Oxygen deprivation-induced injury to isolated rabbit kidney tubules.

Abstract

The utility of freshly isolated suspensions of rabbit tubules enriched in proximal segments for studying the pathogenesis of oxygen deprivation-induced renal tubular cell injury was evaluated. Oxygenated control preparations exhibited very good stability of critical cell injury-related metabolic parameters including oxygen consumption, cell cation homeostasis, and adenine nucleotide metabolism for periods in excess of 2 h. Highly reproducible models of oxygen deprivation-induced injury and recovery were developed and alterations of injury-related metabolic parameters in these models were characterized in detail. When oxygen deprivation was produced under hypoxic conditions, tubules sustained widespread lethal cell injury and associated metabolic alterations within 15-30 min. However, when oxygen deprivation was produced under simulated ischemic conditions, tubules tolerated 30-60 min with only moderate amounts of lethal cell injury occurring, a situation similar to that seen with ischemia in vivo. Like ischemia in vivo, simulated ischemia in vitro was characterized by a fall in pH during oxygen deprivation. No such fall in pH occurred in the hypoxic model. To test whether this fall in pH could contribute to the protection seen during simulated ischemia in vitro, tubules were subjected to hypoxia at medium pHs ranging from 7.45 to 6.41. Striking protection from hypoxic injury was seen as pH was reduced with maximal protection occurring in tubules made hypoxic at pHs below 7.0. Measurements of injury-associated metabolic parameters suggested that the protective effect of reduced pH may be mediated by pHinduced alterations of tubule cell Ca++ metabolism. This study has, thus, defined and characterized in detail a new and extremely versatile model system for the study of oxygen deprivation-induced cell injury in the kidney and has established that pH alterations play a major role in modulating such injury.effects of such injury on nephronal function and renal hemodynamics, and the effects of these factors on renal tubular cell viability and functional integrity (1, 2). Although patterns of injury differ somewhat between animal models and human ischemic acute renal failure (3, 4), tubular cell injury appears to be a common and necessary event for ischemic acute renal failure in each of these settings, and increasing interest has focused on its pathogenesis over the past several years. Furthermore, a variety ofprotective maneuvers with apparent efficacy to ameliorate renal tubular cell injury and the associated acute renal failure including treatment with osmotic agents, calcium channel blockers, and adenine nucleotides have been reported (5-7). However, the structural heterogeneity of the kidney and the reciprocal interactions between tubular cell injury and nephronal and hemodynamic processes occurring in vivo have limited information about pathogenetic mechanisms oftubular cell injury and protection from it that can be obtained from studies in vivo. Isolated tubule preparations studied both as individually ...

Cellular acidification occurs during anoxia in cultured, but not in freshly isolated, rabbit proximal tubular cells

“…Another gradient which is influenced by pH is that of Ca2+ [26]. Under normal physiological conditions, pHj and [Ca2 + ] i appear to be closely linked; a decrease in pH; reduces transmem brane Ca2 + fluxes in cardiomyoctyes and an increase in pi 11 promotes ( 'a2 ' -mediated processes during differ entiation of the cell [26]. Low pHj has also been shown between cultured and freshly isolated PT cells.…”

“…Cytosolic acidosis decreases plasma membrane conductance pathways and, as a re sult, cellular K ' docs not decrease during chemical anoxia in hepatocytes [6], or during anoxia in hepa tocytes and rabbit PTs [19]. Another gradient which is influenced by pH is that of Ca2+ [26]. Under normal physiological conditions, pHj and [Ca2 + ] i appear to be closely linked; a decrease in pH; reduces transmem brane Ca2 + fluxes in cardiomyoctyes and an increase in pi 11 promotes ( 'a2 ' -mediated processes during differ entiation of the cell [26].…”

“…During 1 h of anoxia, cell viability of freshly isolated PT cells de creased significantly to 54% ± 2% (P < 0.05), while no loss in viability was observed in cultured PT cells. Clamping the pH; during anoxia at 6.7 and 6.1 signifi-shown that acidification of hypoxic tissue, resulting from glycolytic lactate production, ATP hydrolysis and C 0 2 accumulation, can enhance the resistance to the damaging effects of O, deprivation in the kidney [7,24,[26][27][28][29], cardiomyocytes [3,16], and hepatocytes [10,11,13,18]. However, the mechanism behind the protec tion offered by lowering the pH is unknown.…”

“…In some cells, such as cardiomyocytes, acidosis has an energyconserving effect caused by a lower energy demand [16]. In addition, intracellular pH (pHj) plays a role in the preservation of ionic gradients across the plasma membrane during ATP depletion, by decreasing membrane conductance pathways of, for example Ca2+, or inhibition of Ca2 + -calmodulin-regulated processes [26]. Moreover, the mechanisms responsible for membrane and cell damage, such as 723 phospholipid and protein degradation by phospholipases and proteases, appear to be pH dependent, with maximal activity at or near physiological pH and minimal activity at acidotic pH [13].…”

“…Moreover, the mechanisms responsible for membrane and cell damage, such as 723 phospholipid and protein degradation by phospholipases and proteases, appear to be pH dependent, with maximal activity at or near physiological pH and minimal activity at acidotic pH [13]. Protection by extracellular acidosis has been shown to be mediated by intracellular acidification in hepatocytes [6], but until now this has not been confirmed in renal cells [7,24,26 Previous studies on anoxia-induced cell injury in proximal tubular (PT) cells revealed a striking differ ence in the sensitivity to anoxia between freshly iso lated and cultured PT cells [20,21]. The fact that cultured cells were more resistant to anoxia could be due to the presence of a protective factor such as, for example, intracellular acidosis.…”