Development and characterization of pituitary GH3 cell clones stably transfected with a human proinsulin cDNA

C. Meoni, F. Bertuzzi, A. E. Pontiroli, L. Falqui, L. Monaco, M. Soria, C. Arcelloni, R. Paroni, C. Foglieni, L. Polastri, F. Galbiati, F. Folli, A. M. Davalli

Research output: Contribution to journalArticle

6 Scopus citations

Abstract

Successful β-cell replacement therapy in insulin-dependent (type I) diabetes is hindered by the scarcity of human donor tissue and by the recurrence of autoimmune destruction of transplanted β cells. Availability of non-β cells, capable of releasing insulin and escaping autoimmune recognition, would therefore be important for diabetes cell therapy. We developed rat pituitary GH3 cells stably transfected with a furin-cleavable human proinsulin cDNA linked to the rat PRL promoter. Two clones (InsGH3/clone 1 and 7) were characterized in vitro with regard to basal and stimulated insulin release and proinsulin transgene expression. Mature insulin secretion was obtained in both clones, accounting for about 40% of total released (pro)insulin-like products. Immunocytochemistry of InsGH3 cells showed a cytoplasmic granular insulin staining that colocalized with secretogranin II (SGII) immunoreactivity. InsGH3 cells/clone 7 contained and released in vitro significantly more insulin than clone 1. Secretagogue-stimulated insulin secretion was observed in both InsGH3 clones either under static or dynamic conditions, indicating that insulin was targeted also to the regulated secretory pathway. Proinsulin mRNA levels were elevated in InsGH3 cells, being significantly higher than in βTC3 cells. Moreover, proinsulin gene expression increased in response to various stimuli, thereby showing the regulation of the transfected gene at the transcriptional level. In conclusion, these data point to InsGH3 cells as a potential β-cell surrogate even though additional engineering is required to instruct them to release insulin in response to physiologic stimulations.

Original languageEnglish (US)
Pages (from-to)829-840
Number of pages12
JournalCell Transplantation
Volume9
Issue number6
DOIs
StatePublished - Jan 1 2000

Keywords

  • Bioengineering
  • Insulin secretion
  • Pituitary cells

ASJC Scopus subject areas

  • Biomedical Engineering
  • Cell Biology
  • Transplantation

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    Meoni, C., Bertuzzi, F., Pontiroli, A. E., Falqui, L., Monaco, L., Soria, M., Arcelloni, C., Paroni, R., Foglieni, C., Polastri, L., Galbiati, F., Folli, F., & Davalli, A. M. (2000). Development and characterization of pituitary GH3 cell clones stably transfected with a human proinsulin cDNA. Cell Transplantation, 9(6), 829-840. https://doi.org/10.1177/096368970000900609