Objective To study the effect and molecular mechanism of vitamin D(VitD)on Tfh cells of MRL/lpr lupus mice.Methods C57/B6 mice and MRL/lpr lupus mice were transfected with siRNA to construct VDR knockout mouse models.Splenic Tfh cells of C57/B6 mice and MRL/lpr lupus mice were divided into control group,lupus group and VDRsiRNA lupus group(treated with vitamin D 0,1 and 10 nmol·L-1,respectively)by siRNA transfection.The percentage of Tfh cells was detected by flow cytometry.MRL/lpr lupus mice Peyer node Tfh cells were randomly divided into 7 groups,blank control group,vitamin D dose groups of 1 and 10 nmol·L-1,paricalcitol group(VitD 10 nmol·L-1+PA),VDRsiRNA control group,VDRsiRNA group(VitD 10 nmol·L-1),CaN inhibitor group(VitD 10 nmol·L-1+CsA),and incubated for 72 h.The concentration of calcium ions in Tfh of each group was detected.The expressions of AT1R,NFAT,CaN and P-CaN in Tfh cells were determined by Western blotting.Results The percentage of Tfh cells decreased significantly with the increase of vitamin D dose.Vitamin D CaN reduce the intracellular calcium concentration of Tfh,up-regulate the expression of AT1 protein in Tfh cells,and down-regulate the expression of CAN,P-CaN and NFAT protein in a dose-dependent manner,and the effect is more obvious when combined with PA.Conclusion Vitamin D may regulate the activation of follicular T helper cells in MRL/lpr mice via the Ca-CaN-NFAT pathway.

Objective To study the effect and molecular mechanism of vitamin D(VitD)on Tfh cells of MRL/lpr lupus mice.Methods C57/B6 mice and MRL/lpr lupus mice were transfected with siRNA to construct VDR knockout mouse models.Splenic Tfh cells of C57/B6 mice and MRL/lpr lupus mice were divided into control group,lupus group and VDRsiRNA lupus group(treated with vitamin D 0,1 and 10 nmol·L-1,respectively)by siRNA transfection.The percentage of Tfh cells was detected by flow cytometry.MRL/lpr lupus mice Peyer node Tfh cells were randomly divided into 7 groups,blank control group,vitamin D dose groups of 1 and 10 nmol·L-1,paricalcitol group(VitD 10 nmol·L-1+PA),VDRsiRNA control group,VDRsiRNA group(VitD 10 nmol·L-1),CaN inhibitor group(VitD 10 nmol·L-1+CsA),and incubated for 72 h.The concentration of calcium ions in Tfh of each group was detected.The expressions of AT1R,NFAT,CaN and P-CaN in Tfh cells were determined by Western blotting.Results The percentage of Tfh cells decreased significantly with the increase of vitamin D dose.Vitamin D CaN reduce the intracellular calcium concentration of Tfh,up-regulate the expression of AT1 protein in Tfh cells,and down-regulate the expression of CAN,P-CaN and NFAT protein in a dose-dependent manner,and the effect is more obvious when combined with PA.Conclusion Vitamin D may regulate the activation of follicular T helper cells in MRL/lpr mice via the Ca-CaN-NFAT pathway.

Objective:To research the correlation of serum levels of IL-21,TGF-β1,TNF-α and IgA1 in children with allergic purpura and the purpura nephritis.Methods:57 cases with allergic purpura who were treated in our hospital from November 2015 to May 2016 were selected and 29 cases were diagnosed with general allergic purpura,and another 28 cases were diagnosed with purpuric nephritis.30 healthy children were selected as the control group.Then the serum levels of IL-21,TGF-β1,TNF-α,IgAl,immunoglobulin A (IgA),C3 and C4 between the three groups were observed and compared.Results:The serum levels of IL-21,TGF-β1,TNF-α,IgA1 and IgA in the purpuric nephritis group were higher than those of the control group and the general allergic purpura group,and the differences were statistically significant (P＜0.05);The serum levels of IL-21,TGF-β1,TNF-α,IgA1 and IgA in the general allergic purpura group were higher than those of the control group,and the differences were statistically significant (P＜0.05);There was no statistically significant difference about the C3 and C4 in the three groups (P＞0.05).Conclusion:The serum levels of IL-21,TGF-β1,TNF-α and IgA1 may be involved in the development ofhenoch-schonlein purpura and purpura nephritis.

Objective:To investigate the baseline levels of 25-hydroxy-vitamin D [25-(OH) D] in 3340 hospitalized children, analyze the 25-( OH) D levels in children with different diseases, age and gender in different seasons, and study the correlation between the 25-( OH) D levels and the clinical indicators. Methods:Totally 3340 hospitalized children were randomly selected, the 25-( OH) D levels were detected by an ELISA method, and Pearson correlation analysis of 25-( OH) D levels and clinical indicators such as liver function, myocardial enzymes, immunoglobulins, lymphocyte subtypes and thyroid function was performed. Results: The serum 25-(OH) D level in 3340 cases (1850 male, 1490 female) was (33. 00 ± 13. 42) ng·m1 -1, and that in those with neonatal diseases was the lowest followed by those with primary nephrotic syndrome, Henoch-schordeinpurpura and juvenile idiopathic arthritis. The ser-um 25-( OH) D levels in the premature was higher than that in full term infant. Except the newborn, the level of serum 25-( OH) D gradually reduced along with the age increase, while the percentage of insufficiency gradually increased. The serum 25-( OH) D level between the male and the female had no significant difference. The 25-( OH) D levels of hospitalized children were the highest in sum-mer. The serum level of 25-( OH) D was positively correlated with body mass index ( BMI) , alanine transaminase ( ALT) , aspartate transaminase (AST), creatine kinase (CK), creatine-phosphokinase (CK-MB), lactate dehydrogenase(LDH), free T4 (FT4) and free T3 (FT3), and negative correlation with alkaline phosphatase(ALP), immunoglobulin E (IgE) and immunoglobulin M (IgM). Conclusion:The incidence of vitamin D insufficiency is high in hospitalized children. The level of vitamin D is different among various diseases. The level of serum 25-( OH) D may have certain relevance with BMI, allergies, myocardial damage and thyroid function.

Objectives To study the protective effects of vitamin D (VitD) on podocyte insulin resistance and its mecha-nisms. Methods Immortalized mouse podocytes in vitro were randomly divided into 4 groups:podocytes+5 mmol/L glucose group (group A);podocytes+5 mmol/L glucose+1 nmol/L propylene glycol group (group B);podocytes+30 mmol/L glucose+1 nmol/L propylene glycol group (group C); podocytes+30 mmol/L glucose+1 nmol/L propylene glycol+1 nmol/L VitD group (group D). The percentage of podocyte apoptosis was determined after 48 h of incubation. Podocyte viability was assessed by MTT assay. The mRNA expressions of vitamin D receptor (VDR) and insulin receptor substrate protein 1 (IRS1) in podocyte were detected by RT-PCR. Western blot analysis was performed to measure the protein levels of p-IRS1/IRS, p-Akt/Akt and p-ERK1/2/ERK1/2. Results There were significant differences in apoptosis percentage, viability and the expression of VDR, IRS1, p-ERK1/2 of podoctyes(P<0.05)among 4 groups. There was no difference in p-Akt/Akt expression among 4 groups(P>0.05). Compared with group A, B , and D, the percentage of podocyte apoptosis in group C was significantly increased, the cell viabi-lity was decreased, the expressions of VDR and IRS1 mRNA and p-IRS1 and p-Akt proteins were down-regulated, whereas p-ERK1/2 was up-regulated in group C. The levels of p-IRS1/IRS1, p-Akt/Akt, p-ERK1/2/ERK1/2 had no statistical differences in group A, B, and D (P>0.05). Conclusions VitD-VDR system alleviates podocyte apoptosis induced by high glucose, and acti-vates insulin signaling pathway and counteracts insulin resistance signal to improve podocyte insulin resistance.

Objective To investigate the protective effect of Vitamin D (VitD) on diabetic rats,and whether the protective mechanism is associated with the expression of nuclear factor kappa B (NF-κB) and insulin resistance (IR).Methods Diabetic Wistar rats were established by intraperitoneal injection of streptozotocin,and randomly divided into diabetic group,VitD treatment group (treatment group).Normal rats were served as normal control group.Treatment group was treated with VitD for 8 weeks.The levels of 24 h urinary protein (24 h UP),fasting plasma glucose (FPG),plasma insulin (p-Ins),plasma adiponectin (p-Adi),plasma glucagon (p-Gln) were measured.Tumor necrosis factor alpha (TNF-α),monocyte chemotactic protein 1 (MCP-1),interleukin-6 (IL-6) were determinated in renal cortical homogenate.The activity of NF-κB was evaluated by electrophoretic mobility shift assay.The mRNA expressions of glucose transporter protein 1 (GLUT1) and GLUT4 in renal cortex were detected by reverse transcriptase (RT)-PCR.Western blot analysis was performed to measure the phosphorylation of NF-κB and its inhibitor I kappa Balpha (IκBα),insulin receptor substrate protein 1 (IRS1),phosphatidylinositol 3 kinase (PI3K),p38 mitogen activated protein kinase (p38MAPK).Results Compared with the normal control group,24 h UP,FPG,p-Ins,p-Gln were significantly increased,and inflammatory markers and the expression of GLUT1 elevated in renal cortex in DM group,there were significant differences(all P ＜0.01).The activity of NF-κB (P ＜0.01) and the phosphorylation of NF-κB p65 and p38MAPK were elevated (all P ＜ 0.01),and phosphorylation of IκBα,IRS1,PI3K were decreased (all P ＜ 0.05,0.01) in diabetic group compared with those of normal control group.VitD treatment could ameliorate urine protein,increase p-Adi,reduce inflammatory markers and NF-κB activity (P ＜ 0.01),maintain GLUT1 expression,but had no effect on GLUT4 expression in renal cortical,attenuate NF-κB p65,p38MAPK phosphorylation (all P ＜ 0.05),partly restore IκBα,IRS1,PI3 K phosphorylation in diabetic rats (all P ＜ 0.05,0.01).Conclusions Protective role of VitD is associated with inhibiting the expression of NF-κB and reducing the insulin resistance in diabetes.

Objective To test the effects of Vitamin D (VitD) on endothelial nitric oxide(NO) production and to study the signal pathway leading to NO release.Methods In vitro cultured human umbilical vein endothelial cells (HUVEC) were treated with various concentrations of VitD(0 mmol/L,0.01 mmol/L,0.10 mmol/L,1.00 mmol/L,10.00 mmol/L) for 60 min,and VitD at concentration of 1.00 mmol/L at different time points (30 min,60 min,90 min,120 min).The effect of VitD on NO production in presence of VitD receptor(VDR) agonist(ZK191784) or antagonist(ZK159222) for 60 min were examined in cell culture supernatant with kit for the detection of nitric oxide fluorescent probe(DAF-FM DA).HUVEC was cultured with VitD in presence of VDR agonist or antagonist for 60 min,and the effect of VitD on NO production with DAF-FM DA and the protein expression and phosphorylation of Caveolin-1 and endothelial nitric oxide synthase(eNOS) were detected by Western blot,respectively.Results VitD caused a concentration-dependent increase in NO production.The maximum effect was observed at a concentration of 1.0 mmol/L and the optimal time of stimulation was 60 min.Effects induced by VitD were enhanced by VDR agonist,and abolished by antagonist.VitD and VDR agonist maintained the expression of Caveolin-1 at the same low phosphorylation level the same as normal,increased the phosphorylated level of eNOS.However,VDR antagonist increased the phosphorylation of caveolin-l,but reduced the level of eNOS phosphorylation,respectively.Conclusions VitD can induce a significant increase in endothelial NO production through VDR.VitD interaction with VDR causes the low phosphorylation of caveolin-1 leading to eNOS activation and NO production.

Objective To explore the possible role of vitamin D in the pathogenesis of IgA nephropathy (IgAN). Me-thods After the IgAN model was successfully induced at 12 weeks, the BALB/C mice were randomly divided into IgAN group (n=15) and IgAN+VitD group (n=15). The nephrosis mice were administrated with 100 μl/d propylene glycol or propyl-ene glycol+1,25(OH)2D, 3 ng/(100g?d), for 6 weeks. The control group was setted (n=15). The level of 24 hour urine protein was determined at week 0, 12 and 18. At week 18, the levels of serum 25(OH)D, ifbroblast growth factor 23 (FGF23) and galactose-deifcient IgA1 (Gd-IgA1) were detected. The mRNA and protein expressions of interleukin-21 (IL-21) in Peyer’s patches (PPs) were detected by lfuorescent quantitative reverse transcription-polymerase chain reaction and western blot respectively. The protein expression of Bcl-6 was detected by western blot. The percentages of Tfh cells/T lymphocytes, B220+IgM+/B lympho-cytes, B220+IgA+/B lymphocytes, B220-IgA+/B lymphocytes in PPs were determined by lfow cytometry. Results Compared with control group, the levels of 24 hour urine protein, FGF23 and Gd-IgA1 were increased, serum 25(OH)D was decreased, the mRNA and protein expressions of IL-21 and the protein level of Bcl-6 were increased, the percentages of Tfh cells/T lym-phocytes, B220+IgM+/B lymphocytes, B220+IgA+/B lymphocytes, B220-IgA+/B lymphocytes were elevated in IgAN group (P<0.05). These indicators were improved in IgAN+VitD group. Compared with the IgAN group, the differences were statisti-cally signiifcant (P<0.05), however compared with control group, some indicators showed no signiifcant differences (P>0.05). Conclusions 1,25(OH)2D may protect the microenvironment of PPs in IgAN through inhibiting the differentiation of Tfh cells and B cells and the generation of Gd-IgA1.

Objective To analyze the clinical and pathological characteristics of Alport syndrome in children. Methods Clinical and pathological information gathered from 62 patients during March 1989 to August 2012 was retrospectively analyzed. Results Four autosomal recessive Alport syndromes (AR-AS) and 58 X-linked Alport syndromes (XL-AS) were analyzed. Of the XL-AS, 47 were boys and 11 were girls. Most of patients induced by upper respiratory tract infections, and onset with hematuria and proteinuria. There was no signiifcant gender difference in family history, impaired renal tubular proteins, hypertension, im-paired renal function, hearing loss, ocular abnormalities or renal pathological changes under light microscopy. However, extensive lamination and split of glomerular basement membrane (GBM) dense layers were found in 83.0%male and 18.2%female patients (P=0.000) and the rest patients were presented with limited distribution of typical GBM changes. Proteinuria progressed signiif-cantly with age in XL-AS males (r=0.501, P=0.000). Five XL-AS patients developed to end stage renal disease (ESRD) between 11 to 16 years old. Conclusions XL-AS is the main inherited type and severe changes of GBM are common in XL-AS males. Proteinuria increases remarkably with age. The detection of type IV collagen in renal tissue or skin is helpful to diagnose Alport syndrome and conifrm inheritance modes.

Fabry disease is an inherited metabolic disease, resulting from a deficiency of the enzyme a-galactosidase A activity. Principal clinical manifestation of Fabry disease was limb pain episodes, angioceratoma, and progressive damage of heart,brain, kidney. Although Fabry disease cannot be cured at present,early diagnosis and enzyme replacement therapy can change the prognosis completely.