Abdominal insufflation for laparoscopy increases intracranial and intrathoracic pressure in human subjects.

BACKGROUND: Laparoscopy has emerged as an alternative to laparotomy in select trauma patients. In animal models, increasing abdominal pressure is associated with an increase in intrathoracic and intracranial pressures. We conducted a prospective trial of human subjects who underwent laparoscopic-assisted ventriculoperitoneal shunt placement (lap VPS) with intraoperative measurement of intrathoracic, intracranial and cerebral perfusion pressures. METHODS: Ten patients undergoing lap VPS were recruited. Abdominal insufflation was performed using CO2 to 0, 8, 10, 12 and 15 mmHg. ICP was measured through the ventricular catheter simultaneously with insufflation and with desufflation using a manometer. Peak inspiratory pressures (PIP) were measured through the endotracheal tube. Blood pressure was measured using a noninvasive blood pressure cuff. End-tidal CO2 (ETCO2) was measured for each set of abdominal pressure level. Pressure measurements from all points of insufflation were compared using a two-way ANOVA with a post hoc Bonferroni test. Mean changes in pressures were compared using t test. RESULTS: ICP and PIP increased significantly with increasing abdominal pressure (both p < 0.01), whereas cerebral perfusion pressure (CPP) and mean arterial pressure did not significantly change with increasing abdominal pressure over the range tested. Higher abdominal pressure values were associated with decreased ETCO2 values. CONCLUSION: Increased ICP and PIP appear to be a direct result of increasing abdominal pressure, since ETCO2 did not increase. Though CPP did not change over the range tested, the ICP in some patients with 15 mmHg abdominal insufflation reached values as high as 32 cmH2O, which is considered above tolerance, regardless of the CPP. Laparoscopy should be used cautiously, in patients who present with baseline elevated ICP or head trauma as abdominal insufflation affects intracranial pressure.

Ethanol promotes new vessel growth in remote nonischemic myocardium.

BACKGROUND: Epidemiologic studies have demonstrated that daily low to moderate alcohol consumption is cardioprotective as compared with abstainers and high alcohol consumption. Our group reported that alcohol consumption improves angiogenesis in chronically ischemic myocardium. We developed a clinically relevant follow-up study to assess the effect of moderate alcohol consumption on new vessel growth in normal myocardium remote from an ischemic territory in a swine model. MATERIALS AND METHODS: Fourteen male Yorkshire swine underwent placement of an ameroid constrictor to induce chronic myocardial ischemia. Postoperatively, animals were supplemented with either 90 mL of ethanol daily (ETOH) or 80 g of sucrose (SUC) of equal caloric value. Seven weeks after ameroid placement, myocardial tissue from a territory remote from the ischemia was harvested for analysis. RESULTS: Both groups had similar microvascular relaxation to endothelial dependent and endothelium-independent vasodilators. Also, both groups had similar myocardial perfusion at rest and with demand pacing. The ETOH group had significantly increased arteriolar and capillary density in the nonischemic myocardium compared with the SUC group. ETOH supplementation also increased expression of pro-angiogenesis proteins vascular endothelial growth factor and vascular endothelial cadherin, and decreased expression of anti-angiogenesis proteins angiostatin and endostatin. CONCLUSIONS: ETOH supplementation increased capillary and arteriolar density, upregulated pro-angiogenesis and pro-survival proteins, and downregulated anti-angiogenesis protein expression. These findings suggest that at moderate doses, ETOH directly promotes new vessel growth in the nonischemic myocardium remote from chronic ischemia.

Atorvastatin regulates apoptosis in chronically ischemic myocardium.

BACKGROUND: We previously demonstrated that atorvastatin upregulates proangiogenic proteins and increases arteriolar density in ischemic myocardium. Despite this, there was a lack of collateral-dependent perfusion, possibly related to apoptosis. We utilized a swine model of metabolic syndrome and chronic myocardial ischemia to investigate the effects of atorvastatin on apoptosis. MATERIALS AND METHODS: Sixteen Ossabaw miniswine were fed a high-cholesterol diet for 14 weeks then underwent surgical placement of an ameroid constrictor to their circumflex artery inducing chronic ischemia. Eight pigs additionally received supplemental atorvastatin (1.5 mg/kg daily). Myocardium was harvested six months later for western blotting and TUNEL staining. RESULTS: Animals supplemented with atorvastatin had significant increases in markers associated with apoptosis including p-38, BAX, and caspase 3 (p < 0.05). Atorvastatin supplementation also resulted in significant increases in expression of cell survival proteins Bcl-2 and P-ERK and an overall decrease in apoptosis demonstrated by TUNEL staining (p < 0.05). CONCLUSIONS: Atorvastatin acts on multiple pathways and its effects on angiogenesis remain unclear. Although there is increased expression in several markers of apoptosis, key anti-apoptotic proteins were also upregulated with an overall decrease in apoptosis. Further investigation of these pathways may provide insight into the role of statins on myocardial protection after ischemia.

Ethanol promotes arteriogenesis and restores perfusion to chronically ischemic myocardium.

BACKGROUND: Moderate alcohol consumption is known to be cardioprotective compared with either heavy drinking or complete abstinence. We assessed the hypothesis that ethanol supplementation would improve myocardial function in the setting of chronic ischemia. METHODS AND RESULTS: Sixteen male Yorkshire swine underwent placement of an ameroid constrictor into the left circumflex artery to induce chronic myocardial ischemia. Postoperatively, animals were supplemented with either 90 mL of ethanol (EtOH) daily (50%/V, EtOH) or 80 g of sucrose of equal caloric value (SUC), serving as controls. Seven weeks after ameroid placement, arteriolar density (1.74 ± 0.210% versus 3.11 ± 0.368% area of arterioles per low-powered field in sucrose (SUC) versus EtOH; P=0.004), myocardial perfusion (ratio of blood flow to the at-risk myocardium compared with the normal ventricle during demand pacing was 0.585 ± 0.107 versus 1.08 ± 0.138 for SUC versus EtOH; P=0.014), and microvascular reactivity were significantly increased in ethanol-treated animals compared with controls in the at-risk myocardium. Analysis of vascular endothelial growth factor and NOTCH pathway signaling suggested proneovascular and proliferative activity in the ischemic area. The average peak blood alcohol level in the treatment group was 40 ± 4 mg/dL, consistent with levels of moderate drinking in humans. CONCLUSIONS: Ethanol supplementation increased arteriolar density and significantly improved myocardial perfusion and endothelium-dependent vasorelaxation in chronically ischemic myocardium. These findings suggest that, at moderate doses, ethanol directly promotes vasculogenesis and improves microvascular function, resulting in significant improvements in myocardial perfusion in the setting of chronic ischemia.

Metformin mitigates apoptosis in ischemic myocardium.

BACKGROUND: Epidemiologic data has shown that metformin confers a survival advantage in patients with cardiovascular disease. Although the underlying cardioprotective mechanism is unclear, it appears to be independent of metformin's insulin-sensitizing effect. The purpose of this study was to evaluate the effect of metformin on the apoptosis pathway in the ischemic and nonischemic cardiac tissue in a swine model of metabolic syndrome. MATERIALS AND METHODS: Ossabaw miniswine were fed either a regular diet (Ossabaw control, n = 8), a high-cholesterol diet (Ossabaw high cholesterol, n = 8), or a high-cholesterol diet supplemented with metformin (Ossabaw high-cholesterol metformin, n = 8). After 9 wk, all animals underwent placement of an ameroid constrictor to the left circumflex coronary artery to induce chronic ischemia. Seven weeks after ameroid placement, animals underwent cardiac harvest. RESULTS: In the chronically ischemic myocardium, metformin significantly upregulates prosurvival proteins: extracellular signal-regulated kinases, nuclear factor κB, phosphorylated endothelial nitric oxide synthase, and P38. Metformin also significantly inhibits or downregulates proapoptosis proteins: FOXO3 and caspase 3. Metformin decreased the percent apoptotic cells in the ischemic and nonischemic myocardium. There was no difference in arteriolar density, capillary density, intramyocardial fibrosis, or collagen deposition in the ischemic or nonischemic myocardium. CONCLUSIONS: Metformin selectively alters the apoptosis pathway by inhibiting FOXO3 and decreasing the active form of caspase 3, cleaved caspase 3. Metformin also upregulates mitogen-activated kinase proteins p38 and extracellular signal-regulated protein kinases 1 and 2, which are considered cardioprotective during ischemic preconditioning. Perhaps, the altered activation of the apoptosis pathway in ischemic myocardium is one mechanism by which metformin is cardioprotective.

Microvascular notch signaling is upregulated in response to vascular endothelial growth factor and chronic myocardial ischemia.

BACKGROUND: Notch signaling is a highly conserved pathway that promotes vascular and myocardial growth. The hypothesis that exogenous vascular endothelial growth factor (VEGF) administration to ischemic myocardium would enhance the neovascular response and upregulate Notch signaling was assessed. METHODS AND RESULTS: Fourteen male Yorkshire swine underwent placement of an ameroid constrictor on the left circumflex artery to induce chronic myocardial ischemia with half of the animals receiving perivascular VEGF to the ischemic area. The remote territory served as the normal ventricle control (NV), while the 2 experimental groups consisted of the area at risk of the non-VEGF animals (AAR) and the area at risk of animals treated with VEGF (VEGF). Capillary and arteriolar density was significantly increased in the VEGF group as compared to both NV and AAR. Expression of Notch receptors and pro-neovascular Notch ligands was significantly higher in the VEGF group. Both Jagged 1 and Notch 3 were the most highly concentrated in the smooth muscle wall of arterioles. CONCLUSIONS: VEGF administration to chronically ischemic myocardium significantly augmented the neovascular response by an increase in both capillary and arteriolar density, and resulted in an upregulation of several Notch receptors and ligands, which were not upregulated with ischemia alone. These findings suggest that the augmented neovascular response seen with VEGF administration was through the VEGF-induced upregulation of Notch signaling.

Does resveratrol improve insulin signaling in chronically ischemic myocardium?

BACKGROUND: Resveratrol is a naturally occurring polyphenol believed to be cardioprotective. We previously demonstrated that resveratrol improves insulin signaling and glucose metabolism in liver and skeletal muscle of swine with metabolic syndrome. Although resveratrol has metabolic benefits in peripheral tissues, its effect on insulin signaling in ischemic myocardium (IM) is unclear. Therefore, we developed a clinically relevant swine model of metabolic syndrome and chronic myocardial ischemia to investigate the effects of resveratrol on insulin signaling in cardiac tissue. MATERIALS AND METHODS: Thirteen male Yorkshire swine were fed a high-cholesterol diet for 4 wk then underwent surgical placement of an ameroid constrictor to their circumflex artery to induce chronic myocardial ischemia. The high-cholesterol control group was given no drug (n = 7). The experimental group was provided the same diet and received supplemental resveratrol (100 mg/kg/d) (n = 6). Tissue was harvested 7 wk after ameroid placement for western blot and histological analyses. RESULTS: In IM, there was no significant difference between the two groups in the insulin signaling markers studied. In nonischemic myocardium, there was a significant decrease in phosphorylated AMP-activated protein kinase α (P = 0.021) in the group treated with resveratrol; otherwise, there were no significant differences between the groups. Immunostaining for glucose transporter 4 and Periodic acid-Schiff staining for myocardial glycogen stores was similar between the groups. CONCLUSIONS: Resveratrol has complex effects on glucose metabolism. Although prior studies demonstrated that resveratrol supplementation improves insulin sensitivity in peripheral tissues, in chronically IM, there are no significant alterations.

Overfed Ossabaw swine with early stage metabolic syndrome have normal coronary collateral development in response to chronic ischemia.

Ossabaw miniswine have been naturally selected to efficiently store large amounts of lipids offering them a survival advantage. Our goal was to evaluate the myocardial response to chronic ischemia of the Ossabaw consuming a hypercaloric, high-fat/cholesterol diet with and without metformin supplementation. At 6 weeks of age animals were fed either a regular diet (OC, n = 9), a hypercaloric high-fat/cholesterol diet (OHC, n = 9), or a hypercaloric high-fat/cholesterol diet supplemented with metformin (OHCM, n = 8). At 9 weeks, all animals underwent ameroid constrictor placement to the left circumflex coronary artery to simulate chronic ischemia. Seven weeks after ameroid placement, all animals underwent hemodynamic and functional measurements followed by cardiac harvest. Both OHC and OHCM animals developed significantly greater weight gain, total cholesterol, and LDL:HDL ratio compared to OC controls. Metformin administration reversed diet-induced hypertension and glucose intolerance. There were no differences in global and regional contractility, myocardial perfusion, capillary and arteriolar density, or total protein oxidation between groups. Myocardial protein expression of VEGF, PPAR-α, γ, and δ was significantly increased in the OHC and OHCM groups. Microvessel reactivity was improved in the OHC and OHCM groups compared to controls, and correlated with increased p-eNOS expression. Overfed Ossabaw miniswine develop several components of metabolic syndrome. However, impairments of myocardial function, neovascularization and perfusion were not present, and microvessel reactivity was paradoxically improved in hypercholesterolemic animals. The observed cardioprotection despite metabolic derangements may be due to lipid-dependant upregulation of the PPAR pathway which is anti-inflammatory and governs myocardial fatty acid metabolism.

Cardioprotective effects of red wine and vodka in a model of endothelial dysfunction.

BACKGROUND: Moderate alcohol consumption is largely believed to be cardioprotective, while red wine is hypothesized to offer benefit in part due to the proangiogenic and antioxidant properties of polyphenols. We investigated the cardiovascular effects of both red wine and vodka in a swine model of endothelial dysfunction. METHODS: Twenty-seven male Yorkshire swine fed a high-fat/cholesterol diet were divided into three groups and received either no alcohol (Control), red wine, or vodka. After 7 wk, myocardial perfusion was measured, and ventricular tissue was analyzed for microvascular reactivity and immunohistochemical studies. RESULTS: There were no differences in myocardial perfusion, in arteriolar or capillary density, or in VEGF expression among groups. Total protein oxidation as well as expression of superoxide dismutase-1 and -2 and NADPH oxidase was decreased in both treatment groups compared to controls. Endothelium-dependent microvessel relaxation, however, was significantly improved only in the red wine-supplemented group. CONCLUSIONS: Supplementation with both red wine and vodka decreased oxidative stress by several measures, implicating the effects of ethanol in reducing oxidative stress in the myocardium. However, it was only in the red wine-supplemented group that an improvement in microvessel function was observed. This suggests that a component of red wine, independent of ethanol, possibly a polyphenol such as resveratrol, may confer cardioprotection by normalizing endothelial dysfunction induced by an atherogenic diet.

Outcomes of Anastomotic Evaluation Using Indocyanine Green Fluorescence during Minimally Invasive Esophagectomy.

BACKGROUND: Limited evidence exists assessing whether anastomotic evaluation using indocyanine green fluorescence (IGF) during minimally invasive esophagectomy (MIE) predicts or improves outcomes. We hypothesized that IGF helps surgeons predict anastomotic complications and reduces anastomotic leaks after MIE. METHODS: In September 2019, our institution began routinely using IGF for intraoperative evaluation of anastomoses during MIE. Data were collected from patients undergoing MIE in the two years before and after this technology began being routinely used. Baseline characteristics and outcomes, including anastomotic leak, in patients who underwent indocyanine green fluorescence evaluation (ICG) and those who did not (nICG) were compared. Outcomes were also compared between ICG patients with normal versus abnormal fluorescence. RESULTS: Overall, 181 patients were included. Baseline demographic and clinical characteristics did not differ between the ICG and nICG groups. ICG patients experienced higher rates of anastomotic leak (10.2% vs. 1.6%, P = .015) and 90-day mortality (8.5% vs. 1.6%, P = .04) compared to nICG patients. Due to lack of equipment availability, 19 nICG patients underwent MIE after the use of IGF became routine, and none developed leaks. ICG patients with abnormal fluorescence had higher rates of anastomotic leak (71.4% vs 1.9%, P < .001) and 30-day mortality (28.6% vs 0%, P = .012) compared to those with normal fluorescence. DISCUSSION: Abnormal intraoperative IGF was associated with increased rate of anastomotic leak, suggesting predictive potential of IGF. However, its use was associated with an increased leak rate and higher mortality. Further studies are warranted to assess possible physiologic effects of indocyanine green on the esophageal anastomosis.

Effects of Alcohol on Postoperative Adhesion Formation in Ischemic Myocardium and Pericardium.

BACKGROUND: Postoperative formation of adhesions increases risk of complications during cardiac reoperations. We previously demonstrated that swine supplemented with vodka had a significant reduction in adhesions at sternotomy after previous thoracotomy. This follow-up study was conducted to determine reproducibility and the mechanism for adhesion reduction in swine supplemented with ethanol. METHODS: An ameroid constrictor was placed in the left circumflex in 14 male Yorkshire swine to induce chronic myocardial ischemia through left minithoracotomy. Animals were supplemented postoperatively with ethanol (45 g ETOH, n = 7) or sucrose (80 g SUC, n = 7) for 7 weeks, followed by a reoperative median sternotomy. RESULTS: The ETOH group had significantly fewer adhesions, thinner pericardial thickness, decreased intramyocardial fibrosis, and decreased myocardial collagen deposition compared with the SUC group. In the myocardium, ETOH animals had decreased expression of proadhesion proteins focal adhesion kinase, paxillin, integrin-ß1, transforming growth factor-ß1 and phosphorylated SMAD and increased expression of adhesion breakdown proteins matrix metalloproteinase (MMP) 1, MMP2, MMP3, and MMP9 compared with SUC animals. However in the pericardium, ETOH animals had increased expression of proadhesion proteins focal adhesion kinase, paxillin, phosphorylated paxillin, vinculin, integrin-ß1, tumor necrosis factor-α, transforming growth factor-ß, and phosphorylated SMAD3, and decreased expression of adhesion breakdown proteins MMP1, MMP3, MMP9, and plasmin compared with SUC animals. CONCLUSIONS: Alcohol supplementation substantially reduced postoperative pericardial adhesion formation, attenuated pericardial thickening, and reduced myocardial fibrosis in response to chronic ischemia. Alcohol supplementation modulates adhesion protein and MMP/tissue inhibitor of metalloproteinase expression, favoring a profile associated with reduced pericardial adhesions. These results suggest that the myocardium is the driving factor in reducing pericardial adhesions and mediating the postoperative healing process.

Alcohol attenuates myocardial ischemic injury.

BACKGROUND: Moderate alcohol consumption is cardioprotective but the mechanism of action remains unclear. Nuclear factor κ-B regulates the expression of genes involved in inflammation, stress, and apoptosis. We used a swine model of diet-induced metabolic syndrome to investigate the effects of red wine and vodka on nuclear factor κ-B signaling and cytokine activity in chronically ischemic myocardium. METHODS: Yorkshire swine were given a high-fat diet for 4 weeks; an ameroid constrictor was then placed on the left circumflex artery. The high-fat diet was continued and the swine were divided into 3 groups for 7 weeks: hypercholesterolemic diet alone (control, n = 8), hypercholesterolemic diet with vodka (vodka, n = 8), and hypercholesterolemic diet with wine (wine, n = 8). Ischemic myocardium was analyzed by Western blot and cytokine array. RESULTS: Administration of alcohol was associated with decreased expression of inhibitor of κ-B kinase complex α, inhibitor of κ-B kinase complex ß, and phosphorylated inhibitor of κ-B ß in the ischemic myocardium compared with the control group. Alcohol administration demonstrated an increase in nuclear factor κ-B in the ischemic myocardium. Both wine and vodka demonstrated a significant decrease in leptin, interleukin-1α, IL-13, IL-15, and interferon-γ. Vodka demonstrated a significant decrease in phosphorylated BCL-2 and caspase-9. CONCLUSION: In ischemic myocardium, alcohol modulates the nuclear factor κ-B pathway, which may contribute to the adaptive response of tissues to the stress of ischemia. Furthermore, both wine and vodka decreased multiple proinflammatory cytokines. This study provides a mechanism by which alcohol may be cardioprotective in ischemic myocardium.

Calpain inhibition decreases inflammatory protein expression in vessel walls in a model of chronic myocardial ischemia.

BACKGROUND: Emerging data suggest a link between calpain activation and the enhanced inflammatory response of the cardiovascular system. We hypothesize that calpain activation associates with altered inflammatory protein expression in correlation with the proinflammatory profile of the myocardium. Our pig hypercholesterolemic model with chronic myocardial ischemia was treated with calpain inhibitors to establish their potential to improve cardiac function. METHODS: Yorkshire swine, fed a high cholesterol diet for 4 weeks then underwent placement of an ameroid constrictor on the left circumflex artery. Two weeks later, animals received either no drug (high-cholesterol control group, n = 8), a low dose of calpain inhibitors (0.12 mg/kg, n = 9), or a high dose of calpain inhibitors (0.25 mg/kg; n = 8). The high-cholesterol diet and calpain inhibitors were continued for 5 weeks, after which the pig was euthanized. The left ventricular myocardial tissue (ischemic and nonischemic) was harvested and analyzed for inflammatory protein expression. Data were statistically analyzed via the Kruskal-Wallis and Dunn post hoc test. RESULTS: Calpain inhibitor treatment coincides with increased expression of IKB-α and decreased expression of macrophages, NFkB, IL-1, and tumor necrosis factor (TNF)-α in the ischemic myocardial tissue as compared with the control group. An NFkB array revealed decreased expression of IRF5, JNK1/2, JNK2, CD18, NFkB p65, c-Rel, Sharpin, TNF R1, TNF R2, and DR5 in the ischemic myocardium of the group treated with a high dose of calpain inhibitors compared with the control. CONCLUSION: Calpain activation in metabolic syndrome is a potential contributor to cardiac dysfunction in metabolic disorders with ischemic background. We suggest that calpain inhibition downregulates NFkB signaling in the vessel walls, which might be useful for improving myocardial blood flow in ischemic conditions.

Calpain inhibition modulates glycogen synthase kinase 3ß pathways in ischemic myocardium: A proteomic and mechanistic analysis.

BACKGROUND: Calpain inhibition has an enhancing effect on myocardial perfusion and improves myocardial density by inhibiting glycogen synthase kinase 3ß (GSK-3ß) and up-regulating downstream signaling pathways, including the insulin/PI3K and WNT/ß-catenin pathways, in a pig model of chronic myocardial ischemia in the setting of metabolic syndrome. METHODS: Pigs were fed a high-fat diet for 4 weeks, then underwent placement of an ameroid constrictor to the left circumflex artery. Three weeks later, the animals received no drug (high-cholesterol controls [HCC]), a high-dose calpain inhibitor (HCI), a low-dose calpain inhibitor (LCI), or a GSK-3ß inhibitor (GSK-3ßI). The diets and drug regimens were continued for 5 weeks and the myocardial tissue was harvested. RESULTS: Calpain and GSK-3ß inhibition caused an increase in myocardial perfusion ratios at rest and during pacing compared with controls. Pigs in the LCI and HCI groups had increased vessel density in the ischemic myocardium, and pigs in the GSK-3ßI group had increased vessel density in the ischemic and nonischemic myocardium compared with the HCC group. Calpain inhibition modulates proteins involved in the insulin/PI3K and WNT/ß-catenin pathways. Quantitative proteomics revealed that calpain and GSK-3ß inhibition significantly modulated the expression of proteins enriched in cytoskeletal regulation, metabolism, respiration, and calcium-binding pathways. CONCLUSIONS: In the setting of metabolic syndrome, calpain or GSK-3ß inhibition increases vessel density in both ischemic and nonischemic myocardial tissue. Calpain inhibition may exert these effects through the inhibition of GSK-3ß and up-regulation of downstream signaling pathways, including the insulin/PI3K and WNT/ß-catenin pathways.

Glycogen Synthase Kinase 3ß Inhibition Improves Myocardial Angiogenesis and Perfusion in a Swine Model of Metabolic Syndrome.

BACKGROUND: Inhibition of glycogen synthase kinase 3ß (GSK-3ß) has been reported to be cardioprotective during stressful conditions. METHODS AND RESULTS: Pigs were fed a high-fat diet for 4 weeks to develop metabolic syndrome, then underwent placement of an ameroid constrictor to their left circumflex artery to induce chronic myocardial ischemia. Two weeks later, animals received either: no drug (high cholesterol control group [HCC]) or a GSK-3ß inhibitor (GSK-3ß inhibited group [GSK-3ßI]), which were continued for 5 weeks, followed by myocardial tissue harvest. Coronary blood flow and vessel density were significantly increased in the GSK-3ßI group compared to the HCC group. Expression levels of the following proteins were greater in the GSK-3ßI group compared to the HCC group: vascular endothelial growth factor receptor 1 , vascular endothelial cadherin, γ-catenin, ß-catenin, protein kinase B, phosphorylated forkhead box O1, and superoxide dismutase 2. CONCLUSIONS: In the setting of metabolic syndrome, inhibition of GSK-3ß increases blood flow and vessel density in chronically ischemic myocardium. We identified several angiogenic, cell survival, and differentiation pathways that include ß-catenin signaling and AKT/FOXO1, through which GSK-3ß appears to improve vessel density and blood flow. These results may provide a potential mechanism for medical therapy of patients suffering from coronary artery disease and metabolic syndrome.

Calpain inhibition improves collateral-dependent perfusion in a hypercholesterolemic swine model of chronic myocardial ischemia.

PURPOSE: Calpain overexpression is implicated in aberrant angiogenesis. We hypothesized that calpain inhibition (MDL28170) would improve collateral perfusion in a swine model with hypercholesterolemia and chronic myocardial ischemia. METHODS: Yorkshire swine fed a high cholesterol diet for 4 weeks underwent surgical placement of an ameroid constrictor to their left circumflex coronary artery. Three weeks later, animals received no drug, high cholesterol control group (n = 8); low-dose calpain inhibition (0.12 mg/kg; n = 9); or high-dose calpain inhibition (0.25 mg/kg; n = 8). The heart was harvested after 5 weeks. RESULTS: Myocardial perfusion in ischemic myocardium significantly improved with high-dose calpain inhibition at rest and with demand pacing (P = .016 and .011). Endothelium-dependent microvessel relaxation was significantly improved with low-dose calpain inhibition (P = .001). There was a significant increase in capillary density, with low-dose calpain inhibition and high-dose calpain inhibition (P = .01 and .01), and arteriolar density with low-dose calpain inhibition (P = .001). Calpain inhibition significantly increased several proangiogenic proteins, including vascular endothelial growth factor (P = .02), vascular endothelial growth factor receptor 1 (P = .003), vascular endothelial growth factor receptor 2 (P = .003), and talin, a microvascular structural protein (P = .0002). There was a slight increase in proteins implicated in endothelial-dependent (nitric oxide mediated) relaxation, including extracellular signal-regulated kinase, phosphorylated extracellular signal-regulated kinase, and inducible nitric oxide synthase with calpain inhibition. CONCLUSIONS: In the setting of hypercholesterolemia, calpain inhibition improved perfusion, with a trend toward increased collateralization on angiography and increased capillary and arteriolar densities in ischemic myocardium. Calpain inhibition also improved endothelium-dependent microvessel relaxation and increased expression of proteins implicated in angiogenesis and vasodilatation.

Alcohol and the Heart: A Proteomics Analysis of Pericardium and Myocardium in a Swine Model of Myocardial Ischemia.

BACKGROUND: Previous studies have demonstrated that moderate alcohol consumption is cardioprotective and reduces postoperative pericardial adhesions; however, the mechanism is not fully understood. Using proteomic analysis, we sought to objectively investigate the effects of daily moderate alcohol consumption in the pericardium and myocardium in a swine model of chronic myocardial ischemia. METHODS: Fourteen swine underwent placement of an ameroid constrictor to induce chronic myocardial ischemia. Animals were supplemented with 90 mL of ethanol daily (ETOH) or 80 g of sucrose of equal caloric value (SUC). After 7 weeks, the ischemic myocardium and pericardium were harvested for proteomics analysis. RESULTS: Pericardial proteomics analysis yielded 397 proteins, of which 23 were unique to SUC and 52 were unique to ETOH. Of the 322 common proteins, 71 were statistically significant and 23 were characterized (p < 0.05). Alcohol supplementation increased structural proteins, and decreased immune protease inhibitors and coagulation proteins in the pericardium (p < 0.01). Myocardial proteomics analysis yielded 576 proteins, of which 32 were unique to SUC and 21 were unique to ETOH. Of the 523 common proteins, 85 were significant, and 32 were characterized (p < 0.05). Alcohol supplementation decreased cardiac remodeling proteins, cell death proteins and motor proteins, and increased metabolic proteins (p < 0.05). CONCLUSIONS: The results suggest that daily moderate alcohol consumption affects numerous pathways that contribute to cardioprotection, including cardiac remodeling, metabolism, and cell death. Our findings reveal the biosignature of myocardial and pericardial protein expression in the setting of chronic myocardial ischemia and daily moderate alcohol consumption.

Atrial Fibrillation, Neurocognitive Decline and Gene Expression After Cardiopulmonary Bypass.

OBJECTIVE: Atrial fibrillation and neurocognitive decline are common complications after cardiopulmonary bypass. By utilizing genomic microarrays we investigate whether gene expression is associated with postoperative atrial fibrillation and neurocognitive decline. METHODS: Twenty one cardiac surgery patients were prospectively matched and underwent neurocognitive assessments pre-operatively and four days postoperatively. The whole blood collected in the pre-cardiopulmonary bypass, 6 hours after-cardiopulmonary bypass, and on the 4th postoperative day was hybridized to Affymetrix Gene Chip U133 Plus 2.0 Microarrays. Gene expression in patients who developed postoperative atrial fibrillation and neurocognitive decline (n=6; POAF+NCD) was compared with gene expression in patients with postoperative atrial fibrillation and normal cognitive function (n=5; POAF+NORM) and patients with sinus rhythm and normal cognitive function (n=10; SR+NORM). Regulated genes were identified using JMP Genomics 4.0 with a false discovery rate of 0.05 and fold change of >1.5 or <-1.5. RESULTS: Eleven patients developed postoperative atrial fibrillation. Six of these also developed neurocognitive decline. Of the 12 patients with sinus rhythm, only 2 developed neurocognitive decline. POAF+NCD patients had unique regulation of 17 named genes preoperatively, 60 named genes six hours after cardiopulmonary bypass, and 34 named genes four days postoperatively (P<0.05) compared with normal patients. Pathway analysis demonstrated that these genes are involved in cell death, inflammation, cardiac remodeling and nervous system function. CONCLUSION: Patients who developed postoperative atrial fibrillation and neurocognitive decline after cardiopulmonary bypass may have differential genomic responses compared to normal patients and patients with only postoperative atrial fibrillation, suggesting common pathophysiology for these conditions. Further exploration of these genes may provide insight into the etiology and improvements of these morbid outcomes.

Investigating the effects of resveratrol on chronically ischemic myocardium in a swine model of metabolic syndrome: a proteomics analysis.

Resveratrol has been shown to improve cardiac perfusion and ventricular function after chronic ischemic injury. Using proteomic analysis, we sought to objectively investigate potential mechanisms, by which resveratrol exerts its cardioprotective effects in the setting of metabolic syndrome and chronic myocardial ischemia. Yorkshire swine were divided into two groups based on diet: high cholesterol (n=7) or a high-cholesterol diet with supplemental resveratrol (n=6). Four weeks later, all animals underwent surgical placement of an ameroid constrictor to their left circumflex artery. Diets were continued for another 7 weeks, and then the ischemic myocardium was harvested for proteomics analysis. Proteomic analysis identified 669 common proteins between the two groups. Of these proteins, 76 were statistically different, of which 41 were characterized (P<.05). Pathway analysis demonstrated that in animals supplemented with resveratrol, there was a downregulation in several proteins involved with mitochondrial dysfunction, cell death, and unfavorable cardiac remodeling. Furthermore, there was an upregulation in proteins involved in free radical elimination. We conclude that resveratrol supplementation significantly alters several critical protein markers in the chronically ischemic myocardium. Further investigation of these proteins may help elucidate the mechanisms by which resveratrol exerts its cardioprotective effects.

Calpain inhibition decreases myocardial apoptosis in a swine model of chronic myocardial ischemia.

INTRODUCTION: Calpain is a family of cysteine proteases that has an important role in the initiation, regulation, and execution of cell death. Our recent studies using a hypercholesterolemic swine model demonstrated that in the setting of the metabolic syndrome, calpain inhibition (CI) improved collateral-dependent perfusion and increased expression of proteins implicated in angiogenesis and vasodilation. In this study, we hypothesized that CI (by MLD28170) would decrease myocardial apoptosis in the same model. METHODS: Yorkshire swine, all fed a high-cholesterol diet for 4 weeks underwent placement of an ameroid constrictor on the left circumflex coronary artery. Three weeks later, animals received either no drug, termed the high-cholesterol control group (HCC; n = 8); low-dose CI (0.12 mg/kg; LCI, n = 9); or high-dose CI (0.25 mg/kg; HCI, n = 8). The high-cholesterol diet and the CI were continued for 5 weeks, after which the pig was humanely killed and the left ventricular myocardium was harvested and analyzed via terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining, oxyblot analysis, and Western blots. Data were analyzed using the Kruskal-Wallis test. RESULTS: The percentage of apoptotic cells to total cells in ischemic myocardial territory was decreased in the LCI and HCI groups compared with the HCC group as shown by TUNEL staining (P = .018). There was a decrease in proapoptotic proteins, including cleaved caspase 3, caspase 9, cleaved caspase 9, Bax, BAD, p-BAD, and Erk 1/2 (P ≤ .049 each), but no decrease in caspase 3 (P = .737). There was also an increase in antiapoptotic proteins, including BCL-2 and p-BCL2 (P ≤ .025 each). In the ischemic myocardium, several proangiogenic proteins were increased in the LCI and HCI groups compared with the HCC group, including p-AKT, p-eNOS, and eNOS (P ≤ .006 each) but there was no increase in AKT (P = .311). CI decreased tissue oxidative stress in both the LCI and HCI groups compared to the HCC group as shown by oxyblot analysis (P = .021). CONCLUSION: In the setting of hypercholesterolemia, CI decreases apoptosis and the expression of proteins in proapoptotic signaling pathways. CI also increased expression of proteins implicated in anti apoptotic pathways and improves oxidative stress in ischemic myocardial tissue.