[Effects of blocking the TGF alpha-EGFR autocrine loop on the growth of human pancreatic carcinoma cells in vitro].
OBJECTIVE
Investigate the effects of blocking the TGF alpha-EGFR autocrine loop on the growth of human pancreatic carcinoma.


METHODS
The pCMV-AS-EGFR, a recombinant vector expressing antisense EGFR under the control of human CMV promotor was constructed and transfected the transformant PC-7/AS-TGF alpha cell line cells, which had been transfected by a recombinant retroviral vector expressing antisense TGF alpha. G418 resistant colonies were isolated and identified as PC-7/AS-TGF alpha/AS-EGFR. The integration and expression of exogenous and endogenous genes were detected by Southern blot and Northern blot analysis. Apoptosis was detected by DNA fragmentation, flow cytometry and in situ cell death detection.


RESULTS
The cells which were double-transfected by the recombinant vectors showed the integration and expression of exogenous genes, and the downregulation of endogenous EGFR and cyclin D1 mRNA. 125I-EGF ligand binding test showed the binding affinity of the EGFR on the cell surface also reduced. The inhibition effect of cotransfecting of antisense TGF alpha and antisense EGFR was remarkable as compared with that of antisense TGF alpha alone. The incorporation rate of 3H-TdR reduced from 25% to 14.5%, and the growth inhibition rate increased from 78.6% to 86.0%. The ability of soft agar colony-formation was completely suppressed.


CONCLUSIONS
These observations strongly support that the blockage of the expression of autocrine growth factor TGF alpha and its receptor EGFR was a potent way in circumventing the malignant properties of the pancreatic carcinoma cells.
Xu
Y
Y
Department of Pathology, PUMC Hospital, CAMS and PUMC, Beijing 100730.
Liu
T
T
Gao
J
J
D002455:Cell Division; D002461:Cell Line, Transformed; D019938:Cyclin D1 / Q000096:biosynthesis* / Q000235:genetics; D066246:ErbB Receptors / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D010190:Pancreatic Neoplasms / Q000378:metabolism / Q000473:pathology*; D016372:RNA, Antisense; D012333:RNA, Messenger / Q000235:genetics; D014162:Transfection; D014170:Transformation, Genetic; D016211:Transforming Growth Factor alpha / Q000096:biosynthesis* / Q000235:genetics; D014407:Tumor Cells, Cultured
D016372:RNA, Antisense; D012333:RNA, Messenger; D016211:Transforming Growth Factor alpha; D019938:Cyclin D1; D066246:ErbB Receptors