The caveolar-mitochondrial interface: regulation of cellular metabolism in physiology and pathophysiology.

The plasma membrane is an important cellular organelle that is often overlooked in terms of a primary factor in regulating physiology and pathophysiology. There is emerging evidence to suggest that the plasma membrane serves a greater purpose than a simple barrier or transporter of ions. New paradigms suggest that the membrane serves as a critical bridge to connect extracellular to intracellular communication particularly to regulate energy and metabolism by forming physical and biochemical associations with intracellular organelles. This review will focus on the relationship of a particular membrane microdomain - caveolae - with mitochondria and the particular implication of this to physiology and pathophysiology.

Ataxia telangiectasia-mutated kinase deficiency exacerbates left ventricular dysfunction and remodeling late after myocardial infarction.

Ataxia telangiectasia-mutated kinase (ATM), a cell cycle checkpoint protein, is activated in response to DNA damage and oxidative stress. We have previously shown that ATM deficiency is associated with increased apoptosis and fibrosis and attenuation of cardiac dysfunction early (1-7 days) following myocardial infarction (MI). Here, we tested the hypothesis that enhanced fibrosis and apoptosis, as observed early post-MI during ATM deficiency, exacerbate cardiac dysfunction and remodeling in ATM-deficient mice late post-MI. MIs were induced in wild-type (WT) and ATM heterozygous knockout (hKO) mice by ligation of the left anterior descending artery. Left ventricular (LV) structural and functional parameters were assessed by echocardiography 14 and 28 days post-MI, whereas biochemical parameters were measured 28 days post-MI. hKO-MI mice exhibited exacerbated LV dysfunction as observed by increased LV end-systolic volume and decreased percent fractional shortening and ejection fraction. Infarct size and thickness were not different between the two genotypes. Myocyte cross-sectional area was greater in hKO-MI group. The hKO-MI group exhibited increased fibrosis in the noninfarct and higher expression of α-smooth muscle actin (myofibroblast marker) in the infarct region. Apoptosis and activation of GSK-3ß (proapoptotic kinase) were significantly lower in the infarct region of hKO-MI group. Matrix metalloproteinase 2 (MMP-2) expression was not different between the two genotypes. However, MMP-9 expression was significantly lower in the noninfarct region of hKO-MI group. Thus ATM deficiency exacerbates cardiac remodeling late post-MI with effects on cardiac function, fibrosis, apoptosis, and myocyte hypertrophy.

Exogenous ubiquitin modulates chronic ß-adrenergic receptor-stimulated myocardial remodeling: role in Akt activity and matrix metalloproteinase expression.

ß-Adrenergic receptor (ß-AR) stimulation increases extracellular ubiquitin (UB) levels, and extracellular UB inhibits ß-AR-stimulated apoptosis in adult cardiac myocytes. This study investigates the role of exogenous UB in chronic ß-AR-stimulated myocardial remodeling. l-Isoproterenol (ISO; 400 µg·kg(-1)·h(-1)) was infused in mice in the presence or absence of UB (1 µg·g(-1)·h(-1)). Left ventricular (LV) structural and functional remodeling was studied 7 days after infusion. UB infusion enhanced serum UB levels. In most parts, UB alone had no effect on morphometric or functional parameters. Heart weight-to-body weight ratios were increased to a similar extent in the ISO and UB + ISO groups. Echocardiographic analyses showed increased percent fractional shortening, ejection fraction, and LV circumferential stress and fiber-shortening velocity in the ISO group. These parameters were significantly lower in UB + ISO vs. ISO. Isovolumic contraction and relaxation times and ejection time were significantly lower in ISO vs. UB + ISO. The increase in the number of TUNEL-positive myocytes and fibrosis was significantly higher in ISO vs. UB + ISO. Activation of Akt was higher, whereas activation of GSK-3ß and JNKs was lower in UB + ISO vs ISO. Expression of MMP-2, MMP-9, and TIMP-2 was higher in UB + ISO vs ISO. In isolated cardiac fibroblasts, UB enhanced expression of MMP-2 and TIMP-2 in the presence of ISO. Neutralizing UB antibodies negated the effects of UB on MMP-2 expression, whereas recombinant UB enhanced MMP-2 expression. UB activated Akt, and inhibition of Akt inhibited UB + ISO-mediated increases in MMP-2 expression. Thus, exogenous UB plays an important role in ß-AR-stimulated myocardial remodeling with effects on LV function, fibrosis, and myocyte apoptosis.

Lack of ataxia telangiectasia mutated kinase induces structural and functional changes in the heart: role in ß-adrenergic receptor-stimulated apoptosis.

Ataxia telangiectasia mutated kinase (ATM) is involved in cell cycle checkpoints, DNA repair and apoptosis. ß-Adrenergic receptor (ß-AR) stimulation induces cardiac myocyte apoptosis. Here we analysed basal myocardial structure and function in ATM knockout (KO) mice and tested the hypothesis that ATM modulates ß-AR-stimulated myocyte apoptosis. Left ventricular (LV) structure and function, myocyte apoptosis, fibrosis and expression of fibrosis-, hypertrophy- and apoptosis-related proteins were examined in wild-type (WT) and KO mice with or without l-isoprenaline treatment for 24 h. Body and heart weights were lower in KO mice. M-Mode echocardiography showed reduced septal wall thicknesses and LV diameters in KO mice. Doppler echocardiography showed an increased ratio of early peak velocity (E wave) to that of the late LV filling (A wave) in KO mice. Basal fibrosis and myocyte cross-sectional area were greater in KO hearts. Expression of fibrosis-related genes (connective tissue growth factor and plasminogen activator inhibitor-1) and hypertrophy-related gene (atrial natriuretic peptide) was higher in KO hearts. ß-Adrenergic receptor stimulation increased myocyte apoptosis to a similar extent in both groups. Activation of c-Jun N-terminal kinases and expression and phosphorylation of p53 in response to ß-AR stimulation were only observed in the WT group. Akt phosphorylation was lower in KO sham-treated animals and remained lower following ß-AR stimulation in the KO group. ß-Adrenergic receptor stimulation activated glycogen synthase kinase-3ß to a similar extent in both groups. Thus, lack of ATM induces structural and functional changes in the heart, with enhanced myocardial fibrosis and myocyte hypertrophy. ß-Adrenergic receptor-stimulated apoptosis in WT hearts is associated with a p53- and JNKs-dependent mechanism, while decreased Akt activity may play a role in increased myocyte apoptosis in the absence of ATM.

Β-adrenergic receptor stimulation induces endoplasmic reticulum stress in adult cardiac myocytes: role in apoptosis.

Accumulation of misfolded proteins and alterations in calcium homeostasis induces endoplasmic reticulum (ER) stress, leading to apoptosis. In this study, we tested the hypothesis that ß-AR stimulation induces ER stress, and induction of ER stress plays a pro-apoptotic role in cardiac myocytes. Using thapsigargin and brefeldin A, we demonstrate that ER stress induces apoptosis in adult rat ventricular myocytes (ARVMs). ß-AR-stimulation (isoproterenol; 3h) significantly increased expression of ER stress proteins, such as GRP-78, Gadd-153, and Gadd-34, while activating caspase-12 in ARVMs. In most parts, these effects were mimicked by thapsigargin. ß-AR stimulation for 15 min increased PERK and eIF-2α phosphorylation. PERK phosphorylation remained higher, while eIF-2α phosphorylation declined thereafter, reaching to ~50% below basal levels at 3 h after ß-AR stimulation. This decline in eIF-2α phosphorylation was prevented by ß1-AR, not by ß2-AR antagonist. Forskolin, adenylyl cyclase activator, simulated the effects of ISO on eIF-2α phosphorylation. Salubrinal (SAL), an ER stress inhibitor, maintained eIF-2α phosphorylation and inhibited ß-AR-stimulated apoptosis. Furthermore, inhibition of caspase-12 using z-ATAD inhibited ß-AR-stimulated and thapsigargin-induced apoptosis. In vivo, ß-AR stimulation induced ER stress in the mouse heart as evidenced by increased expression of GRP-78 and Gadd-153, activation of caspase-12, and dephosphorylation of eIF-2α. SAL maintained phosphorylation of eIF-2α, inhibited activation of caspase-12, and decreased ß-AR-stimulated apoptosis in the heart. Thus, ß-AR stimulation induces ER stress in cardiac myocytes and in the heart, and induction of ER stress plays a pro-apoptotic role.

Ataxia telangiectasia mutated kinase plays a protective role in ß-adrenergic receptor-stimulated cardiac myocyte apoptosis and myocardial remodeling.

ß-Adrenergic receptor (ß-AR) stimulation induces cardiac myocyte apoptosis and plays an important role in myocardial remodeling. Here we investigated expression of various apoptosis-related genes affected by ß-AR stimulation, and examined first time the role of ataxia telangiectasia mutated kinase (ATM) in cardiac myocyte apoptosis and myocardial remodeling following ß-AR stimulation. cDNA array analysis of 96 apoptosis-related genes indicated that ß-AR stimulation increases expression of ATM in the heart. In vitro, RT-PCR confirmed increased ATM expression in adult cardiac myocytes in response to ß-AR stimulation. Analysis of left ventricular structural and functional remodeling of the heart in wild-type (WT) and ATM heterozygous knockout mice (hKO) 28 days after ISO-infusion showed increased heart weight to body weight ratio in both groups. M-mode echocardiography showed increased percent fractional shortening (%FS) and ejection fraction (EF%) in both groups 28 days post ISO-infusion. Interestingly, the increase in %FS and EF% was significantly lower in the hKO-ISO group. Cardiac fibrosis and myocyte apoptosis were higher in hKO mice at baseline and ISO-infusion increased fibrosis and apoptosis to a greater extent in hKO-ISO hearts. ISO-infusion increased phosphorylation of p53 (Serine-15) and expression of p53 and Bax to a similar extent in both groups. hKO-Sham and hKO-ISO hearts exhibited reduced intact ß1 integrin levels. MMP-2 protein levels were significantly higher, while TIMP-2 protein levels were lower in hKO-ISO hearts. MMP-9 protein levels were increased in WT-ISO, not in hKO hearts. In conclusion, ATM plays a protective role in cardiac remodeling in response to ß-AR stimulation.

Osteopontin: role in extracellular matrix deposition and myocardial remodeling post-MI.

Remodeling after myocardial infarction (MI) associates with left ventricular (LV) dilation, decreased cardiac function and increased mortality. The dynamic synthesis and breakdown of extracellular matrix (ECM) proteins play a significant role in myocardial remodeling post-MI. Expression of osteopontin (OPN) increases in the heart post-MI. Evidence has been provided that lack of OPN induces LV dilation which associates with decreased collagen synthesis and deposition. Inhibition of matrix metalloproteinases, key players in ECM remodeling process post-MI, increased ECM deposition (fibrosis) and improved LV function in mice lacking OPN after MI. This review summarizes--1) signaling pathways leading to increased expression of OPN in the heart; 2) the alterations in the structure and function of the heart post-MI in mice lacking OPN; and 3) mechanisms involved in OPN-mediated ECM remodeling post-MI.

Role of osteopontin in heart failure associated with aging.

Cardiovascular disease is one of the leading causes of death in the elderly. Much of the morbidity and mortality in the elderly is attributable to acute ischemic events leading to myocardial infarction (MI) and death of cardiac myocytes. Evidence has been provided that aging associated with adverse remodeling post MI as demonstrated by less effective myocardial repair, greater infarct expansion, and septal hypertrophy. Expression of osteopontin (OPN) increases in the heart post MI. Transgenic mice studies suggest that increased expression of OPN plays a protective role in post-MI LV remodeling by modulating collagen deposition and fibrosis. OPN, a multifunctional protein, has the potential to influence the molecular and cellular changes associated with infarct healing. The post-MI infarct healing process involves temporarily overlapping phases that include the following--(1) inflammation with migration and adhesion of neutrophils and macrophages, phagocytosis and inflammatory gene expression; (2) tissue repair with fibroblast adhesion and proliferation, myofibroblast differentiation, extracellular matrix deposition and scar formation; and (3) structural and functional remodeling of infarcted and non-infarcted myocardium through cardiac myocyte apoptosis, hypertrophy and myocardial angiogenesis. This review is focused on the expression of OPN in the heart post MI and its role in various phases of infarct healing.

Deficiency of ataxia telangiectasia mutated kinase delays inflammatory response in the heart following myocardial infarction.

BACKGROUND: Ataxia­telangiectasia results from mutations in ataxia telangiectasia mutated kinase (ATM) gene. We recently reported that ATM deficiency attenuates left ventricular (LV) dysfunction and dilatation 7 days after myocardial infarction (MI) with increased apoptosis and fibrosis. Here we investigated the role of ATM in the induction of inflammatory response, and activation of survival signaling molecules in the heart acute post­MI. METHODS AND RESULTS: LV structure, function, inflammatory response, and biochemical parameters were measured in wild­type (WT) and ATM heterozygous knockout (hKO) mice 1 and 3 days post­MI. ATM deficiency had no effect on infarct size. MI­induced decline in heart function, as measured by changes in percent fractional shortening, ejection fraction and LV end systolic and diastolic volumes, was lower in hKO­MI versus WT­MI (n=10 to 12). The number of neutrophils and macrophages was significantly lower in the infarct LV region of hKO versus WT 1 day post­MI. Fibrosis and expression of α­smooth muscle actin (myofibroblast marker) were higher in hKO­MI, while active TGF­ß1 levels were higher in the WT­MI 3 days post­MI. Myocyte cross­sectional area was higher in hKO­sham with no difference between the two MI groups. MMP­9 protein levels were similarly increased in the infarct LV region of both MI groups. Apoptosis was significantly higher in the infarct LV region of hKO at both time points. Akt activation was lower, while Bax expression was higher in hKO­MI infarct. CONCLUSION: ATM deficiency results in decreased dilative remodeling and delays inflammatory response acute post­MI. However, it associates with increased fibrosis and apoptosis.

Deficiency of ataxia telangiectasia mutated kinase modulates cardiac remodeling following myocardial infarction: involvement in fibrosis and apoptosis.

UNLABELLED: Ataxia telangiectasia mutated kinase (ATM) is a cell cycle checkpoint protein activated in response to DNA damage. We recently reported that ATM plays a protective role in myocardial remodeling following ß-adrenergic receptor stimulation. Here we investigated the role of ATM in cardiac remodeling using myocardial infarction (MI) as a model. METHODS AND RESULTS: Left ventricular (LV) structure, function, apoptosis, fibrosis, and protein levels of apoptosis- and fibrosis-related proteins were examined in wild-type (WT) and ATM heterozygous knockout (hKO) mice 7 days post-MI. Infarct sizes were similar in both MI groups. However, infarct thickness was higher in hKO-MI group. Two dimensional M-mode echocardiography revealed decreased percent fractional shortening (%FS) and ejection fraction (EF) in both MI groups when compared to their respective sham groups. However, the decrease in %FS and EF was significantly greater in WT-MI vs hKO-MI. LV end systolic and diastolic diameters were greater in WT-MI vs hKO-MI. Fibrosis, apoptosis, and α-smooth muscle actin staining was significantly higher in hKO-MI vs WT-MI. MMP-2 protein levels and activity were increased to a similar extent in the infarct regions of both groups. MMP-9 protein levels were increased in the non-infarct region of WT-MI vs WT-sham. MMP-9 protein levels and activity were significantly lower in the infarct region of WT vs hKO. TIMP-2 protein levels similarly increased in both MI groups, whereas TIMP-4 protein levels were significantly lower in the infarct region of hKO group. Phosphorylation of p53 protein was higher, while protein levels of manganese superoxide dismutase were significantly lower in the infarct region of hKO vs WT. In vitro, inhibition of ATM using KU-55933 increased oxidative stress and apoptosis in cardiac myocytes.