title: Glucocorticoid regulation of chloroquine nonsensitive insulin degradation in cultured fetal rat hepatocytes.
abstract: The influence of cortisol and other culture conditions on insulin degradation by the chloroquine-sensitive pathway and the chloroquine-nonsensitive pathway (CNP) was investigated in fetal rat hepatocytes during 3 days of culture. The proportions of the chloroquine nonsensitive release of 125I-insulin degradation products into the conditioned medium/h increased from the 1st to the 3rd day of culture, i.e. from 19 to 50% by cells grown in the presence of cortisol and from 17 to 82% by those grown in the absence of cortisol. Replacement of the conditioned medium with the respective fresh medium dramatically enhanced cellular insulin degradation by CNP, i.e. from 22 to 58%, and 19 to 85% in cells grown for 2 days in the presence and absence of cortisol, respectively. Thus, the conditioned medium contained some factor(s) that inhibited CNP. Therefore, we used the inhibited insulin and alpha-casein degradation by papain in vitro as an assay to investigate the nature of the putative anti-(insulin) protease. Cycloheximide completely prevented the appearance of anti-papain activity in the medium. Conditioned medium obtained from cells grown in the presence of cortisol contained about 2-fold more anti-papain activity than the medium that was obtained in the absence of the steroid. The release of anti-papain activity also declined with time from 1 to 3 days of culture and showed an inverse relationship with the magnitude of cellular insulin degradation by CNP. The inhibition of papain-mediated insulin degradation by the anti-(insulin) protease was noncompetitive. The anti-(insulin) protease was nondialyzable (up to the 10-kDa exclusion limit) and inactivated by heat treatment at 50 degrees C for 30 min. These results suggest that fetal hepatocytes synthesize and secrete a glucocorticoid-regulated heat-labile low molecular mass (less than 25 kDa) anti-(insulin) protease, which may contribute to the suppression of insulin degradation caused by the enzymes involved in CNP.
authors.affiliation: Laboratoire Interactions Cellulaires, U.E.R. Odontologie, Université Paris, France.
authors.forename: M
authors.initials: M
authors.lastname: Ali
authors.forename: C
authors.initials: C
authors.lastname: Plas
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D002738:Chloroquine / Q000494:pharmacology*; D003470:Culture Media; D005333:Fetus; D006854:Hydrocortisone / Q000494:pharmacology*; D007328:Insulin / Q000378:metabolism*; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D010206:Papain / Q000378:metabolism; D051381:Rats
chemical_list: D003470:Culture Media; D007328:Insulin; D002738:Chloroquine; D010206:Papain; D006854:Hydrocortisone