We show here that the apposition of plasma membrane caveolae and mitochondria (first noted in electron micrographs >50 yr ago) and caveolae-mitochondria interaction regulates adaptation to cellular stress by modulating the structure and function of mitochondria. In C57Bl/6 mice engineered to overexpress caveolin specifically in cardiac myocytes (Cav-3 OE), localization of caveolin to mitochondria increases membrane rigidity (4.2%; P<0.05), tolerance to calcium, and respiratory function (72% increase in state 3 and 23% increase in complex IV activity; P<0.05), while reducing stress-induced generation of reactive oxygen species (by 20% in cellular superoxide and 41 and 28% in mitochondrial superoxide under states 4 and 3, respectively; P<0.05) in Cav-3 OE vs. TGneg. By contrast, mitochondrial function is abnormal in caveolin-knockout mice and Caenorhabditis elegans with null mutations in caveolin (60% increase free radical in Cav-2 C. elegans mutants; P<0.05). In human colon cancer cells, mitochondria with increased caveolin have a 30% decrease in apoptotic stress (P<0.05), but cells with disrupted mitochondria-caveolin interaction have a 30% increase in stress response (P<0.05). Targeted gene transfer of caveolin to mitochondria in C57Bl/6 mice increases cardiac mitochondria tolerance to calcium, enhances respiratory function (increases of 90% state 4, 220% state 3, 88% complex IV activity; P<0.05), and decreases (by 33%) cardiac damage (P<0.05). Physical association and apparently the transfer of caveolin between caveolae and mitochondria is thus a conserved cellular response that confers protection from cellular damage in a variety of tissues and settings.

Introduction Natriuretic peptides modulate cardiac hypertrophy and are potential therapeutic options for patients with heart failure. Caveolae, microdomains in the plasma membrane that contain caveolin proteins and natriuretic peptide receptors, have been implicated in cardiac hypertrophy and natriuretic peptide localization. We hypothesized that cardiac myocyte-specific overexpression of caveolin-3, a muscle-specific caveolin, would alter natriuretic peptide signaling and attenuate cardiac hypertrophy. Methods and Results We generated transgenic mice with cardiac myocyte-specific overexpression of caveolin-3 (Cav-3 OE) and also used an adenoviral construct to increase Cav-3 in cardiac myocytes. Cav-3 OE mice subjected to transverse aortic constriction had increased survival, reduced cardiac hypertrophy and maintenance of cardiac function compared to control mice. In left ventricle at baseline, mRNA for atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) were increased 7-fold and 3-fold, respectively in Cav-3 OE mice compared to controls and were accompanied by increased protein expression for ANP and BNP. In addition, ventricles from Cav-3 OE mice had greater cGMP levels, less NFAT nuclear translocation, and more nuclear Akt phosphorylation than did ventricles from controls. Cardiac myocytes incubated with Cav-3 adenovirus showed increased expression of Cav-3, ANP, and Akt phosphorylation. Incubation with methyl-β-cyclodextrin, which disrupts caveolae, or with wortmannin, a PI3K inhibitor, blocked the increase in ANP expression. Discussion These results imply that cardiac myocyte-specific overexpression of Cav-3 is a novel strategy to enhance natriuretic peptide expression, attenuate hypertrophy and possibly exploit the therapeutic benefits of natriuretic peptides in cardiac hypertrophy and heart failure.

Background Caveolae are small, flask-like invaginations of the plasma membrane. Caveolins are structural proteins found in caveolae that have scaffolding properties to allow organization of signaling. We tested the hypothesis that delayed cardiac protection induced by volatile anesthetics is caveolae/caveolin-dependent. Methods An in vivo mouse model of ischemia-reperfusion injury with delayed anesthetic preconditioning was tested in wild-type, caveolin-1 knockout, and caveolin-3 knockout mice. Mice were exposed to 30 min oxygen or isoflurane and allowed to recover for 24 h. After 24 h recovery, mice underwent 30 min coronary artery occlusion, followed by 2 h of reperfusion at which time infarct size was determined. Biochemical assays were also performed in excised hearts. Results Infarct size as a percent of the area at risk was reduced by isoflurane in wild-type (24.0 ± 8.8% vs. 45.1 ± 10.1%) and caveolin-1 knockout mice (27.2 ± 12.5%). Caveolin-3 knockout mice did not show delayed anesthetic preconditioning (41.5 ± 5.0%). Microscopically distinct caveolae were observed in wild-type and caveolin-1 knockout mice but not caveolin-3 knockout mice. Delayed anesthetic preconditioning increased the amount of caveolin-3 protein but not caveolin-1 protein in discontinuous sucrose gradient buoyant fractions. Additionally, glucose transporter-4 was increased in buoyant fractions and caveolin-3/glucose transporter-4 colocalization was observed in wild-type and caveolin-1 knockout mice after anesthetic preconditioning. Conclusions These results demonstrate that delayed anesthetic preconditioning involves translocation of caveolin-3 and glucose transporter-4 to caveolae resulting in delayed protection in the myocardium.

Ischemic damage is recognized to cause cardiomyocyte (CM) death and myocardial dysfunction, but the role of cell-matrix interactions and integrins in this process has not been extensively studied. Expression of α7β1D integrin, the dominant integrin in normal adult CMs, increases during ischemia/reperfusion (I/R), while deficiency of β1 integrins increases ischemic damage. We hypothesized that the forced overexpression of integrins on the CM would offer protection from I/R injury. Tg mice with CM-specific overexpression of integrin α7β1D exposed to I/R had a substantial reduction in infarct size compared with that of α5β1D-overexpressing mice and WT littermate controls. Using isolated CMs, we found that α7β1D preserved mitochondrial membrane potential during hypoxia/reoxygenation (H/R) injury via inhibition of mitochondrial Ca 2+ overload but did not alter H/R effects on oxidative stress. Therefore, we assessed Ca 2+ handling proteins in the CM and found that β1D integrin colocalized with ryanodine receptor 2 (RyR2) in CM T-tubules, complexed with RyR2 in human and rat heart, and specifically bound to RyR2 amino acids 165-175. Integrins stabilized the RyR2 interdomain interaction, and this stabilization required integrin receptor binding to its ECM ligand. These data suggest that α7β1D integrin modifies Ca 2+ regulatory pathways and offers a means to protect the myocardium from ischemic injury.

Background Volatile anesthetics have a dual effect on cell survival dependent on caveolin expression. The effect of volatile anesthetics on cancer cell survival and death after anesthetic exposure has not been well investigated. The authors examined the effects of isoflurane exposure on apoptosis and its regulation by caveolin-1 (Cav-1). Methods The authors exposed human colon cancer cell lines to isoflurane and proapoptotic stimuli and assessed what role Cav-1 plays in cell protection. They evaluated apoptosis using assays for nucleosomal fragmentation, cleaved caspase 3 expression, and caspase activity assays. To test the mechanism, they used pharmacologic inhibitors (i.e., pertussis toxin) and assessed changes in glycolysis. Results Apoptosis as measured by nucleosomal fragmentation was enhanced by isoflurane (1.2% in air) in HT29 (by 64% relative to control, P < 0.001) and decreased in HCT116 (by 23% relative to control, P < 0.001) cells. Knockdown of Cav-1 in HCT116 cells increased the sensitivity to apoptotic stimuli but not with scrambled small interfering RNA (siRNA) treatment (19.7 ± 0.4 vs. 20.0 ± 0.6, P =0.7786 and 19.7 ± 0.5 vs. 16.3 ± 0.4, P =0.0012, isoflurane vs. control in Cav-1 small interfering RNA vs. scrambled small interfering RNA treated cells, respectively). The protective effect of isoflurane with various exposure times on apoptosis was enhanced in HT29 cells overexpressing Cav-1 (P <0.001 by two-way ANOVA). Pertussis toxin effectively blocked the antiapoptotic effect of isoflurane exhibited by Cav-1 in all cell lines. Cav-1 cells had increased glycolysis with isoflurane exposure; however, in the presence of tumor necrosis factor-related apoptosis-inducing ligand, this increase in glycolysis was maintained in HT29-Cav-1 but not control cells. Conclusion Brief isoflurane exposure leads to resistance against apoptosis via a Cav-1–dependent mechanism.