Objective To investigate the efficiency and mechanism of differentiation of reprogrammed adipose-derived stem cells (ADSCs) to neurons in vitro.Methods ADSCs from rats were cultured in vitro and then purified and identified.ADSCs at the third passage were divided into three groups:ADSCs without lentivirus-mediated gene transfection (blank group),ADSCs transfected with lentivirus carrying no neurogenin2 (Ngn2) (empty virus group) and ADSCs with lentivirus-mediated transfection of Ngn2 (Ngn2 group).All groups were induced in the medium containing cell growth factor for 15 days.The positive expression of neuron-specific nuclear protein (NeuN) in three groups was detected using immunofluorescence method so as to observe the efficiency of neuron differentiation.Expression variances of Mash1,Hes1 and Dll1 in each group were detected by Western blot analysis and the mechanism of differentiation was also discussed.Results After 15 days of induction,positive expression rate of NeuN in Ngn2 group,empty virus group,blank group was 90.12％,45.34％ and 40.26％ respectively,with significant differences among groups (P ＜ 0.01).Western blot analysis showed that Ngn2 group had a significantly higher expression of Dll1 (P ＜0.01) and obvious lower expressions of Hes1 and Mash1 (P ＜0.01),as compared with empty virus group and blank group.However,there were no significant differences of expression levels of Dll1,Mash1 and Hes1 between empty virus group and blank group (P ＞ 0.05).Conclusions After induction,the ratio of neuron differentiation of reprogrammed ADSCs is increased by almost 99％,as compared with simple ADSCs.The increased dfferentiation of reprogrammedADSCs to neurons may be associated with the inhibition of notch signaling through up-regulating Dll1 and down-regulating Mash1 and Hesl.

Objective To test the expression level of IL-37 in peripheral blood mononuclear cells (PBMCs)of patients with primary biliary cirrhosis (PBC)and further explore its clinical significance in the pathological process of PBC.Methods Pe-ripheral blood samples were collected from 42 patients diagnosed as PBC and 38 health individuals examined at the same time during June 2013 to August 2015 in Changhai Hospital.PBMCs were separated by sucrose density gradient centrifugation, qualified Real Time-Polymerase Chain Reaction (qRT-PCR)was used to measure IL-37 mRNA expression level in PBMCs. Enzyme-Linked Immuno Sorbent Assay (ELISA)was to measure the protein level of IL-37,IL-6,IL-17,TNF-α,TGF-β,IL-18 and IL-23 in plasma.Meanwhile,the pathological stages of PBC cases were recorded.Pearson correlation analysis was performed on IL-37 and IL-6,TNF-α,IL-17,TGF-β,IL-18 and IL-23.Spearman rank correlation analysis was on IL-37 and pathological stages of PBC.Results The mRNA and protein level of IL-37 in experimental and controlled group were 2.81 ±0.94 vs 1.09±0.56,356.14±169.36 pg/ml vs 86.68±48.23 pg/ml separately(t=9.811,9.462,P<0.000 1),with sta-tistical differences.The correlation analysis showed that IL-37 was positively related with IL-17,TNF-α,IL-6 and TGF-β(r=0.561 2,0.661 9,0.672 1,0.765 3,P<0.001),and disease stages (Ⅰ~Ⅳ)(rs=0.348 9,P<0.05).Conclusion IL-37 might involve in the pathological process of PBC,and it is significant for disease prediction and diagnosis.

Objective To analyze the affection and clinical significance of CD26/DPP4 on CD4+T cells and its cytokines in patients withCrytococcalMeningitis.Methods Peripheral blood was collected from 36 patients diagnosed withCrytococcal Meningitis in Changhai Hospital and Changzheng Hospital,Shanghai from August,2011 to December,2015,meanwhile 36 health controls’was also acquired.Peripheral blood mononuclear cell (PBMC)was separated by density gradient centrifuga-tion,CD26+CD4+T and CD26-CD4+T cell groups were classified by Flow Cytometry,the expression level of cytokines was tested by reverse transcriptase-polymerase chain reaction (RT-PCR).The correlation between DPP4 activity,CD26+CD4+T (%)and APACHE II score,IL-17,TNF-α,IL-4,IFN-γwas measured by Pearson coefficient.Results CD26+CD4+T(%)between experimental and control groups was 13.35±3.83 vs 8.39±2.14 (t=6.78,P<0.000 1).DPP4 activity was 50.89±17.21 mU/ml vs 73.83±20.24 mU/ml (t=5.18,P<0.000 1),with statistically significant differences.In ex-perimental groups,CD26+CD4+T (%)was positively related with APACHE II score,IL-17,TNF-α(r=0.431,0.564, 0.688,P=0.003 8,0.001,0.004 6).DPP4 activity was negatively interrelated with APACHE II score,IL-17,TNF-α,IFN-γ(r=-0.544,-0.489,-0.678,-0.734;P<0.001).Conclusion CD26/DPP4 may be involved in the pathogenesis of Crytococcal Meningitis through regulation of Th subgroups,and it was the potential therapeutic target and the predicted marker of the disease.

Objective To explore the change of microRNA let-7b in heat acclimatization/heat stroke rat models and its relation with HSP70,IL-6,TNF-α,TGF-3 and IL-12,and analyze its clinical significance.Methods 60 male rats with almost the same anal temperature,weight,and weeks' age were selected from Laboratory Animal Center in the Second Military Medical University.They were randomly divided into control,heat acclimatization,heat stroke groups averagely.Heat acclimatization/heat stroke rat models were built in hot climate simulated animal tank,blood was collected from cordis apex and serum was separated.Real-time fluorescent quantitative polymerase chain reaction (qRT-PCR) was used to test microRNA let-7b in serum,enzyme-linked immunosorbent reaction (ELISA) was to measure the protein level of HSP70,IL-6,TNF-α,TGF-β and IL-12 in serum.Kruskal-Wallis H test was used to analyze quantitative data in the three groups,Spearman rank correlation coefficient was used to reveal relation between two variables in heat stroke group.Results M values of miRNA let-7b in control,heat acclimatization,heat stroke groups were 0.99,1.04 and 1.93 separately,Q values were 0.30,0.25 and 0.44 (x2 =38.95,P＜0.001),separately,with statistical significance.The results of pairwise comparison showed no statistical difference in control and heat acclimatization (P＞0.05),but there were differences in control and heat stroke,heat acclimatization and heat stroke groups statistically (P＜0.05).Spearman correlation analysis showed that in heat stroke group,let-7b was positively related with HSP70,TNF-α and IL-6 (rs =0.579,0.498 and 0.609,P＜0.05) with statistical significance.Conclusion miR let-7b might involve in the pathology of heat stroke,it provided a potential biomarker for monitoring patients in high temperature and humidity clinically.

Objective To investigate the expression level of Gas6 in patients with Immune Thrombocytopenia (ITP) and its clinical significance.Methods The experimental group was peripheral blood samples collected from 35 cases diagnosed with ITP in hematology department of Changhai Hospital in Shanghai from October 2013 to December 2015.Control group was peripheral blood from 35 healthy examined individuals at the same time.After separating plasm from the two group samples,the protein level of Gas6,IFN-α,IL-4,IFN-γ and IL-17 were measured by Enzyme-linked immunosorbent assay (ELISA),comparison of expressional level of the two groups was measured by t test.Pearson correlation analysis was used to decide the relation between Gas6 and cytokines such as IFN-α.Results The expression level of Gas6 in experimental and control groups was 27.28±7.56 ng/ml vs 20.51±5.39 ng/ml (t=4.314,P＜0.000 1);IFN-γ was 221.67±57.64 pg/ml vs 45.32 ±16.79 pg/ml (t=17.38,P＜0.000 1);IL-4 was 113.86±26.48 pg/ml vs 49.87±14.98 pg/ml (t=12.44,P＜0.000 1);IL-17 was 168.96±47.88 pg/ml vs 109.56±28.97 pg/ml (t=6.28,P＜0.000 1);IFN-α was 34.83±8.12 pg/ml vs 29.89 ± 5.76 pg/ml (t=2.936,P=0.004 5),all with statistical differences (P＜0.05).Pearson analysis showed that Gas6 was positively related with IL-17,IL-4,IFN γ (r=0.564,0.486,0.449,P＜0.05) and there was difference statistically,but Gas6 was not correlated with IFN-α.Conclusion Gas6 may participate in the disease formation of ITP through affection on cytokines secreted by Th cell subsets,and it was the potential therapeutic and predicted target for this disease clinically.