Adenovirus-mediated gene expression in isolated rat pancreatic acini and individual pancreatic acinar cells.
In this study we have examined the feasibility of using replication-deficient recombinant adenoviral vectors to transfer and express genes in pancreatic acinar cells in vitro. We infected primary cultures of both isolated pancreatic acini and individual acinar cells with a recombinant adenovirus containing the coding sequence for beta-galactosidase. Our data demonstrate that recombinant adenoviruses readily infect pancreatic acinar cells in vitro. Close to 100% infection and maximal beta-galactosidase expression were obtained, when acini or acinar cells were infected with 5x10(6) or 10(6) plaque-forming units (pfu) of virus per millitre of acini or acinar cell suspension, respectively. Examination of the time-course of beta-galactosidase expression showed that there was a lag of approximately 6 h before beta-galactosidase levels increased. Thereafter beta-galactosidase expression increased rapidly. By 20 h post-infection beta-galactosidase activity had increased from undetectable levels to 2.5-3.0 units/mg of cellular protein. Acini/acinar cells maintained a robust secretory response after adenoviral infection. The cholecystokinin-octapeptide (CCK8) dose/response curves for amylase secretion for acini and acinar cells infected with 5x10(5) and 1x10(5) pfu/ml of virus, respectively, were biphasic, with maximal amylase secretion being stimulated by 1 nM CCK8. In addition, the dose/response curves were identical to those obtained from control, sham-infected, acini/acinar cells. Our findings indicate that replication-deficient recombinant adenoviral vectors will be excellent tools to transfer and express genes in isolated pancreatic acini or acinar cells.
Padfield
P J
PJ
Department of Pathology, Saint Louis University Health Sciences Center, St. Louis, MO, USA. padfield@wpogate.slu.edu
Elliott
A C
AC
Baldassare
J J
JJ
D000256:Adenoviridae / Q000235:genetics*; D000681:Amylases / Q000032:analysis / Q000187:drug effects / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D002766:Cholecystokinin / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005089:Exocytosis; D015870:Gene Expression / Q000235:genetics*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D008954:Models, Biological; D010179:Pancreas / Q000166:cytology* / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection / Q000379:methods; D019076:Transgenes / Q000235:genetics; D001616:beta-Galactosidase / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics
D011994:Recombinant Proteins; D002766:Cholecystokinin; D000681:Amylases; D001616:beta-Galactosidase