| ObjectiveTo investigate the changes of puromycin on glomerular podocyte apoptosis and Park7 m RNA and protein expression,and the protective mechanism of tacrolimus in podocyte injury.MethodMouse glomerular podocyte were cultured in vitro,and three groups were set up:control group,PAN group and FK506 group,respectively.After 12 h,24h and 48 h treatment,the cell morphology was observed and the apoptosis rate was detected.and Real-Time PCR,Western Blot and immunofluorescence were performed to detect the m RNA and protein expression of Park7.The cells were collected for Real-Time PCR,Western Blot and immunofluorescence to detect the m RNA and protein expression of Park7.Result(1)The cell body area of PAN group at 12 h,24h and 48 h was significantly smaller than that of control group,while the cell body area of FK506 group at that corresponding time point was larger than that of PAN group,and the tightness of intercellular junctions was also better than that of PAN group.(2)The apoptosis rate of foot cells in the control group was at a low value at all times.The apoptosis rate of 12 h and 24 h cells was significantly higher in the PAN group compared with the control group,and the increase in the apoptosis rate of 48 h cells was more significant(P<0.01).In the FK506 group,the apoptosis rate of both 24 h and 48 h pedunculated cells was decreased compared with the PAN group(P<0.01).(3)The expression of Park7 m RNA was relatively low at all time points in the control group.Park7 m RNA expression was significantly higher in the PAN group at 12 h,24h,and 48 h compared with the control group(P<0.01).The expression of Park7 m RNA in the FK506 group decreased at 12 h,24h and 48 h compared with the PAN group(P<0.01).(4)The expression of Park7 protein at each time point of Western blot was relatively low.The expression of Park7 protein was significantly higher in the PAN group at 12 h,24h and48 h compared with the control group,and the expression of Park7 protein in the FK506 group decreased at 12 h and 24 h compared with the PAN group.With the extension of time,at 48 h,the expression of Park7 protein in PAN group was significantly higher than that in control group and FK506 group,and the difference between groups was statistically significant(P<0.01).(5)Immunofluorescence staining showed that Park7 in the control group was uniformly distributed on the surface of the cell membrane and in the cytoplasm in small amounts,and granular distribution was also seen in the perinuclear area.After PAN stimulation,Park7 was significantly increased on the cell membrane and in the cytoplasm compared to the control group,with dense clusters distributed.Park7 was more evenly distributed in the cell membrane after FK506 intervention,coarse granular distribution was visible around the nucleusand,and the distribution was significantly improved compared with the PAN group.ConclusionFK506 effectively inhibited the damaging effect of PAN on podocytes and stabilized the expression of Park7,providing a basis for the treatment of glomerular diseases with FK506.ObjectiveTo analyze clinical data,pathology and genetic testing of Alport syndrome with steroid-resistant nephrotic syndrome onset to raise awareness of AS.MethodsFrom January 2015 to December 2019,15 cases were diagnosed with primary nephrotic syndrome in the Department of Pediatrics,Guangzhou First People’s Hospital,affiliated Guangzhou Medical University,after ineffective treatment,from initial SDNS to SRNS,and finally confirmed genetically AS,combined Analysis of clinical features,pathology,extrarenal manifestations and genetic testing.Results(1)In 15 children with AS,all had microscopic hematuria with varying degrees of edema,massive proteinuria,hypoproteinemia,and hyperlipidemia.There were 3 cases of proteinuria with visual hematuria and 2 cases of significant elevation of urea nitrogen/blood creatinine.(2)13 cases underwent renal biopsy pathology,and the results of renal pathology in children with AS were diverse: 3 cases of moderate mesenteric hyperplastic glomerulonephritis,3 cases of focal segmental glomerulosclerosis(FSGS),3 cases of Ig A nephropathy,2 cases of thin basement membrane nephropathy,1 case of mild glomerulopathy,and 1 case of mild mesenteric hyperplastic glomerulonephritis.(3)15 children with PNS were genetically tested and 13 cases of COL4A5 mutation and one case each of COL4A4 and COL4A3 mutation were found.ConclusionsIf the clinical treatment of SRNS is not effective,family history should be asked actively and AS genetic test should be performed as soon as possible.Early diagnosis can help determine the prognosis of the child,avoid unnecessary medication and achieve precise treatment. |
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