ASYNCHRONOUS REFORMATION OF INDIVIDUAL KALLIKREIN-RELATED SECRETORY PROTEINASES IN RAT SUBMANDIBULAR GLANDS FOLLOWING DEGRANULATION BY CYCLOCYTIDINE

Time scales for the reformation of the secretory granules in granular tubules and their constituent proteinases were assessed after inducing a massive degranulation by intraperitoneal injection of cyclocytidine in conscious animals. The minimum working dose of cyclocytidine to pr oduce the maximum degranulation and depletion of proteinase activity, at 3h after injection, was 75 mg/kg. Histologically, although most gra nular tubule cells then appeared to be extensively degranulated, isola ted individual cells showing little or no degranulation always persist ed. Acinar cells also showed some depletion of secretory material. At 15 h after injecting cyclocytidine the formation of new granules had b egun in the granular tubule cells, but it was not extensive or uniform in adjacent r-ells; however, the acinar cells already appeared to be regranulated. The pattern of granule reformation in granular tubule ce lls progressed gradually, so that 7-10 days after cyclocytidine-induce d degranulation the cells were mostly packed with granules and showed similar appearances to those of normal resting control glands. Individ ual proteinases in extracts of the glands were assayed specifically us ing fluorogenic oligopeptide amidase substrates, with and without appr opriate inhibitors. This revealed a 95% reduction in total proteinase activity 3 h after cyclocytidine (75 mg/kg). In the same extracts, aci nar peroxidase was reduced by 28%. Peroxidase levels recovered to cont rol values within 15 h after cyclocytidine but recovery of proteinases progressed more gradually and did not occur uniformly for the differe nt constituent proteinases. Tissue kallikrein (rK1) showed the most ra pid recovery and had reached levels approaching normal within 3 days. The activities of other kallikrein-related proteinases took much longe r to recover and T-kininogenase (rK10) was still below normal levels a fter 7 days.