Objective To establish a stable HCV full-length genome replication cell model which is labeled with reporter gene and easyly to quantify intracellular HCV proteins and RNA level. Methodsneo gene was inserted into Luc-JC1 to make Luc-JC construct. Luc-JC RNA was obtained by in vitro transcription and then delivered into Huh7 cells by transfection. G418-resistant clones of Huh7 cells were obtained by selection. Clones of HCV full-length genome replication cell were confirmed by luciferase activity assay, Western blot and cleaveage of eYFP-MAVS by HCV NS3/4A protease. Then, HCV replication cell colonies were treated by different dose IFN-α in order to observe the change of luciferase activity, HCV protein and RNA level. Results At 3-4 weeks post-transfection, visible colonies were selected and stained by crystal violet. Luciferase activity and HCV NS3, NS5A protein were detected by luciferase activity assay and Western blot, respectively. Subcellular localization of eYFP-MAVS transferred from mitochondria to cytoplasms by cleavage of NS3/4A protease in cell colonies. Luciferase activity, HCV protein and RNA diminished obviously after IFN-α treatment. Conclusion A stable HCV full-length genome replication cell model labeled by reporter gene was successfully established and reporter activity can be used to indicate level of HCV proteins and RNA in cells. This cell model is a useful tool for the study on HCV pathogenesis and the screening of antiviral drugs.

STIM1 is recognized as an ER Ca2+ sensor of calcium release-activated calcium (CRAC) channel that is constructed by membrane protein Orai1, However, this regulatory system may also be regulated by other proteins. Reticulocalbin 2 (RCN2) was purified and identified from STIM1-Orai1 complex. Confocal microscopy revealed that RCN2 co-localized with STIM1 in ER before and after Ca2+ store depletion. Single cell [Ca2+]I measurements of RCN2 EF hands mutant showed slight influence on SOC electrophysiological characters. Furthermore, a novel collar form aggregation of RCN2 surrounding STIM1 clusters suggested that RCN2 potentially plays a role of structure maintenance in STIM1 clustering.

This study was aimed to explore the expression of 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2) in 3 different lymphoblastic cell lines with relation to their glucocorticoid (GC) sensitivity. The 11β-HSD2 expressions in acute lymphoblastic leukemia Jurkat cells, lymphoma Daudi and Raji cells, and peripheral blood T cells of a healthy volunteer were analyzed by real time PCR and Western blot. Glucocorticoid (GC)-induced apoptosis in 3 different cell lines was detected by flow cytometry. Cell growth in Jurkat cells treated with cortisol was analyzed by trypan blue dye exclusion. Flow cytometry was performed to observe GC-induced apoptosis in Jurkat cells treated by combination of GC with 11β-HSD2 inhibition 18β-glycyrrhetinic acid (18β-GA). The results demonstrated that 11β-HSD2 highly expressed in Jurkat cells, but not in Daudi, Raji cells and normal blood T cells. Compared to Daudi and Raji cells, Jurkat cells were more resistant to GC-induced apoptosis. Furthermore, the inhibition of 11β-HSD2 by 18β-GA resulted in increased cellular sensitivity to GC as shown by elevated induction of apoptosis. it is concluded that 11β-HSD2 is at least partly responsible for GC resistance in Jurkat cells. 11β-HSD2 may be a potential target for reduction of GC-resistance in therapeutic applications.
11-beta-Hydroxysteroid Dehydrogenase Type 2 ; metabolism ; Cell Line, Tumor ; Glucocorticoids ; pharmacology ; Glycyrrhetinic Acid ; analogs & derivatives ; pharmacology ; Humans ; Jurkat Cells ; Lymphocytes ; drug effects ; metabolism

AIM: To investigate the regulatory effects of microRNA(miR)-195 on the biological behaviors, such as viability,apoptosis and migration, of lung cancer A549 cells, and to explore the related mechanisms.METH-ODS:After miR-195 mimics were transfected into the A549 cells,the cell viability, cell cycle distribution and apoptosis were measured by CCK-8 assay and flow cytometry.Transwell assay was used to detect cell migration ability.Furthermore, the protein levels of cyclin D1,CDK2,Bcl-2 and p-Rb/Rb were determined by Western blot.Dual-luciferase reporter as-say was used to screen and identify the possible target genes of miR-195.RESULTS: Over-expression of miR-195 in the A549 cells inhibited the cell viability and induced cell cycle arrest,accompanied with the decrease in the cell migration a-bility and the increase in the apoptotic rate(P<0.05).Furthermore,the protein levels of cyclin D1,CDK2,Bcl-2 and p-Rb were significantly decreased(P<0.05).Dual-luciferase reporter assay demonstrated that MYB was a potential target gene of miR-195.Over-expression of MYB in the A549 cells partially reversed the effects of miR-195 on the cell viability, apoptosis and migration.CONCLUSION: miR-195 inhibits lung cancer A549 cell growth and migration, and promotes cell apoptosis by targeting MYB gene.

OBJECTIVETo evaluate the effect of artificial and bioartificial liver support systems for management of acute and acute-on-chronic liver failure.

METHODSArticles documenting randomized clinical trials concerning any liver support systems vs standard conservative therapy, published between January, 1970 and June, 2008, were retrieved by database searching. Of the 1134 articles retrieved, 12 randomized trials involving 479 patients were included. The data were extracted and the trial quality was assessed by 2 independent reviewers. The primary outcome measure was all-cause mortality, and the results were combined on the risk ratio (RR) scale.

RESULTSOf the 12 trials included, 10 assessed artificial liver support systems for acute or acute-on-chronic liver failure, and 2 assessed bioartificial systems for acute liver failure. Overall, the liver support systems had moderate effect on mortality compared with standard conservative therapy (RR=0.80; 95% CI 0.664-0.969, P=0.022). Meta-regression indicated that the effect of the support systems depended on the type of liver failure (P=0.00). In stratified meta-analyses, the support systems appeared to reduce the mortality by 43% in acute-on-chronic liver failure (RR=0.57; 95% CI 0.39-0.84, P=0.004), but not in acute liver failure (RR=0.899; 95% CI 0.72-1.12, P=0.361).

CONCLUSIONArtificial liver support systems reduce the mortality of acute-on-chronic liver failure as compared with standard conservative therapy, but have no significant effect on the mortality of acute liver failure. Bioartificial liver support systems lower the mortality rates in both acute and acute-on-chronic liver failure, and should be the future focus of development.

Chronic Disease ; therapy ; Clinical Trials as Topic ; Databases, Factual ; Humans ; Liver Failure, Acute ; therapy ; Liver, Artificial ; Regression Analysis

OBJECTIVETo explore the effect of recombinant human erythropoietin (rhEPO) on expression of brain-derived neurotrophic factor (BDNF) in different brain regions of aging rats.

METHODSForty male SD rats were randomized equally into negative control group, D-galactose group, EPO treatment group, and positive control group. Rat models of subacute aging were established by continuous subcutaneous injection of 5% D-galactose. Immunohistochemical staining was used to analyze the variation of BDNF expressions in different brain regions of the aging rats with different treatments.

RESULTSSignificant brain region-specific differences in BDNF expression were found among the rats in different groups. Compared with those in the negative control group, the numbers of BDNF-positive cells in the hippocampal CA1 region, CA3 region, dentate gyrus (DG) and frontal cortex were all decreased obviously in D-galactose group (P<0.05) but increased in both EPO group and the positive control group (P<0.05) without significant differences between the latter two groups. In the rats in the same group, the number of BDNF-positive cells varied markedly in different brain regions (P<0.05), and the expression level of BDNF was the highest in the frontal cortex followed by the hippocampal CA3 region and the dentate gyrus, and was the lowest in the hippocampal CA1 region.

CONCLUSIONTreatment with rhEPO enhances the expression of BDNF in rat neural cells, suggesting that rhEPO may protect the nervous system from aging by regulating the BDNF pathway.

Aging ; Animals ; Brain-Derived Neurotrophic Factor ; metabolism ; CA1 Region, Hippocampal ; metabolism ; CA3 Region, Hippocampal ; metabolism ; Dentate Gyrus ; metabolism ; Erythropoietin ; pharmacology ; Frontal Lobe ; metabolism ; Galactose ; Humans ; Male ; Neurons ; drug effects ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Recombinant Proteins ; pharmacology

Objective To investigate the clinical impact of neutrophil to lymphocyte ratio (NLR) in the diagnosis of ulcerative colitis.Methods The study included 97 patients with ulcerative colitis (UC) and 95 patients with irritable bowel syndrome (IBS) from June 2009 to June 2016.The differences of NLR between the two groups were analyzed and the result of sensitivity and specificity were calculated.The differencesn of NLR were analyzed between the groups of different severity and in different lesionsseparately.Results NLR were significantly higher in ulcerative colitis than in irritable bowel syndrome (t=2.327,P＜0.021).Sensitivity was 69.1 ％ and specificity was 75.8 ％.There was statistic different in groups of different severity determination of newly diagnosed ulcerative colitis (F=8.221,P=0.001)and there was no statistic different between different lesions (F=0.737,P=0.483).Conclusion NLR is valuable in the diagnosis of UC and IBS can determine the degree of inflammation of ulcerative colitis.

Objective To evaluate the effectiveness of Lingshao-Zaoren granule in the treatment of female overactive bladder. Methods A total of 60 female OAB patients who met the inclusion criteria were randomly divided into 2 groups, 30 cases in each group. The control group recieved the Tolterodine Tartrate Sustained Release Tablets and the Lingshao-Zaoren granule placebo, and the treatment group used the Tolterodine Tartrate Sustained Release Tablets and the true Lingshao-Zaoren granule treatment. Both groups were treated for 4 weeks. Overactive bladder symptom scale (OABSS) was used to determine the severity of OAB.Results After treatment 14,28 d,OABSS scores of the treatment group(5.3 ± 2.3,1.4 ± 1.2 vs.8.4 ± 2.4,F=137.209),and control group(7.8 ± 1.9,6.8 ± 1.4 vs.8.6 ± 2.6,F=8.927),were significantly lower than the baseline of each group respectively (P<0.01). Besides, OABSS scores of the treatment group after 14 and 28 d were significantly lower than the control group (t=4.668, 15.678, P<0.01). The pain scores (5.9 ± 1.9, 2.7 ± 1.1 vs.9.5 ±2.3,F=108.819)of treatment group at 14 and 28 d were significantly lower than the baseline(P<0.01);and the pain scores of treatment group at 14 and 28 d were significantly lower than the control group (t=6.342, 14.812,P<0.01).The lower abdomen discomfort scores at 14,28 d in treatment group(1.9 ± 1.4,1.1 ± 1.0 vs. 3.3 ±1.1,F=28.762),and control group(2.7 ±1.0,2.4 ±0.8 vs.3.4 ±1.2,F=12.103)were significantly lower than the baseline of each group (P<0.01); and the abdominal discomfort scores of treatment group at 14, 28 d were significantly lower than the control group (t=2.521, 5.041, P<0.05 or P<0.01). Conclusions The Lingshao-Zaoren granule could decrease OABSS score,pain score,abdominal discomfort symptoms,improve clinical symptoms of the female patients with OAB.

Objective To evaluate the application of novel fluorescent staining method with hypersensitivity and enhancement for microscopic examination of superficial fungal.Methods A total of 200 cases with clinically suspected superficial fungal infection were screened by the hypersensitive,enhanced fluorescent staining,calcofluor white staining and microscopic examination with 1.78 mol/L potassium hydroxide (KOH) based smear assay,respectively.The positive percentages of the different methods were calculated and compared.Results In the developed fluorescent staining examination,clean background was shown in the view field.The hyphae and spores displayed in bright blue,thus fungal morphology was so distinct that they were easy to be distinguished.Compared with calcofluor white staining and KOH based smear,the fluorescent staining showed high positive rate with significant difference respectively (x2 =17.984,P ＜ 0.05;x2 =32.063,P ＜ 0.05).Conclusion The novel fluorescent staining should be a rapid,accurate method for microscopic examination of fungi,which is worthy to be widely used in clinical laboratories for early diagnosis of fungal infection.

Objective To investigate the cognitive, intelligent and cranial MRI status of 4 patients with myotonic dystrophy type 1 (DM1). Methods A pedigree of 4 patients,admitted to our hospital from September 2011 to December 2011 and diagnosed as having DM1, were chosen in our study; Chinese edition tests of Wechsler adult intelligence scale and Wechsler memory scale were used to test the intelligence of these patients; Montreal cognitive assessment (MoCA) and Neurobehavioral cognitive status examination (NCSE) were applied to evaluate the patients' cognitive function.All of the 4 patients received a cranial MRI scan. Results Except for 1 patient,the other 3 patients had low intelligent quotient (IQ) and memory quotient (MQ),and their verbal IQs were all lower than performance IQs.Three patients showed impaired visual space/executive skills,calculation,memory,and conceptual thinking abilities; 1 patient showed impaired sentence repetition and verbal fluency on MoCA,though his final scores were within normal range. Three patients had brain atrophy to some extent on MRI scan,prominently in the frontal lobe and temporal lobe; white matter lesions and thickened skull were founded in all 4 patients. Conclusion Multiple cognition disorders and brain involvements can be found in DM1 patients and deserve further investigation.