Clinical evaluation of assisted diagnosis of early diabetic kidney disease by serum C1q,NGAL and CysC / 中华检验医学杂志

Objective To investigate the diagnostic value of serum complement component 1 q (C1q), cystatin C (CysC), neutrophil gelatinase-associated lipocalin (NGAL), creatinine (SCr) and urea (SUr) for early diabetic kidney disease.Methods A retrospective study was conducted in 530 subjects who visited or received physical examination in Mianyang Central Hospital from March 2017 to November 2017. Of these, 229 patients with diabetes mellitus (DM group) were divided into simple diabetes mellitus group (SDM group,n=100) and early diabetic kidney disease group (EDKD group,n=129) respectively, and 301 subjects were selected as the healthy control (HC group).The levels of serum C1q, CysC, NGAL, SCr and SUr were detected in all subjects .Then, the diagnostic values of single and combined detection of these indexes for early diabetic kidney disease were evaluated .The statistical difference among the observed indexes of each group were compared by one-way ANOVA, and the diagnostic performance of those indexes were evaluated by receiver operating characteristic curve ( ROC) analysis.Results The median of serum C1q,CysC,NGAL and SCr in early diabetic kidney disease group were 187.80 mg/L, 1.02 mg/L, 148.70μg/L and 72.00μmol/L.The simple diabetes mellitus group were 177.00 mg/L, 0.80 mg/L, 116.20μg/L and 55.35 μmol/L.The healthy control group were 163.60 mg/L, 0.75 mg/L, 104.20μg/L and 59.40μmol/L. Compared with simple diabetes group ( t1 values were 6.50, 9.44, 3.87,9.19 and 4.34, P<0.05)and healthy control group(t2 values were 7.26, 12.54, 5.86,6.16 and 6.49, P<0.05),the levels of serum C1q, CysC, NGAL, SCr and SUr in diabetic kidney disease group were significantly increased . According to the ROC curve analysis , the diagnostic performance of CysC ( AUC=0.837 ) was much higher than those of C1q(AUC=0.731), NGAL(0.742)and SCr(AUC=0.720)for early diabetic kidney disease by using single detection.Otherwise, combined the diagnostic performance of joint detection reached the better when combined C1q and CysC (AUC =0.890,Se =79.1％, Sp =86.5％, YI =0.656).While increase detection of NGAL (z=1.817, P>0.05) or NGAL+SCr (z=1.806,P>0.05) did not further improve the diagnosis performance of early diabetic kidney disease .Conclusions C1q, CysC and NGAL with equal diagnostic performances are superior to SCr and SUr in diagnosis of early diabetic kidney disease . Among them, the combined detection of C1q and CysC has better diagnostic performance for early diabetic kidney disease.

Recent advances of metabolomics techniques for diagnosis and treatment of kidney disease / 中华检验医学杂志

Kidney disease(KD)is a major challenge for global public healthcare systems. Traditional biomarkers(such as serum creatinine and urea)for measuring kidney function are not sufficient sensitivity and specificity.They are not only affected by many factors,but also were only increase significantly in severe KD.Therefore,more sensitive biomarkers for evaluation of KD is essential. Metabolomics is a promising tool that identify non-targeted,global small-molecule metabolite profiles of complex samples,such as biofluids and tissues extract.The application of metabolomics in KD studies has developed rapidly in the past years.Many benefits have been shown from the use of metabolomics in identifying biomarkers for KD.In particular,metabolomic approaches have the potential to diagnose KD with a higher accuracy than traditional diagnostic methods.Metabolomics in KD research has been expanded from experimental study into clinical applications.

Determination of Sulfurous Anhydride in Rhizoma Dioscoreae by Ion Chromatography-Direct Extraction / 中国药师

Objective:To establish a quantitative analysis method to determine the content of sulfurous anhydride in Rhizoma di-oscoreae by ion chromatography-direct extraction. Methods:Sulfurous anhydride was extracted by KOH solution (25 mmol·L-1). An IonPac? AS11-HC column(250 mm × 4 mm, 9. 0 μm) was used. The column temperature was 20℃, the eluent was KOH solution (20 mmol·L-1 ) at flow rate of 1. 00 ml·min-1 and the conductivity temperature was 20℃. Results:There was a good linear rela-tionship between the injection quantity (1.160-29.100 μg)and the peak area of sulfite(r =0.999 9). The average recovery was 98. 9%(RSD=0. 6%, n=9). The quantitation limit was 1. 38 ng·ml-1. Conclusion: The method is simple, accurate and rapid, which is appropriate for the quantitative analysis of sulfite anhydride in Rhizoma dioscoreae.

Current problems and solution for urinary albumin measurements / 中华检验医学杂志

Measurement of urine albumin plays a vital part in the early detection and appropriate management of chronic kidney disease. However, urinary albumin measurements have yet not been standardized due to lack of a reference method and its reference material.It is very important for laboratory technicians to be aware of biochemical characteristics of urinary albumins, laboratory techniques, urine sampling methods, reference interval setting, and units of reporting.

Effectiveness analysis of determining the content of notopterol and isoimperatorin in notopterygium / 重庆医学

Objective To establish the effect of determining the content of notopterol and isoimperatorin in notopterygium from different habitat and harvest time by HPLC method .Methods HPLC instrument was used ,Waters symmetry C18(4 .6 mm × 250 mm ,5 μm) was adopted with a mobile phase of acetonitrile‐water (45∶55) ,the separation was carried out at a flow rate of 1 mL/min ,column temperature was 30 ℃ and the detection wavelength was 310 nm .Results Notopterol and isoimperatorin showed good linearity in the ranges of 0 .240 8-1 .505 0 μg and 0 .117 6-0 .735 0 μg with average recoveries rate (n=6) of 0 .98% and 1 .42%respectively .The content of notopterol and isoimperatorin in crude drugs of notopterygium from different habitat marked differ‐ence ,and harvest period and growth years made effect to content of two components .Conclusion HPLC method is easy and fast , and can be used as a basis for confirming the best habitat and harvest period of notopterygium .

Biomarkers of acute kidney injury in clinical laboratory / 中华检验医学杂志

Acute kidney injury ( AKI) is a common and serious clinical problem.Serum creatinine (SCr) and urine output are still used as the most common biomarkers for diagnosis of AKI.However, neither of them is an ideal indicator for diagnosis of AKI.In recent years , a number of potential biomarkers of AKI with more favorable test characteristics than SCr have been identified and studied in a variety of experimental and clinical settings.This review describes the novel biomarkers of AKI , including NGAL , IL-18, KIM-1, Cys C and L-FABP, which have now progressed to the clinical phase.

Validation study of chronic kidney disease epidemiology collaboration predictive equations based on serum creatinine and cystatin C in Chinese chronic kidney disease patients / 中华检验医学杂志

Objective To validate the four chronic kidney disease epidemiology collaboration (CKD-EPI) predictive equations based on serum creatinine (SCr) and cystatin C (Cys C) in Chinese CKD patients,and try to develop the GFR predictive equations for Chinese CKD patients.Methods254 CKD patients were randomly selected from four Grade ⅢA hospitals in different regions in China from September 2007 to December 2010.Clearance of dual plasma sampling 99mTc-DTPA was used to measure glomerular filtration rate (rGFR) in 254 CKD patients.The serum concentration of Cr and Cys C were measured.CKD-EPI SCr equation,Cys C equation,Cys C equation adjusted for age,sex and race,SCr/Cys C combinated equation adjusted for age,sex and race were used to estimate GRF ( labeled as eGFR1,eGFR2,eGFR3 and eGFR4,respectively).The correlation,bias and precision of eGFRs were compared with rGFR by Wilcoxon signed rank test,intraclass correlation coefficient (ICC) and Spearman correlation analysis.The deviation degree between rGFR and different eGFRs was compared via Bland-Altman graph.The accuracy within 15％,25％,30％ ( P15,P25,P30) and the staging correctness of eGFR against CKD at different stages was calculated.ResultsThe rGFR in 254 CKD participants was [ 48.07 (26.19 -92.97 )] ml · min -1·(1.73 m2) -1.The Spearman correlatiou coefficients (CC) of eGFR and rGFR varied within the range of 0.873 - 0.896 ( P =0.000 ).The intra-class CC ( ICC ) varied within the range of 0.920 - 0.942.The correlation of eGFR4 was the best.The absolute deviations of 4 eGFRs and deviation precision were eGFR4 ＜eGFR3 ＜ eGFR2 ＜ eGFR1.The 95％ confidence intervals for the regression line of 4 eGFRs shown by Bland-Altman graphs were 92.5,87.3,83.0 and 76.1 ml · min-1 · ( 1.73 m2 ) -1,respectively,with the best result of eGFR4.For P30,the correctness of 4 eGFRs were eGFR4 ＞ eGFR3 ＞ eGFR2 ＞ eGFR1,but no significant difference was found by Chi square test (x2 =6.448,P =0.092).The overall correctness rate in 4 eGFRs against CKD stages were 48.4％ -57.5％,with the highest consistency of eGFR4,but their staging correctnessratewerenotideal(Kappa values were 0.405,0.348,0.366 and 0.463,respectively).Conclusions Compared with CKD-EPI SCr equation,no advantage was found in CKD-EPI Cys C equation.The Cys C equation adjusted by age and sex shows a little advantages over CKD-EPI Cys C equation in bias,precision,correlation and accuracy.The CKD-EPI SCr/Cys C combinated equation adjusted by age,sex and race has advantage over other three equations not only in bias,precision,correlation and accuracy,but also in staging correctness.However,the validation of this equation is still not fairly ideal for Chinese CKD patients.Based on these findings,it is essential for the Chinese CKD patients to develop SCr/Cys C combined predictive equation which adjusted by age,sex or other factors.(Chin J Lab Med,2012,35:798-804)

Yictims'Serum electrolyte disturbances in sufferers during the Wenchuan earthquake in China / 国际检验医学杂志

Objective To study the electrolyte disturbances in the sufferers during the Wen-chuan earthquake in China,and investigate their possible pathomechanism. Methods Serum concentra-tions of potassium (K), sodium (Na), chlorine (Cl), calcium (Ca), magnesium (Mg) and inorganic phosphate (P) were determined in four hundred sufferers. Their values were analyzed via statistical method. Results Significant differences of all determined six electrolytes were observed among various groups. Pearson correlation analysis showed that correlative changes among electrolytes were similar in various groups except the correlation coefficients of Na-Mg in dialysis group (r= 0.066, P = 0.546) and non-dialysis group (r=-0.433, P= 0.044). Multivariate logistic analysis revealed the highest risk factor was K+,which led to crush injury (OR= 28.037, P= 0.000) and crush syndrome (OR=6.434,P=0.000). The receiver operating characteristic (ROC) curve revealed that K+ was the most specific (specificity: 98.5%; cutoff value: 5.8 mmol/L),Ca2+ was the most sensitive (sensitivity: 81.1%; cutoff value: 1.75 mmol/L) for crush injury diagnosis. The specificity of serum K+ , Mg2+ amd P was comparative in diagnosis of crush syndrome. The sensitivity of Ca2+ was the highest,ac-counting for 75.3%. Conclusion Electrolyte disturbances of suffers occur during a natural disaster; serum electrolyte measurement contributes to correct diagnosis and severity assessment.

Establishment of enzymatic kinetic assay of serum argininosuccinate lyase activity and its preliminary clinical application / 中华检验医学杂志

Objective To establish a continuous monitoring assay of serum argininosuccinate lyase (ASL) activity with automatic biochemistry analyzer, and perform methodology validation and preliminary clinical application.Methods According to the chemical reaction catalyzed by ASL and the working characteristics of automatic biochemistry analyzer, an enzyme coupled reaction system with high specificity was set up, and the methodology validation was performed.Three hundred and nine patients with various liver diseases, 269 non-liver disease patients and 40 healthy controls were enrolled in this study.Serum ASL, ALT, and AST level were determined in all subjects.Results A new kinetics assay of ASL activity was set up with automatic biochemistry analyzer.The methodological validation demonstratod that inter-assay and intra-assay coefficient of variation were 4.0% and 5.9% respectively and the mean recovery was 100.5%.The linear range was 0-167.7 U/L.The lowest detection limit was approximately 0 U/L.The interference test showed that there is no significant interferences while the concentration of bilirubin is less than 342 μmoL/L or commonly used anticoagulants is employed at their routine concentrations.However,interference was significant when Hb level is more than 0.06 g/L.Preliminary study of clinical application showed that there was no significant difference of serum ASL level between non-liver disease group and healthy group ( q = 0.027, P = 0.979 ), but there was significant differences for both serum ALT and AST levels (ALT:q =6.461,P =0.000;AST:q =6.481,P =0.000).Conclusions A continuous monitoring assay for the determination of serum ASL activity is successfully established. Serum ASL may be a good biomarker for liver injury.

Correlation between oxidative stress and trace elements in blood of patients with cancer / 中国组织工程研究

BACKGROUND: The occurrence and development of tumor may result from the reaction of a series of factors. Up to now, earlier detection, monitoring treatment, and prognosis assessment for tumor are very difficult. It can provide the theoretical foundation of rehabilitation and survival for cancer patients to study the mechanisms of occurrence and development of tumors, or investigate the changes of blood level and tissue distribution.OBJECTIVE: To investigate the changes of levels of some trace elements and serous prooxidant-antioxidant system of cancer patients.DESIGN: A controlled observation trialSETTING: Department of Laboratory Medicine, Mianyang CentralHospital.PARTICIPANTS: From September 1999 to December 2000,111 cancer patients were selected from the Third People's Hospital of Zigong, Yibin First People's Hospital of Sichuan Province, and Longchang County people's Hospital of Neijiang City, Sichuan Province.Of them, 21 cases were of liver cancer, 16 of gastric carcinoma, 15 of colorectal cancer, 11 of breast cancer, 13 of lung cancer, 13 of esophageal cancer, 7 of brain cancer and 15 of other cancers. At the same time; 36 control subjects were recruited from those who came to the hospital for health examination.METHODS :Blood samples of subjects after overnight fasting were pre pared by collecting venous blood in 5-mL Vacutainer tubes (Becton Dickinson, American). After 2-3 hours, the blood samples were centrifuged at 3 000 r/min for 15 minutes. 2.0 to 3.0 mL of serum was separated. The sera were stored in the refrigerator until analysis. Serum glutathione(GSH)level was determined by dithiobis-2-nitrobenzoic acid colorimetry. Xanthine oxidase (XOD) concentration was determined with xanthine-nitrotetrazole colorimetry. Glutahione oxidase (GSHPx) activity was determined with dinitrobenzoic acid colorimetry. Malondialdehyde(MDA) concentration in serum was measured with thiobarbituric acid reaction. Vitamin C (Vit C),vitamin E(Vit E) and total antioxidant power(TAOP) levels were measured with phenanthroline colorimetry. Albumin concentration was determined with bromcresol purple. Serum transferrin (Trf) and ceruloplasmin (CER)concentrations were determined with immunonephelometry. The concentrations of Cu, Zn, Fe, and Se were measured with flame atomic absorption spectroscopy.MAIN OUTCOME MEASURES: MDA (as lipid peroxidation marker),trace elements copper, zinc, iron, and selenium and their transport proteins (albumin, transferrin and ceruloplasmin); antioxidation status markers: the concentrations of XOD, GSH, GSHPx, vitamin C, and vitamin E, and total antioxidant power levels.RESULTS: It was shown that lipid peroxidation ( measured as MDA)was significantly higher in cancer patients than in healthy controls [(5.21±1.05) nmol/L vs (4.04±0.68) nmol/L,P ＜ 0.001], and the TAOP level was significantly decreased in cancer patients than in the health controls[(4.34±0.980) U/L vs (5.87±0.93) U/L,P ＜ 0.001]. There were not obvious changes of antioxidation components (XOD, GSH, GSHPx, vitamin C,and vitamin E). Serum albumin concentration was found to be significantly lower in the cancer patients than in the health controls [(34.19±6.94)g/L vs (42.34±4.89) g/L ,P ＜ 0.001], and serum ceruloplasmin concentration was found to be significantly higher in the cancer patients than in the health controls [(0.371 ±0.031) g/L vs (0.346±0.026) g/L,P ＜ 0.05] butserum transferrin concentration remained unaltered (P ＞ 0.05). As com pared with the healthy controls, serum copper level was significantly increased[(19.27±4.74) μmol/L vs (14.29±2.71) μmol/L, P ＜ 0.001], serum selenium levels was significantly decreased[(1.175±0.333 0 μmol/L vs (1.413±0.446) μmol/L,P ＜ 0.001)]. However, the concentrations of zinc and iron remained unchanged. Correlation was observed between copper and MDA levels (r=0.281, P=0.003) in the cancer patients but not in the healthy controls. Moreover, a correlation was also observed between serum iron and MDA levels in the patients with liver cancer (r=0.680,P=0.001).CONCLUSION: The presence of an association between oxidative stress and some trace elements was found in cancer patients; however, the results are possibly inconsisteut because of different cancer types, cancer grades,or other characteristics of the patients engaged in the test.

Alteration of serum Cystatin C concentrations in patients with hepatopathy infected with hepatitis B or C virus / 中华检验医学杂志

Objective To investigate the clinical application of Cystatin C as a biological marker for monitoring hepatic pathological change in patients with virus hepatitis.Methods Two hundred and seven patients infected with hepatitis B or C virus(HBV, HCV)were divided into cirrhosis group(group A),chronic HBV group(group B),chronic HCV group(group C),and liver cancer group(group D). 32 healthy controls(group H) were recruited . The serum TIMP-1,TIMP-2,and Cystatin C as well as some traditional markers for monitoring liver function and renal function including creatinine, creatinine clearance rate, alanine transaminase, and aspartate transaminase were determined.Results In these groups, serum Cystatin C(F=28.334, P

Prokaryotic expression, purification, identification of human cystatin C and preparation of its antiserum / 第三军医大学学报

Objective To construct a prokaryotic expression vector of cystatin C (Cys C), purify Cys C protein produced by the expression system, and prepare its antiserum. Methods Total RNA was isolated from HL-60 cells, and human Cys C gene was amplified with RT-PCR. The cDNA fragment was cloned into pMD18-T vector and which was confirmed by sequencing. The enzyme-digested target fragment was cloned into PET-32(a) expression vector and transfected into E.coli. BL 21(DE3), in which Cys C expression was induced. After the inclusion body protein was purified through Ni2+ affinity chromatography, processed by dialysis, identified by Western blotting, a rabbit was immunized with the fusion protein, and the antiserum was obtained. Results The result of DNA sequence analysis showed that the cloned Cys C gene sequence was completely corresponding to GenBank data. SDS-PAGE and Western blotting showed that the expressed Cys C fusion protein was about 35?103, mainly existing in the inclusion body of E.coli., that could be purified through Ni2+ affinity chromatography. The titer of the antiserum to the purified protein was 1∶8 000 by ELISA, and Western blotting confirmed that the antiserum reacted specifically to the Cys C protein. Conclusion A recombinant Cys C protein and the specific polyclonal antibody have been obtained, which provides a basis for establishment of immunoassays of human Cys C.

Preparation,characterization and preliminary application of monoclonal antibody against cystatin C / 中国免疫学杂志

Objective:To prepare monoclonal antibodies(McAb)against cystatin C(Cys C)and to establish the particle enhanced turbidimetric immunoassay(PETIA)for determining human serum Cys C.Methods:The prokaryotic expression vector pET32a(+)/Cys C was constructed and Cys C expression was induced.McAbs against Cys C were prepared with the hybridoma technique after mice were immunized with the purified recombinant protein.Then the McAbs were covalently attached to uniform microparticles,PETIA method for determination of human serum Cys C was established,and primary evaluation tests of methodology were performed.Results:Three hybridoma cell lines were obtained successfully,the secreted antibodies were isotype of IgG1,and Western blot confirmed that the antibodies reacted specifically to the Cys C protein.After one of the hybridoma cell lines was injected into mice abdominal cavity,the ascites abundant for McAb was obtained.The titer of the McAb against the purified protein was 1∶4?106.With the self-made McAb,PETIA for human serum Cys C was established.The primary evaluation tests of methodology revealed that self-established PETIA method had a satisfactory performance,which was equal to the import kit.Conclusion:The prepared McAb against Cys C is prepared,which could be used to establish PETIA for determining human serum Cys C.