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Compensated heart failure predisposes to outer medullary tubular injury: Studies in rats.BackgroundHeart failure (HF) is considered a putative factor predisposing to acute renal failure (ARF). Since outer medullary hypoxic injury may play an important role in the pathogenesis of acute tubular necrosis, we explored the impact of experimental HF on the propensity to develop ARF with hypoxic medullary injury following the inhibition of prostaglandin and nitric oxide synthesis.MethodsCompensated, high-output HF was induced in Sprague-Dawley rats by aorto-caval fistula. At the eighth to ninth postoperative day, the rats were injected with indomethacin and Nω nitro-L-arginine methyl ester (L-NAME; ARF protocol) and were sacrificed 24 hours later for morphologic evaluation.ResultsKidney function comparably declined in HF-ARF rats and in control sham operated animals (CTR-ARF). Nevertheless, outer medullary hypoxic damage with medullary thick ascending limb (mTAL) necrosis occurred almost exclusively in the HF-ARF group (11 ± 4% vs. 0.2 ± 0.2% of tubules in CTR-ARF, P < 0.03). In a third group of HF animals subjected to vehicles only (HF-Nil), kidney function was preserved and renal morphology remained intact. Papillary-tip necrosis was consistently found in all animals subjected to indomethacin and L-NAME, irrespective of preconditioning. Morphometric evaluation disclosed that HF was not associated with mTAL hypertrophy.ConclusionsIncipient HF predisposes to hypoxic outer medullary injury, probably reflecting the impact of regional vasoconstrictive stimuli rather than tubular hypertrophy when protective local vasodilating mechanisms are hampered. The presence and extent of outer medullary hypoxic damage cannot be predicted from the functional derangement, which in the experimental settings may also represent prerenal azotemia or papillary damage.

Nitric oxide (NO) exerts profound effects on renal physiology [1—41. Renal NO generation occurs in endothelial cells by the constitutive isoform of nitric oxide synthase (cNOS) and in macrophages, smooth muscle and mesangial cells by the inducible isoform (iNOS). The former system plays an important role in basal renal vascular tone and hemodynamic regulation, in mesangial contraction, renin release and tubuloglomerular feedback. The inducible NOS, widely distributed in the rat kidney, predominantly in the outer medulla and in cortical glomeruli [5], counterbalances vasoconstrictive, thrombogenic and proliferative stimuli associated with cytokine stimulation and inflammatory responses [6]. Intact nitrovasodilation is particularly important at the outer medulla, where tissue oxygen tension as low as 30 mm Hg normally exists [7]. Inhibition of NOS results in profound renal hypoperfusion, both at the cortex and outer medulla [8, 9], associated with substantial aggravation of medullary hypoxia [10]. Blockade of NO synthesis potentiates medullary hypoxic injury from radiocontrast and non-steroidal anti-inflammatory agents [9—111 and augments medullary hypoperfusion and damage during acute ureteral obstruction [121. Recently developed Clark-type selective NO microelectrodes enable real-time monitoring of NO concentrations in vitro in cell lines and incubated tissues [13—17], in organs maintained ex vivo [18] or in the living organism [19, 20]. With concomitant determination of intrarenal blood flow it could be a potent experimental tool for the evaluation of the role of NO in renal hemodynamics. In preliminary studies with the NO electrode, a paradoxical increase in medullary NO reading was noted following NOS inhibition. This led us to evaluate potential artifacts of NO monitoring, such as fluctuations in tissue oxygenation (since oxygen and reactive oxygen species scavenges NO with the production of NO2/NO3 , not sensed by the electrode) and temperature (which markedly affect the electrode current) [21]. Methods

Loop diuretics reduce hypoxic damage to proximal tubules of the isolated perfused rat kidney. The straight portion (S3) of the proximal tubule lies in close proximity to the thick ascending limbs (TALs) at the cortico-medullary junction. Since a delicate balance exists between oxygen demand and the limited oxygen supply in this region, we hypothesized that reduction of thick limb metabolic activity might augment oxygen availability to S3 segments, which depend heavily upon aerobic metabolism, and prevent hypoxic damage. The degree of functional deterioration and morphological damage to S3 was assessed in isolated rat kidneys perfused with an erythrocyte-free medium. Bumetanide (10−5M) and furosemide (10−4M) reduced S3 fragmentation from 9.8 ± 3.9% of tubules in controls to 0 and 1.4 ± 0.9%, respectively (P < 0.0005). Tubular glucose reabsorption was better preserved in kidneys exposed to loop diureties than in control kidneys (P < 0.01), and urinary alkaline phosphatase (P < 0.05) and the total amount of LDH released into the perfusate and urine (P < 0.01) were lower in the treatment groups. Morphological damage to S3 was closely correlated with medullary TAL necrosis (r = 0.66, P < 0.001), urinary alkaline phosphatase excretion (r = 0.89, P < 0.001) and glycosuria (r = 0.83, P < 0.001). We conclude that under hypoxic conditions TALs and S3 segments may compete with each other for a limited oxygen supply. Reduction of active transport in the mTAL might augment oxygen availability to S3 segments and improve their survival.

Hypercalcemie nephropathy: Chronic disease with predominant medullary inner stripe injury. Because of the recently observed augmentation of medullary hypoxic injury by calcium in isolated perfused rat kidneys (Kidney Int 34:186–194, 1988), renal morphology of chronic, prolonged hypercalcemia was investigated in vivo. Rats were treated with repeated injections of vitamin D2 (400,000 units/week) for two to eight weeks. Chronic elevation of plasma calcium from 2.1 to 2.8 mmol/liter (P < 0.001) was associated by a fall in maximal urine osmolality with no change in glomerular filtration rate. The most significant morphological alterations occurred in the inner stripe of the outer medulla; these changes were characterized by a sequence of active injury with subsequent destruction and atrophy of the medullary thick ascending limbs, fibroblastic and lymphocytic infiltration, and secondary dilatation of collecting ducts. Similar changes occurred in the medullary rays. These alterations were accompanied by increased renal prostanoid production and a predisposition to acute kidney failure from indomethacin. Because of its selective occurrence in zones of poorest oxygen supply, this inner stripe injury may derive from vulnerability to hypoxia and may play a role in some chronic nephropathies.

To study the interaction between calcium and the medullary hypoxic lesions found in isolated perfused rat kidneys, the acute effects of high extracellular calcium upon renal function and morphology were evaluated in kidneys perfused with cell-free medium at a total calcium concentration of 8 to 9 mg/dl (controls), 13 to 14 and 19 to 20 mg/dl (high Ca++). High Ca++ increased hypoxic damage to medullary thick ascending limbs from 58.2 +/- 4.0% of tubules in controls to 80.2 +/- 4.0% (P less than 0.005) in the deepest area of the outer medulla. Morphological changes in the cortex were minimal. The increase in damage to medullary thick limbs induced by high Ca++ was prevented by the calcium channel blocker verapamil. Addition of the calcium ionophore A23187 to controls reproduced the effects of high Ca++ with an increase in the proportion of damaged thick limbs to 92.1 +/- 4.1% (P less than 0.001 vs. controls). Addition of equimolar amounts of magnesium chloride did not reproduce the effect of high calcium perfusions. When transport activity was reduced with ouabain, high calcium perfusions were no longer associated with structural damage. In kidneys perfused with a medium enriched with amino acids, the proportion of tubules with severe, irreversible damage increased from 12 +/- 3 to 43 +/- 10% (P less than 0.01) after high calcium perfusion, and to 75 +/- 12% (P less than 0.001) after perfusion with the calcium ionophore. High extracellular and intracellular calcium appear to act in concert with hypoxia to increase the susceptibility of the renal medulla to injury by mechanisms potentially operative in hypercalcemic and ischemic nephropathy.(ABSTRACT TRUNCATED AT 250 WORDS)

Early medullary hypoxic injury from radiocontrast and indomethacin. We evaluated the acute changes in cortical and outer medullary oxygen tension and the alterations in renal function and morphology within the first 90 minutes after the administration of indomethacin and iothala-mate to anesthetized Sprague-Dawley rats. Both agents were found to produce marked and protracted outer medullary hypoxia averaging 12 ± 4 and 9 ± 2 mm Hg, respectively (mean ± SE). Given together to salt depleted uninephrectomized rats they produced an early hypoxic injury localized selectively in the outer medulla. This lesion progressed from 3 ± 1% of medullary thick ascending limbs (mTALs) at 15 minutes to 22 ± 7% at 24 hours. Condensed “dark” cells were observed at 15 minutes, probably representing a type of early injury. Residual red cell mass, quantified in the outer medullary vasculature of perfusion-fixed kidneys and presumably reflecting stasis, was substantially increased in iothalamate treated rats. Red cell mass in the interbundle zone correlated with mTAL necrosis. Taken together, these results show an early period of medullary hypoxia, accompanied by a selective injury to mTALs in the central interbundle zone with apparent stasis. These findings contrast sharply with the ischemia-reflow pattern of renal damage and emphasize the important role of medullary hypoxia in the genesis of acute renal failure in this model.