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Objective:To explore the early predictive value of urinary nephrin in acute kidney injury (AKI) for critically ill neonates.Methods:A prospective study was conducted to neonates who were admitted to Neonatal Intensive Care Unit (NICU) Children′s Hospital of Soochow University, from July to October 2016.According to whether AKI occurred during the NICU′s hospitalization, neonates were divided into AKI group and non-AKI group.Urinary nephrin levels were detected at the first 24 h of NICU, and the score for neonatal acute physiology (SNAP) was assessed within 24 hours of NICU.Multivariate linear analyses were applied to analyze potential variables that were asso-ciated with urinary nephrin level.Multivariate Logistic regression analysis was adopted to evaluate the relationship between urinary nephrin and AKI after adjusting for confounding factors.A receiver operating characteristic(ROC) curve and the area under the ROC curve (AUC) were calculated to assess the early predictive value of urinary nephrin for neonatal AKI. Results:Among the 156 neonates enrolled in the study, 16 cases(10.2%) developed AKI.The median of urinary nephrin, urinary albumin and SNAP scores were 0.27 μg/mg uCr, 0.48 g/g uCr and 9 scores with AKI group, while the median of urinary nephrin, urinary albumin and SNAP scores were 0.16 μg/mg uCr, 0.16 g/g uCr and 7 scores with non-AKI group.When compared with non-AKI neonates, urinary nephrin ( Z=-3.201, P=0.001), urinary albumin ( Z=-2.652, P=0.008) and SNAP score ( Z=-2.611, P=0.009) were significantly higher in AKI neonates.Multiple linear regression analysis proved that urinary nephrin levels were significantly correlated with urinary albumin ( B=0.488, SE=0.117, P<0.001). Multivariate Logistic regression analysis revealed that urinary nephrin remained significantly associated with AKI ( P=0.018) after adjusting for confounding factors, including gestational age, birth weight, gender, SNAP score, mechanical ventilation and apnea.Urinary nephrin achieved AUC of 0.746 (95% CI: 0.606-0.886, P=0.001). Conclusions:As a biomarker of glomerular injury, urinary nephrin is significantly related to the occurrence of AKI and has early predictive value for AKI in critically ill neonates.
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Objective To investigate the effects of WNK3 kinase on the regulation of large-conductance calcium-activated potassium channels (Maxi K channels) on African green monkey kidney fibroblast-like cells (Cos-7 cells) and its mechanisms.Methods (1) Cos-7 cells were transfected with 0,0.6,1.2,1.8 μg WNK3 plasmid+0.5 μg Maxi K plasmid.The total protein expression of Maxi K channel and the phosphorylation of mitogen-activated protein kinase (MAPK) extracellular regulated kinase-1 and-2 (ERK1/2) were detected by Western blotting.(2) Cos-7 cells were divided into the control group (2.5 μg Maxi K plasmid) and the experimental group (2.5 μg WNK3 plasmid+2.5 μg Maxi K plasmid).Cell surface biotinylation was used to investigate the cell surface protein expression of Maxi K channel in Cos-7 cells.Immunoprecipitation and Western blotting were used to detect the ubiquitination of Maxi K channel protein.(3) WNK3 kinase was knocked down by WNK3 siRNA.The lysosomal degradation pathway was blocked by the proton pump inhibitor (Baf-A1).Cos-7 cells were divided into Maxi K+negative control siRNA group,Maxi K+WNK3 siRNA group and Maxi K+WNK3 siRNA+Baf-A1 group.The protein expression of Maxi K channel protein was detected by Western blotting.Results (1) Compared with those in 0 μg WNK3 plasmid groups,in 0.6,1.2,1.8 μg WNK3 plasmid groups the total protein expression of the Maxi K channel increased and the phosphorylation level of MAPK ERK1/2 reduced on a dose-dependent manner (all P ＜ 0.01).(2)Compared with those in the control group,the total protein expression and cell surface membrane protein expression of the Maxi K channel increased in the experimental group (P ＜ 0.01),while the ubiquitination of the Maxi K channel protein reduced (P ＜ 0.01).(3) Compared with the Maxi K +negative control siRNA group,the expression of Maxi K protein reduced in the Maxi K+WNK3 siRNA group (P ＜ 0.01),but did not change in the Maxi K+WNK3 siRNA + Bar-A1 group (P ＞ 0.05).The expression of Maxi K protein in Maxi K+WNK3 siRNA+Baf-A1 group was higher than that in Maxi K+WNK3 siRNA group (P ＜ 0.01).Conclusions WNK3 kinase inhibits the lysosomal degradation pathway of Maxi K channel protein by reducing the ubiquitination of Maxi K channel,and promotes the expression of Maxi K channel protein in cells and on cell membrane.These effects may be achieved by suppressing MAPK ERK1/2 signal transduction pathway.
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Objective To investigate the expression of inducible co-stimulator (ICOS) and inducible co-stimulator ligand (ICOSL) on PBMCs,and the plasma concentrations of soluble forms of ICOSL and their clinical relationship with systemic lupus erythematosus (SLE) patients.Methods Peripheral blood samples were collected from 45 SLE patients and 39 healthy subjects (HC).The expressions of ICOS and ICOSL on peripheral blood mononuclear cells (PBMCs) were detected by flow cytometry.The concentrations of soluble ICOSL were assessed by measured by enzyme linked immunosorbent assay (ELISA).And the relationship between their expression levels and patients' clinical manifestations were analyzed.Levene F test was used for statistical analysis,the comparison between groups was conducted using t test,and the correlation between two variables were tested by Pearson correlation analysis.Results The expression of ICOS on CD4+ and CD8+ T cells were significantly higher than that of the HC [(19.1±1.7)％ vs (14.0±1.5)％,t=2.156,P=0.035],[(10.0± 1.0)％ vs (6.4±1.0)％,t=2.587,P=0.012].The expression of ICOSL on CD14+ mononuclear cells in SLE patients was significantly higher than that in the HC group [(2.94±0.88)％ vs (0.89 ±0.21)％,t=2.152,P=0.04].Plasma ICOSL concentrations in patients with active SLE were significantly higher than those of patients with inactive SLE [(362±25) ng/ml vs (278±15) ng/ml,t=2.356,P=0.025].We also found a significant negative correlation between the soluble ICOSL expression and the surface ICOSL expression on both mononuclear cells and B cells (r=-0.4243,P=0.022;r=-0.4099,P=0.025).MMPI induced an evident reduction in sICOSL levels released from the cells,which was statistically significant in comparison with untreated cells (P＜0.05).Conclusion The up-regulated expressions of ICOS and ICOSL on peripheral lymphocytes and the high levels of plasma concentration of soluble ICOSL are closely correlated with the severity of the disease,suggesting that ICOS/ICOSL pathway may play a critical role in the pathogenesis of SLE.
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Objective:To investigate the expression of inducible costimulatory ( ICOS) and inducible costimulatory ligand ( ICOSL) on peripheral blood mononuclear cells ( PBMCs ) and their clinical relationship with rheumatoid arthritis ( RA ) patients.Methods:Peripheral blood samples were collected from 85 RA patients and 50 HC in this study.Expression of ICOS and ICOSL on PBMC from the subjects were detected by flow cytometry and real-time polymerase chain reaction( RT-PCR).The alteration of ICOS and ICOSL were observed after hormone therapy in 15 patients with RA and the relationship between their expression level and patients′clinical manifestations were analysed.Results:The ICOS and ICOSL mRNA level of RA patients′PBMCs were significantly higher than that in HC.The expression level of ICOS on CD4+T cells was higher than than that in HC[(7.08±4.72)% vs (3.01+1.39)%,P<0.0001].The expression of ICOSL on monocytes[(5.77±3.45)%vs (3.64±1.43)%,P<0.05] and B cells [(5.78± 4.52)%vs (3.97±1.63)%,P<0.05] were significantly elevated in RA patients.In RA patients with active disease,however,ICOSL expression on monocytes and B cells were increased as compared with those in inactive RA patients [ ( 5.45 ±3.50 )% vs ( 4.04 ± 1.55)%,P=0.036],[(6.59 ±5.74)%vs (5.63±4.30)%,P=0.016].Furthermore,after receiving immunosuppressive therapy, the expressions of ICOS and ICOSL were notably reduced as compared with pre-therapy levels on PBMCs from patients [ ( 5.45 ±3.50)%vs (4.04±1.55)%,P=0.036],[(6.59 ±5.74)%vs (5.63±4.30)%,P=0.016].Conclusion:The high levels of ICOS and ICOSL expression were closely correlated with the degree of disease and therapeutic response,suggesting that ICOS/ICOSL pathway may play a critical role in pathogenesis of RA.
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Objective To investigate the relationship between single nucleotide polymorphism (SNP) of susceptibility genes CD-KAL1 ,CDKN2A-CDKN2B ,TCF7L2 and HHEX-IDE of type 2 diabetes mellitus and gestational diabetes mellitus (GDM) in Jiangsu region .Methods 185 healthy pregnant women without GDM tendency (control group) and 176 pregnant women with GDM (GDM group) were enrolled .Case-control method and Multiplex SNaPshot SNP genotyping technology were utilized to detect genotypes at SNP loci of CDKAL1 ,CDKN2A-CDKN2B ,TCF7L2 and HHEX-IDE of pregnant women both in the two groups , and all SNP loci were subject to continuous disequilibrium analysis .Results Compared the allele frequencies of pregnant women in GDM group and the control group ,C allele of CDKAL1(rs7754840) was a risk factor for GDM .The risk of suffering GDM of CC genotype carriers was 1 .73 times the non-CC genotype carriers[P<0 .01 ,OR=1 .73 ,95% CI:1 .28-2 .33] .No statistical difference wasfound in gene distribution of detection loci of CDKN2A-CDKN2B(rs10811661),TCF7L2(rs7903146) and HHEX-IDE (rs1111875) .The genotype frequencies of CDKAL1(rs7754840) ,CDKN2A-CDKN2B(rs10811661) ,TCF7L2(rs7903146) and H HEX-IDE(rs1111875) were all accordance with the law of genetic equilibrium .Conclusion CDKAL1(rs7754840) is closely re-lated to GDM susceptibility in Jiangsu region .
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Objective To evaluate the performance of electrolyte module in OLYMPUS AU5421 Automatic Biochemical Analy-zer in K+ ,Na+ ,Cl- detection .Methods Electrolyte module in OLYMPUS AU5421 Automatic Biochemical Analyzer was employed to detect K+ ,Na+ ,Cl- ,and their precision ,accuracy ,linearity range ,cross-contamination and the test results of instruments were compared .The linear regression equation and regression coefficients were calculated according to the methods of Clinical and Labo-ratory Standards Institute(CLSI) EP6-A ,EP9-A2 documents .Results The within-run precisions of K + ,Na+ ,Cl- were 0 .328% , 0 .306% ,0 .343% ,respectively ,and their between-run precisions were 0 .780% ,1 .012% ,0 .985% ,respectively .Their regression coefficients of linear equation were all greater than 0 .999 ,and their cross-contamination rate were 1 .066% ,0 .812% and 0 .858% , respectively .Conclusion Electrolyte module in OLYMPUS AU5421 Automatic Biochemical Analyzer modules has advantages of good precision ,accuracy ,linear range ,and low cross-contamination rate .