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Objective This study was to observe the effects of different test conditions on the qualitative and quantitative detection of cryoglobulin .Methods We prepared 5 blood samples of different types of cryoglobulinemia . We detect the cryoglobulin qualitatively and quantitatively at different temperatures (37 ℃and room temperature of 20-25 ℃), and with different observation time (3 days and 7 days) and with different amount of blood (5 ml and 20 ml) .Further we will categorize the type of cryoglobulin and detect the components of cryoglobulin by immunofixation electrophoresis ( IFE) and other laboratory tests.Results (1) Blood samples from two groups were clotting and the serums were separated at 37 ℃ and room temperature respectively , and cryoglobulins of two groups were all qualitatively positive . Quantitative detection of cryoglobulins showed that the concentrations of cryoglobulins of room temperature group are lower than that of 37℃group;(2) Compared with 7 days, observing for only 3 days may lead to false-negative results in qualitative detection of cryoglobulin , and concentrations of cryoglobulin are also decreased;(3) Compared with 20 ml blood sample,5 ml blood sample is not enough for qualitative and quantitative detection of cryoglobulins .It may lead to false-negative results;(4) After purification, IFE and other laboratory tests can be used to categorize the types and find the components of cryoglobulins .Such examinations are helpful for finding the potential causes of cryoglobulinemia .Conclusions The positive of serum cryoglobulin is a key indicator of cryoglobulinemia .Detection of cryoglobulin can be affected by temperature, observed time and the blood volume for measurement .In addition, IFE and other laboratory tests are helpful for finding the type and the components of cryoglobulin .
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Objective To investigate the relationship of α-klotho protein and obesity related glomerulonephritis.Methods The chronic kidney disease (CKD) patients with or without ORG were diagnosed by renal biopsy.The normal and abdominal obesity control people were enrolled from the physical examination center.Propensity scoring analysis was done to balance the four groups of people in important clinical characteristics.The α-klotho levels in blood and urine were detected by ELISA.ORG mouse model was established and the mRNA and protein expression of klotho protein were detected by real-time quantitative PCR and Western blotting.Results (1) The plasma α-klotho levels decreased in ORG patients,CKD patients and abdominal obesity control people compared with normal control people [(251.7 ± 124.1) ng/L,(336.3 ± 126.1) ng/L,(377.1 ± 120.4) ng/L vs (472.3 ± 204.2)ng/L,all P ＜ 0.05].The ORG patients had the lowest plasma α-klotho levels (P ＜ 0.05).(2) ORG patients also had the lowest urine α-klotho levels compared with CKD patients,abdominal obesity and normal control people [(24.7±11.4) mg/mol vs (82.5±33.8) mg/mol,(74.5±32.5) mg/mol,(100.8±51.1)mg/mol,all P ＜ 0.05].There was no difference in urine α-klotho levels of CKD patients,abdominal obesity and normal control people.(3) Compared with the normal control mouse,ORG model mouse showed decreased mRNA and protein expression of α-klotho protein in renal tissue.Conclusion The lower plasma and urine α-klotho levels in ORG patients may be due to the reduced expression of α -klotho protein in kidney.
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Objective To examine whether aldosterone contribute to obesity related glomerular disease. Methods C57BL/6J mice were randomly divided into three groups: a control group (low?fat?diet, n=10), a model group (high?fat?diet, n=10) and a intervention group (high?fat?diet, n=12). After 8 weeks intervention group were treated with a mineralocorticoid receptor antagonist, spirolactone (SPL).The physicochemical indexes and the renal pathology of the three groups were all detected. The mRNA and protein expressions of podocyte marker protein were determined by real?time PCR and Western blotting, respectively. Results Compared with the control group, body weight, kidney weight, Lee ’s index, fat index, blood cholesterol, blood triglyceride, creatinine clearance rate, urinary protein excretion, glomerular average diameter, glomerular foot process average width were significantly up ? regulated (P<0.05); The mRNA and protein expression of nephrin, podocin, podoplanin and podocalyxin were significantly down?regulated in model group (P<0.05). Meanwhile, these changes were attenuated by SPL. Conclusion Aldosterone can participate in the process of obesity? related renal injury, and these can be attenuated by mineralocorticoid receptor antagonist, spirolactone. This gives us preliminary clues to treat ORG.
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Objective To explore whether the glomerular podocytes can be damaged by aristolochic acid. Methods Thirty-two male SD rats were equally divided into the following 2 groups:model group in which the rats received the extract of Aristolochia manshuriensis Kom (AmK) by gavage; control group only received tap water by gavage.24 h urinary protein excretion was measured at the end of the 1st and 4th week,and SDS-PAGE gel electrophoresis was performed to detect the protein in urine.At the end of the 4th week,all the rats were sacrificed and the glomeruli were isolated by laser capture microdissection technique.The mRNA expression of nephrin,podocin,CDA2P,podocalyxin and podoplanin in isolated glomeruli was determined by RT-PCR,and the average width of glomerular foot process was measured by electron microscopy and image analysis. Results At the end of the 4th week,24 h urinary protein excretion in the model group was significantly higher than that in the control group (P＜0.01) and the urinary albumin content in model group was also obviously increased.The average width of glomerular foot process in the model group was significantly larger than that in control group (P＜0.01).The mRNA expressions of nephrin,podocin,CDA2P,podocalyxin and podoplanin in glomeruli were significantly down-regulated in the model group compared with the control group,which decreased by 34％,62％,56％,50％(P＜0.01) and 27％ (P＜0.05),respectively. Conclusions Aristolochic acid can damage the glomerular podocytes,resulting in the down-regulation of nephrin,podocin,CD2AP,podoplanin and podocalyxin mRNA expression, the segmental widening of foot process, and increased urinary protein excretion.
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Objective To investigate the protective effects of Wnt-7a protein on renal interstitial fibrosis in mice of unilateral ureteral obstruction (UUO)model.Methods Eighteen male C57BL/6 mice were randomly divided into 3 groups:sham-operation group,the UUO model group and Wnt-7a treatment group.The body weight of mice was measured everyday.All the mice were sacrificed at thc seventh day after the operation.The left kidney was taken for histology evaluation and molecular biology assay.Masson's stain was performed as a main indicator of interstitial fibrosis.The expression of vimentin,α-smooth muscle actin,and E-adherin in renal tissue was detected by immunohistochemistry staining and the expression of α-smooth muscle actin and E-cadhe(nn) in renal tissue was detected by Western blotting.Results Compared with sham-operation group,body weight of the (,)odel group was significantly lower (P＜0.05),and the relative area of interstitial fibrosis was significantly larger (P＜0.05).Furthermore,the expression of vimentin and α-SMA was significantly up-regulated (P＜0.05),and the expression of E-cadherin was significantly down-regulated (P＜0.05).Compared with model group,all the above-mentioned abnormalities were restored to some extent and showed significant differences (P＜0.05) in Wnt-7a treatment group.Conclusion Wnt-7a protein can decrease the interstitial fibrosis by inhibiting epithelial to mesenchymal transition in UUO mice.
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Objective To investigate the effects of adiponectin on angiotensin Ⅱ-induced extracellular matrix production of mesangial cells(MCs) and its possible signaling pathway.Methods RT-PCR and indirect immunofluorescence examination were performed to detect the adiponectin receptors in MCs.Quantitative real-time PCR and enzyme-linked immunosorbent assay (ELISA) were used to observe the effects of adiponectin on angiotensin Ⅱ-induced transforming growth factor β1(TGF-β1) and fibronectin production of MCs.Western blotting was used to measure the ratio of p-AMPK to total AMPK.Results(1)Adiponectin receptors 1 and 2 were found in MCs. (2)The up-regulated mRNA and protein expression of TGF-β1 and fibronectin in MCs induced with 10-7 mol/L angiotensin Ⅱ (Ang Ⅱ) was significantly inhibited by 10 mg/L adiponectin (P＜0.05).(3)The p-AMPK/AMPK ratio was significantly increased after incubation with adiponectin for 15 min and 30 min(vs 0 min, P＜0.05), which suggested that adiponectin could activate the AMPK signaling pathway in MCs.The activation of AMPK signaling pathway was blocked by 40 μmol/L compound C, a specific inhibitor of AMPK. (4)The inhibitory effects of adiponectinonangiotensin Ⅱ-upregulatedTGF-β1andfibronectinexpressioninMCswere significantly relieved by 40 μmol/L compound C (P＜0.05).Conclusions There are adiponectin receptors 1 and 2 in MCs.Adiponectin has inhibitory effects on the angiotensin Ⅱ-upregulated TGF-β1and fibronectin expression in MCs.AMPK signaling pathway may play an important role in the effects of adiponectin above-mentioned.
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Objective To investigate the prevalence and risk factors of chronic kidney disease (CKD) in the general adult population in the Hulunbeir Prefecture, Inner Mongolia Autonomous Region where many minorities of north China live. Methods Sampling surveywas performed in the residents aged 20 years and older in the Hulunbeir Prefecture. All the investigated subjects were tested for urinary albumin to creatinine ratio (ACR); hematuria by microscopy of urinary sediment; and GFR estimated by modified MDRD equation for Chinese adults (eGFR). The related risk factors of CKD were also investigated. Results A total of 4522 subjects were enrolled in the study. The prevalence of albuminuria was 7.11％, hematuria was 2.64％ and reduced eGFR [60 ml-min-1·(1.73 m2)-1] was 2.75％. The prevalence of hypertension was 38.90％; hyperglycemia 6.61％; hyperlipidemia 2.72％; increased waist 24.79％ and metabolic syndrome 15.02％. After the subjects with combined microalbuminuria, hematuria and reduced eGFR were excluded, the prevalence of CKD was 12.95％. Logistic regression analysis and stratified analysis showed increased age, increased waist, elevated systolic pressure, hyperglycemia,hypertriglyceridemia and metabolic syndrome were independently associated with albuminuria;increased age, elevated systolic pressure and hyperglycemia were independently associated with reduced eGFR; increased age was independently associated with hematuria. Conclusions The prevalence of adult CKD is 12.95％ in the Hulunbeir Prefecture, Inner Mongolia Autonomous Region. Independent risk factors of CKD include increased age, increased waist, hypertension,abnormal blood glucose or lipid, and metabolic syndrome.
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Objective To study the pathogenesis of anemia in chronic aristolochic acid nephmpathy(CAAN) rats. Methods The hemoglobin(Hb)values of sixty-two male SD rats were assayed to determine its normal range.Among them,24 rats with normal Hb value were randomly divided into 2 groups:model group (MG)in which rats received the extract of Aristololochia manshuriensis Kom (AmK) by gavage,and control group (CG) received tap water only by gavage.Body weisht(BW),Hb,24 h urinary protein excretion(UP)and creatinine clearance (Ccr)of 6 rats in each group were measured before administration and at the end of the 8th week, respeetively.then these rats were sacrificed.The relative area of renal interstitial fibrosis was measured by microscopy.The mRNA expression of erythropoietin (EPO)in kidney tissue Was determined by real-time RT-PCR;protein expression of type I collagen(Coll),aminopeptidase P (APP),hypoxia indHeible factor let and 2α(HIF-1α and HIF-2α)in kidney tissue Was examined by immunohistochemistry staining. Results Hb values of normal rats presented normal distribution. The normal Hb was (155.9±16.5) g/L. Rat anemia was diagnosed when Hb was below 123.6 g/L. There was no difference in all the examination results between CG and MG before administration (P>0.05). Compared with CG, the Hb and Cer in MG were significantly decreased [(121.66±15.68) g/L vs (169.00±12.89) g/L, (0.63±0.13) ml/min vs (1.27±0.18) ml/min, P< 0.01], and the UP in MG was significantly increased at the end of the 8th week [(27.04±9.40) mg/d vs (6.11±0.84) mg/d, P<0.01]; the relative areas of fibrosis and Col l in renal interstitium of MG were significantly enlarged [(12.89±2.33)% vs (0.55±0.10)%, (13.92±2.92)% vs (1.32±0.84)%, P<0.01]; the protein expression of APP and the mRNA expression of EPO in the kidney tissue of MG were significantly down-regulated [(0.55±0.23)% vs (3.77±1.06)%, 0.005±0.001 vs 0.032±0.013, P<0.01]; the protein expression of HIF-lα and HIF-2α in the kidney tissue of MG was significantly up-regulated (2.55±0.16 vs 1.12±0.46, 2.33±0.33 vs 1.15±0.27, P<0.01), at the end of the 8th week. Conclusions The pathogenesis of anemia in CAAN may be due to the decreased production of EPO caused by the destruction of peritubular capillary. The compensatory up-regulation of HIF-lα and HIF-2α expression can not prevent the anemia development.
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Objective To study whether Hirsutella sinensis (HS) can antagonize aristolochic acid (AA) induced fibrogenesis on human proximal tubular epithelial cells (HKC). Methods HKC were incubated with medium alone, medium with HS 10 mg/L, medium with AA-Na 40 mg/L or medium with AA-Na 40 mg/L and HS 10 mg/L, respectively. After 12 h (for mRNA) or 36 h (for protein) ,cells were lysed,and the mRNA and protein expression level of transforming growth factor-?1 (TGF-?1), connective tissue growth factor (CTGF), tissue inhibitor of metalloproteinase-1 (TIMP-1)and plasminogen activator inhibitor-1 (PAI-1) of HKC were measured by RT-PCR, ELISA (for TGF-?1, TIMP-1 and PAI-1) and Western blotting (for CTGF), respectively. Results The mRNA and protein expression of TGF-?1, PAI-1, CTGF and TIMP-1 were significandy up-regulated by AA-Na 40 mg/L. Compared with the control group, the mRNA expression of TGF-?1,.CTGF, TIMP-1 and PAI-1 was up-regulated to 1.24,1.31,1.27 and 1.36 times,respectively (P