Optimal protective hypothermia in arrested mammalian hearts.

Many therapeutic hypothermia recommendations have been reported, but the information supporting them is sparse, and reveals a need for the data of target therapeutic hypothermia (TTH) from well-controlled experiments. The core temperature ≤35°C is considered as hypothermia, and 29°C is a cooling injury threshold in pig heart in vivo. Thus, an optimal protective hypothermia (OPH) should be in the range 29-35°C. This study was conducted with a pig cardiopulmonary bypass preparation to decrease the core temperature to 29-35°C range at 20 minutes before and 60 minutes during heart arrest. The left ventricular (LV) developed pressure, maximum of the first derivative of LV (dP/dtmax), cardiac power, heart rate, cardiac output, and myocardial velocity (Vmax) were recorded continuously via an LV pressure catheter and an aortic flow probe. At 20 minutes of off-pump during reperfusion after 60 minutes arrest, 17 hypothermic hearts showed that the recovery of Vmax and dP/dtmax established sigmoid curves that consisted of two plateaus: a good recovery plateau at 29-30.5°C, the function recovered to baseline level (BL) (Vmax=118.4%±3.9% of BL, LV dP/dtmax=120.7%±3.1% of BL, n=6); another poor recovery plateau at 34-35°C (Vmax=60.2%±2.8% of BL, LV dP/dtmax=28.0%±5.9% of BL, p<0.05, n=6; ), which are similar to the four normothermia arrest (37°C) hearts (Vmax=55.9%±4.8% of BL, LV dP/dtmax=24.5%±2.1% of BL, n=4). The 32-32.5°C arrest hearts showed moderate recovery (n=5). A point of inflection (around 30.5-31°C) existed at the edge of a good recovery plateau followed by a steep slope. The point presented an OPH that should be the TTH. The results are concordant with data in the mammalian hearts, suggesting that the TTH should be initiated to cool core temperature at 31°C.

Moderate hypothermia (30 degrees C) maintains myocardial integrity and modifies response of cell survival proteins after reperfusion.

Hypothermia preserves myocardial function, promotes signaling for cell survival, and inhibits apoptotic pathways during 45-min reperfusion. We tested the hypothesis that signaling at the transcriptional level is followed by corresponding proteomic response and maintenance of structural integrity after 3-h reperfusion. Isolated hearts were Langendorff perfused and exposed to mild (I group; n = 6, 34 degrees C) or moderate (H group; n = 6, 30 degrees C) hypothermia during 120-min total ischemia with cardioplegic arrest and 180-min 37 degrees C reperfusion. Moderate hypothermia suppressed anaerobic metabolism during ischemia and significantly diminished left ventricular end-diastolic pressure at the end of ischemia from 52.7 +/- 3.3 (I group) to 1.8 +/- 0.9 (H group) mmHg. Unlike the I group, which showed poor cardiac function and high left ventricular pressure, the H group showed preservation of myocardial function, coronary flow, and oxygen consumption. Compared with normal control hearts without ischemia (n = 5), histological staining in the I group showed marked disarray and fragmentation of collagen network (score 4-5), while the H group showed preserved collagen integrity (score 0-1). The apoptosis-linked tumor suppressor protein p53 was expressed throughout the I group only (score 4-5). The H group produced elevated expression for hypoxia-inducible factor 1alpha and heme oxygenase 1, but minimally affected vascular endothelial growth factor expression. The H group also elevated expression for survival proteins peroxisomal proliferator-activated receptor-beta and Akt-1. These results show in a constant left ventricular volume model that moderate hypothermia (30 degrees C) decreases myocardial energy utilization during ischemia and subsequently promotes expression of proteins involved in cell survival, while inhibiting induction of p53 protein. These data also show that 34 degrees C proffers less protection and loss of myocardial integrity.

Short-cycle hypoxia in the intact heart: hypoxia-inducible factor 1alpha signaling and the relationship to injury threshold.

Hypoxia-inducible factor 1alpha (HIF-1alpha) transcriptionally activates multiple genes, which regulate metabolic cardioprotective and cross-adaptive mechanisms. Hypoxia and several other stimuli induce the HIF-1alpha signaling cascade, although little data exist regarding the stress threshold for activation in heart. We tested the hypothesis that relatively mild short-cycle hypoxia, which produces minimal cardiac dysfunction and no sustained or major disruption in energy state, can induce HIF-1alpha activation. We developed a short-cycle hypoxia protocol in isolated perfused rabbit heart to test this hypothesis. By altering cycling conditions, we identified a specific cycle with O(2) content and duration that operated near a threshold for causing functional injury in these rabbit hearts. Mild short-cycle hypoxia for 46 min elevated HIF-1alpha mRNA and protein within 45 min after reoxygenation. Expression also increased for multiple HIF-1alpha target genes, such as VEGF and heme oxygenase 1. After mild hypoxia, VEGF protein accumulation occurred, although HIF-1alpha and VEGF protein accumulation were suppressed after more severe hypoxia, which also caused depletion of ATP and nondiffusible nucleotides. In summary, these results indicate that mild near-threshold hypoxia induces HIF-1alpha cascade, but more severe hypoxia suppresses protein accumulation for this transcription factor and the target genes. Posttranscriptional suppression of these proteins occurs under conditions of altered energy state, exemplified by ATP depletion.

Hypothermia preserves myocardial function and mitochondrial protein gene expression during hypoxia.

Hypothermia before and/or during no-flow ischemia promotes cardiac functional recovery and maintains mRNA expression for stress proteins and mitochondrial membrane proteins (MMP) during reperfusion. Adaptation and protection may occur through cold-induced change in anaerobic metabolism. Accordingly, the principal objective of this study was to test the hypothesis that hypothermia preserves myocardial function during hypoxia and reoxygenation. Hypoxic conditions in these experiments were created by reducing O2 concentration in perfusate, thereby maintaining or elevating coronary flow (CF). Isolated Langendorff-perfused rabbit hearts were subjected to perfusate (Po2 = 38 mmHg) with glucose (11.5 mM) and perfusion pressure (90 mmHg). The control (C) group was at 37 degrees C for 30 min before and 45 min during hypoxia, whereas the hypothermia (H) group was at 29.5 degrees C for 30 min before and 45 min during hypoxia. Reoxygenation occurred at 37 degrees C for 45 min for both groups. CF increased during hypoxia. The H group markedly improved functional recovery during reoxygenation, including left ventricular developed pressure (DP), the product of DP and heart rate, dP/dtmax, and O2 consumption (MVo2) (P < 0.05 vs. control). MVo2 decreased during hypothermia. Lactate and CO2 gradients across the coronary bed were the same in C and H groups during hypoxia, implying similar anaerobic metabolic rates. Hypothermia preserved MMP betaF1-ATPase mRNA levels but did not alter adenine nucleotide translocator-1 or heat shock protein-70 mRNA levels. In conclusion, hypothermia preserves cardiac function after hypoxia in the hypoxic high-CF model. Thus hypothermic protection does not occur exclusively through cold-induced alterations in anaerobic metabolism.

Hypothermic protection of the ischemic heart via alterations in apoptotic pathways as assessed by gene array analysis.

Hypothermia improves resistance to ischemia in the cardioplegia-arrested heart. This adaptive process produces changes in specific signaling pathways for mitochondrial proteins and heat-shock response. To further test for hypothermic modulation of other signaling pathways such as apoptosis, we used various molecular techniques, including cDNA arrays. Isolated rabbit hearts were perfused and exposed to ischemic cardioplegic arrest for 2 h at 34 degrees C [ischemic group (I); n = 13] or at 30 degrees C before and during ischemia [hypothermic group (H); n = 12]. Developed pressure, the maximum first derivative of left ventricular pressure, oxygen consumption, and pressure-rate product (P < 0.05) recovery were superior in H compared with in I during reperfusion. mRNA expression for the mitochondrial proteins, adenine translocase and the beta-subunit of F1-ATPase, was preserved by hypothermia. cDNA arrays revealed that ischemia altered expression of 13 genes. Hypothermia modified this response to ischemia for eight genes, six related to apoptosis. A marked, near fivefold increase in transformation-related protein 53 in I was virtually abrogated in H. Hypothermia also increased expression for the anti-apoptotic Bcl-2 homologue Bcl-x relative to I but decreased expression for the proapoptotic Bcl-2 homologue bak. These data imply that hypothermia modifies signaling pathways for apoptosis and suggest possible mechanisms for hypothermia-induced myocardial protection.