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Aim: To explore the expression of triggering receptor expressed on myeloid cells (TREM-1) mRNA in acute pancreatitis (AP)., Methods: Using the reverse transcription polymerase chain reaction (RT-PCR), we examined the expression of TREM-1 mRNA in 10 cases of mild acute pancreatitis (MAP), 8 cases of severe acute pancreatitis (SAP), and 10 cases of healthy control subjects. And we also examined the expression of TREM-1 mRNA in 14 cases of AP (including 10 MAP and 4 SAP) before treatment, after successful therapy and clinically cured., Results: The expression of TREM-1 mRNA in the groups of MAP, SAP patients and healthy control subjects was 0.771+/-0.274, 1.092+/-0.331 and 0.459+/-0.175, respectively; there was a significant difference among the three groups (P<0.05). And there was also a significant difference between the AP patients (0.914+/-0.341) and healthy control subjects (0.459+/-0.175) (P<0.05). Moreover, in the 14 cases of AP, before treatment, after successful therapy and clinically cured, the expression of TREM-1 mRNA was 0.905+/-0.226, 0.739+/-0.169 and 0.633+/-0.140, respectively, and there was a significant difference among the three stages (P<0.05)., Conclusion: The expression of TREM-1 mRNA in the patients with AP increases obviously, and correlates with the degree of AP. Furthermore, the expression of TREM-1 mRNA is distinctly different at the different stages of AP. It indicates TREM-1 may play an important role in the occurrence and development of AP.

Objective: To investigate the antitumor effect of somatostatin receptor subtype 2 (SSTR2) gene transfection into pancreatic cancer cell line PC-3, and the mechanisms involved in this effect., Methods: The full-length human SSTR2 cDNA was introduced into the pancreatic cancer cell line PC-3 by lipofect amine-mediated transfection. Positive clones were screened by G418 and stable expression of SSTR2 was detected by immunocytochemistry SABC method and RT-PCR. Fifteen athymic mice were randomly divided into 3 groups of 5 mice to be xenografted with SSTR2-expressing cells (experimental group), empty vector control group, and control group cells respectively. The weight of mice and the size of tumor were measured every week. Eight weeks later the mice were killed and the tumors were taken out. TUNEL assay was used to determine the apoptotic index (AI) in these tumors. Immunohistochemistry SP method was used to determine the expressions of apoptosis regulating genes, Bcl-2 and Bax, and intratumoral. microvessel density (MVD). Moreover, the tumor volume and weight were compared among these three groups., Results: The AI was significantly higher in the experimental group (3.39% +/- 0.84%) compared with the empty vector control group (0.69% +/- 0.08%) and control group (0.68% +/- 0.09%) (both P < 0.05). The significant decrease in Bcl-2, and increase in Bax protein expressions were detected in the experimental group compared with the empty vector control group and control group (both P < 0.05). MVD was significantly lower in the experimental group (6.3 +/- 1.7) than those in the empty vector control group (12.6 +/- 1.7) and control group (13.5 +/- 1.9) (P < 0.05). Moreover, the tumor volume and weight were significantly lower in the experimental group as compared with the empty vector control group and control group (P < 0.05). However, no significant differences in all the indicators were observed between the empty vector control group and the control group (both P > 0.05)., Conclusions: Re-expression of SSTR2 gene, the expression of which is frequently lost in human pancreatic adenocarcinoma, can induce apoptosis which may be mediated via down-regulation of Bcl-2 and up-regulation of Bax, and inhibit tumor angiogenesis in pancreatic carcinoma, resulting in inhibition of tumor growth.

Aim: To investigate the anti-angiogenic effect of somatostatin receptor subtype 2 (SSTR2) gene transfer into pancreatic cancer cell line PC-3, and the mechanisms involved in this effect., Methods: The full length human SSTR2 cDNA was introduced into pancreatic cancer cell line PC-3 by lipofectamine-mediated transfection. Positive clones were screened by G418 and stable expression of SSTR2 was detected by immunohistochemistry SABC methods and RT-PCR. Enzyme-linked immunosorbent assay (ELISA) was used to detect vascular endothelial growth factor (VEGF) levels in the cell culture supernatants of SSTR2-expressing cells, vector control and mock control cells. Furthermore, the expressions of VEGF and matrix metalloproteinase-2 (MMP-2) were detected by immunohistochemistry SABC methods and RT-PCR in these cells., Results: VEGF levels in the cell culture supernatants were significantly reduced in the SSTR2-expressing cells (first week, 172.63+/-21.2 ng/L and after two months, 198.85+/-26.44 ng/L) compared with the vector control (first week, 790.39+/-86.52 ng/L and after two months, 795.69+/-72.35 ng/L) and mock control (first week, 786.42+/-90.62 ng/L and after two months, 805.32+/-84.36 ng/L) (P<0.05). The immunohistochemical assay showed a significant reduction of the integral optical density of VEGF and MMP-2 in the SSTR2-expressing cells (42.25+/-8.6 and 70.5+/-6.25, respectively) compared with the vector control (85.75+/-12.9 and 110.52+/-13.5, respectively) and mock control (82.6+/-9.28 and 113.56+/-9.62, respectively) (P<0.05). Conversely, the average gray value of VEGF and MMP-2 was significantly increased in the SSTR2-expressing cells (121.56+/-8.43 and 134.46+/-19.95, respectively) compared with the vector control (55.72+/-5.6 and 62.26+/-12.68, respectively) and mock control cells (58.48+/-6.2 and 65.49+/-9.16, respectively) (P<0.05). Moreover, the expressions of VEGF mRNA and MMP-2 mRNA were significantly reduced in the SSTR2-expressing cells (0.1384+/-0.017 and 0.2343+/-0.070, respectively) compared with the vector control (1.024+/-0.117 and 0.806+/-0.119, respectively) and mock control (1.085+/-0.105 and 0.714+/-0.079, respectively) (P<0.05)., Conclusion: The expression of reintroduced human SSTR2 gene exerts its antiangiogenic effects by down-regulating the expressions of the factors involved in tumor angiogenesis and metastasis, suggesting SSTR2 gene transfer as a new strategy of gene therapy for pancreatic cancer.

Aim: To evaluate the roles and mechanisms of celecoxib in inducing proliferation inhibition and apoptosis of human cholangiocarcinoma cell lines., Methods: Cyclooxygenase-2-overexpressing human cholangiocarcinoma cell line QBC939 and cyclooxygenase-2-deficient human cholangiocarcinoma cell line SK-CHA-1 were used in the present study. The anti-proliferative effect was measured by methabenzthiazuron (MTT) assay; apoptosis was determined by transferase-mediated dUTP nick end labeling (TUNEL) detection and transmission electron microscopy (TEM). Cell cycle was analyzed by flow cytometry (FCM). The PGE(2) levels in the supernatant of cultured cholangiocarcinoma cells were quantitated by enzyme-linked immunoabsordent assay (ELISA)., Results: Celecoxib suppressed the production of PGE(2) and inhibited the growth of QBC939 cells. Celecoxib at 10, 20, and 40 micromol/L inhibited PGE(2) production by 26 %, 58 %, and 74 % in QBC939 cells. The PGE(2) level was much lower constitutively in SK-CHA-1 cells (18.6+/-3.2) compared with that in QBC939 (121.9+/-5.6) cells (P<0.01) and celecoxib had no significant influence on PGE(2) level in the SK-CHA-1 cells. The PGE(2) concentration in SK-CHA-1 cells also reduced but not significantly after treatment with celecoxib. The PGE(2) concentration in SK-CHA-1 cells was (16.5+/-2.9) ng/well, (14.8+/-3.4) ng/well, (13.2+/-2.0) ng/well and (12.6+/-3.1) ng/well respectively, when pre-treated with 1 micromol/L, 10 micromol/L, 20 micromol/L and 40 micromol/L of celecoxib for 48 h (P>0.05, vs control). The anti-proliferation effect of celecoxib (20 micromol/L) on QBC939 cells was time-dependent, it was noticeable on day 2 (OD490=0.23+/-0.04) and became obvious on day 3 (OD490=0.31+/-0.07) to day 4 (OD490= 0.25+/-0.06), and the OD490 in the control group (day 1) was 0.12+/-0.03 (P<0.01, vs control). The anti-proliferation effect of celecoxib could be abolished by the addition of 200 pg/mL PGE(2). The proliferation of SK-CHA-1 cells was inhibited slightly by celecoxib, the cell density OD490 in the presence of celecoxib and in control group was 0.31+/-0.04 and 0.42+/-0.03 respectively on day 2 (P>0.05), 0.58+/-0.07 and 0.67+/-0.09 respectively on day 3 (P>0.05), and 0.71+/-0.08 and 0.78+/-0.06 respectively on day 4 (P>0.05). Celecoxib induced proliferation inhibition and apoptosis by G(1)-S cell cycle arrest: the percentage of QBC939 cells in G(0)-G(1) phase after treatment with 40 micromol/L (74.66+/-6.21) and 20 micromol/L (68.63+/-4.36) celecoxib increased significantly compared with control cells (54.41+/-5.12, P<0.01). The percentage of SK-CHA-1 cells in G(0)-G(1) phase after treatment with various concentrations of celecoxib didn't change significantly compared with control cells. The TUNEL index was much higher in QBC939 cells treated with 20 micromol/L celecoxib for 2 d (0.063+/-0.018) and for 4 d (0.102+/-0.037) compared with control cells (0.017+/-0.004, P<0.01)., Conclusion: The current in vitro study indicates that inhibition of proliferation and induction of apoptosis in human cholangiocarcinoma cells by cyclooxygenase-2 specific inhibitor celecoxib may involve in COX-dependent mechanisms and PGE(2) pathway. Celecoxib as a chemopreventive and chemotherapeutic agent might be effective primarily on COX-2-expressing cholangiocarcinoma.

Aim: To explore the effects of COX-2 gene in the proliferative activity induced by bile from anomalous pancreaticobiliary ductal union (APBDU) on human cholangiocacinoma cell line., Methods: Bile sample from APBDU and normal bile sample were used for this study. The proliferative effect of bile was measured by methabenzthiazuron (MTT) assay; COX-2 mRNA was examined by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). Cell cycle was analyzed by flow cytometry (FCM), and the PGE(2) levels in the supernatant of cultured cholangiocarcinoma cells were quantitated by enzyme-linked immunoabsordent assay (ELISA)., Results: Bile from APBDU can significantly promote the proliferation of human cholangiocarcinoma QBC939 cells compared with normal bile (P=0.005) and up-regulated remarkably their COX-2 mRNA expression (P=0.004). The proliferative activity of APBDU bile can be abolished by addition of cyclooxygenase-2 specific inhibitor celecoxib., Conclusion: Bile from APBDU can promote the proliferation of human cholangiocarcinoma QBC939 cells via COX-2 pathway.

Aim: To explore the potential carcinogenicity of bile from congenital choledochal cyst (CCC) patients and the mechanism of the carcinogenesis in congenital choledochal cyst patients., Methods: 20 bile samples from congenital choledochal cyst patients and 10 normal control bile samples were used for this study. The proliferative effect of bile was measured by using Methabenzthiazuron (MTT) assay; Cell cycle and apoptosis were analyzed by using flow cytometry (FCM), and the PGE(2) levels in the supernatant of cultured cholangiocarcinoma cells were quantitated by enzyme-linked immunoabsorbent assay (ELISA)., Results: CCC bile could significantly promote the proliferation of human cholangiocarcinoma QBC939 cells compared with normal bile (P=0.001) and negative control group (P=0.002), and the proliferative effect of CCC bile could be abolished by addition of cyclooxygenase-2 specific inhibitor celecoxib (20 microM). The QBC939 cells proliferative index was increased significantly after treated with 1 % bile from CCC patient (P=0.008) for 24 h, the percentage of S phase (29.48+/-3.27)% was increased remarkably (P<0.001) compared with normal bile (11.72+/-2.70) %, and the percentage of G0/G1 phase (54.19+/-9.46) % was decreased remarkably (P=0.042) compared with normal bile (69.16+/-10.88) %, however, bile from CCC patient had no significant influence on apoptosis of QBC939 cells (P=0.719)., Conclusion: Bile from congenital choledochal cyst patients can promote the proliferation of human cholangiocarcinoma QBC939 cells via COX-2 and PGE(2) pathway.

Objective: To investigate the expression of cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) in extra-hepatic cholangiocarcinoma and the relationship between their expression and clinicopathological parameters., Methods: COX-1 and COX-2 were detected in 56 extra-hepatic cholangiocarcinomas, including 31 matched tissues originating from non-tumorous bile ductal tissue adjacent to tumours and 6 normal bile ductal tissues, by immunohistochemistry strept avidin-biotin complex using isozyme selective antibodies., Results: There was no difference in expression of COX-1 between carcinomas (96%, 54/56) and noncancerous specimens (94%, 29/31, P>0.05) or normal bile ductal tissues (100%, 6/6, P>0.05). The positive rate of COX-2 expression in extra-hepatic cholangiocarcinomas (86%, 48/56) was significantly higher than their matched tissues (39%, 12/31, P<0.01) and normal bile ductal tissues (0%, 0/6, P<0.01). Overexpression of COX-2 in extra-hepatic cholangiocarcinoma was related to the metastasis of lymph nodes, distant organs or tissues (P<0.05) as well as the degree of tumour differentiation (P<0.05)., Conclusions: The overexpression of COX-2 plays a crucial role in the carcinogenesis and development of extra-hepatic cholangiocarcinoma, indicating that COX-2 may serve as a target for chemoprevention of extra-hepatic cholangiocarcinoma.