Marius Locke

Heat shock protein (HSP 72) expression in patients undergoing cardiac operations

The major mammalian stress-inducible protein, heat shock protein 72, protects cells from certain stresses and rapidly accumulates in cells after ischemia. Heat shock protein 72 is rapidly synthesized in the myocardium of various species in response to ischemia, but it has not been investigated in human heart. To determine if heat shock protein 72 accumulated in the ischemic myocardium of patients undergoing cardiac operations, we obtained sequential right atrial biopsy specimens from 12 patients undergoing repair at three intervals: before bypass, after reperfusion, and after bypass. Immunoblot analysis for heat shock protein 72 demonstrated a high expression in the human heart compared with other mammalian hearts, p (Binomial) = 0.01. Compared with before bypass, heat shock protein 72 contents after reperfusion and after bypass were 98.2% +/- 8.9%, p (signed-rank) = 0.65, and 87.6% +/- 17.1%, p (signed-rank) = 0.28, respectively. Although heat shock protein 72 concentration was unchanged in hearts after reperfusion and after bypass, the initial prebypass level of heat shock protein 72 was high. The high heat shock protein 72 level detected in human hearts may reflect preoperative disease and drug therapy, or inherently high levels may be usual in the human myocardium. These findings indicate that the myocardium of patients undergoing cardiac operations contains relatively high concentrations of heat shock protein 72, which are not increased during the surgical procedure.

Preservation of the Latissimus Dorsi Muscle During Cardiomyoplasty Surgery

Abstract Background: Cardiomyoplasty surgery has been shown to be associated with damage and degeneration of the assisting skeletal muscle. The purpose of this study was to use ischemic (short‐term) and thermal (long‐term) preconditioning to protect the muscle during surgery and the subsequent ischemia. Methods: Three 10‐minute cycles of ischemia‐reperfusion were accomplished noninvasively on goat latissimus dorsi muscle (LDM) immediately prior to surgery. In another experiment, LDM was noninvasively heat shocked for 20 minutes at 42°C 24 hours prior to surgery. LDM damage was evaluated 5 days postsurgery using enzyme activities (β‐glucuronidase, β‐GLN; citrate synthase), hydroxyproline, morphology, and blood flow. Results: The lysosomal enzyme, β‐GLN, was significantly increased (43%, p < 0.05) by surgical dissection and remained high in the ischemic preconditioned LDM (58%, p < 0.05) and in the heat shocked LDM (57%, p < 0.05). Conclusion: These findings show that these two protective protocols do not reduce the muscle damage that occurs during surgical preparation of the LDM for cardiomyoplasty.

Cardiomyoplasty: Preservation of the Latissimus Dorsi Muscle

ABSTRACT Muscle necrosis has been frequently observed in cardiomyoplasty patients and in experimental animal studies. The purpose of this study was to determine if heat shock could provide protection to skeletal muscle as has been shown in cardiac muscle. A 15‐minute heat shock at 42°C resulted in an immediate increase in HSP72 mRNA and was followed within 3 hours by a two‐fold increase in HSP72. Surgical dissection of the latissimus dorsi muscle (LDM) followed by an ischemic period resulted in a two‐fold increase in HSP72 in control LDM, whereas the already high levels in the heat‐shocked LDM increased only slightly with surgery and ischemia. Citrate synthase activity and tissue histology indicated that heat shock did not protect the LDM from the imposed surgical trauma and ischemic insults used in this study.

Cardiomyoplasty: Comparison of Latissimus Dorsi Muscles of Three Large Mammals with that of Human

Abstract Cardiomyoplasty has the potential to become an alternative therapy for congestive heart failure patients and is presently in Phase III clinical trials. In experimental studies, it is necessary to use an animal with muscle characteristics that resemble those of humans. Therefore, the purpose of this study was to compare morphological and biochemical characteristics of the latissimus dorsi muscle (LDM) of three common large mammals with those of human. Of the three mammals studied, the goat had the most overall similarities to the human when comparing mitochondrial capacity, percent fiber types, fiber areas, myofibrillar (MF)‐AT‐Pase activity, and 72‐kDa heat shock protein (HSP) content. The pig was dissimilar to the human in its fiber‐type arrangement, glycolytic capacity, percent fiber type, MF‐ATPase activity, and HSP‐72 content. The dog differed from the human in that it had high‐mitochondrial enzyme activity, a fiber‐type profile consisting of all high‐aerobic fibers, and fiber cross‐sectional areas that were nearly half those of humans. These findings show that the LDM of the goat most resembles that of the human.

Myocardial Self-Preservation: Absence of Heat Shock Factor Activation and Heat Shock Proteins 70 mRNA Accumulation in the Human Heart During Cardiac Surgery

Following myocardial ischemia, heat shock proteins (HSPs) have been found to be associated with a reduction in infarct size and enhanced postischemic functional recovery. Stress‐induced regulation of the HSPs is mediated by the activation and binding of the heat shock transcription factor (HSF) to a specific DNA sequence located in front of all HSP genes, known as the heat shock element (HSE). To determine whether HSPs were induced in the human heart following the ischemic stress experienced during cardiac surgery, biopsies were performed of the right atrium at three sequential times: prior to establishing cardiopulmonary bypass; immediately after aortic declamping; and following termination of bypass. These samples from the atria of patients undergoing coronary bypass surgery were assessed for HSF activation using mobility shift gels, and analyzed for HSP 72 mRNA by Northern blot. Although a high level of the HSP 72 protein was noted at all intervals, no HSF activation was detected, nor was an accumulation of HSP 72 mRNA observed at any time during surgery. These data suggest that HSPs are not induced during cardiac surgery and that the high “constitutive” level of the HSP 72 protein detected in these hearts may not be secondary to an HSF‐HSE interaction, but rather, the result of other transcription factors acting at alternative regions of the HSP 70 promoter.

DIMINISHED HEAT SHOCK RESPONSE IN THE AGED MYOCARDIUM 845

Induction of heat shock proteins (Hsps), Hsp72 in particular, has been associated with myocardial protection. Since a decreased Hsp response has been reported to occur with aging, it was of interest to determine if hearts from aged animals also demonstrate an altered heat shock response and subsequent myocardial protection. Adult (6 months old) and aged (22 months old) Fischer 344 rats were heat stressed by raising their rectal temperatures to 41 degrees C for 10 min. At selected times following heat stress (0-24 h) hearts were examined for heat shock transcription factor trimerization and DNA-binding activity (Hsf1 activation), Hsp72 mRNA accumulation, Hsp72 and Hsf1 protein content, as well as, protection from ischemia using the Langendorff isolated heart model. Following heat stress, hearts from aged animals demonstrated a 47% reduction in Hsf1 activation, a reduction in Hsp72 mRNA and a 35% reduction in Hsp72 protein content, compared to hearts from adults. Interestingly, myocardial Hsf1 protein content was similar between aged and adult animals. Hearts from heat stressed adult animals (24-h prior) demonstrated an enhanced postischemic recovery as indicated by a greater recovery of left ventricular pressure and rate of contraction (P < 0.05), while hearts from heat stressed aged animals failed to demonstrate an enhanced postischemic recovery. These results suggest that hearts from aged animals exhibit an impaired ability to produce the protective Hsps and thus, may explain, at least in part, the increased susceptibility of aged hearts to stress.