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Objective To investigate the effect and mechanism of renal fibrosis after macrophage depletion in C3 - deficient unilateral ureteral obstruction mice. Methods Renal interstitial fibrosis model was established by unilateral ureteral obstruction (UUO) in male C3-deficient mice and age-matched C57BL/6 WT mice (8-12 weeks of age). Mice were randomly divided into 4 groups, including sham operation in wild type group(WT/sham)(n=18), UUO operation in wild type group(WT/UUO)(n=18), sham operation in C3-deficient group(C3KO/sham)(n=18), and UUO operation in C3-deficient group(C3KO/UUO)(n=18). The expression of complement C3 was detected by immunohistochemical staining and renal interstitial macrophages were assessed by immunofluorescence staining. Tubulointerstitial fibrosis was observed by both HE staining and Masson staining after 14 days of UUO. Collagen accumulation and score of tubulointerstitial injury were obtained. Wild type and C3-deficient UUO mice were treated by liposome clodronate in early or late stage respectively and then interstitially infiltrated macrophages and renal fibrosis were analysed. Mice were sacrificed randomly at 3,7,14 days after UUO and obstructed kidneys were collected. Macrophage phenotype was detected by double-labeling immunofluorescence with F4/80 and iNOS for the M1, F4/80 and CD206 for the M2 macrophage subpopulation. iNOS, Arg-1 and CD206 were also detected by western blot. Results C3 deficient mice exhibited attenuated renal fibrosis, reduced collagen accumulation and tubulointerstitial injury score compared with WT mice (P<0.01). Meanwhile, macrophage depletion in early or late stage of UUO reduced renal fibrosis in WT mice, but had no effect on C3-deficient UUO mice. Decreased accumulation of M1 macrophages and expression of iNOS, increased accumulation of M2 macrophages and expression of Arg-1, CD206 were found in C3 deficient mice compared with WT mice in early stage of UUO (P<0.01). Conclusion Renal fibrosis is not reduced after depletion of macrophages in C3 deficient UUO mice due to the altered macrophage polarization.
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Objective@#To investigate the effect and mechanism of renal fibrosis after macrophage depletion in C3-deficient unilateral ureteral obstruction mice.@*Methods@#Renal interstitial fibrosis model was established by unilateral ureteral obstruction (UUO) in male C3-deficient mice and age-matched C57BL/6 WT mice (8-12 weeks of age). Mice were randomly divided into 4 groups, including sham operation in wild type group (WT/sham) (n=18), UUO operation in wild type group (WT/UUO) (n=18), sham operation in C3-deficient group (C3KO/sham) (n=18), and UUO operation in C3-deficient group (C3KO/UUO) (n=18). The expression of complement C3 was detected by immunohistochemical staining and renal interstitial macrophages were assessed by immunofluorescence staining. Tubulointerstitial fibrosis was observed by both HE staining and Masson staining after 14 days of UUO. Collagen accumulation and score of tubulointerstitial injury were obtained. Wild type and C3-deficient UUO mice were treated by liposome clodronate in early or late stage respectively and then interstitially infiltrated macrophages and renal fibrosis were analysed. Mice were sacrificed randomly at 3,7,14 days after UUO and obstructed kidneys were collected. Macrophage phenotype was detected by double-labeling immunofluorescence with F4/80 and iNOS for the M1, F4/80 and CD206 for the M2 macrophage subpopulation. iNOS, Arg-1 and CD206 were also detected by western blot.@*Results@#C3 deficient mice exhibited attenuated renal fibrosis, reduced collagen accumulation and tubulointerstitial injury score compared with WT mice (P<0.01). Meanwhile, macrophage depletion in early or late stage of UUO reduced renal fibrosis in WT mice, but had no effect on C3-deficient UUO mice. Decreased accumulation of M1 macrophages and expression of iNOS, increased accumulation of M2 macrophages and expression of Arg-1, CD206 were found in C3 deficient mice compared with WT mice in early stage of UUO (P<0.01).@*Conclusion@#Renal fibrosis is not reduced after depletion of macrophages in C3 deficient UUO mice due to the altered macrophage polarization.
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Objective@#To analyze the clinicopathological features of IgA nephropathy (IgAN) patients with anemia and the influencing factors of prognosis.@*Methods@#The clinical and pathological data of patients diagnosed with primary IgAN at the First Affiliated Hospital of Fujian Medical University from January 1, 2006 to December 31, 2016 were retrospectively analyzed. The patients were divided into anemia group and non-anemia group according to whether the patient was anemia or not. The clinical and pathological data of the two groups were collected. All of them were followed up from the date of renal biopsy to January 1, 2018. Survival curves of the two groups were drawn by Kaplan-Meier method, and compared by Log-rank test. Multivariate Cox proportional hazards regression model was adopted to explore the influencing factors of prognosis in IgAN patients.@*Results@#A total of 231 subjects were enrolled, including 122 males (52.8%), and the male-female ratio was 1.12∶1. Their age was (34.8±10.1) years (15-68 years). There were 70 patients (30.3%) in anemia group, 161 cases (69.7%) in non-anemic group. Compared with non-anemia group, anemia group had higher proportion of females, lower serum albumin, higher proportion of tubular atrophy/interstitial fibrosis (T1/2), endothelial cell proliferation (E1) and crescent formation (C1/2), which were statistically significant (all P<0.05). The patients had a median follow-up time as 6.3 years (0.3-12.9 years). Survival analysis showed that patients in anemia group had lower cumulative renal survival rate than that in non-anemia group (χ2=15.234, P<0.001). Multivariate Cox hazards regression analysis revealed that anemia (HR=3.820, 95%CI 1.674-8.719, P=0.001), tubular atrophy/interstitial fibrosis (T1/2) (HR=3.770, 95%CI 1.026-13.852, P=0.046), glomerular segmental sclerosis/adhesion (S1) (HR=4.211, 95%CI 1.139-15.576, P=0.031), hypertension (HR=2.988, 95%CI 1.276-6.999, P=0.012), increased 24 h urinary protein (HR=1.103, 95%CI 1.046-1.163, P<0.001) and estimated glomerular filtration (eGFR)<60 ml·min-1·(1.73 m2)-1 (HR=3.725, 95%CI 1.639-8.462, P=0.002) were the independent risk factors for poor renal prognosis in patients with IgAN.@*Conclusions@#The clinicopathological features of IgAN patients with anemia are relatively serious, and the renal cumulative survival rate is lower. Anemia, tubular atrophy/interstitial fibrosis (T1/2), glomerular segmental sclerosis/adhesion (S1), hypertension, increased urinary protein and eGFR<60 ml·min-1·(1.73 m2)-1 are the independent risk factors for poor renal prognosis in patients with IgAN.
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Objective To analyze the clinicopathological features of IgA nephropathy (IgAN) patients with anemia and the influencing factors of prognosis. Methods The clinical and pathological data of patients diagnosed with primary IgAN at the First Affiliated Hospital of Fujian Medical University from January 1, 2006 to December 31, 2016 were retrospectively analyzed. The patients were divided into anemia group and non-anemia group according to whether the patient was anemia or not. The clinical and pathological data of the two groups were collected. All of them were followed up from the date of renal biopsy to January 1, 2018. Survival curves of the two groups were drawn by Kaplan-Meier method, and compared by Log-rank test. Multivariate Cox proportional hazards regression model was adopted to explore the influencing factors of prognosis in IgAN patients. Results A total of 231 subjects were enrolled, including 122 males (52.8％), and the male-female ratio was 1.12:1. Their age was (34.8 ± 10.1) years (15-68 years). There were 70 patients (30.3％) in anemia group, 161 cases (69.7％) in non-anemic group. Compared with non-anemia group, anemia group had higher proportion of females, lower serum albumin, higher proportion of tubular atrophy/interstitial fibrosis (T1/2), endothelial cell proliferation (E1) and crescent formation (C1/2), which were statistically significant (all P<0.05). The patients had a median follow-up time as 6.3 years (0.3-12.9 years). Survival analysis showed that patients in anemia group had lower cumulative renal survival rate than that in non-anemia group (χ2=15.234, P<0.001). Multivariate Cox hazards regression analysis revealed that anemia (HR=3.820, 95％CI 1.674-8.719, P=0.001), tubular atrophy/interstitial fibrosis (T1/2) (HR=3.770, 95％CI 1.026-13.852, P=0.046), glomerular segmental sclerosis/adhesion (S1) (HR=4.211, 95％CI 1.139-15.576, P=0.031), hypertension (HR=2.988, 95％CI 1.276-6.999, P=0.012), increased 24 h urinary protein (HR=1.103, 95％CI 1.046-1.163, P<0.001) and estimated glomerular filtration (eGFR)<60 ml·min-1·(1.73 m2)-1 (HR=3.725, 95％CI 1.639-8.462, P=0.002) were the independent risk factors for poor renal prognosis in patients with IgAN. Conclusions The clinicopathological features of IgAN patients with anemia are relatively serious, and the renal cumulative survival rate is lower. Anemia, tubular atrophy/interstitial fibrosis (T1/2), glomerular segmental sclerosis/adhesion (S1), hypertension, increased urinary protein and eGFR<60 ml·min-1· (1.73 m2)-1 are the independent risk factors for poor renal prognosis in patients with IgAN.
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Objective To investigate the efficacy of leflunomide combined with prednisone in the induction therapy of proliferative lupus nephritis (LN).Methods A prospective,multicenter,randomized controlled clinical trial was conducted in patients with biopsy-proved proliferative lupus nephritis recruited from 15 renal centers from 2013 to 2015.Patients were randomized to two groups.Oral leflunomide or intravenous cyclophosphamide was given to patients in each group.Both groups received a tapering course of oral prednisone therapy.All patients were followed up for 24 weeks.The blood biochemistry,urine index,clinical curative effect and adverse reaction were recorded and analyzed statistically.Results A total of 100 patients were enrolled in this clinical trial,including 48 patients in leflunomide group and 52 patients in cyclophosphamide group.After 24 weeks,the overall response rate was 79％ (95％ CI 67％-90％) in the leflunomide group and 69％ (95％ CI 56％-82％) in the cyclophosphamide group.23％ (95％CI 11％-35％) of patients in leflunomide group showed complete remission compared with 27％ (95％CI 24％-30％) in cyclophosphamide group (P=0.35).The levels of 24-hr urine protein excretion,SLEDAI and anti-dsDNA antibody titers were decreased in patients treated with leflunomide group after 24-weeks treatment.And the levels of serum albumin and complement 3 after treatment were significantly higher compared with these before treatment.There was also no significant difference in changes of 24-hr urine protein excretion,SLEDAI score,anti-dsDNA antibody titers,serum albumin and complement C3 levels after treatment between two groups.Incidence of adverse events did not differ between the leflunomide and cyclophosphamide group.Conclusions Leflunomide combined with prednisone showed same efficacy compared with cyclophosphamide as induction therapy for lupus nephritis.Leflunomide might be an useful medicine in the induction therapy of lupus nephritis.
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Objective To investigate the effect of complement C3a on mouse podocytes phenotype transformation.Methods Purified C3a recombinant protein was used to stimulate mature mouse podocytes.The expression of the mature podocyte markers synaptopodin,podocin,nephrin,CD2-associated protein (CD2AP) and the mesenchymal cell markers fibroblast specific protein 1 (FSP-1),α-smooth muscle actin (α-SMA) were detected by RT-PCR,Western blotting,immunochemistry and immunofluorescence,respectively.Some podocytes were transfected with integrin-linked kinase (ILK) siRNA before the administration of C3a,the expression of nephrin and α-SMA were accessed by Western blotting,and the expression of Snail and α-actinin 4 were accessed by Western blotting and immunochemical method.The migration ability of podocytes was observed by scratch test.Results Immunocytochemistry and immunofluorescence analysis showed that synaptopodin,podocin,nephrin,CD2AP were highly expressed by mature mouse podocytes.The expression of these podocyte markers could be markedly inhibited after 24 h of C3a (0.1 μmol/L) treatment,and accompanied by the induction of mesenchymal markers FSP-1 and α-SMA.Compared with control group,the mRNA levels of synaptopodin,podocin,CD2AP and nephrin were significantly repressed by the administration of C3a in a dose-dependent manner,whereas the transcription of FSP-1 and α-SMA were remarkably up-regulated by C3a treatment (P ＜ 0.05,respectively).Western blotting analysis also confirmed the decrease of synaptopodin,podocin,nephrin and CD2AP protein and the increase of FSP-1 and α-SMA protein were closely depend on the C3a concentration (P ＜ 0.05,respectively).To further assess the downstream of C3a,some podocytes were transfected with ILK siRNA before the administration of C3a.Compared with C3a group,the protein levels of nephrin and α-SMA were significantly changed by the administration of ILK siRNA (P ＜ 0.05,respectively).The expression of α-actinin 4 and Snail induced by C3a were inhibited by ILK knockdown (P ＜ 0.05,respectively),accompanied by a decline of cell migration potency.Conclusion Complement fragment C3a can induce transformation of mouse podocytes to mesenehymal cells,and ILK signaling pathway is involved in this cell type transformation.
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Objective To explore the risk factors of bone density disorder and vascular calcification in non-dialysis chronic kidney disease (CKD) patients.Methods Clinical data of nondialysis CKD patients who were admitted to the First Affiliated Hospital of Fujian Medical University between January 2013 and June 2014 were retrospectively analyzed.Using dual energy X-ray absorptiometry to evaluate their bone mineral density (BMD) and T value.Patients were divided into normal BMD group (T≥-1),osteopenia group (-2.5 ＜ T ＜-1) and osteoporosis group (T≤-2.5).The vascular calcification was evaluated by pectoral computed tomography.Multi-factor stepwise logistic regression analysis was used to assess the risk factors for low bone density and vascular calcification in non-dialysis CKD patients.Results A total of 337 non-dialysis CKD patients were enrolled.There were 110 (32.6％) patients with normal BMD,and 146(43.3％) patients with osteopenia,and 81(24.0％) patients with osteoporosis.Gender,history of hypertension,25-hydroxy vitamin D and N-terminal osteocalcin shown statistical differences among three groups (all P ＜ 0.05).The incidence rate of 25-hydroxy vitamin D deficiency shown statistical difference among three groups (P=0.012).Further,the rates were increased with the decreased bone mass (x2=7.100,P=0.008).The other mineral bone disorders,such as hypocalcemia,hyperphosphatemia,low intact parathyroid hormone (iPTH) and high iPTH had no statistical difference among three groups (all P ＞ 0.05).Multi-factor stepwise logistic regression analysis revealed that increased iPTH (OR=1.938),and low bone density (OR=1.724) were independent risk factors for CKD patients with vascular calcification (all P ＜ 0.05),while women (OR=3.312) and vascular calcification (OR=1.742) were independent risk factors for CKD patients with low bone density (all P ＜ 0.05).Conclusion Increased iPTH and low bone density are independent risk factors for non-dialysis CKD patients with vascular calcification,while women and vascular calcification are independent risk factors for non-dialysis CKD patients with low bone density.
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Objective To follow up the long-term prognosis of acute kidney injury (AKI) patients with normal basic renal function,and to further identify the clinical features as well as risk factors associated with the prognosis of AKI patients.Methods Clinical date of 166 patients who occurred AKI episode during hospitalization from Jan 1 2011 to Dec 31 2014 in The First Affiliated Hospital of Fujian Medical University were retrospectively analyzed.All these patients had normal basic renal function and had follow-up of more than two years after discharge.According to their renal function after two years,patients were divided into recover and non-recover group.The clinical features and risk factors associated with the prognosis of AKI patients were identified using multivariate logistic regression,and the proportion of renal function progression was calculated during follow-up period.Results One hundred and sixty-six patients were enrolled in this observational study,including 114 male,52 female with an average age of 58.1± 16.6.Eighty-seven patients were AKI stage 1,39 AKI stage 2,and 40 AKI stage 3.Thirty-seven patients were caused by pre-renal factors,113 patients by renal causes and 16 patients by post-renal causes.Renal function when discharged (P=0.002,OR=2.980) and infection (P=0.003,OR=2.786) were the risk factors of failing to restore after two years.Eighty-four patients' renal function returned to normal when discharged,but the number of patients whose renal function progressed to CKD 3 stage and even worse 1 year and two years later were 12 (14.3％) and 20 (23.8％) respectively.Fifty-four patients were diagnosed as partial recovery and 28 patients as non-recovery when discharged.One year later 22 (40.7％) and 12 (42.9％) patients' renal function progressed to CKD 3 stage and more,while those numbers became 28 (51.9％) and 16 (57.1％) two years later.Conclusions The risk factors of AKI long-term outcome include unrecovered renal function when discharged and infection.After AKI episode,even with fully recovered renal function,patients are still possible to progress to CKD,highlighting the importance of follow-up observation.
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Objective To investigate the role of the group IB secretory phospholipase A2 (sPLA2-IB) and M-type phospholipase A2 receptor (PLA2R) in human podocyte injury and its possible signal transduction pathway.Methods Differentiated human podocytes were exposed to PBS or different sPLA2-IB concentration conditions (10-9,10-7,10-5 mol/L) for 2 hours.The wound healing assay was used to measure cell migration rate;Apoptosis in cultured human podocytes was assessed by Hoechst 33342 staining and flow cytometry;Western blot was used to analyze the protein expression of p-cPLA2α,p-p38,and p53.Then control siRNA or PLA2R siRNA were transfected to podocytes.Podocytes were divided into normal control group,negative control siRNA group and PLA2R siRNA group.Twenty four hous later,the cells were stimulated by 105 mol/L sPLA2-IB for 2 hours.The protein expression of p-cPLA2α,p-p38,and p53 were detected by Western blot.Results Compared to PBS control group,the migration ability of podocytes decreased when stimulated with sPLA2-IB (10-7mol/L-10-5 mol/L),and the apoptosis of podocytes increased in a concentration-dependent manner,the protein level of p-cPLA2α,p-p38 and p53 protein increased too.After the knockdown of PLA2R by PLA2R siRNA transfection,stimulated the podocytes with the same dosage of sPLA2-IB,the protein expression of p-cPLA2α,p-p38 and p53 all decreased.Conclusion sPLA2-IB stimulation can increase human podocyte apoptosis and decrease its migration ability.The possible mechanism might be through p38-cPLA2α-p53 pathway.
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Objective To investigate the role of the group IB secretory phospholipase A2 (sPLA2-IB) and M-type phospholipase A2 receptor (PLA2R) in human podocyte injury and its possible signal transduction pathway.Methods Differentiated human podocytes were exposed to PBS or different sPLA2-IB concentration conditions (10-9,10-7,10-5 mol/L) for 2 hours.The wound healing assay was used to measure cell migration rate;Apoptosis in cultured human podocytes was assessed by Hoechst 33342 staining and flow cytometry;Western blot was used to analyze the protein expression of p-cPLA2α,p-p38,and p53.Then control siRNA or PLA2R siRNA were transfected to podocytes.Podocytes were divided into normal control group,negative control siRNA group and PLA2R siRNA group.Twenty four hous later,the cells were stimulated by 105 mol/L sPLA2-IB for 2 hours.The protein expression of p-cPLA2α,p-p38,and p53 were detected by Western blot.Results Compared to PBS control group,the migration ability of podocytes decreased when stimulated with sPLA2-IB (10-7mol/L-10-5 mol/L),and the apoptosis of podocytes increased in a concentration-dependent manner,the protein level of p-cPLA2α,p-p38 and p53 protein increased too.After the knockdown of PLA2R by PLA2R siRNA transfection,stimulated the podocytes with the same dosage of sPLA2-IB,the protein expression of p-cPLA2α,p-p38 and p53 all decreased.Conclusion sPLA2-IB stimulation can increase human podocyte apoptosis and decrease its migration ability.The possible mechanism might be through p38-cPLA2α-p53 pathway.
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Objectives To investigate the risk factors of acute renal injury (acute kidney injury) in patients with acute left heart failure.Methods Clinical data of 188 patients with acute left heart failure who were admitted to our hospital were retrospectively analyzed.Logistic regression analysis was used to assess the risk factors for AKI.Results Among 188 patients with acute left heart failure,incidence of acute kidney injury was 33.51％.Univariate and Multivariable logistic regression analyses showed that the independent predictors of acute kidney injury were lower baseline eGFR (OR=4.294,P ＜ 0.001) and anemia (OR=3.573,P=0.006).Conclusions The incidence of acute left heart failure complicated with AKI was high.Basic state of renal function and anemia were the independent risk factors for AKI.
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Objective To observe the effects of bone marrow?derived mesenchymal stem cells (BMSC) on glomerular podocyte injured by lipopolysaccharide (LPS) and the expression of related protein. Methods Podocytes are divided into control group, BMSC group, LPS group and LPS plus BMSC group. After 24 hours of intervention, observing each experimental group podocyte form under inverted phase contrast microscope;detecting the expressions of mRNA and protein of nephrin, CD2AP, synaptopodin, and TRPC6 by RT?PCR and Western?blot. Results Compared with control group, expressions of nephrin, CD2AP, and synaptopodin in LPS group decreased (P<0.05) while that of TRPC6 increased (P<0.05); compared with LPS group, expressions of nephrin, CD2AP, and synaptopodin in LPS+MSC group increased (P<0.05) while that of TRPC6 decreased (P<0.05). Conclusion BMSC may relieve LPS?induced podocyte injury.
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Objective To observe the effect of fasudil on cytoskeleton reconstruction in mouse podocytes induced by angiotensin Ⅱ (Ang Ⅱ),as well as to study the protective mechanism of fasudil in the pathological changes of podocytes.Methods Conditionally immortalized mouse podocytes were treated with Ang Ⅱ (10-7 mol/L).The podocytes were pre-incubated for 30 min or 60 min with various concentrations of fasudil (10-8,10-7,10-5 mol/L),then Ang Ⅱ (10-7 mol/L) were added and furtherincubated for 24 hours.The cytoskeleton distribution of podocyte as indicated by F-actin and synaptopodin was observed by fluorescence microscopy and Western blotting.At the same time,the activity of Rho signal pathway that mediates actin filament polymerization was analyzed by measuring the extent of Rho-associated coiled-coil-containing protein kinase 1 (ROCK-1) and myosin phosphatase-1 (MYPT1).Results Compared to the control group,AngⅡ disrupted the podocyte actin cytoskeleton and significantly decreased the expression of synaptopodin (P ＜ 0.05),while fasudil stabilized the actin filaments,and improved the synaptopodin expression (P ＜ 0.05).The expression enhancement of ROCK-1 and MYPT1 by Ang Ⅱ were reduced significantly by fasudil (P＜0.05).Conclusion The cytoskeleton reconstruction of podocytes can be induced by Ang Ⅱ and inhibited by fasudil,which suggests that the protective effect of fasudil may be partially contributed to the Rho/ROCK signaling pathway.
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Objective To investigate the effects of erythropoietin (EPO) on complement 3a (C3a)-induced renal tubular epithelial to mesenchymal transition. Methods The HK-2 cells were divided into 6 groups namely control group,EPO group,TGF-β group,TGF-β+EPO group,C3a group and EPO+C3a group.The mRNA and protein expressions of α-SMA,E-cadherin and C3 were investigated by RT-PCR,Western blot and immunofluorescence respectively. Results Compared with control group and EPO group,the mRNA and protein expressions of α-SMA in HK-2 cells were up-regulated after the intervention of C3a or TGF-β (all P＜0.05).On the contrast,the mRNA and protein expressions of E-cadherin were down-regulated(P＜0.05),the mRNA and protein expressions of C3 were enhanced (all P＜0.05).However,all those above effects of C3a or TGF-β were inhibited after the intervention of EPO (all P＜0.05). Conclusion EPO is capable of suppressing the epithelial to mesenchymal transition induced by C3a.
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In recent years, multi-walled carbon nanotubes (MWCTs) are very favorable to the adsorption of middle molecular substances in the hemoperfusion because of their multiporous structure, large surface area and high reactivity, which are beneficial to the excellent absorption properties. The purpose of this study was to study the MWCTs on the adsorption capacity of the middle molecular substances. Vitamin B12 (VB12) was selected as a model of the middle molecular substances. The morphologies of MWCTs and activated carbon from commercial "carbon kidney" were observed with scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The adsorption behavior of VB12 was compared to each other with UV-visible absorption spectra. The MWCTs formed a sophistaicate gap structure, and compared to the activated carbon, MWCTs had a larger surface area. By Langmuir equation and Freundlich equation fitting analysis, VB12 adsorption on MWCTs is fit for multi-molecular layer adsorption, and the adsorption type of activated carbon is more inclined to the model corresponding to Langmuir monolayer adsorption. The adsorption rate of MWCTs is faster than that of the activated carbon and the adsorption capacity is greater, which could be expected to become the new adsorbent in the hemoperfusion.
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Adsorption , Charcoal , Chemistry , Nanotubes, Carbon , Chemistry , Porosity , Toxins, Biological , Chemistry , Vitamin B 12 , ChemistryABSTRACT
Objective To investigate the effects of erythropoietin(EPO)on the function of glomerular endothelial cells in rats with chronic renal failure(CRF). Methods The CRF model was established by a two stage 5/6 nephrectomy procedure in rats.Experimental rats were randomly divided into four groups:sham operation group (control group),CRF group,CRF rats treated with 30 U/kg EPO(low-dosage group)and with 50 U/kg EPO (high-dosage group).CRF rats received EPO by hypodermic injection for 6 weeks and then were sacrificed.Serum creatinine(Scr),blood urea nitrogen fBUN),urine protein,haematoglobin (Hb) and blood pressure were measured.The renal morphologie changes were evaluated on periodic acid-schiff (PAS) stained sections.The CD34 and CD31 expressions in glomerulus were detected by immunohistochemistry method.The mRNA of endothelin 1(ET-1),endothelial nitric oxide synthase (eNOS) and vascular endothelial growth factor (VEGF) were detected by RT-PCR. Results The expressions of CD34 and CD31 protein in glomerulus,and the expressions of eNOS and VEGF roRNA in renal tissue were higher in EPO treatment group than those in CRF model group(all P<0,05).The expression of ET-1 mRNA in renal tissue was lower in EPO treatment group than that in CRF model group.In addition,the Scr,BUN,urine protein and blood pressure in EPO treatment group were significantly lower than those in CRF model group (all P<0.05).Haematoglobin in EPO treatment group was higher than that in CRF model group (P<0.05).Reanl pathological injury wss improved by EPO treatment in dose-dependent manner. Conclusion EPO can ameliorate renal pathological injury and renal function in rats with chronic renal failure,maybe through promoting the renovation of glomerular capillary endothelium and improving the function of glomerular endothelial cells.
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Objective To investigate the effects of erythropoietin (EPO) on the number and function of peripheral blood endothelial progenitor cells (EPCs) from rats with chronic renal failure (CRF). Methods The model of chronic renal failure was established by a two-stage 5/6nephrectomy procedure in rats. Experimental rats were randomly divided into four groups (n =7,respectively): sham operation group, CRF group, CRF rats treated with 30 U/kg EPO (low-dosage group) and with 50 U/kg EPO (high-dosage group). CRF rats were given EPO by hypodermic injection for 6 weeks, then EPCs were isolated by density gradient centrifugation from peripheral blood mononuclear cells. The ability of cell proliferation, adhesion and vasculogenesis in vitro was further observed. Results Compared to sham operation group, the ability of cell proliferation,adhesion and vasculogenesis in vitro in CRF rats was remarkably decreased (P＜0.05, respectively).Such ability was promoted significantly in dose-dependent manner by EPO treatment (P＜0.05,respectively). Conclusion EPO can improve the number and ability of endothelial progenitor cells from rats with chronic renal failure.
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Objective To investigate the differentiation of rat bone marrow mesenchymal stem cells (MSCs) to renal tubular epithelial-like cells under different conditions. Methods MSCs were obtained from rat marrow. MSCs were isolated by gradient density centrifugation and plastic adherence and then purified. Surface markers were identified with flow cytometry after amplification in vitro. The purified MSCs of the third passage were cultured respectively as follows: (1) control group: DMEM medium with fetal bovine serum(FBS). (2) all-trans retinoic acid (ATRA) group: DMEM medium with FBS, ATRA and ischemic reperfusion-injured kidney tissue homogenate. (3)combination group: DMEM medium with FBS, ATRA, ischemic reperfusion-injured kidney tissue homogenate, epidermal growth factor (EGF) and bone morphogenetic protein 7 (BMP-7). After 7 days, the MSCs were collected for alkaline phosphatase (AKP) staining, cytokeratin-18 and E-cadherin immunocytochemical analysis. Results The positive rates of the third passage MSCs in CD44, CD90 and CD29 were 97.8%±0.9%, 96.8%±1.4% and 97.6%±2.4%,respectively, but in CD11b/c and CD34 were only 13.2%±0.6% and 1.2%±0.5%. The MSCs in control group were spindle. The MSCs in ATRA group were round and elliptic. The MSCs in combination group became cobblestone-like cells after 7 days. AKP staining showed that tubular epithelial-like cells from MSCs in control group were negative, some above cells in ATRA group were positive and number of above cells increased in combination group. Compared with negative control group, the ratios of cytokeratin-18 positive cells in ATRA group and combination group were respectively increassed by 29.47%±1.08% and 47.52%±2.13% (all P<0.05), the ratios of E-cadherin positive cells in ATRA group and combination group were respectively increased by 14.88%±2.46% and 36.15%±1.13% (all P<0.05). Conclusion MSCs may differentiate by renal tubular epithelial-like cells under the induction of ischemic reperfusion-injured kidney tissue homogenate and ATRA in vitro, which are further differentiated under the combined induction of EGF and BMP-7.
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Aim To investigate the effects of bone morphogenic protein(BMP)-7 on the expression of extracelluar matrix components(ColⅠand FN)in human renal proximal tubular cells(HK-2)induced by transforming growth factor-?1(TGF-?1),and to explore the inhibition of BMP-7 on renal interstitial fibrosis.Methods HK-2 cells were treated with TGF-?1,BMP-7,or a combination of both for 48 h.The expression of FN was assessed by indirect immunofluorescence.RT-PCR and Western blot were used to determine the mRNA and protein expression of FN and ColⅠ?1.Results Indirect immunofluorescence staining showed that FN staining in 3 ?g?L-1 TGF-?1 group was considerably stronger than that in control group.By an addition of 200 ?g?L-1 BMP-7,the expression of FN was significantly inhibited.RT-PCR and Western blot showed the mRNA and protein expression of FN and ColⅠ?1 were up-regulated by TGF-?1 after 48 hours significantly(P
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Aim To investigate the effects of fluvastatin on the cellular proliferation and the expression of connective tissue growth factor(CTGF)and type Ⅳ collagen(Col Ⅳ)in rat mesangial cells(MCs)induced by transforming growth factor-?1(TGF-?1).Methods MCs are divided into the following groups according to different factors:control group,TGF-?1 group,TGF-?1 plus fluvastatin(Flu,different concentrations)groups.The influence of Flu on rat proliferation of MCs was detected by MTT.The expression of CTGF was measured with RT-PCR and Western blot respectively.The secretion of Col Ⅳ protein was quantitated by ELISA.Results 5 ?g?L-1 TGF-?1 could stimulate proliferation of MCs and the expression of CTGF.Col Ⅳ in MCs significantly.Flu could inhibit proliferation of MCs and the expression of CTGF,Col Ⅳ in MCs induced by TGF-?1 in a dose-dependent manner:1 ?mol?L-1 Flu could suppress proliferation of MCs and downregulate the expression of CTGF,Col Ⅳ significantly,while 10 ?mol?L-1 Flu could just suppress the expression of CTGF protein significantly.Simultaneously exogenous CTGF could promote the expression of Col Ⅳ in a dose-dependent manner:2.5 ?g?L-1 CTGF could just upregulate the expression of Col Ⅳ.Conclusion Fluvastatin could inhibit proliferation of MCs and expression of CTGF-mediated Col Ⅳ in MCs.