Proteomic Study on the Effect of Wenshen Tongdu Formula （温肾通督方） on Spleen B Cells in Spinal Cord Injury Model Mice / 中医杂志

ObjectiveTo explore the effects and possible mechanism of Wenshen Tongdu Formula （温肾通督方， WTF） on spinal cord injury. MethodsThirty-six C57BL/6 female mice were randomly divided into sham operation group， model group and WTF group， with 12 mice in each group. The spinal cord injury model was established in the model group and the WTF group using the modified Allen's method， while in the sham operation group the spinal cord was only exposed. Since the 1st day after surgery， 50 g/（kg·d） of WTF solution was given to the WTF group by gavage， while 20 ml/（kg·d） of normal saline was given to the sham operation and model group by gavage， all for 14 days. Before surgery and on the 1st， 7th， and 14th days after surgery， the motor function of the mice was evaluated using the inclined plane test and hind limb motor function score （by BMS）. On the 3rd day after surgery， the nerve electrophy-siology was detected through electromyography and motor evoked potential； the spleen length was measured， and B cells in the spleen were sorted by magnetic beads； the differential expression of proteins were detected through proteomics technology； and the protein expression of mitochondrial outer membrane transport porin 20 （Tom20） and downstream cleaved caspase-3 in spleen B cells were measured using Western blotting. On the 14th day after surgery， MRI was used to observe the recovery of the spinal cord. ResultsCompared to those in the sham operation group at the same time， the BMS scores and subscores and the inclined plane test angle in the model group were reduced on the 1st， 7th and 14th days after surgery； the peak value of electromyogram and motor evoked potential were reduced， and the spleen length was shortened， while the expression of Tom20 and cleaved caspase-3 increased in splenic B cells increased （P＜0.05）. Compared to those in the model group at the same time， the BMS subscores on the 14th day and the angle of the inclined plane test on the 7th and 14th days after surgery increased in the WTF group； the peak value of electromyography and motor evoked potential， as well as the length of spleen increased， and the expression of Tom20 and cleaved caspase-3 decreased （P＜0.05）. The proteomics results showed that there were 100 differential proteins in the WTF group versus the model group， of which 37 were up-regulated and 63 were down-regulated. GO enrichment analysis showed that differential proteins mainly played their roles in oxygen binding， exogenous apoptosis negative feedback， zinc ion response， and oxygen transport. KEGG enrichment analysis showed that differential proteins were mainly concentrated in metabolic pathways， Huntington's disease， oxidative phosphorylation and other pathways. Subcellular localization showed that differential proteins were associated with mitochondria. Magnetic resonance imaging on the 14th day after surgery showed that the spinal cord structure of the mice in the sham operation group was intact， and the segments were clear， with normal spinal cord signal； the low signal area in the spinal cord injury area increased in the model group， and the spinal cord became significantly thinner； the injured segment had obvious depression in the WTF group， but the structure was more complete than that in the model group. ConclusionWTF may promote spinal cord injury repair by regulating immune function， and its mechanism may be related to inhibiting pyroptosis of spleen B cells.

Efficacy of conformal intensity-modulated radiotherapy with paclitaxel liposome versus cisplatin in the treatment of locally advanced cervical cancer / 中国基层医药

Objective:To investigate the efficacy of conformal intensity-modulated radiotherapy combined with paclitaxel liposome versus cisplatin in the treatment of locally advanced cervical cancer. Methods:Sixty patients with cervical cancer who received treatment in Rushan People's Hospital from February 2014 to September 2016 were included in this study. They were randomly assigned to undergo either conformal intensity-modulated radiotherapy combined with cisplatin (control group, n = 30) or conformal intensity-modulated radiotherapy combined with paclitaxel liposome (observation group, n = 30) for five cycles. Clinical efficacy and incidence of adverse reactions were compared between the two groups. Results:Total response rate in the observation group was slightly, but not significantly, higher than that in the control group [93.3% (28/30) vs. 90.0% (27/30), P > 0.05]. The incidences of myelosuppression and gastrointestinal reactions in the observation group were 26.7% (8/30) and 53.3% (16/30), respectively, which were significantly lower than those in the control group [46.7% (14/30), 76.7% (23/30), χ2 = 5.16, 7.17, both P < 0.05]. After radiotherapy and chemotherapy, the score of quality of life in the observation group was significantly higher than that in the control group ( t = 3.19, 4.12, 3.22, 4.53, 3.63, 3.08, 3.78, all P < 0.05). Conclusion:Conformal intensity-modulated radiotherapy combined with paclitaxel liposome for treatment of locally advanced cervical cancer can effectively reduce myelosuppression and gastrointestinal reactions and improve quality of life. Conformal intensity-modulated radiotherapy combined with paclitaxel liposome has certain clinical significance in the treatment of locally advanced cervical cancer.

Fludarabine and intravenous busulfan conditioning with post-transplantation cyclophosphamide for allogeneic peripheral stem cell transplantation for adult patients with lymphoid malignancies: a prospective single-arm phase II study / 医学前沿

Post-transplantation cyclophosphamide (PT-Cy) alone or in combination with other immunosuppressive drugs has emerged as a promising strategy in the setting of allogeneic hematopoietic stem cell transplantation. Improved survival rate was reported in lymphoid malignancies following PT-Cy strategy compared with myeloid disease in non-myeloablative bone marrow transplant setting. Thus, we aimed to evaluate the safety and efficacy of PT-Cy combined with cyclosporine as graft-versus-host disease (GVHD) prophylaxis after myeloablative conditioning and T cell-replete peripheral stem cell transplantation in lymphoid malignancies. This single-arm phase II clinical trial (NCT01435447) involving 31 adult patients was conducted from January 2013 to June 2018. The donor-type neutrophil engraftment rate was 100%, and the overall incidence of grade II to IV and grade III to IV acute GVHD was 39% and 24%, respectively. The cumulative incidence rates of chronic GVHD (35%), including moderate to severe forms (10%), were reduced compared with those of the historical group (P = 0.03 and P = 0.04, respectively). With a median follow-up of 18 months, the estimated 2-year overall and event-free survival was 64.8% (95% confidence interval: 47.8%-86.7%) and 58.4% (95% CI: 41.9%-81.7%), respectively. The 2-year cumulative incidence rate of relapse was 19.5% (95% CI: 9.0%-35.8%), whereas the non-relapse mortality rate was 21.8% (95% CI: 11.3%-38.1%). These results demonstrated the feasibility of PT-Cy as GVHD prophylaxis in this clinical setting. This strategy could significantly reduce the incidence of chronic GVHD and its moderate to severe forms but not of acute GVHD and results in similar survival outcomes compared with the historical group. A prospective study with additional patients is warranted to confirm the role of PT-Cy in lymphoid malignancy.

Research progress on signaling molecules involved in articular cartilage repair / 生物医学工程学杂志

After the articular cartilage injury, the metabolic level is increased during the progressive degeneration, the chondrocytes secrete a variety of inflammatory factors, and the original cell phenotype is gradually changed. For a long time, a large number of researchers have done a lot of researches to promote anabolism of chondrocytes and to maintain the stability of chondrocyte phenotype. There are many molecular signaling pathways involved in the process of promoting cartilage repair. This review focuses on the key signaling molecules in articular cartilage repair, such as transforming growth factor-beta and bone morphogenetic protein, and reveals their roles in the process of cartilage injury and repair, so that researchers in related fields can understand the molecular mechanism of cartilage injury and repair widely and deeply. Based on this, they may find promising targets and biological methods for the treatment of cartilage injury.

MRI classification and imaging findings of dysembryoplastic neuroepithelial tumors / 中华放射学杂志

Objective To investigate the MRI classifications and imaging findings of dysembryoplastic neuroepithelial tumor(DNET). Methods MR images of 34 patients with pathologic confirmed DNET of Beijing Sanbo Brain Hospital were retrospectively reviewed in this study. The classification was made according to the number of pseudocysts, scope of involvement, morphology and location. Results MRI appearances of DNET were divided into three subtypes: cystic‐like, polycystic‐like and diffuse type. Twelve cases had cystic cortical, including front lobe (5 cases), temporal lobe (5 cases), parietal lobe (2 cases). These cases presented quasi‐circular or oval shape, with hypointense on T1WI and strongly hyperintense on T2WI. T2‐FLAIR was observed hyperintense ring sign in the tumor periphery and the cystic content was close to CSF but having the largest difference to that of CSF, which signal was higher than CSF. Twenty cases were polycystic‐like, front lobe (7 cases), temporal lobe (7 cases), parietal lobe (5 cases), occipital lobe (1 case). In these 20 cases, they had slightly hypointense on T1WI and strongly hyperintense on T2WI. Located in the cortex and subcortical matter, with wedge shape, gyriform or triangle shape.On T2‐FLAIR, internal septation and hyperintense"ring sign"were observed. Two cases were diffuse type, bilateral (1 case), unilateral (1 case). In these 2 cases, diffuse lesions involving multiple areas with hyperintense ring and internal septation on FLAIR, including subcortical white matter, deep nucleus and periventricular area. Conclusions The MR appearances of DNET are variable. Understanding the MR imaging type of DNET might improve the MR diagnosis of DNET.

Mechanism of Sclerostin/Lrp4 in the regulation of vascular smooth muscle cell calcification / 中华肾脏病杂志

Objective To investigate the possible mechanism of sclerostin/Lrp4 in calcification of VSMC induced by high phosphorus and the protective effect of Ginkgo biloba extract.Methods Aortic vascular smooth muscle cells (VSMCs) of SD rats were extracted and identified.VSMCs were divided into normal control group,high phosphorus induced calcification group (10 mmol/L β-glycerophosphate+50 μg/ml ascorbic acid),and high phosphorus induced calcification+Ginkgo biloba extract intervention group (10 mmol/L β-glycerophosphate+50 μg/ml ascorbic acid+0.5 mg/ml GBE),cultured in different mediums for 14 days.Vonkossa staining and alizarin red staining were used to detect the calcification of VSMCs.The mRNA level of BGP was detected by real time PCR,and the protein expressions of sclerostin and Lrp4 were detected by Western blot.Results Compared with normal control group,vonkossa staining and alizarin red staining showed significant calcium deposition in calcification group.Compared with calcification group,calcium salt deposition was significantly reduced in GBE treatment group.Real time PCR results showed β-catenin and BGP mRNA expressions in VSMC calcification group were higher than those in normal control group (P＜ 0.05).mRNA expressions of β-catenin and BGP in GBE treatment group were lower than those in calcification group (all P ＜ 0.05).Compared with normal control group,the protein expression of sclerostin was increased,but the protein expression of Lrp4 was decreased in calcified group (all P ＜ 0.05).Compared with calcification group,the protein expression of sclerostin decreased and the protein expression of Lrp4 increased in GBE treatment group (all P ＜ 0.05).Conclusions High phosphorus can induce VSMC calcification by activating Wn/β-catenin signaling pathway.Sclerostin/Lrp4 is involved in hyperphosphine-induced VSMC calcification.GBE can reduce the high phosphorus induced VSMC calcification by regulating the Wnt/β-catenin signaling pathway.

Advanced glycation end products induce mitochondrial pathway apoptosis in glomerular mesangial cells / 中华肾脏病杂志

Objective To investigate whether advanced glycation end products (AGEs) can induce the expression of Ros,JC-1 and its apoptosis-related proteins in glomerular mesangial cells under high glucose environment,induce apoptosis and injury of glomerular mesangial cells.Methods Rat glomerular mesangial cell line HBZY-1 was cultured in vitro.The cells were cultured with different concentrations of AGEs for 0,12,24 and 48 hours respectively.MTT assay was used to observe the cell proliferation ability.After the optimal time and concentration of AGEs were selected,the caspase enzyme inhibitor Z-VAD-fmk and reactive oxygen species (ROS) scavenger N-acetyl-L-cysteine (NAC) were cultured and the apoptosis rate was detected by cell death detection apoptosis ELISA plus and Annexin V-FITC/PI kit.JC-1 staining was used to detect the changes of mitochondrial membrane potential (MMP).Cell ROX deep red flow cytometry was used to detect the total ROS level.The expression of anti-apoptotic protein Bcl-2,pro-apoptotic protein BAX,caspase-9,caspase-3 and poly ADP-ribose polymerase (PARP)-activated fragments was detected by Western blotting.Results AGEs could decrease the activity of glomerular mesangial cells in a time and concentration-dependent manner,and induce cell death.The percentage of apoptotic cells in glomerular mesangial cells was significantly increased after treatment with 250 mg/L AGEs for 24 h (P ＜ 0.01),and Z-VAD-fmk could significantly alleviate AGEs-induced glomerular mesangial cell apoptosis (P ＜ 0.01).Compared with the control group,AGEs increased the level of intracellular reactive oxygen species and decreased MMP in a time-dependent manner,and the two time points that AGEs significantly caused the change were 1 h and 2 h (all P ＜ 0.01).AGEs also reduced the expression of antiapoptotic protein Bcl-2 and increased the expression of pro-apoptotic protein Bax,cleaved caspase-9,cleaved caspase-3 and cleaved PARP (all P ＜ 0.01).Compared with AGEs group,NAC could significantly stabilize MMP (P ＜ 0.01),increase Bcl-2 expression (P ＜ 0.01),and decrease the expression of BAX,cleaved caspase-9,cleaved caspase-3 and cleaved PARP (all P ＜ 0.01).Conclusion AGEs induce mitochondrial pathway apoptosis in glomerular mesangial cells by increasing intracellular ROS level and destroying MMP.

Abated microRNA-21 attenuates high glucose-induced autophagy inhibition in rat mesangial cells by PTEN/Akt/mTOR pathway / 中华肾脏病杂志

Objective@#To investigate the effects of abated microRNA-21 (miRNA-21) on phosphatase and tensin homologue on chromosome ten protein (PTEN) and PI3K/Akt/mTOR pathway, as well as their further influence on the autophagy in high glucose (HG, 25.0 mmol/L) induced rat glomerular mesangial cells.@*Methods@#MiRNA-21 inhibitor and negative control were transfected by liposome 2000 into rat glomerular mesangial cells (HBZY-1). The cells were divided into normal glucose (5.5 mmol/L) group, normal glucose+negative control group, normal glucose+miRNA-21 inhibitor group, HG group, HG+negative control group and HG+miRNA-21 inhibitor group. Cell proliferation and hypertrophy were assayed by MTT and the ratio of total protein to cell number respectively. The miRNA-21 expression was detected using real time PCR. The expressions of PTEN/Akt/mTOR signaling signatures, autophagy-associated protein (p62 and LC3 Ⅱ) and collagen Ⅰ was detected by Western blotting and real time PCR. Autophagosomes were observed using electron microscopy.@*Results@#Compared with those in normal glucose group, in HG group cells had hypertrophy and proliferation, up-regulated miRNA-21 expression, and down-regulated PTEN protein and mRNA expressions (all P<0.01). Also there were and up-regulated p-Akt, p-mTOR, p62 and collagen Ⅰ expression, and lower LC3 Ⅱ expression and autophagosomes (all P<0.01). Further, compared with those in HG group, cells hypertrophy and proliferation in HG+miRNA-21 inhibitor group were reduced, expressions of p-Akt, p-mTOR, p62 and collagen Ⅰ were down-regulated, while expressions of PTEN and LC3 Ⅱ and autophagosomes were up-regulated (all P<0.01).@*Conclusions@#MiRNA-21 inhibitor up-regulates PTEN expression, which inhibits the activation of Akt/mTOR signaling pathway, ameliorates cell hypertrophy, proliferation and enhances autophagy to reduce extracellular matrix accumulation.

Risk Factors and Correlation Analysis between the Oxford Classification and Clinical Indicators of IgA Nephropathy / 中国医科大学学报

Objective To analyze the risk factors and correlation between clinical indicators and the four main pathological lesions of IgA ne?phropathy in the Oxford classification:mesangial hypercellularity(M0/1),endocapillary proliferation(E0/1),segmental sclerosis or adhesion(S0/1), and tubular atrophy/interstitial fibrosis(T0/1/2). Methods Clinical and pathological data were collected from 514 patients with biopsy?proven IgA nephropathy admitted in our hospital from February 17,2006 to October 11,2011. These patients were all above 18 years old. Cases with sec?ondary causes of mesangial IgA deposition were excluded,such as Henoch?chonlein purpura,ankylosing spondylitis and psoriasis et al. The inde?pendent risk factors affecting the pathological classification were analyzed by Spearman rank correlation analysis and two?category and multi?classi?fication logistic regression using SPSS 17.0 statistical software. Results In 514 IgAN patients,the ratio of males to females was 1.06:1. The aver?age age was 35.70±11.99 years,and the average disease duration was 18.31±30.42 months. M0E0S0T0 was the major pathologic classification of isolated hematuria. Chronic kidney disease(CKD)stage,24 hours proteinuria,albuminuria,urine transferrin and IgG levels were positively corre? lated with M lesion;serum albumin,C3 and PLT showed a negative correlation with M lesion. Twenty four hours proteinuria and blood platelet count were the independent risk factors for M lesion. As shown by stratified analysis ,the proportion of M1 in cases with 24 hours proteinuria≥3.5 g/d is much higher than that in cases with non?nephrotic range proteinuria. Age,systolic blood pressure,uRBC,24 hours proteinuria,albuminuria urine transferrin and IgG levels were positively correlated with E lesion,Duration,serum albumin showed a negative correlation with E lesion. Age and duration of nephritis were independent risk factors for E lesion. 73.3%of patients that above 60 years old showed endothelial proliferation. CKD stage,24 hours proteinuria were positively correlated with S lesion. Age,CKD stage,systolic blood pressure,diastolic blood pressure,C4,TC, LDL?C,CRP,Fib,UA,Cys?C and 24 hours proteinuria,urineβ2?microglobulin,albumin,transferrin and IgG levels were positively associated with T lesion;hemoglobin,serum albumin,serum IgG showed a negative correlation with T lesion. Infection history,high CRP levels,DBP more than 90 mmHg,hypoalbuminemia,high low density lipoproteinemia,and anemia were independent risk factors for T lesion. Conclusion Twenty four hours proteinuria,blood platelet count,age,duration of nephritis,hypoalbuminemia,anemia,hyperlipidemia,DBP≥90 mmHg and high CRP lev?els were risk factors for the Oxford classification of IgA nephropathy. Renal biopsy should be carried out in time to make clear the pathological clas?sification and individual treatment,so as to improve the prognosis.

Glomerular C4d Deposition Is a Prognostic Factor for IgA Nephropathy / 中国医科大学学报

Objective To investigate the significance of glomerular deposition of C4d in accessing the severity and prognosis of IgA nephropathy. Methods A total of 131 patients were recruited for the study. Immunofluorescence was used to detect the deposition of C4d in renal tissue of pa?tients with IgA nephropathy,and the relationship between C4d deposition and clinical and pathological parameters and renal remission was ana?lyzed. Results Totally 30 patients had glomerular deposition of C4d. Compared with the patients without C4d deposition,the patients with C4d deposition had significantly higher levels of serum creatinine,urinary protein excretion and C4d and higher prevalence of hypertension,but had sig?nificantly decreased levels of glomerular filtration rates. With the histopathological phenotypes segregated by Lee 's classification,the ratios of C4d deposition presented an increase(P=0.005). The patients with C4d deposition had more severe mesangial proliferation,endocapillary hypercellu?larity,segmental glomerulosclerosis and tubular?interstitial injury. The rates of renal remission were significantly lower in IgA nephropathy patients with C4d deposition than those without C4d deposition(P<0.001). Conclusion IgA nephropathy patients with C4d deposition have more se?vere clinical and pathological manifestations and lower rate of renal remission. Glomerular C4d deposition is expected to be an important pathologi?cal prognostic factor for predicting the prognosis of IgA nephropathy.

Transient high glucose induces persistent inflammatory status in rat glomerular mesangial cell via histone methylation modification / 中华肾脏病杂志

Objective To investigate whether the effect of transient high glucose on inflammatory factors expression could be continuous in rat glomerular mesangial cell,and its relation with histone methylation modification.Methods Rat glomerular mesangial cells (HBZY-l) were divided into three groups:the high glucose group (25.0 mmol/L glucose),the hypertonic group (MA,5.5 mmol/L glucose+ 19.5 mmol/L mannitol) and the normal-glucose control group (5.5 mmol/L glucose),which were cultured for 24 h respectively.All 3 groups were then changed with normal-glucose medium to culture for 24 h,48 h and 72 h.Their protein,mRNA and supernatant were harvested.The protein expressions of mono-methylation of H3 lysine 4 (H3K4mel) was measured by Western blotting,and the mRNA expressions of NF-κB subunit p65 and set7/9 were determined by real timequantitative PCR.The expression of monocyte chemoattractant protein 1 (MCP-1) and vascular cell adhesion molecule 1 (VCAM-1) were detected by enzyme-linked immunosorbent assay.Results (1)Compared with those in normal control group,the expressions of H3K4mel protein and set7/9 mRNA were first up-regulated in high glucose group,then gradually down-regulated in the following 48 h normal-glucose medium (as compared with those at 0 h,all P ＜ 0.05).At 72 h there was no statistic difference between high glucose group and normal control group (all P ＞ 0.05).(2) Compared with those in normal control group,the up-regulated p65 mRNA,VCAM-1 and MCP-1 sustained at least for 72 h in high glucose group.Conclusions Transient high glucose can induce persistent inflammatory factors expression in rat glomerular mesangial cells,which may via histone modification.

Abated microRNA-21 attenuates high glucose-induced autophagy inhibition in rat mesangial cells by PTEN/Akt/mTOR pathway / 中华肾脏病杂志

Objective To investigate the effects of abated microRNA-21 (miRNA-21) on phosphatase and tensin homologue on chromosome ten protein (PTEN) and PI3K/Akt/mTOR pathway,as well as their further influence on the autophagy in high glucose (HG,25.0 mmol/L) induced rat glomerular mesangial cells.Methods MiRNA-21 inhibitor and negative control were transfected by liposome 2000 into rat glomerular mesangial cells (HBZY-1).The cells were divided into normal glucose (5.5 mmol/L) group,normal glucose + negative control group,normal glucose +miRNA-21 inhibitor group,HG group,HG+negative control group and HG+miRNA-21 inhibitor group.Cell proliferation and hypertrophy were assayed by MTT and the ratio of total protein to cell number respectively.The miRNA-21 expression was detected using real time PCR.The expressions of PTEN/ Akt/mTOR signaling signatures,autophagy-associated protein (p62 and LC3 Ⅱ) and collagen Ⅰ was detected by Western blotting and real time PCR.Autophagosomes were observed using electron microscopy.Results Compared with those in normal glucose group,in HG group cells had hypertrophy and proliferation,up-regulated miRNA-21 expression,and down-regulated PTEN protein and mRNA expressions (all P ＜ 0.01).Also there were and up-regulated p-Akt,p-mTOR,p62 and collagen Ⅰ expression,and lower LC3 Ⅱ expression and autophagosomes (all P ＜ 0.01).Further,compared with those in HG group,cells hypertrophy and proliferation in HG+miRNA-21 inhibitor group were reduced,expressions of p-Akt,p-mTOR,p62 and collagen Ⅰ were down-regulated,while expressions of PTEN and LC3 Ⅱ and autophagosomes were up-regulated (all P ＜ 0.01).Conclusions MiRNA-21 inhibitor up-regulates PTEN expression,which inhibits the activation of Akt/mTOR signaling pathway,ameliorates cell hypertrophy,proliferation and enhances autophagy to reduce extracellular matrix accumulation.

A phase IV study of homoharringtonine, cytarabine, aclacinomycin and G-CSF (HCAG) regimen compared with traditional IA regimen in the treatment of newly diagnosed elderly acute myeloid leukemia patients / 上海交通大学学报（医学版）

Objective · To compare the efficacy and prognostic factors of HCAG regimen with traditional IA regimen in the treatment of newly diagnosed elderly acute myeloid leukemia (AML) patients. Methods · Forty-one patients with AML (aged 55-71 years) were randomly divided into two groups (Group HCAG and Group IA) between 2014 and 2016 for induction and consolidation therapy. Multivariate analysis was applied to identify prognostic factors for relapse-free survival (RFS). Results · A total of 29 patients (70.7%) achieved complete remission (CR). The estimated 2-year overall survival (OS) was 66.8% in Group HCAG and 75.4% in Group IA (P=0.913). The estimated 2-year RFS was 61.8% in Group HCAG and 49.1% in Group IA (P=0.411). Age remained as the unfavorable prognostic factor, leading to significant differences in OS and RFS. In addition, RFS was influenced by cytogenetic/molecular risk stratification. Conclusion · Although HCAG seemed not to particularly benefit the group, the dose reduction of anthracyclines may be applied in elderly patients with comparable short-time outcome. Furthermore, the introduction of homoharringtonine resulted in an improvement of treatment response for more than 20% compared with CAG regimen.

MicroRNA-148b influences high glucose-induced endoplasmic reticulum stress in rat mesangial cell by targeting AMPKα1 / 中华肾脏病杂志

Objective To observe the expression of microRNA-148b (miR-148b) induced by high glucose in rat mesangial cells,and to explore its effect on its target gene AMP-activated protein kinase α1 (AMPKα1) and extracellular matrix excretion.Methods Rat mesangial cells were divided ino 3 groups:normal glucose (NG,5.5 mmol/L glucose) group,hypertonic (MA,5.5 mmol/L glucose+19.5 mmol/L mannitol) group and high-glucose (HG,25.0 mmol/L glucose) group.MiR-148b expression was detected by real time PCR.Then miR-148b inhibitor was transfected to rat mesangial cells.Their protein expressions of AMPKα1,glucose regulated protein 78 (GRP78),C/EBP homologous protein (CHOP),fibronectin (FN) and collagen Ⅳ were detected by Western blotting.The expression of AMPKα1 mRNA was detected by real time PCR.The expression of collagen Ⅳ was also detected by immunofluorescence.Results Compared with NG group,HG group showed up-regulated miR-148bexpression,down-regulated AMPKαl mRNA and protein expressions,and up-regulated CHOP,GRP78,collagen Ⅳ and FN expressions (all P ＜ 0.05).HG-induced mesangial cells with miR-148binhibitor had up-regulated AMPKα1 mRNA and protein expressions,and down-regulated CHOP,GRP78,collagen Ⅳ,FN expressions as compared with HG-induced cells without miR-148b inhibitor (all P ＜ 0.05).Conclusions HG can up-regulate miR-148b expression and down-regulate AMPKα1 expression in rat mesangial cells,then activate endoplasmic reticulum stress to induce extracellular matrix excretion.MiR-148b inhibitor up-regulates AMPKα1 expression,inhibits endoplasmic reticulum stress and reduces extracellular matrix excretion.

MicroRNA-503 regulates high-glucose induced apoptosis of renal tubular epithelial cells by targeting Bcl-2 / 中华肾脏病杂志

Objective To investigate the expression vibration of microRNA-503(miR-503) and its effect on target gene Bcl-2,caspase enzyme activity and apoptosis of human renal tubular epithelial cells (HK-2) induced by high glucose,and to clarify the pathogenesis of renal tubular injury induced by high glucose.Methods HK-2 cells were cultured in normal glucose group (NG),mannitol hypertonic control group (MA),and high glucose group (HG).The morphology of apoptotic cells was observed using inverted microscope.The expression of miR-503 was determined using realtime quantitative PCR.The apoptosis rate of HK-2 cells was detected by Annexin V-FITC double dye using flow cytometry instrument.The expression of Bcl-2 and cleaved caspase-9 were detected by Western blotting.Results In the high glucose and mannitol groups HK-2 cell,an obviously increased apoptotic rate was observed under inverted microscope compared with normal glucose group (P ＜ 0.05).MA and HG up-regulated miR-503 expression (P ＜ 0.01),down-regulated anti-apoptotic protein Bcl-2 expression (P ＜ 0.05) and up-regulated cleaved caspase-9 (P ＜ 0.05).Conclusions The expression of miR-503 increases in HK-2 cells cultured by high glucose and mannitol.MiR-503 promotes apoptosis of HK-2 cells via activating mitochondrial apoptotic pathways and enhancing cleaved caspase-9 for Bcl-2 insufficiency.The tubular toxicity of high glucose is partly due to osmotic pressure.The miR-503 may be involved in diabetic tubular injury and may be a new therapeutic target of DN.

The efficiency of serum lncRNA GAS5/miR-21 as biomarkers in patients with diabetes and diabetic nephropathy / 中华肾脏病杂志

Objective To elucidate the efficiency lncRNA GAS5 and miR-21 as biomarkers in diabetes mellitus and diabetic nephropathy.Methods The patients were divided into three groups,diabetic nephropathy group (DN group proven by renal biopsy,n=25,14 males and 11 females),diabetes group (DM group,with normal urine albumin creatinine ratio,n=10,4 males and 6 females),and normal control group (NC group,n=9,4 males and 5 females).The expressions of lncRNA GAS5 and miR-21 in serum samples were detected by real-time quantitative PCR.The correlation between serum lncRNA GAS5 and miR-21 expressions and the clinical parameters was analyzed by T-test,Pearson,Spearman test and multivariate linear regression analysis.Differences of lncRNA GAS5 and miR-21 in different groups were analyzed by one-way analysis of variance.The ROC curve was used to analyze the diagnostic efficacy of lncRNA GAS5 and miR-21 in diabetes and diabetic nephropathy.All data were analyzed by SPSS 20.0 and GraphPad software,with P ＜ 0.05 as considered statistically significant.Results (1) The expression of serum lncRNA GAS5 was significantly down-regulated and serum miR-21 was significantly up-regulated in both diabetes mellitus and diabetic nephropathy patients compared to the NC group all (P ＜ 0.05).(2) In DN patients,the expression of serum lncRNA GAS5 was gradually up-regulated along with the increment of 24 h urinary protein.The expression of serum miR-21 was gradually up-regulated along with renal biopsy stage Ⅱb-Ⅲ of DN (P ＜ 0.05).(3)FBG and HbA1c were all negatively correlated with serum lncRNA GAS5 (P ＜ 0.05),and FBG was independently correlated with serum lncRNA GAS5 (P ＜ 0.05).Urine microalbumin,Total cholesterol (TC),Scr,Urea and SBP were all positively correlated with serum miR-21(P ＜ 0.05).Albumin (ALB)and estimated GFR (eGFR) were negatively correlated with serum miR-21(P ＜ 0.05),and ALB was independently correlated with serum miR-21 (P ＜ 0.05).(4) The diagnostic efficiency of serum lncRNA GAS5,miR-21 and lncRNA GAS5/miR-21 as "diagnostic signature" for DM were was good (P ＜ 0.05).(5) The diagnostic efficiency of serum miR-21 and lncRNA GAS5/miR-21 as "diagnostic signature" for DN were was good (P ＜ 0.05).Conclusions (1) Serum lncRNA GAS5 had good diagnostic efficiency in diabetes mellitus.The sensitivity of lncRNA GAS5/miR-21 for diagnosis of diabetes was 85.71％,and specificity was 88.89％.(2) The level of serum miR-21 can be used as a noninvasive diagnostic marker for diabetic nephropathy.

Correlation between quality of life and mineral metabolism in maintenance hemodialysis patients / 中华肾脏病杂志

Objective With multi?center investigation, to assess the life quality of patients with maintained hemodialysis (MHD) in Liaoning Province and to explore the relationship among the mineral metabolism, the life quality of the patients with MHD, and the repeated hospitalization within the latest three years. Methods 1192 patients with hemodialysis (at least 3 months) from January to March in 2015 at ten blood purification centers in Liaoning Province were selected for the cross?————————sectional survey. The Kidney Health?related Quality of Life (HRQOL) version 1.3 was used to evaluate the MHD patients' life quality. The total length of hospitalization was divided into four groups: 0 days, 3 to 15 days, 16 to 30 days and above 30 days. Results When serum calcium value ranged from 2.1 to 2.5 mmol/L, kidney?disease component summary (KDCS), mental component summary (MCS), physical component summary (PCS) and SF?36+KDCS corresponded to a higher value (P<0.05). When serum phosphorus value ranged from 1.13 to 1.78 mmol/L, KDCS and SF?36+KDCS corresponded to a higher value (P<0.05). When the calcium phosphorus product value ranged from 40.68 to 49.94, MCS corresponded to a higher value (P<0.05). KDCS showed a linear correlation with age (P<0.001), dialysis age, serum calcium (less than or equal to 2.5 mmol/L) (P<0.05); PCS showed a linear correlation with age (P<0.001) and dialysis age (P<0.05); SF?36+KDCS showed a linear correlation with age (P<0.001), and serum calcium (less than or equal to 2.5 mmol/L) (P<0.05), while age and dialysis age were negatively correlated. The hospitalization days showed a linear correlation with age, dialysis age (P<0.001) and serum phosphorus, calcium phosphorus product value (P<0.05), while dialysis age and calcium phosphorus product value were negatively correlated. Among different groups of total hospitalization days in three years, age, hemodialysis age, serum calcium, serum phosphorus, calcium?phosphorus product value and quality of life values were all statistically significant (P<0.05). Conclusions The life quality of patients with MHD were correlated with serum calcium, phosphorus, calcium and phosphorus product value, iPTH, dialysis age and age, while age and dialysis age were of negative correlation. The total number of hospitalization days in 3 years was closely linearly correlated with age and dialysis age, significantly correlated with serum phosphorus, calcium and phosphorus product value, while dialysis age, calcium and phosphorus product value were in a negative correlation. The total number of hospitalization in 3 years was correlated with the patients' age, dialysis age, serum calcium, serum phosphorus, calcium and phosphorus product value and quality of life.

Role of STAT3 in the regulation of autophagy in the glomerular mesangial cells of diabetic nephropathy / 中华肾脏病杂志

Objective To observe the changes of STAT3 signaling transduction pathway and autophagy activity in human glomerular mesangial cells cultured in high glucose, as well as the effect of STAT3 on autophagy, exploring whether SAT3 further influence extracellular matrix proteins type IV collagen secretion through the regulation of autophagy. Methods Culture human renal mesangial cells under different conditions, STAT3 pathway was inhibited with specific blocking agent S3I?201 and siRNA respectively. The experiment was divided into: (1) Control group: normal glucose concentration; (2) High glucose group: divided into 12 h, 24 h, 48 h, 72 h incubation group. (3) High glucose+S3I?201 group: pretreated cells with 30 μmol/L S3I?201 (Selleck S1155) for 1 h, then incubation with high glucose for another 24 hours. (4) High glucose+STAT3?siRNA group: siRNA transfection firstly, then incubation with high glucose for 24 hours. (5) High glucose+S3I?201+3?MA group: pretreated cells with 2 mmol/L 3?MA (Selleck S2767) and 30 μmol/L S3I?201 for 1 h, then incubation with high glucose for another 24 hours. Western blot was employed to detect the protein of STAT3, p?STAT3 and autophagy related protein LC3, p62 expressions. The changes of autophagosome quantity was observed with transmission electron microscope. The extracellular matrix protein collagen IV expression was measured with ELISA. Results Compared with the control group, glomerular mesangial cells cultured with high glucose for 24h, the expressions of STAT3 and p?STAT3 increased (P<0.01), while the expression of autophagy related proteins LC3II/LC3I decreased. The expression of p62 increased and the number of autophagosome reduced under transmission electron microscope, which all indicated the decrease of autophagy activity (P<0.05). Blocking STAT3 signaling pathway with S3I?201 and STAT3?siRNA respectively, compared with high glucose group, LC3II/LC3I was up?regulated and p62 was down?regulated, and the number of autophagosome was increased significantly, which all indicated the increase of autophagy activity (P<0.05). Extracellular matrix proteins collagen IV expression of cells cultured with high glucose was higher than the control group (P<0.05), and the application of S3I?201 blocking STAT3 pathway caused type IV collagen expression to decrease (P<0.05). The application of the autophagy inhibitor 3?MA could convert the result and lead to an increase of type IV collagen expression (P<0.01). Conclusions High glucose could active STAT3 signaling pathway of human renal mesangial cell and increase STAT3, p?STAT3 expression. High glucose could inhibit autophagy activity of human renal mesangial cells. Inhibition of STAT3 pathway activation may reduce the inhibitory effect of high glucose on autophagy of human renal mesangial cells. High glucose leads to an increase of type IV collagen secretion of human glomerular mesangial cells. The activation of STAT3 pathway may increase type IV collagen secretion through negative regulation of autophagy, which eventually leads to diabetic nephropathy.

Renal ultrasound elastography can reflect clinical-pathological changes in chronic kidney disease patients / 中华肾脏病杂志

Objective To analyze how is the elastography of renal tissue correlated to clinical biochemical indexes and pathological changes in patients with chronic kidney disease (CKD), and toexplore the potential of renal elastography to become a new noninvasive method available for the dynamic monitoring of renal disease progression, as well as its efficacy assessment and prognosis evaluation. Methods Patients admitted to the department of nephrology of the First Affiliated Hospital of China Medical University and received renal biopsy from August 2014 to January 2015 were selected. One hundred and thirteen cases of CKD patients, 61 males and 52 females were enrolled, including 23 cases of IgA nephropathy, 39 cases of membranous nephropathy, 15 cases of minimal change nephropathy and 7 cases of focal segmental glomerulosclerosis. The Young modulus of renal cortex and medulla (YMcortex and YMmedul a) were detected by Aix Plorer type full digital color Doppler ultrasound. The correlations between the YMs and clinical biochemical indicators in blood and urine, and the difference of YMs among different pathological changes in patients with CKD were analyzed by statistics. Results The YMcortex and YMmedul a in CKD patients were higher than those in the control group (all P<0.05); and with the progression of CKD, the YMcortex and YMmedul a gradually increased. The YMcortex in CKD G5 patients was higher than that in CKD G1?3 patients (all P<0.05). The YMmedul a in CKD G3?5 patients was higher than that in CKD G1?2 patients (all P<0.05). The YMcortex was correlated with systolic pressure, serum creatinine, cystatin C, serum albumin, serum phosphorus, calcium and phosphorus product, uric acid, intact parathyroid hormone (iPTH), urinary NAG, estimate glomerular filtration rate (eGFR) and hemoglobin (all P<0.05). The YMmedul a was correlated with systolic pressure, serum creatinine, serum albumin, uric acid, iPTH, urine microalbumin (MA), urinary NAG and hemoglobin (all P<0.05). Serum cystatin C (β=0.485, P=0.018) and uric acid (β=0.418, P=0.039) were independently correlated with the YMcortex. Serum creatinine (β=0.380, P=0.019), uric acid (β=0.482, P=0.004) and smoking (β=0.337, P=0.009) were independently correlated with YMmedul a. The YMcortex and YMmedul a in different pathological types were statistically significant (P<0.001, P=0.003). The YMcortex and YMmedul a in patients with membranous nephropathy and IgA nephropathy were higher than those in the patients with minimal change nephropathy (all P<0.05). The YMmedul a in patients with focal segmental glomerulosclerosis was higher than that in the patients with minimal change nephropathy (P<0.05). The YMcortex in the patients with phases Ⅳ and Ⅴ based on the Lee grading system of IgA nephropathy was higher than that in the patients with phases Ⅱ andⅢ (P<0.05). According the Oxford classification for IgA nephropathy, the YMcortex and YMmedul a in the T1 and T2 patients were higher than those in the T0 patients (P<0.05). The YMcortex and YMmedul a showed no statistically significant differences among different stages of membranous nephropathy. Conclusions The YMcortex and YMmedul a are associated with the progress of renal insufficiency, which may become new indicators for determining CKD progression. The renal ultrasound elastography may become a new non?invasive method for early diagnosing CKD, dynamic monitoring disease progression, and assessing efficacy and prognosis.

Ursolic acid inhibits high glucose-induced epithelial mesenchymal transition of podocyte by mediating Wnt/β-catenin signaling pathway / 中华肾脏病杂志

Objective To observe the epithelial mesenchymal transition (EMT) of podocyte induced by high glucose,and to explore the potential protective mechanism of ursolic acid (UA).Methods The podocytes cultured in vitro were divided into four groups:normal group (glucose 5.5 mmol/L),mannitol group (glucose 5.5 mmol/L+mannitol 19.5 mmol/L),high glucose group (glucose 25 mmol/L) and UA group (glucose 25 mmol/L + UA 5 μmol/L).Podocyte morphology changes were observed by inverted phase contract microscope.The expression of zonula occludens-1 (ZO-1) and α-smooth muscle actin (α-SMA) were detected by immunofluorescence.The expressions of β-catenin and glycogen synthesis kinase-3β (GSK3β) were detected by Western blotting.The expressions of Wnt1,Wnt3a,Wnt5a,Wnt5b and GSK3β were detected by real-time PCR.Results Podocytes showed irregular arborization shape in normal glucose and transited to longer cobblestone-like shape as mesenchyme cell by high glucose culture.Compared with normal group,the expression of ZO-1 protein was down-regulated and the expression of α-SMA was up-regulated by high glucose culture (P ＜ 0.05).The expression of Wnt5a mRNA was down-regulated;β-catenin mRNA and protein were up-regulated (P ＜ 0.05);and GSK3β protein was down-regulated by high glucose culture (P ＜ 0.05).Compared with high glucose group,ursolic acid inhibited podocyte EMT,up-regulated the expression of ZO-1 protein,Wnt5a mRNA,GSK3β (P ＜ 0.05),and down-regulated the expressions of α-SMA protein,β-catenin mRNA and protein (P ＜ 0.05).Conclusion Ursolic acid attenuates high glucose induced epithelial mesenchymal transition of podocyte by inhibiting Wnt/β-catenin signaling pathway.