Assessment of in vitro differentiation of bovine pancreatic tissue in insulin-expressing cells.

CONTEXT

Expansion and culture of beta cell progenitors in vitro may represent an alternative to the use of differentiated beta cells from donor pancreata.

OBJECTIVE

The aim of our study was to investigate to what extent exocrine or endocrine pancreatic cells can be differentiated in insulin-producing cells in vitro.

SETTING

Bovine exocrine tissue (n=4) and islets (n=4) were cultured in DMEM with serum.

INTERVENTIONS

After 7 days, the cells were trypsinized and cultured in the same medium for cell proliferation, or in DMEM/F-12 containing growth factors to induce cell differentiation.

MAIN OUTCOME MEASURE

Proliferating capacity after 4 weeks in culture. In addition, insulin expression was evaluated by RT-PCR and by immunohistochemical staining.

RESULTS

After 4 weeks of culture, cells from exocrine tissue showed a 69.5+/-10.0 fold increase, while cells from islets showed a 31.2+/-11.4 fold increase (P=0.059). In differentiating medium, monolayers from exocrine and islet tissue were organized into islet-like structures containing cells which stained positively for insulin. Morphometrical analysis and RT-PCR confirmed the presence of insulin in the cells at the protein and the mRNA level.

CONCLUSIONS

In our experimental conditions, cells from pancreatic tissue proliferated and differentiated in insulin-containing cells. However, the level of insulin as well as mRNA expression is only a small fraction of that shown by fresh islets. Only selective identification of cell precursors may allow efficient generation of insulin-producing cells in vitro.