ABSTRACT
Objective To explore the significance and the change of miRNAs expression profile in different period of HBV infection.Methods Establish the detection method of microRNA(miRNA) by RNA DNA probe liquid chip (flexible multi-analyte profiling,xMap) and estimate the specificity,repeatability and accuracy of this method.From October 2010 to October 2011,collect HBV infected patients' periphcral blood of the First affiliated hospital of SooChow University,including acute hepatitis B,chronic hepatitis B,hepatits B liver cirrhosis,liver cancer patients and health control,each group contains 40 cases.The levels of miR-191,-223,-222,-145,-21,-31,-126,-20a,-372 of peripheral blood mononuclear cell were detected by xMap liquid chiptechnology.The level of miR-103 was taken as reference.The ratio of (the mean fluorescence intensity of target miRNAs-corresponding backgroud mean fluorescence intensity)/( the mean fluorescence intensity of miR-103-corresponding backgroud mean fluorescence intensity) as a valid data and analysis the characteristics of miRNAs expression.The SPSS 17.0 was used as statistical software.The single factor analysis of variance was used as the method to analysis group comparison,and make multiple comparison by the LSD-t method if the result with a significant difference.Results The specificity of Xmap liquid chip method to detect miRNAs was 100％ ;The repeated experiment proved that the CV value was less than 5％ in the high value reference miRNA test,less than 10％ in the low value reference miRNAs test;the accruracy experiment proved that the recovery rate was ( 100 ± 5)％ in the nine miRNAs.There were no statistically differences with miR-222 ( F =1.32,P ＞ 0.05),-191 ( F =1.98,P ＞ 0.05),-145 ( F =0.78,P＞0.05),-21(F=0.64,P ＞0.05),-31 (F =0.83,P ＞0.05),-372(F =1.75,P ＞0.05)in different groups; There was statistically significant differences in miR-223 ( F =14.56,P ＜ 0.05) among different groups,with the highest expression level in actue hepatitis B group (15.37 ± 4.01),and the lowest expression level in liver cancer group (6.91 ±3.18) ; There was statistically significant differences in miR126 (F =17.43,P ＜ 0.05)among different groups,with the highest expression level in health control group (6.33 ±2.75) and the lowest expression level in liver cancer group (2.38 ± 1.07).There were statistically significant differences in miR-20a ( F =19.484,P ＜ 0.05) among different groups,with the highest expression level in health control group (0.33 ±0.18) and the lowest expression level in liver cancer group (0.81 ±0.24).Conclusion The detection method of miRNA by Xmap liquid chip has strong specificity,high accuracy and good repeatability,suitable for large throughput clinical testing.The study for miRNA in HBV infected diseases provides a new clue to the research of chronic progress mechanism.
ABSTRACT
<p><b>OBJECTIVE</b>To evaluate the neuroprotective effect of memantine, a non-competitive antagonist at the N-methyl-D-aspartate receptor, against hypoxic ischemia (HI) by exploring its regulation on the expression and synthesis of heat shock protein 70 (HSP70) gene in neonatal rat models with cerebral HI.</p><p><b>METHODS</b>Memantine was intraperitoneally injected at a dose of 20 mg/kg in neonatal rat models either before (PRE group) or after (POST group) induction of HI. The expression and synthesis of the HSP70 gene and its corresponding product were determined by rapid competitive PCR and immunohistochemistry, respectively.</p><p><b>RESULTS</b>There was an increase in the expression of HSP70 mRNA two hours after induction of HI, which reached its peak at 48 hours, then decreased gradually. The same expression occurred at relatively low levels in the control group. Also, HSP70 synthesis was detected as early as 2h after HI, reached its peak between 48 and 72 hours, then declined over time. After memantine administration, the expression of the gene and its synthesis of the corresponding product decreased significantly during the time intervals 24 - 72 h for the gene and 48 - 72 h for the product compared to the HI group.</p><p><b>CONCLUSION</b>It was shown that HI is very sensitive to the expression of the HSP70 gene and synthesis of its corresponding product, which could be regulated by memantine. The latter may have the ability to reduce brain damage; thus decreased HSP70 mRNA expression could be a marker for HI. It is suggested that memantine can be a promising agent for neuroprotection against HI, although an overall and objective assessment of memantine is required to see if it can be used on neonates clinically later on.</p>