Renal cell apoptosis in chronic obstructive uropathy: The roles of caspases

Background. Apoptosis of tubular and interstitial cells is well documented in kidneys with chronic obstructive uropathy (COU) and probably plays an im portant role in the pathogenesis of this condition. The molecular control o f apoptosis in COU remains poorly understood. Apoptosis in general is known to proceed initially along distinct pathways, which later converge into a common arm characterized by orderly activation of caspases. Caspases are cy tosolic enzymes that belong to a 12-member family and serve as effector mol ecules for apoptosis. The role of individual caspases in mediating renal ce ll apoptosis in kidneys with COU is studied. Methods. Kidneys were harvested from sham-operated mice and mice with COU c reated by left ureter ligation at days 4, 7, 15, 20, and 30. The following studies were performed: (1) determination of dried kidney weight; (2) in si tu end labeling of fragmented DNA to detect apoptotic tubular and interstit ial cells; (3) ribonuclease protection assay with specific anti-sense RNA p robes for caspases 1, 2, 3, 6, 7, 8, 9, 11, and 12 to detect the expression of individual caspases; (4) immunostaining for caspases; and (5) assay for caspase 3. To assess the role of caspases in COU-associated renal cell apo ptosis, the frequencies of apoptotic tubular and interstitial cells were se parately quantitated for each experimental time point, and their patterns o f variation were correlated with those of individual caspases. Results. The obstructed kidneys showed progressive tissue loss (60% of cont rol at day 15). Apoptosis of both tubular and interstitial cells was seen i n obstructed kidneys. Tubular cell apoptosis peaked at four days after uret er ligation (13-fold of control), remained high between days 4 to 15, and t hereafter decreased rapidly. Apoptotic interstitial cells were scanty initi ally, but gradually increased throughout the entire experiment. Apoptosis w as minimal throughout the experiment in control and contralateral kidneys. In control and contralateral kidneys, caspases 2, 3, 6, 7, 8, and 9 mRNAs w ere expressed at low levels, whereas those for caspases 1, 11, and 12 were not detected. The obstructed kidneys displayed increased expression of all tested caspases. Caspases 1, 11, and 12 mRNAs were detected in obstructed k idneys in a common pattern characterized by a sharp increase at day 4, foll owed by a decrease until day 20, and a subsequent sharp increase until the end of the study at day 30. A similar pattern was noted for other caspases (2, 3, 6, 7. 8, and 9), which maximally reached twofold to fourfold that of controls. Immunostaining for caspases 1, 2, 3, 6, 7, 8, and 9 showed the s ame pattern characterized by focal and weak expression in proximal tubules of control or contralateral kidney, contrasting with increased staining in atrophic or dilated tubules of obstructed kidneys. Interstitial cells also displayed staining for several caspases, which paralleled the increasing de nsity of interstitial cells toward the end of the experiment. Caspase-3 ass ay showed a marked increased activity in obstructed kidneys that reached fo urfold and sevenfold of control at days 4 and 30, respectively. The rise an d fall of caspase mRNAs between days 4 and 30 paralleled a similar fluctuat ion in tubular cell apoptosis. The subsequent increase of mRNAs was correla ted with a continuous rise of interstitial cell apoptosis. Conclusions. Urinary obstruction in mice induces apoptosis of both tubular and interstitial cells in the affected kidney in a distinctive pattern that parallels an increased expression of caspases. This correlation suggests t hat these caspases mediate COU-associated renal cell apoptosis. Among the e valuated caspases, increased renal caspase 3 activity implies its central r ole in renal cell apoptosis associated with urinary obstruction.