The expression of Transgelin in pancreatic cancer accompanied with diabetes and its effects on the migration and invasion of SW1990 cell line / 中华消化杂志

Objective To investigate the expression of actin-associated protein Transgelin in pancreatic cancer with or without diabetes and its effects on migration and invasion in SW1990 cell line. Methods The expression of Transgelin in 92 pancreatic cancer tissue specimens (45 cases accompanied with diabetes) and adjacent tumor-free tissue specimens (over 5cm from the edge of the tumor) was detected by immunohistochemistry, and their association with clinical pathological characteristics were also analyzed. Transgelin siRNA was designed and transfected into pancreatic cancer cell line SW1990. The changes of migration and invasion before and after transfection were observed through Transwell test.Results The positive percentage of Transgelin expression in pancreatic cancer was 68.5 ％ (63/92), which was significantly higher than that of adjacent tumor-free tissues[33.7％ ( 31/92), P＜ 0.05]. The positive percentage of Transgelin expression in pancreatic cancer accompanied with diabetes was 84.4％ (38/45), which was significantly higher than that without diabetes[53.2％ (25/47), P＜0.05]. The expression of Transgelin in pancreatic cancer tissues was associated with lymph nodes metastasis and TNM staging (both P＜0.05), but not related with gender, age, site, differentiation and portal vein or nerve invasion (P＞0.05). After Transgelin was interfered for 48 hours, the migration ability was significantly lower (migration cell number 49.2 ±9.5 cells) than negative control group (61.9±7.5 cells) and blank group (65.3±10.6 cells) (both P＜0.05), and the invasion of SW 1990 cells (48.0 ± 8.6 cells) also significantly lower than negative control group (63.5±11.4 cells) and blank group (67.5±9.6 cells) (both P＜0. 05). Conclusion Transgelin may involve in the metastasis of pancreatic cancer accompanied with diabetes through promoting pancreatic cancer cell migration and invasion.

Activated protein C improves the severity of severe acute pancreatitis via up-regulating the expressions of endothelial cell protein C receptor and thrombomodulin.

BACKGROUND AND AIMS: Activated protein C (APC) is increasingly understood to have diverse regulatory functions in inflammation. However, the exact mechanism of action remains unclear in severe acute pancreatitis (SAP). The aim of this study was to demonstrate the effects of APC on expressions of thrombomodulin (TM) and endothelial cell protein C receptor (EPCR), and its subsequent effect on the severity of SAP. METHODS: Sprague-Dawley rats were randomly divided into four groups. The rats were given intravenous injections of APC (50, 10 microg/kg, respectively, treated groups) or saline (SAP group) just before induction of SAP. One group of rats underwent only sham operation as control group. Experimental samples were harvested at 16 h after induction. The protein and mRNA levels of matrix metalloprotease 9 (MMP-9), TM, and EPCR in pancreatic tissue were investigated. Serum tumor necrosis factor alpha (TNF-alpha) and interleukin-8 (IL-8) levels were determined. The severity of disease was evaluated by histological score of pancreatic injury, wet/dry weight ratio of pancreatic tissue, and serum amylase. RESULTS: In the APC 50 microg/kg-treated group, serum TNF-alpha, IL-8, and pancreatic MMP-9 levels were decreased and the levels of pancreatic EPCR and TM were up-regulated compared with the SAP group. A significant dose-dependent relationship was found between the decreased levels of serum IL-8 and the APC-treated dosage. Furthermore, the severity of SAP was ameliorated by APC treatment. CONCLUSIONS: APC could augment the anti-coagulation and anti-inflammatory activity by up-regulating EPCR and TM expressions, thus attenuating the severity of SAP.

Study of the role of activated protein C in the apoptosis of endothelial cell / 中华急诊医学杂志

Objective To study the protective effects of activated protein C (AFC) on the apoptosis of endothelial cells induced by lipopolysaccharide (LPS) in order to clarify the mechanisms associated with the expression of some genes related to apoptosis. Method The human umbilical vein endothelial cells were incubated with LPS (1.0 μg/mL) for one hour to make the models of cell apoptosis, and then the different concentrations of AFC (10 ng/mL and 50 ng/mL) were added to the models of cell apoptosis as treatment group. Therefore, there were two groups, model group and APC treated group. The factors related with apoptosis such as P53, Bax, Bcl-2, and caspase-3 mRNA or protein level were measured by using RT-PCR and Western blotting. Results Compared with LPS stimulated cells, the expressions of P53, Bax and caspase-3 mRNA and levels of protein were decreased and the expression of Bcl-2 mRNA and protein level were increased in APC treated cells particularly in APC 50 ng/mL treated cells (P <0.05). Conclusions The APC inhibits the apoptosis of HUVECs induced by LPS via regulating the mitochondrial-dependent apoptosis pathway, and it may become a novel therapeutic agent for infection disease.

Expression of janus kinase 1/signal transducer and activator of transcription 1 signaling pathway in cerulean stimulated pancreatic acinar cells / 中华胰腺病杂志

Objective To investigate the relationship between the activity of Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling pathway in cerulein-induced pancreatic acinar cell line AR42J. Methods The in vivo model of AP was induced by cerulean treated pancreatic acinar cell line AR42J, then RPM and AG490 were given for intervention. Western blot was used to determine theexpressions of JAK1 and phosphorylation JAK1 ( P JAK1 ) , STAT1, PSTAT1 and TNFα, IL-1β, IL-6. The expressions of IL-6, IL-1 β, and TNFα mRNA were measured by RT-PCR. Survival rate of cells was evaluated by trypan blue stain. Results The relative expressions of JAK1, P JAK1, STAT1, P STAT1 and TNF-o, IL-1β, IL 6 without cerulean treatment were 0.09 ±0.04,0.14 ±0.08,0.21 ±0.09,0.12 ±0.12,0.10 ±0.02,0.08 ± 0.03,0.02 ± 0.02. After cerulean treatment, the expressions of abovementioned protein increased in a time-dependant manner, the expressions at 24h were 0.53 ± 0.09,0.53 ± 0.13,0.56 ± 0.09,0.55 ± 0.10,0.25 ± 0.04,0.25 ±0.09,0.27 ±0.07, which were significantly higher than those in the control group (P ＜0.05). 2 4 h after RPM and AG 4 9 0 inhibition, the expressions of TNF-α, IL-1 β, IL-6 proteins significantly decreased to 0.17 ± 0.03and 0.17 ± 0.01,0.15 ± 0.05 and 0.14 ± 0.07,0.19 ± 0.04 and 0.19 ± 0.05; their expressions of mRNA significantly decreased ( P ＜ 0.05 ). The cell survival rates in RPM and AG490 treatment group were (72.4 ± 11.2) %, (69.7 ± 9.8 ) %, and in cerulein-stimulated cells (42.2 ± 12.3 ) % ( P ＜ 0.05 ).Conclusions The JAK1/STAT1 signaling pathway was involved in pancreatic inflammatory response with cerulein stimulation. Early treatment with inhibitors to the JAK1/STAT1 signaling pathway might control the inflammatory response in acute pancreatitis.

Effect of PIAS1 gene silencing on inflammatory response of pancreatic acinar cell / 中华胰腺病杂志

Objective To investigate the effect of protein inhibitor of activated signal transducer and activator of transcription 1 ( PIAS1 ) gene silencing on the inflammatory response of rat pancreatic acinar cell lines AR42J with cerulein stimulation, to study its role in the pathogenesis of acute pancreatitis.Methods The siRNA targeting PIASI was designed, synthesized, transfected into AR42J cells by lipofectmine 2000.24 h later, cerulean was added and cultured for another 24 h.Subsequent AR42J cells with cerulein stimulation were divided into 4 groups: cerulein, liposome, negative-siRNA and PIAS1-siRNA groups.In addition, a group with PBS was as control group.The activity of p38 mitogen- activated protein kinase (p38MAPK) was detected by western blotting.TNF-α, IL-1β, IL-6, matrix metalloproteinase (MMP) 9 expression were analyzed by RT-PCR and western blotting, respectively.Results The expression of p38MAPK in PIAS1-siRNA, negative-siRNA, liposome, cerulein,and control group was 1.93 ±0.11, 1.22 ±0.10, 1.30 ±0.17,1.32 ± 0.21, 0.12 ± 0.02;while the expression of phosphorylated p38MAPK was 2.10 ± 0.25, 1.36 ± 0.20,1.26 ±0.15, 1.23 ±0.25, 0.58 ±0.48, the expression in PIAS1-siRNA group was significantly increased when compared with other groups (P＜0.05).The levels of TNF-α, IL-1β, IL-6, MMP-9 mRNA were 1.66 ±0.15,1.66 ± 0.15,1.90 ±0.01, 1.56 ±0.20 in PIAS1-siRNA group, while the expression of protein was 2.06 ±0.37,2.20 ±0.34, 1.80 ±0.10, 1.17 ±0.05, which was markedly higher than those in other group (P ＜0.05).Conclusions PIAS1 gene silencing could enhance p38MAPK activity, and improve inflammatory mediator expression in pancreatic acinar cells with cerulein stimulation.

Role of connexin 43 in apoptosis of pancreatic cancer cell line BxPC3 / 中华胰腺病杂志

Objective To investigate the role of connexin 43(Cx43)in the apoptosis of pancreatic cancer cell line BxPC and its possible mechanism.Methods pcDNA-Cx43,pcDNA-Cx43N,pcDNA3.0,siRNA-Cx43 and siRNA-NC were transfected into BxPC3 cells via liposome method.Cx43 protein and Cytochrome C(Cyt C)concentration was determined by Western blot,and the apoptosis was analyzed by Annexin V/PI binding assay.The mitechondria apoptosis pathway involved in Cx43 associated apoptosis was examined which contains the depolarization of mitechondrial membrane potential (MMP);fluorospectrophotometer was used to measure the activities of caspase-3 and caspase-9. Gap junction intercellular communication(GJIC) was determined by dye-transfer method.Results Cx43 protein expression increased after BxPC3 transfeetion,apoptosis rate increased from(6.35±0.43)%in empty vector transfection group to(14.29±1.24)%;after H202 treatment,apoptosis rate increased from(20.34±2.47)%to(31.27±2.56)%(P<0.05).Meanwhile,mitochondrial membrane potential was decreased,Cyt C was increasingly released from mitochondria,caspases activities were increased;after siRNA43 interference,apoptosis rate decreased from(7.42±0.47)% to(5.19±1.37)%,after H_2O_2 treatment,apoptosis rate decreased from (19.43±1.71)%to(11.67±1.97)%(P<0.05).Decreased mitochondrial membrane potential and Cyt C release were observed,caspases activities were decreased.GJIC of pcDNA-Cx 43 transfection group increased from 14.52±0.57 to 23.05±3.84.and it increased from 1.70 ±0.24 to 3.84 ±0.45 in the presence of β-GA(P<0.05).But the apoptosis rate was not significantly different.Conclusions Cx43 could promote BxPC3 apoptosis via mitochondrial apoptotic signal pathway,and the possible mechanism included signal pathway other than GJIC.

Effects of ginsenoside Rg3 on growth and apoptosis of gastric cancer cell lines in vitro / 上海交通大学学报（医学版）

Objective To investigate the effects of ginsenoside Rg3 on growth and apoptosis of gastric cancer cell line MKN-45 and SGC-7901 in vitro. Methods MKN-45 and SGC-7901 cells at logarithmic growth phase were obtained, and were cultured with ginsenoside Rg3 of different concentrations (20, 30, 40, 50 μg/mL) for 24, 48 h or 24, 48 and 72 h. Cells cultured without ginsenoside Rg3 were served as controls. The inhibition rates of ginsenoside Rg3 on MKN-45 and SGC-7901 cells were detected by MTT assay, apoptosis rate of SGC-7901 cells was determined by Annexin V/PI double staining flow cytometry, cell cycles of SGC-7901 cells were analysed by flow cytometry, and morphological changes of SGC-7901 cells in 50 μg/mL ginsenoside Rg3 treatment group were observed by transmission electron microscopy. Results The inhibition rates on MKN-45 and SGC-7901 cells in each ginsenoside Rg3 treatment group were significantly higher than those in control group (P < 0.05), and the inhibition rates increased with the concentrations of ginsenoside Rg3 and time of culture ( P < 0.05). Compared with control group, the apoptosis rates of SGC-7901 cells and percentages of cells in G_1/G_1 cell cycle in each ginsenoside Rg3 treatment group were significantly increased in a concentration and time dependent manner. Typical morphology of SGC-7901 cell apoptosis was observed by transmission electron microscopy in 50 μg/mL ginsenoside Rg3 treatment group. Conclusion Ginsenoside Rg3 has significant inhibition effect on gastric cancer cell lines in vitro with a concentration and time dependent manner, the mechanism of which may involve the induction of gastric cell line apoptosis.

Study on X-linked inhibitor of apoptosis associated factor-1 suppressing xenograft growth in nude mice with hepatocellular carcinoma / 上海交通大学学报（医学版）

Objective To investigate the inhibitory effect of X-linked inhibitor of apoptosis associated factor-1(XAF1) on xenograft growth in nude mice with hepatocellular carcinoma. Methods The models of xenografted nude mice with human hepatocellular carcinoma cell line SMMC7721 were established. Intratumor injection was performed on three tumor sites in each group of mice (n=5) with recombinant adenovirus Ad5/F35-XAF1, control virus Ad5/F35-Null at the same infective titre or PBS of the same volume every two days for two weeks. The volumes of xenografts in all nude mice were measured every three days, and the differences between Ad5/F35-XAF1 group and the other two groups were compared. The apoptosis of tumor cells was determined by in situ end-labeling TUNEL method, the expression of XAF1 protein and microvessel density (MVD) were detected by immunohistochemistry. Results Intratumoral injection of Ad5/F35-XAF1 significantly inhibited the growth of tumor xenografts with smaller tumor size, less tumor weight and lower MVD compared with those injected with control virus Ad5/F35-Null and PBS (P<0.05 or P<0.01). However, the apoptosis index and expression of XAF1 protein in Ad5/F35-XAF1 group were significantly increased compared with the other two groups (P<0.01). Conclusion Ad5/F35-XAF1 significantly inhibits xenograft growth in nude mice with hepatocellular carcinoma, which is probably associated with the effects of XAF1 inducing hepatocellular carcinoma cell apoptosis and suppressing tumor angiogenesis.

Experimental study on relationship between Smad4 and angiogenesis related factors in human gastric cancer cell line / 中华消化杂志

Objective To study the effect of Smad4 gene on angiogenesis related factors in human gastric cancer cell line.Methods Recombinant eukaryotie expressing plasmid pcDNA3.1 (-)-Smad4 containing Smad4 gene and empty vector pcDNA3.1 (-) were introduced into human gastric cancer cell line MKN28 using lipofectam and selected by G418,respectively.Two cell lines were obtained as follows:Smad4+-MKN28 cell line which was MKN28 transfected with a stable hybrid containing Smad4 gene and Smad4--MKN28 cell line with empty plasmid as control.The transcription and expression of VEGF and TSP1 were investigated by RT-PCR and Western blot.Results The mRNA expression of TSP1 in Smad4+-MKN28 cells was higher (P＜0.05) than that in control cells,while VEGF was lower(P＜0.05).Western blot showed the consistent results as measurement by RT-PCR.Conclusion Smad4 restoration in gastric cancer cells reduced angiogenesis rates through down-regulation of angiogenesis activitor and up-regulation of angiogenesis inhibitor.

Effect of human telomerase antisense oligodeoxyribonucleotides on the growth in hibition of gastric cancer cell lines with well, moderate and poor differentiati on / 中华消化杂志

Objective To investigate the effect of specialized human tel omerase antisense oligodeoxyribonucleotides (AS-ODN) on the growth inhibition o f well, moderate and poor differentiated gastric cancer cell lines, and to explore its inhibitory mechanism and the correlation between the inhibition ratio of gastric cancer cells and differentiation of the tumor cells. Methods Under the given circumstances, three distinct differentiated gastric cancer cell lines were treated with AS-ODN. The telomerase a ctivities were measured by modified telomeric repeat amplification protocal assa y. The cell viability was detected by Trypan blue test, and the cell apoptosis was determined by cell morphological observation under light and electromicroscope, flow cytometry an d TUNEL assay. Results The telomerase activity and cell growth were apparently suppressed in MK N45 and SGC7901 cells, under defined concentrations of AS-ODN. Whilst, in MKN28 cells, only telomerase activity was suppressed at same concentration . There were no obvious changes in non-antisense oligomers treated group. The apoptotic features of MKN45 and SGC7901 were noticed by microscopic observa tion, TUNEL assay, after three distinct gastric cancer cell lines being continuo usly exposed to 10 ?mol/L AS-ODN for 96 h. Furthermore, the flow cytometric analy sis verified that the average apoptotic rate of MKN45 and SGC7901 was 44.75% and 33.56% respectively, but there were no obvious changes in non-antisense oli gomers treated group (P

Possible signal pathway of the apoptosis in gastric cancer cell lines induced by arsenic trioxide / 中华消化杂志

Objective To investigate the possible signal pathway of the apoptosis in gastric cancer cell lines(SGC 7901 and MKN 45)induced by arsenic trioxide. Methods TUNEL method was used to observe the influence of calcium antagonist, inhibitors of protein kinase C (PKC) and protein tyrosine kinase(PTK) on the apoptosis of gastric cancer cells induced by arsenic trioxide. Levels of cAMP, PKC and PTK were detected before and after the treatment with arsenic trioxide. Results Both PKC and PTK inhibitors could induce apoptosis of gastric cancer cell lines, also both of them had a cooperative action with arsenic trioxide in inducing apoptosis of gastric cancer cells, while calcium antagonist had no any effect on the apoptosis of gastric cancer cell lines. PKC and PTK levels decreased but cAMP level increased during the apoptosis of gastric cancer cells induced by arsenic trioxide ( P

Experimental study on apoptosis induced by pcDNA3-survivin-mutant in gastric cancer cell lines / 中华消化杂志

Objective Survivin is overexpressed in gastric cancer. However it not expressed in normal gastric mucosa. The expression of survivin is tightly related to the prognosis of gastric cancer.By gene reconstruction we generated pcDNA3 survivin mutant(Cys84Ala) plasmid, and observed its effect on the gastric carcinoma cell lines. Methods The survivin mRNA and protein expression levels were determined by reverse transcription polymerase chain reaction(RT PCR) analysis,Western blot and immunohistochemical staining respectively . Flowcytometry and acridine orange staning were employed to detect apoptosis. Results Overexpression of survivin mRNA and protein were detected in the gastric cancer cell lines. Inhibition of survivin by survivin mutant cDNA induced apoptosis,activated caspase 3 activity,cleaved PARP and promoted cytochrome C releasing in gastric cancer cells,and effectively sensitized gastric cancer cells to chemotherapeutic agents. Conclusion Inhibition of survivin may induce apoptosis in gastic cancer and sensitize gastric cancer cells to chemotherapeutic agents.Survivin targeted therapeutic protocol may potentially benefit gastric cancer therapy.

Effect of activated protein C on apoptosis in human umbilical vein endothelial cells induced by lipopolysaccharide / 中国病理生理杂志

AIM: To study the effect of activated protein C(APC) at different concentrations on apoptosis of human umbilical vein endothelial cells(HUVECs) induced by lipopolysaccharide(LPS).METHODS: The HUVECs were induced by LPS(1.0 mg/L) as apoptotic model that was administered by different concentration of APC(10 ?g/L or 50 ?g/L).Meanwhile,the control group and induced apoptosis group induced by LPS(1.0 mg/L) stimulation were also set up.The changes of cellular ultrastructures were observed under electron microscope.The DNA ladder and TUNEL fluorescent staining were measured in cells.Annexin-Ⅴ/PI double staining was used to measure the cell apoptosis rate by flow cytometry.Cell survival rate was measured by MTT assay.The proliferating cell nuclear antigen(PCNA) expression levels in cells were also measured by Western blotting to reflect the proliferation of the cells.RESULTS: There were significant apoptotic changes in the cells induced by LPS,but the apoptotic changes were reduced and apoptosis rates were decreased in the cells treated with APC.Meanwhile,cell survival rate and the protein levels of PCNA were increased after APC treatment,particularly at the concentration of 50 ?g/L,which showed difference when compared with those induced apoptosis group by LPS(P