Endopin serpin protease inhibitors localize with neuropeptides in secretory vesicles and neuroendocrine tissues

Background/aims: The endopin serpin protease inhibitors have been identified by molecular studies as components of secretory vesicles that produce neuropeptides. Endopin 1 inhibits trypsin-like serine proteases, and endopin 2 inhibits cathepsin L that produces neuropeptides in secretory vesicles. To assess the secretory vesicle and neuroendocrine tissue distribution of these endopins, the goal of this study was to define specific antisera for each endopin isoform and to examine their localization with neuropeptides and in neuroendocrine tissues.

Methods: This study utilized methods consisting of Western blots, immunoelectron microscopy, and immunofluorescence microscopy for evaluation of the localization of endopin protease inhibitors in neuroendocrine tissues.

Results: Immunoelectron microscopy with these selective antisera demonstrated the localization of endopins 1 and 2 within secretory vesicles of adrenal medulla (bovine). Cellular immunofluorescence confocal microscopy illustrated the high level of colocalization of endopins 1 and 2 with enkephalin and NPY neuropeptides that are present in secretory vesicles of adrenal medullary chromaffin cells in primary culture. Tissue distribution studies (by Western blots) showed the expression of endopins 1 and 2 in bovine brain, pituitary, adrenal medulla, and other neuroendocrine tissues.

Conclusions: These results implicate endopins 1 and 2 as endogenous protease inhibitors in neuropeptide-containing secretory vesicles and neuroendocrine tissues.