A novel pancreatic model: the snip method of pancreatic isolation for in vitro study.
We aimed to devise a method of preparing the pancreas for in vitro study that would provide tissue that is viable and functional for 24 h, with preservation of integral functional cell-membrane structures. Pancreata of NIH Swiss mice were excised, gently insufflated with media, and carefully snipped into portions of < 0.5 mm. Snips were incubated in cell culture for 0, 8, 24, and 48 h, with viability measured by 3-[4,5-dimethylthiazol-2-yl]-2,3-diphenyltetrazolium bromide (MTT) assay and amylase production quantified after stimulation with cerulein. Recombinant tumor necrosis factor-alpha (TNF-alpha) was added to cell culture, and apoptosis demonstrated by Hoechst staining at 0, 24, and 48 h. At 0, 8, and 24 h, pancreatic snips were determined to be viable by MTT assay. They also were functional with intact cell-membrane apparatuses at these same time points, as evidenced by amylase production in response to a cholecystokinin analogue. We were able to induce apoptosis with TNF-alpha ligand in these pancreatic snips in support of viability and overall cellular function. We conclude that the snip method provides an effective in vitro pancreatic model. Acinar cells are viable and functional for > or = 24 h, with evidence that their cell-surface receptors are preserved in their operational state.
Jaffrey
C
C
Department of Surgery, University of South Florida, Tampa 33601, USA.
Eichenbaum
D
D
Denham
D W
DW
Norman
J
J
D000681:Amylases / Q000096:biosynthesis; D000818:Animals; D017209:Apoptosis; D002470:Cell Survival; D002478:Cells, Cultured; D002108:Ceruletide / Q000494:pharmacology; D008297:Male; D051379:Mice; D008954:Models, Biological; D010179:Pancreas / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D011994:Recombinant Proteins / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D002108:Ceruletide; D000681:Amylases