Characterization of cultured human nesidioblasts and their associated macromolecules: cross-reactivity with a cloned rat pancreatic beta-cell line.

Abstract

Near-total pancreatectomy in a neonate presenting with persistent hypoglycemia offered an unusual opportunity to grow preparations enriched in beta-cells. Morphology, chromosomal analysis, and immunohistochemistry were used to characterize a subculture (Nesi B) that remained stable through passage 11. Insulin secretion of Nesi B was constitutive at 38-74 nU/microliters of medium/24 h, increasing modestly in the presence of isobutylmethylxanthine, an inhibitor of cyclic AMP phosphodiesterase. Endocrine cells, exclusive of those in islet regions, were widely distributed throughout the original tissue section, and immunostaining of the Nesi B subculture demonstrated well-differentiated heavily granulated insulin-positive cells, each with a normal modal number of chromosomes (46). Nesi B cell-associated macromolecules were isolated in 3 x 10(-3) ethylenedinitrilotetraacetate/phosphate-buffered saline and were found to be reactive with a heterologous immune serum elicited to a cloned rat pancreatic beta-cell line (RIN-5F). Western (immuno) blotting showed this immunoreactivity to reside primarily in a 95-kDa fraction of Nesi B-derived components. These results indicate that human nesidioblasts can be cultured for a sufficient number of passages to allow isolation and immunochemical characterization of pancreatic beta-cell macromolecules shared between rat and human and that may serve as organ-specific antigens for inflammatory disorders of the pancreas.