Effects of ischemia and perfusion temperature on the synthesis of stress-induced (heart shock) proteins in isolated and perfused rat hearts

Abstract

Isolated and perfused rat hearts were examined by 2-dimensional gel electrophoresis and liquid scintillation counting to determine the effect of ischemia or perfusion temperature on protein synthesis. Isolated hearts were subjected to ischemia at either 4 degrees, 20 degrees, or 30 degrees C and then perfused at 37 degrees C and radio-labeled with [3H]-leucine from 0.5 to 2.5 h of perfusion. Following 0.5 h of 4 degrees C or 0.2 h of 20 degrees C ischemia, minimal or no effect was seen by fluorography on the patterns of protein synthesis and there was no apparent synthesis of the stress-induced (heat shock) protein with Mr = 71,000 (SP71). After 4.0 h of 4 degrees C or 0.5 h of 20 degrees C ischemia, SP71 was detectable on fluorograms. After 17 h of 4 degrees C or 1 h of 20 degrees C or 30 degrees C ischemia, SP71 was a prominent spot on fluorograms; the percentage incorporation of precursor into SP71 was significantly increased and overall incorporation of precursor into protein appeared depressed. Following 17 h at 4 degrees C, during which time the buffer was continuously oxygenated, percentage incorporation of precursor into SP71 was only moderately increased, suggesting that hypoxia was responsible for the high level of SP71 synthesis. The effect of perfusion temperature was initially examined using a perfusion method which results in a moderate synthesis of SP71 at 37 degrees C between 2.5 and 4.5 h of perfusion. Fluorograms revealed synthesis of some normal proteins at all temperatures, with little or no synthesis of SP71 at 31 degrees and 34 degrees C, moderate synthesis at 37 degrees C, and intense synthesis at 40 degrees C. The percentage incorporation of precursor into SP71 and total incorporation of precursor appeared depressed at 31 degrees C and 34 degrees C, from that at 37 degrees C, while at 40 degrees C, percentage incorporation of precursor into SP71 was significantly increased but overall incorporation of precursor into protein appeared depressed. When SP71 synthesis was induced by 17 h of 4 degrees C ischemia, hearts were sensitive to a lower perfusion temperature (34 degrees C); the percent incorporation of precursor into SP71 was significantly reduced from that seen at 37 degrees C. Interestingly, the effect of prolonged ischemia on protein synthesis (increased synthesis of SP71; suppression of overall synthesis) is the same as the heat shock response seen after perfusion at 40 degrees C.