Iterative exposure of clonal BRIN-BD11 cells to ninhydrin enables selection of robust toxin-resistant cells but with decreased gene expression of insulin secretory function.
OBJECTIVE
Prevention of pancreatic beta-cell destruction combined with preservation of insulin secretory function is an important goal for cell-based diabetes therapy. This study describes the generation and characteristics of toxin-resistant beta-cells.


METHODS
By using iterative exposures to ninhydrin, a new class of robust ninhydrin-tolerant insulin-secreting BRIN-BD11 ninhydrin-tolerant (BRINnt) cells was generated. Low- and high-passage BRINnt cells were used to evaluate beta-cell function and tolerance against toxins in comparison with native BRIN-BD11 cells. Differences in viability, gene expression, insulin secretory function, antioxidant enzyme activity, DNA damage, and DNA repair efficiency were compared.


RESULTS
BRIN-BD11 ninhydrin-tolerant cells exhibited resistance toward ninhydrin and hydrogen peroxide but not streptozotocin (STZ). Both total superoxide dismutase (SOD) and catalase enzyme activities of BRINnt cells were significantly enhanced, and ninhydrin-induced DNA damage was decreased. BRIN-BD11 ninhydrin-tolerant cells also exhibited enhanced DNA repair efficiency. However, this was accompanied by loss of secretagogue-induced insulin release, decreased cellular insulin content, and deficits in insulin and glucose transporter 2 gene expression. Prolonged culture of BRINnt cells in the absence of ninhydrin reversed the degenerated function of BRINnt cells but restored ninhydrin susceptibility.


CONCLUSIONS
These data illustrate dissociation between beta-cell toxin resistance and secretory function, indicating difficulties in generation of robust and well-functioning cells using this approach.
Liu
Hui-Kang
HK
School of Biomedical Sciences, University of Ulster, Coleraine, Northern Ireland, UK.
McCluskey
Jane T
JT
McClenghan
Neville H
NH
Flatt
Peter R
PR
D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D002374:Catalase / Q000378:metabolism; D002999:Clone Cells; D004249:DNA Damage; D004260:DNA Repair / Q000187:drug effects; D004351:Drug Resistance; D015870:Gene Expression / Q000187:drug effects; D051273:Glucose Transporter Type 2 / Q000235:genetics; D006861:Hydrogen Peroxide / Q000633:toxicity; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology*; D009555:Ninhydrin / Q000494:pharmacology*; D051381:Rats; D013311:Streptozocin / Q000633:toxicity; D013482:Superoxide Dismutase / Q000378:metabolism
D000975:Antioxidants; D051273:Glucose Transporter Type 2; D007328:Insulin; C495683:Slc2a2 protein, rat; D013311:Streptozocin; D006861:Hydrogen Peroxide; D002374:Catalase; D013482:Superoxide Dismutase; D009555:Ninhydrin