ABSTRACT
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.
ABSTRACT
Objective To establish a recombinant retroviral vector containing insulin-like growth factor-1(IGF-1) gene and to provide the basis for the application of IGF-1 in treating nervous system diseases such as stroke.Methods The plasmid pcDNA3.1-IGF-1 was cut by EcoR I/Xho I,and subcloned to retroviral vector pLXSN,resulting in the recombinant plasmid(pLXSN-IGF-1.)The recombinant IGF-1 expression vector was evaluated by using enzyme cutting and sequencing.By the Lipofectamine 2000,pLXSN-IGF-1 was transferred to packaging cell line-pA317.Culture supernatant of these cells was detected for titration of the recombinant virus.Results The two fragments from recombined IGF-1 eukaryotic expression vector by EcoR I and Xho I represented 400?bp and 6.0?kb by agarose electrophoresis,and PCR showed positive fragment which was about 400?bp long,and sequence analysis showed the same sequence as expected.The cell line pA317-IGF-1 was established,and average virus titer of the recombinant virus in the culture supernatant was about 6.5?10~5 CFU/ml.Conclusion A recombinant retroviral containing IGF-1 gene was successfully constructed.
ABSTRACT
Objective To observe the influences of ?-amyloid peptide(A?) on the proliferation and apoptosis of neural stem cells(NSCs) and to investigate the protective mechanism of insulin-like growth factor-1(IGF-1) from toxicity of A?.Methods NSCs were isolated from E14 SD rats brains and then treated with IGF-1,A? and IGF-1 plus A? respectively.The cells were dyed with trypanblue for the count of dead cells and the death rate.BrdU was used to label cells and to evaluate cell proliferation.NSCs and newborn cells were identified by immunocytochemistry and apoptosis was detected by TUNEL technique.Results Cell death rate was low and there were many BrdU-positive cells,but no TUNEL-positive cells in the medium containing IGF-1.Cell death rate increased strikingly 6-48 hours after A? was added,and a great many TUNEL-positive cells were observed in the group treated with A?.Death rate of cells and TUNEL-positive cells decreased more markedly in the medium containing both IGF-1 and A? than that containing A? only.Conclusion A? promotes death and apoptosis of NSCs,while IGF-1 accelerates the proliferation of NSCs and inhibits NSCs apoptosis induced by A?.