Rat serum improves rat pseudoislet formation and insulin gene expression.
Rat islet cells in culture are able to form tridimensional aggregates with an architecture and functional activity similar to native islets: pseudoislets. Pseudoislets represent an alternative source for islet transplantation, because their transplant results in a long term allograft acceptance without immunosuppression of the host. Use of pseudoislets has been limited by their reduced yield and by poor reaggregation mass. Since culture conditions have been reported to affect reaggregation, the aim of this study was to evaluate the effects of different concentrations of two sera (Fetal Bovine Serum [FBS] and Rat Serum [RS]) on reaggregation and insulin gene expression in pseudoislets. Islets were isolated from male Lewis rat by means of histopaque gradient centrifugation. The day after islets were disrupted into single cells and cultured in RPMI 1640 5.6 mM glucose with 2%, 5% and 10% solutions of both FBS and RS. Cells spontaneously reaggregated to form pseudoislets. After seven days of culture, pseudoislets were counted and analysed for insulin secretion and insulin gene expression using RT-PCR. Rat serum increased the number of aggregates and their diameters. Insulin gene expression of pseudoislets cultured with RS showed a ten fold increase in comparison to those cultured with FBS. These data show that the culture medium supplemented with RS improves total reaggregate volume and increases insulin gene expression. With the perspective of pseudoislets' use in transplantation RS is better indicated than FBS for the production of rat pseudoislets.
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DI.M.I., Dipartimento di Medicina Interna, Università di Genova.
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D000818:Animals; D002449:Cell Aggregation; D018929:Cell Culture Techniques / Q000379:methods*; D002478:Cells, Cultured; D003470:Culture Media; D015870:Gene Expression; D007328:Insulin / Q000235:genetics*; D007515:Islets of Langerhans / Q000166:cytology* / Q000378:metabolism; D051381:Rats
D003470:Culture Media; D007328:Insulin