Temperature effects on ischemic and hypoxic renal proximal tubular injury.

UNLABELLED: Body temperature (T) profoundly alters the severity of ischemic acute renal failure. Therefore, the present study evaluated T effects on: (a) in vitro proximal tubular cell killing during hypoxia/reoxygenation to assess when it exacerbates injury; (b) renal ATP losses and metabolic rate (O2 consumption) during hemorrhagic shock (55-60 mm Hg); and (c) membrane deacylation (assessed by free fatty acid, FFA, release) to determine if T modifies this pathway of ischemic renal damage. Hypoxic cell kill (45 minutes) was 20 +/- 1% 47 +/- 4%, and 61 +/- 2% at 32 degrees C, 37 degrees C, and 40 degrees C respectively (by lactate dehydrogenase release; p less than 0.001). During reoxygenation (15 minutes), minimal lactate dehydrogenase was released, irrespective of T. ATP decrements during shock were profoundly T dependent (% loss of ATP; 15%, 30%, 58% at 32.5 degrees C, 37 degrees C, and 39.5 degrees C, respectively; p less than 0.001), reflecting T-dependent increments in renal metabolic rate, not decreased O2 delivery (arterial O2 content; renal blood flow). ATP losses during shock correlated with the extent of S3 proximal tubular morphologic damage. O2 deprivation dramatically increased FFA levels both in vivo and in vitro but the increments were only slightly T dependent. In vitro, % lactate dehydrogenase release and FFA levels did not significantly correlate and bovine serum albumin, a FFA binder, conferred no protection. CONCLUSIONS: (a) T dramatically accentuates hypoxic, not reoxygenation, injury; (b) changes in membrane deacylation do not appear to underlie this effect; and (c) T has a profound impact on renal ATP losses during shock, thereby affecting the severity of ischemic renal damage.

Regional responses within the kidney to ischemia: assessment of adenine nucleotide and catabolite profiles.

UNLABELLED: Renal cortex (C) has predominantly aerobic metabolism, whereas inner medulla (IM) has both aerobic and anaerobic capacities. This study was undertaken (1) to assess how well rat IM anaerobic metabolism maintains this region's ATP content during ischemia; and (2) to determine whether regional variations in adenylate pool/catabolite responses to ischemia exist, obscuring interpretation of cellular energetics in rat studies of acute renal failure (ARF). Adenine nucleotides/catabolites were measured in rat C, IM and outer medulla (OM) after 15 and 45 min of ischemia. After 15 min, all regions showed profound ATP depletion, although the IM maintained slightly higher (by 0.23 mumol/g) absolute ATP levels than C/OM tissues (normal ATP value = 8.7 mumol/g). By 45 min, significant differences in regional ATP levels did not exist. Striking regional catabolite differences were apparent at both 15 and 45 min. Most prominent were: (1) intrarenal purine base/inosine gradients, levels falling approx. 22-50% from C to IM; and (2) preferential OM AMP/IMP/adenosine accumulation. To assess whether more homogeneous results might be found in rabbit kidney, possibly making this animal preferable to rats for studies of renal ischemia, rabbit C, OM and IM adenylate pools were analyzed after 15 min of ischemia. C vs. IM ATP differences were greater (approx. 1.3 mumol/g) and large catabolite concentration differences were still apparent. CONCLUSIONS: (1) anaerobic mechanisms support IM ATP levels during ischemia but, in terms of normal concentrations, the impact is small, particularly in the rat; and (2) marked regional differences in adenylate catabolite levels exist within ischemic kidneys. These need to be recognized when analyzing adenylate pool responses in ischemic ARF.

Degree and time sequence of hypothermic protection against experimental ischemic acute renal failure.

The purpose of this study was to assess the degree, time sequence, and biochemical correlates of hypothermic protection against ischemic acute renal failure. Rats subjected to 40 minutes of bilateral renal artery occlusion (RAO) were made mildly hypothermic (32 degrees-33 degrees C, by cold saline peritoneal lavage) during the following time periods: 1) RAO only, 2) reperfusion only (beginning at 0, 15, 30, or 60 minutes after RAO and maintained for 45 minutes), or 3) during and after (0-45 minutes) RAO. Continuously normothermic (37 degrees C) RAO rats served as controls. The control rats developed severe acute renal failure (blood urea nitrogen [BUN], 95 +/- 4 mg/dl; creatinine, 2.2 +/- 0.1 mg/dl; and extensive tubular necrosis at 24 hours). Hypothermia confined to RAO was highly protective (BUN, 33 +/- 5 mg/dl; creatinine, 0.62 +/- 0.07 mg/dl; and minimal necrosis). Hypothermia partially preserved ischemic renal adenylate high-energy phosphate (ATP and ADP), increased AMP and inosine monophosphate concentrations, and lessened hypoxanthine/xanthine buildup (assessed at end of RAO). Hypothermia confined to the reflow period (beginning at 0, 15, and 30 minutes) was only mildly protective (e.g., BUN, 58-63 mg/dl); the degree of protection did not differ according to the time of hypothermic onset. Lowering reflow temperature to 26 degrees C had no added benefit. Hypothermia that started at 60 minutes after RAO conferred no protection. Combining ischemic and postischemic hypothermia abolished all renal failure (assessed at 24 hours). This study offers the following conclusions: Mild hypothermia can totally prevent experimental ischemic acute renal failure. Hypothermia is highly effective during ischemia, and it is mildly protective during early reflow; these benefits are additive. During early reflow, hypothermic protection is not critically time dependent. By 60 minutes of reflow, no effect is elicited; this absence of effect possibly signals completion of the reperfusion injury process. Hypothermia's protective effects may be mediated, in part, by improvements in renal adenine nucleotide content and, possibly, by decreasing postischemic oxidant stress.

Plasma iron and transferrin iron-binding capacity evaluated by colorimetric and immunoprecipitation methods.

We evaluated plasma iron (PI) and total iron-binding capacity (TIBC) or transferrin in normal individuals and in patients with iron imbalance. The standard colorimetric measurements of PI and TIBC and the standard isotope-dilution measurement of TIBC were compared with an immunoprecipitation method and also with immunoelectrophoresis of transferrin. PI concentrations as measured by the standard and immunoprecipitation methods agreed closely for all individuals except those with saturated transferrin, where nontransferrin iron increased the results in the standard assay. This excess iron in saturated plasma may be derived from either free iron or iron-bearing ferritin. There were also differences in TIBC between the two methods. Iron-deficient sera gave higher values for transferrin when measured by immunoelectrophoresis. Unsaturated iron-binding capacity was increased in the isotope-dilution method in some iron-saturated plasma, compounding errors when added to erroneously high PI values to compute TIBC. Perhaps some exchange of iron occurred between added iron and transferrin iron in the isotope-dilution method. These measurements confirm the accuracy of the standard colorimetric method of measuring PI and TIBC except in iron-saturated plasma. However, the greater specificity of a polyclonal immunoprecipitation method of measuring PI and TIBC makes it particularly useful in differentiating transferrin-bound iron from nontransferrin iron.

The effect of transferrin saturation on internal iron exchange.

Radioiron was introduced into the intestinal lumen to evaluate absorption, injected as nonviable red cells to evaluate reticuloendothelial (RE) processing of iron, and injected as hemoglobin to evaluate hepatocyte iron processing. Redistribution of iron through the plasma was evaluated in control animals and animals whose transferrin was saturated by iron infusion. Radioiron introduced into the lumen of the gut as ferrous sulfate and as transferrin-bound iron was absorbed about half as well in iron-infused animals, and absorbed iron was localized in the liver. The similar absorption of transferrin-bound iron suggested that absorption of ferrous iron occurred via the mucosal cell and did not enter by diffusion. The decrease in absorption was associated with an increase in mucosal iron and ferritin content produced by the iron infusion. An inverse relationship (r = -0.895) was shown between mucosal ferritin iron and absorption. When iron was injected as nonviable red cells, it was deposited predominantly in reticuloendothelial cells of the spleen. Return of this radioiron to the plasma was only 6% of that in control animals. While there was some movement of iron from spleen to liver, this could be accounted for by intravascular hemolysis. Injected hemoglobin tagged with radioiron was for the most part taken up and held by the liver. Some 13% initially localized in the marrow in iron-infused animals was shown to be storage iron unavailable for hemoglobin synthesis. These studies demonstrate the hepatic trapping of absorbed iron and the inability of either RE cell or hepatocyte to release iron in the transferrin-saturated animal.