ObjectiveTo investigate the effects of MMI-166 on the proliferation and apoptosis of human pancreatic cancer SW1990 cells.MethodsMMI-166 of different concentrations (25,50,100 μg/ml) were used to treat human pancreatic cancer SW1990 cell for 24,48 h.Effect of MMI-166 on cell proliferation wasdetected by 3- (4,5-dimethyl-2-thiazole) -2-5-biphenly-tetrazole bromide ( MTT ) method and effect on cell apoptosis was tested by Annexin V-PI method and flow cytometry (FCM).ResultsTwenty-four hours after MMI-166 treatment of different concentrations (25,50,100 μg/ml),the inhibitory rates of the cells were (34.23±3.87)％,(44.81 ±2.01)％,(53.91 ±1.74)％,and the corresponding values were (39.95 ± 1.83) ％,( 52.26 ± 3.46 ) ％,( 63.20 ± 2.48 ) ％ at 48 h,which suggested a time-and concentrationdependent manner.The cell's apoptosis rates were (11.19 ±0.47)％,(23.01 ±0.53)％,(28.10 ± 0.52) ％ at 24 h,and the corresponding values were ( 11.19 ± 0.47 ) ％,( 23.01 ± 0.53 ) ％,( 28.10 ± 0.52)％ at 48 h,which were significantly higher than those in control group [ (0.09 ±0.12)％,P ＜0.05].ConclusionsMMI-166 can inhibit proliferation and induce apoptosis of human pancreatic SW1990 cell in a time- and concentration-dependent manner.

Objective To investigate the effects of MMI-166 on apoptosis and apoptosis-related protein expression of human pancreatic cancer SW1990 cells and its transplanted tumor,and explore possible mechanism.Methods The human pancreatic cancer xenograft model was constructed by using human pancreatic cancer SW1990 cells.Tumor-bearing nude mice were randomly divided into control and MMI-166 groups,and they were treated with normal saline or MMI-166 (200 mg · kg-1 · d-1) for 28 days.Apoptosis index (AI),p53,c-Myc,Bax,Bcl-2,Survivin,Caspase-1,Fas proteins were detected by deoxynucl-eotidyl transferase-mediated nick end labeling (TUNEL method) and Western blot.MMI-166 of different concentrations (0,50,100 μg/ml) were used to treat human pancreatic cancer SW1990 cell for 24 h.The c-Myc,Survivin proteins expressions were measured by Western blotting.Results Apoptosis index in MMI-166 group was 81.1 ±7.9,which was significantly higher than that in control group (21.3 ±2.2,P =0.000) ; the expressions of c-Myc,Survivin were 7715 ± 2229,4594 ± 1240,which were significantly higher than those in control group (16870 ± 2446,15208 ± 1903,P =0.000) ; the expressions of p53,Bax,Bcl-2,Caspase-1,Fas were not significantly different from those in control group.After 50,100 μg/ml MMI-166 treatment,the expression of c-Myc was significantly down-regulated (0.098 ± 0.003,0.073 ± 0.008 vs.0.169 ± 0.007,F =189.361,P ＜ 0.05) ; and the expression of Survivin was not significantly changed.Conclusions MMI-166 may induce cell apoptosis of SW1990 by down-regulating the expression of c-Myc.

Objective To investigate the expressions of TRIM59,Twist and E-cadherin in hepatocellular carcinoma and their clinical significance. Methods The expressions of TRIM59,Twist and E-cadherin protein were tested by immunohistochemistry in 80 cases of hepatocellular carcinomas and the adjacent paracancerous tissues. Results The positive rate of TRIM59 in hepatocellular carcinoma was significantly higher than that in the adjacent paracancerous tissue(76.3%vs. 8.0%,P<0.05). Significant difference was also observed in the expres-sion rate of Twist between the hepatocellular carcinomas and the paracancerous tissue(66.3%vs. 6.0%,P<0.05). The positive rate of E-cadherin in hepatocellular carcinoma was significantly lower than that in the adjacent para-cancerous tissue(27.5%vs. 90.0%,P<0.05). The differences of the expression of TRIM59 in hepatocellular carci-noma of pathological grading,tumor differentiation,vascular invasion and clinical TNM stage were significant(P<0.05). The differences of the expression of Twist in hepatocellular carcinoma of pathological grading ,differentia-tion,vascular invasion and clinical TNM stage was also significant(P<0.05,respectively). The differences of the expression of E-cadherin in hepatocellular carcinoma of pathological grading,differentiation,vascular invasion and clinical TNM stage were also significant(P<0.05,respectively). Significantly positive correlation was also found between TRIM59 and Twist by using spearman correlation analysis(P<0.05). Negative correlations were observed between TRIM59 and E-cadherin(P < 0.05),and between Twist and E-cadherin(P < 0.05). Survival analysis showed that TRIM59 expression was an independent prognostic factor in hepatocellular carcinoma. Conclusion TRIM59,Twist and E-cadherin protein expression might be associated with the development,invasion,and metas-tasis of hepatocellular carcinoma. TRIM59 may become a new target gene for the treatment of human hepatocellular carcinoma.

Objective To investigate the effect of intermittent hypoxia (IH) with pulmonary emphysema on the ex-pression of hypoxia inducible factor-1α(HIF-1α),Bax and Bcl-2, and the mechanism underlying the role of HIF-1αin he-patocyte apoptosis thereof. Methods Sixty rats were randomly divided into four groups: normal control group, rats were treated normally;IH group, rats were treated by 30 s nitrogen and then 90 s air, and rats were treated by from 9:00-17:00 daily;pulmonary emphysema group, rats were treated by smudging for half an hour, twice a day (8:00 and 18:00);IH with pul-monary emphysema group, rats were treated by 30 s nitrogen and then 90 s air from 9:00-17:00 daily. After exposure four-teen weeks, rats were killed. qRT-PCR assay was conducted to detect the expression of HIF-1α mRNA, Bax mRNA and Bcl-2 mRNA in live tissues. Results The expressions of HIF-1αmRNA, Bax mRNA and Bax/Bcl-2 were significantly higher in IH with pulmonary emphysema group than those in control group,IH group and pulmonary emphysema group (P<0.05). The expression level of Bcl-2 mRNA was significantly lower in IH with pulmonary emphysema group than that of con-trol group and pulmonary emphysema group (P < 0.05), but no significant difference compared with that of IH group (P >0.05). The levels of HIF-1αand Bax were positively correlated with the level of Bax/Bcl-2 (r=0.732 and 0.699),but the lev-els of HIF-1αand Bax were negatively correlated with the level of Bcl-2 (r=-0.705). Conclusion The expression levels of HIF-1αmRNA, Bax mRNA and Bcl-2 mRNA were over-regulated in hepatocytes induced by intermittent hypoxia with pul-monary emphysema. The HIF-1αexpression was correlated with Bax and Bcl-2, suggesting that HIF-1αmay promote the hepatocyte apoptosis through transcriptional co-activators, Bax and Bcl-2.

Objective To observe the effect of exposure of emphysema with intermittent hypoxia on oxidative stress injury of myocardial cells in rats. Methods Sixty male Wistar rats were divided randomly into four experimental groups(each n=15). The normal control group was bred normally. The emphysema group was exposed to cigarette smoke twice a day(once 30 minutes). The intermittent hypoxia(IH)group was exposed to intermittent hypoxia circumstance 8 hours/day,and the emphysema with IH group was exposed to cigarette smoke twice a day (once 30 minutes)and intermittent hypoxia circumstance 8 hours/day. Each group was exposed for 8 weeks. At the beginning of 9 weeks,the blood gas analysis was performed in 5 rats selected randomly from each group,and the rest rats were sacrificed and their hearts and lungs were taken. Under light microscope,the lung tissues stained with hematoxylin-eosin(HE)were examined. The lung pathology and the results of blood gas analysis showed that the emphysema with IH rat model was established successfully. The levels of malonaldehyde(MDA)and superoxide dismutase(SOD)in rat myocardium were measured by enzyme-linked immunosorbent assay(ELISA),and the subunit p22phox mRNA expressions of nicotinamide adenine dinucleotide phosphate(NADPH)-oxidase were detected by real-time reverse transcription-polymerase chain reaction(RT-PCR). Results Compared with the normal group, the MDA levels and p22phox mRNA expressions were increased obviously in emphysema group, IH group and emphysema with IH group〔MDA(μmol/g):2.93±0.54, 3.58±0.63, 4.51±0.72 vs. 1.75±0.56, p22phox mRNA:0.043±0.004,0.067±0.015,0.123±0.016 vs. 0.018±0.002,all P<0.05〕,but the activities of SOD were decreased significantly(U/mg:36.07±4.79,33.51±7.12,24.29±5.36 vs. 46.08±5.12,all P<0.05). In emphysema with IH group,the increase of MDA levels and p22phox mRNA expressions and decrease of SOD levels were more remarkable compared with those in emphysema group and IH group(all P<0.05). The expression of p22phox mRNA was positively correlated with MDA level(r=0.734,P<0.001). Conclusion The myocardial tissue oxidative stress injury in rats induced by emphysema with intermittent hypoxia exposure is more serious than that induced by exposure of either emphysema or intermittent hypoxia alone,NADPH oxidase possibly being the important medium of myocardial cell response to oxidative stress.

ObjectiveTo study the expression of matrix metallopro-teinase-2 (MMP-2),matrix metalloproteinase-9 (MMP-9) in the pancreatic carcinomas. Methods MMP-2,MMP-9 expression were detected by immunohistochemistry in surgically resected specimens ( cancer tissues,cancer-adjacent tissues and normal tissues) from 30 PC patients,11 pancreatitis patients and 6 normal patients.the results were analyzed combined with clinical pathologic characteristics.The data was analyzed by Chi-Square test.ResultsThere was no expression of MMP-2,MMP-9 in normal pancreatic tissues.Both of MMP-2 and MMP-9 expressions were 18.2％ in chronic pancreatitis titssues.Expression of MMP-2,MMP-9 were higher in cancer tissues than in cancer-adjacent tissues(63.3％ vs 23.3％,56.7％ vs 40.0％,P ＜0.05).There were positive correlation between MMP-2,MMP-9 expression and lymph nodal metastasis,tumor sizes,clinical stage and differentiation of tumor(P ＜0.05).Conclusions The expression of MMP-2 and MMP-9 was high,and linked to its unfavorable prognosis.

Objective To investigate of the MMI-166 on the expression of MMP-2,MMP-9 and the cell apoptosis of nude mouse xenografts of SW1990 human pancreatic cancer cells.Methods Establishment of control and experimental groups,randomly,the human pancreatic cancer xenograft model of SW1990 was constructed.The control group was treated with normal saline,and experimental group was treated with MML-166 (200 mg · kg-1 · d-1).The tumor volume and tumor inhibition rate was measured by vernier caliper through length and short diameter.The expression of MMP-2 and MMP-9 protein was observed using immunohistochemistry in the tumor tissues.Apoptosis index was detected by deoxynucleotidyl transferase-mediated nick end labeling (TUNEL method).Results The tumor volume of MMI-166 group (1252.30± 464.84) mm3 was less than the control group (2241.82±208.06) mm3,significantly.The inhibition rate was 34.47％ between the experimental groups (treat with MMI-166) (1.42±0.15) g and control group (2.17±0.20) g.The expression of MMP-2 (2.80 ± 1.10) ％ and MMP-9 (2.60 ± 1.52) ％ protein was significantly downregulated in MMI-166 group,compared with the control group.Apoptotic index in the experimental group (75.60±9.71) ％ was higher than the control group (17.40 ± 10.14) ％,significantly.Conclusion The mechanism of MMI-166 inhibiting pancreatic tumor growth and inducing apoptosis may be related to the suppression of MMP-2 and MMP-9 protein expression.

Objective To explore the mechanism of Lkb1 regulated epithelial regeneration in asthma by airway organoid culture.Methods Lkb1f/f(the control group,n=10)and Scgb1a1CreER;Lkb1f/fmice(the Lkb1 knockout group,n=9)were taken to establish allergic asthma models by aerosol inhalation of ovalbumin(OVA).Bronchial lavage fluid(BALF)and lung tissue were collected.The number of inflammatory cells in BALF were counted.The amount of CLCA3 positive cells was compared by immunofluorescence staining of lung tissue sections.Club cells were selected by flow cytometry for organoid culture.The average diameter of organoids and organoid formation rate were calculated.Expression levels of goblet cell marker CLCA3,cilia cell markers FOXJ1 and AMPK in Club cells were detected by RT-PCR.Results There were no significant differences in the number of macrophages,eosinophils,neutrophils and lymphocytes in BALF between the control group and the Lkb1 knockout group.The number of CLCA3 positive cells were decreased after Lkb1 knockout.Results of organoid culture showed that the average diameter of organoids derived from Club cells and organoid formation rate were decreased after the absence of Lkb1.The expression of FOXJ1 was reduced.After Lkb1 deletion,the expression of AMPKα in Club cells were decreased and the proliferation of Club cells was inhibited.Activation of AMPK,the downstream signaling pathway of Lkb1,could attenuate the effect of Lkb1 deficiency on the regeneration of Club cells.Conclusion Lkb1 promotes the proliferation of airway progenitor cells by AMPK pathway.