and high dose CTGF-treated (rhCTGF, 5.0 ng/ml). Then the expression of α-smooth muscle actin (α-SMA) were assessed by indirect immuno-fluorescence, and the percentage of α-SMA positive cells were assessed by flow cytometry. RT-PCR were also performed to examine the mRNA level of α-SMA. Upon the stimulation of different concentrations of rhCTGF, the expression of α-SMA were markedly stronger than that in negative controls. The percentages of α-SMA positive cells were significantly higher in the stimulated groups than that of negative controls (38.9 %, 65.5 % vs 2.4 %, P<0.01) .α-SMA mRNA levels were also up-regulated by the stimulation of rhCTGF (P<0.01). These results suggest that CTGF can promote the transdifferentiation of human renal tubular epithelial cells towards myofibroblast (Myo-F).
文摘Objective To analyze and identify the differentially expressed proteins in human renal tubular epithelial ceils ( HK-2) after injury caused by oxalic acid and calcium oxalate monohydrate ( COM ) crystal,and to explore the potential role of renal tubular cell injury in kidney stone formation. Methods Normal HK-2 cells
文摘Objective To investigate the effect and mechanism of adrenomedullin ( AM ) on apoptosis of renal tubular epithelial cell in rats induced by renal ischemia reperfusion injury. Methods Thirty-two Wistar rats were randomly divided into 4 groups: control group,IRI group, empty plasmid group and AM group. One week after re-
基金supported by grants from the National Natural Science Foundation of China(Nos.31171087,30970662,and 82100715)the Henan Medical Science and Technology Research Program(Joint Construction,No.LHGJ20190246).
文摘Objective Renal fibrosis is the ultimate pathway of various forms of acute and chronic kidney damage.Notably,the knockout of transient receptor potential channel 6(TRPC6)has shown promise in alleviating renal fibrosis.However,the regulatory impact of TRPC6 on renal fibrosis remains unclear.Methods In vivo,TRPC6 knockout(TRPC6−/−)mice and age-matched 129 SvEv(WT)mice underwent unilateral renal ischemia-reperfusion(uIR)injury surgery on the left renal pedicle or sham operation.Kidneys and serum were collected on days 7,14,21,and 28 after euthanasia.In vitro,primary tubular epithelial cells(PTECs)were isolated from TRPC6−/−and WT mice,followed by treatment with transforming growth factorβ1(TGFβ1)for 72 h.The anti-fibrotic effect of TRPC6−/−and the underlying mechanisms were assessed through hematoxylin-eosin staining,Masson staining,immunostaining,qRT-PCR,and Western blotting.Results Increased TRPC6 expression was observed in uIR mice and PTECs treated with TGFβ1.TRPC6−/−alleviated renal fibrosis by reducing the expression of fibrotic markers(Col-1,α-SMA,and vimentin),as well as decreasing the apoptosis and inflammation of PTECs during fibrotic progression both in vivo and in vitro.Additionally,we found that the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(AKT)/glycogen synthase kinase 3 beta(GSK3β)signaling pathway,a pivotal player in renal fibrosis,was down-regulated following TRPC6 deletion.Conclusion These results suggest that the ablation of TRPC6 may mitigate renal fibrosis by inhibiting the apoptosis and inflammation of PTECs through down-regulation of the PI3K/AKT/GSK3βpathway.Targeting TRPC6 could be a novel therapeutic strategy for preventing chronic kidney disease.
