Inhibition Effect Of Somatostatin Receptor Subtype 2 (SSTR2) Transfection On The Human Pancreatic Adenocarcinoma Cells

Background:Pancreatic carcinoma is the fourth leading cause of cancer related deaths in Western countries, Despite advances in surgery, chemotherapy, and radiation therapy in recent decades, the mortality rate for pancreatic cancer remains equal to its incidence, with an overall 1-year survival rate of 12% or so. It have been demonstrated that somatostatin inhibits the growth of pancreatic cancers expressing somatostatin receptor subtype 2 (SSTR2) but not receptor-negative cancers. SSTR2 is expressed in normal human endocrine pancreas but not in exocrine pancreatic carcinomas and their metastasis as well as in most of human pancreatic cancer cell lines. SSTR2 expression may be an important tumor suppressor pathway that is lost in human pancreatic cancer.Objectiv:We corrected the SSTR2 defect in human pancreatic cancer BxPC-3 cells by stable transfection with adenoviral vector containing the SSTR2, investigated its effect on tumorigenicity of human pancreatic cancer, elucidated the underlying mechanisms. We evaluated transductional efficiency of adenoviral vector bearing COX-2 promoter and SSTR2 gene in pancreatic cancer. Matrine was combined with SSTR2 gene expression to observe whether it can potentiate the inhibition of cell proliferation and to elucidate the underlying mechanisms.Methods:We used AdCAG-SSTR2 and empty vector transfected so- matostatin receptor-negative human pancreatic cancer cells (BxPC-3) respectively. The expression of SSTR2 were detected by RT-PCR and Western blot. Negative contral, vector control, and SSTR2 transfected cells were cultured and the rate of cell growth was determined by direct cell counting using a hemacytometer. The apoptosis was assessed by flow cytometric Annexin V/PI labelling assay. The expression of PCNA, Fas, Bax, Bcl-xl, VEGF and MMP9 were detected by Real Time PCR.AdCOX-2L-SSTR2 and AdCAG-SSTR2 transfected human pancreatic cancer cells (BxPC-3) respectively. The expression of SSTR2 mRNA and protein was detected by RT-PCR and Western blot. The expression of Bax, VEGF were detected by Real Time PCR.The apoptosis was assessed by flow cytometric Annexin V/PI labelling assay in negative control, AdCAG-SSTR2 transfection, administration of Matrine and AdCAG-SSTR2 transfection combining administration of Matrine respectively. The expression of Bax, Bcl-xl were detected by Real Time PCR.Results:The stable expression of SSTR2 was detected in the cells transfected by AdCAG-SSR2. The significant decrease of pancreatic cancer cell growth was detected in experimental group compared with control groups(P< 0.01).The ratio of apoptosis significantly increased in cells re-expressing SSTR2 (9.45%) than control group (P<0.01).The increase of Fas, Bax(1.99,3.18fold) and decrease of Bcl-xl, VEGF, MMP9(0.34,0.23 & 0.30fold) were detected by Real Time PCR.The stable expression of SSTR2 wasn't detected in the cells transfected by AdCOX-2L-SSTR2, and no effect on the expression of Bax and VEGF was found.The ratio of apoptosis was 5.86±1.01%, 9.45±0.87%,11.92±0.56% and 0.33±0.02% in administration of Matrine group, AdCAG-SSR2 transfection group, administration of Matrine combining with AdCAG-SSR2 trans- fection group and control group respectively. There was significant di- fference between administration of Matrine group and AdCAG-SSR2 transfection group; the ratio of apoptosis in administration of Matrine combining with AdCAG-SSR2 transfection group was significantly higher than in administration of Matrine group. The decrease of Bcl-xl was detected in administration of Matrine group (0.67 fold) and administration of Matrine combining with AdCAG-SSR2 transfection group (0.19 fold).The expression of Bax hadn't changed in administration of Matrine group (1.23 fold). There was no significant different expression of Bax between administration of Matrine group and administration of Matrine combining with AdCAG-SSR2 transfection group (3.18 vs 3.21 fold).Conclusions:SSTR2 reversed tumorigenicity of human pancreatic cancer cells. The mechanisms of inducing apoptosis may involved in up- regulating the expression of Fas,Bax and down-regulating the ex- pression of Bcl-xl. The decrease of VEGF and MMP9 may predict response to therapy of anti-angiogenesis in pancreatic carcinoma with SSTR2. COX-2 promoter havn't represented advantage in promoting the expression of SSTR2 in BxPC-3. Administration of Matrine elevated the ratio of apoptosis and potentiated the effect of AdCAG-SSR2, The mechanisms of inducing apoptosis may be due to the down-regulating the expression of Bcl-xl. Administration of Matrine showed potential therapy effect in pancreatic carcinoma.