The guinea pig is unique in its physiologic and pancreatic morphological responses to high doses of diabetogen alloxan. Pancreatic B cells of alloxan-treated guinea pigs degenerate, with large numbers of islet cells demonstrating pyknotic nuclei, loss of over 50% of immunostainable B cells, and a marked reduction in serum insulin levels. This is followed, within 72 hrs, by reappearance of a full complement of immunohistochemically staining B cells, and, within 14 days, by restoration of normal serum in- sulin levels. The proposed research will explore the mechanism of B cell regeneration in the alloxan-treated guinea pig. 3H- thymidine labeling combined with immunohistochemistry will be employed to determine: A) whether the regenerated B cells arise mitotically; B) the nature of the progenitors of B cells; C) conditions (including age) which modulate the B cell regeneration. Since we have previously demonstrated that guinea pig B cells do not regenerate following alloxan treatment in vitro, we will also attempt to identify serum and pancreatic factors which can stimulate B-cell regeneration following alloxan-induced B-cell loss in cultures of guinea pig islet cells. Monoclonal antibodies will be raised to cell surface antigens of islets isolated from alloxan- treated regenerating guinea pig pancreas to determine if there is a unique cell population which serves as the source of the regenerated cells, and whether such cells are present in normal and diabetic human pancreas. Alloxan toxicity is due at least in part to free radical damage to B cells. Since guinea pig B cells are relatively resistant to low doses of alloxan, we will use in vitro techniques to determine whether they are resistant to free radical damage via elevated superoxide dismutase levels. Studies of B-cell regeneration and replication of B cells are of great importance to diabetes research. The cultured islet cell system provides both a useful model for the understanding of B-cell differentiation and regeneration following injury, and a sensitive in vitro bioassay for humoral and/or paracrine factors which may stimulate such processes.
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