ANGIOTENSIN-II-INDUCED CELL HYPERTROPHY - POTENTIAL ROLE OF IMPAIRED PROTEOLYTIC ACTIVITY IN CULTURED LLC-PK1 CELLS

Background. Angiotensin II-induced hypertrophy of both mesangial and t ubular cells has been shown to be caused by enhanced protein synthesis . There are no data about its role on protein breakdown. Therefore, pr otein turnover and proteolytic activities were investigated in LLC-PK1 cells. Methods. Protein turnover was measured by determining the inco rporation and release of [C-14]phenylalanine; collagenolytic and gelat inolytic activities were assayed by using fluorogenic peptidyl substra tes. Results. Angiotensin II (10(-8)-10(-6)M) exerted a dose-dependent inhibition of collagenolytic and gelatinolytic activities, associated with reduction of protein degradation rate. In addition angiotensin I I stimulated protein synthesis in the cells. These combined effects on protein turnover resulted in an increase in both cell size and cell p rotein content (31.7% after 48 h). However, the rise of cell protein c ontent was only partly (48.0%) prevented by the protein synthesis inhi bitor cycloheximide (10(-5)M), which supports the role of decreased pr otein degradation in the angiotensin-II-induced cell hypertrophy. The angiotensin-II-induced effects on proteolytic activities as well as on cell protein content could be abolished by coincubation with the angi otensin II type I-receptor antagonist DuP 753 (10(-6)M). The calcium-c hannel blocker verapamil (10(-6)M) ameliorated the impairment of colla genolytic activity. On the contrary the calcium ionophore A23187 (10(- 6)M) mimicked the action of angiotensin II on this enzyme activity (co ntrol 34.5 +/- 1.9; angiotensin II 24.0 +/- 2.0; A23187 23.0 +/- 2.2 a nd angiotensin II + verapamil, 33.8 +/- 2.6 pmol/min/mu g DNA). The ro le of cytosolic [Ca2+] in the actions of angiotensin II could be final ly shown by a dose-dependent rise which was partly blunted by verapami l. Conclusion. The angiotensin-II-induced hypertrophy in LLC-PK1 cells is caused not only by enhanced protein synthesis but also by reduced protein degradation. The concomitant decline of collagenolytic and gel atinolytic activities may contribute to the accumulation of extracellu lar matrix, and presumably also to cell hypertrophy. These effects are obviously mediated via angiotensin II type I receptors and seem to be [Ca2+] dependent.