Protein kinase C βII (PKCβII) levels increase in the myocardium of patients with end-stage heart failure (HF). Also targeted over-expression of PKCβII in the myocardium of mice leads to dilated cardiomyopathy associated with inflammation, fibrosis and myocardial dysfunction. These reports suggest a deleterious role of PKCβII in HF development. Using a post-myocardial infarction (MI) model of heart failure in rats, we determined the benefit of chronic inhibition of PKCβII on the progression of heart failure over a period of 6 weeks after the onset of symptoms and the cellular basis for these effects. Four weeks after MI, rats with HF signs that were treated for 6 weeks with the PKCβII selective inhibitor (βIIV5-3 conjugated to TAT47-57 alone) (3mg/kg/day) showed improved fractional shortening (from 21% to 35%) compared to control (TAT47-57 alone). Formalin-fixed mid-ventricle tissue sections stained with picrosirius red, hematoxylin-eosin and toluidine blue dyes exhibited a 150% decrease in collagen deposition, a two-fold decrease in inflammation and a 30% reduction in mast cell degranulation, respectively, in rat hearts treated with the selective PKCβII inhibitor. Further, a 90% decrease in active TGFβ1 and a significant reduction in SMAD2/3 phosphorylation indicated that the selective inhibition of PKCβII attenuates cardiac remodeling mediated by the TGF-SMAD signaling pathway. Therefore, sustained selective inhibition of PKCβII in a post-MI HF rat model improves cardiac function and is associated with inhibition of pathological myocardial remodeling.
Protein kinase; PKCβII inhibitor peptide; cardiac remodeling; heart failure; myocardial infarction; mast cells, myocardial fibrosis; inflammation
Genetic variants near/within the ALDH2 gene encoding the mitochondrial aldehyde dehydrogenase 2 have been associated with blood pressure and hypertension in several case–control association studies in East Asian populations.
Three common tag single nucleotide polymorphisms (tagSNP) in the ALDH2 gene were genotyped in 1,134 subjects of Chinese origin from the Stanford Asia-Pacific Program for Hypertension and Insulin Resistance (SAPPHIRe) family cohort. We examined whether the ALDH2 SNP genotypes predicted the development of hypertension in the prospective SAPPHIRe cohort.
Over an average follow-up period of 5.7 years, carriers homozygous for the rs2238152 T allele in the ALDH2 gene were more likely to progress to hypertension than were non-carriers (hazard ratio [HR], 2.88, 95% confidence interval [CI], 1.06-7.84, P = 0.03), corresponding to a population attributable risk of ~7.1%. The risk associated with the rs2238152 T allele were strongest in heavy/moderate alcohol drinkers and was reduced in non-drinkers, indicating an interaction between ALDH2 genetic variants and alcohol intake on the risk of hypertension (P for interaction = 0.04). The risk allele was associated with significantly lower ALDH2 gene expression levels in human adipose tissue.
ALDH2 genetic variants were associated with progression to hypertension in a prospective Chinese cohort. The association was modified by alcohol consumption.
ALDH2; Hypertension; SNP; Chinese
Myocardial remodeling and heart failure (HF) are common sequelae of many forms of cardiovascular disease and a leading cause of mortality worldwide. Accumulation of damaged cardiac proteins in heart failure has been described. However, how protein quality control (PQC) is regulated and its contribution to HF development are not known. Here, we describe a novel role for activated protein kinase C isoform βII (PKCβII) in disrupting PQC. We show that active PKCβII directly phosphorylated the proteasome and inhibited proteasomal activity in vitro and in cultured neonatal cardiomyocytes. Importantly, inhibition of PKCβII, using a selective PKCβII peptide inhibitor (βIIV5-3), improved proteasomal activity and conferred protection in cultured neonatal cardiomyocytes. We also show that sustained inhibition of PKCβII increased proteasomal activity, decreased accumulation of damaged and misfolded proteins and increased animal survival in two rat models of HF. Interestingly, βIIV5-3-mediated protection was blunted by sustained proteasomal inhibition in HF. Finally, increased cardiac PKCβII activity and accumulation of misfolded proteins associated with decreased proteasomal function were found also in remodeled and failing human hearts, indicating a potential clinical relevance of our findings. Together, our data highlights PKCβII as a novel inhibitor of proteasomal function. PQC disruption by increased PKCβII activity in vivo appears to contribute to the pathophysiology of heart failure, suggesting that PKCβII inhibition may benefit patients with heart failure. (218 words)
Renin released by ischemia/reperfusion (I/R) from cardiac mast cells activates a local renin-angiotensin system (RAS). This exacerbates norepinephrine release and reperfusion arrhythmias (VT/VF), making RAS a new therapeutic target in myocardial ischemia.
Methods and Results
We investigated whether ischemic preconditioning (IPC) prevents cardiac RAS activation in guinea-pig hearts ex-vivo. When I/R (20-min ischemia/30-min reperfusion) was preceded by IPC (2×5-min I/R cycles), renin and norepinephrine release and VT/VF duration were markedly decreased, a cardioprotective anti-RAS effect. Activation and blockade of adenosine A2b/A3-receptors, and activation and inhibition of PKCε, mimicked and prevented, respectively, the anti-RAS effects of IPC. Moreover, activation of A2b/A3-receptors, or activation of PKCε, prevented degranulation and renin release elicited by peroxide in cultured mast cells (HMC-1). Activation and inhibition of mitochondrial aldehyde dehydrogenase type-2 (ALDH2) also mimicked and prevented, respectively, the cardioprotective anti-RAS effects of IPC. Furthermore, ALDH2 activation inhibited degranulation and renin release by reactive aldehydes in HMC-1. Notably, PKCε and ALDH2 were both activated by A2b/A3-receptor stimulation in HMC-1, and PKCε inhibition prevented ALDH2 activation.
The results uncover a signaling cascade initiated by A2b/A3-receptors, which triggers PKCε-mediated ALDH2 activation in cardiac mast cells, contributing to IPC-induced cardioprotection by preventing mast-cell renin release and the dysfunctional consequences of local RAS activation. Thus, unlike classical IPC where cardiac myocytes are the main target, cardiac mast cells are the critical site at which the cardioprotective anti-RAS effects of IPC develop.
Renin; Ischemia; Reperfusion; Norepinephrine; Arrhythmia
Heart failure (HF) in which the blood supply does not match the body's needs, affects 10% of the population over 65 years old. The protein kinase C (PKC) family of kinases has a key role in normal and disease states. Here we discuss the role of PKC in HF and focus on the use of specific PKC regulators to identify the mechanism leading to this Pathology and potential leads for therapeutics.
Cardiac mitochondria, the main source of energy as well as free radicals, are vital organelles for normal functioning of the heart. Mitochondrial number, structure, turnover and function are regulated by processes such as mitochondrial protein quality control, mitochondrial fusion and fission and mitophagy. Recent studies suggest that abnormal changes in these mitochondrial regulatory processes may contribute to the pathology of heart failure (HF). Here we discuss these processes and their potential as therapeutic targets.
Acute administration of ethanol can reduce cardiac ischemia/reperfusion injury. Previous studies demonstrated that the acute cytoprotective effect of ethanol on the myocardium is mediated by protein kinase C epsilon (PKCε). We recently identified aldehyde dehydrogenase 2 (ALDH2) as an PKCε substrate, whose activation is necessary and sufficient to confer cardioprotection in vivo. ALDH2 metabolizes cytotoxic reactive aldehydes, such as 4-hydroxy-2-nonenal (4-HNE), which accumulate during cardiac ischemia/reperfusion. Here, we used a combination of PKCε knockout mice and a direct activator of ALDH2, Alda-44, to further investigate the interplay between PKCε and ALDH2 in cardioprotection. We report that ethanol preconditioning requires PKCε, whereas direct activation of ALDH2 reduces infarct size in both wild type and PKCε knockout hearts. Our data suggest that ALDH2 is downstream of PKCε in ethanol preconditioning and that direct activation of ALDH2 can circumvent the requirement of PKCε to induce cytoprotection. We also report that in addition to ALDH2 activation, Alda-44 prevents 4-HNE induced inactivation of ALDH2 by reducing the formation of 4-HNE-ALDH2 protein adducts. Thus, Alda-44 promotes metabolism of cytotoxic reactive aldehydes that accumulate in ischemic myocardium. Taken together, our findings suggest that direct activation of ALDH2 may represent a method of harnessing the cardioprotective effect of ethanol without the side effects associated with alcohol consumption.
Impaired mitochondrial fusion/fission plays a causal role in neuronal death. This study delineated a PKCδ-related signaling cascade in which excessive mitochondrial fission is induced during oxidative stress. Moreover, a selective peptide inhibitor of PKCδ inhibits impaired mitochondrial fission under these pathological conditions.
Neuronal cell death in a number of neurological disorders is associated with aberrant mitochondrial dynamics and mitochondrial degeneration. However, the triggers for this mitochondrial dysregulation are not known. Here we show excessive mitochondrial fission and mitochondrial structural disarray in brains of hypertensive rats with hypertension-induced brain injury (encephalopathy). We found that activation of protein kinase Cδ (PKCδ) induced aberrant mitochondrial fragmentation and impaired mitochondrial function in cultured SH-SY5Y neuronal cells and in this rat model of hypertension-induced encephalopathy. Immunoprecipitation studies indicate that PKCδ binds Drp1, a major mitochondrial fission protein, and phosphorylates Drp1 at Ser 579, thus increasing mitochondrial fragmentation. Further, we found that Drp1 Ser 579 phosphorylation by PKCδ is associated with Drp1 translocation to the mitochondria under oxidative stress. Importantly, inhibition of PKCδ, using a selective PKCδ peptide inhibitor (δV1-1), reduced mitochondrial fission and fragmentation and conferred neuronal protection in vivo and in culture. Our study suggests that PKCδ activation dysregulates the mitochondrial fission machinery and induces aberrant mitochondrial fission, thus contributing to neurological pathology.
In approximately one billion people, a point mutation inactivates a key detoxifying enzyme, aldehyde dehydrogenase (ALDH2). This mitochondrial enzyme metabolizes toxic biogenic and environmental aldehydes, including the endogenously produced 4-hydroxynonenal (4HNE) and the environmental pollutant, acrolein. ALDH2 also bioactivates nitroglycerin, but it is best known for its role in ethanol metabolism. The accumulation of acetaldehyde following the consumption of even a single alcoholic beverage leads to the Asian Alcohol-induced Flushing Syndrome in ALDH2*2 homozygotes. The ALDH2*2 allele is semi-dominant and heterozygotic individuals exhibit a similar, but not as severe phenotype. We recently identified a small molecule, Alda-1, which activates wild-type ALDH2 and restores near wild-type activity to ALDH2*2. The structures of Alda-1 bound to ALDH2 and ALDH2*2 reveal how Alda-1 activates the wild-type enzyme and how it restores the activity of ALDH2*2 by acting as a structural chaperone.
Mitochondrial aldehyde dehydrogenase 2 (ALDH2) is emerging as a key enzyme involved in cytoprotection in the heart. ALDH2 mediates both the detoxification of reactive aldehydes such as acetaldehyde and 4-hydroxy-2-nonenal (4-HNE) and the bioactivation of nitroglycerin (GTN) to nitric oxide (NO). In addition, chronic nitrate treatment results in ALDH2 inhibition and contributes to nitrate tolerance. Our lab recently identified ALDH2 to be a key mediator of endogenous cytoprotection. We reported that ALDH2 is phosphorylated and activated by the survival kinase protein kinase C epsilon (PKCε) and found a strong inverse correlation between ALDH2 activity and infarct size. We also identified a small molecule ALDH2 activator (Alda-1) which reduces myocardial infarct size induced by ischemia/reperfusion in vivo. In this review, we discuss evidence that ALDH2 is a key mediator of endogenous survival signaling in the heart, suggest possible cardioprotective mechanisms mediated by ALDH2, and discuss potential clinical implications of these findings.
Protein kinase C epsilon (PKCε) is critical for cardiac protection from ischaemia and reperfusion (IR) injury. PKCε substrates that mediate cytoprotection reside in the mitochondria. However, the mechanism enabling mitochondrial translocation and import of PKCε to enable phosphorylation of these substrates is not known. Heat shock protein 90 (HSP90) is a cytoprotective protein chaperone that participates in mitochondrial import of a number of proteins. Here, we investigated the role of HSP90 in mitochondrial import of PKCε.
Methods and results
Using an ex vivo perfused rat heart model of IR, we found that PKCε translocates from the cytosol to the mitochondrial fraction following IR. Immunogold electron microscopy and mitochondrial fractionation demonstrated that following IR, mitochondrial PKCε is localized within the mitochondria, on the inner mitochondrial membrane. Pharmacological inhibition of HSP90 prevented IR-induced interaction between PKCε and the translocase of the outer membrane (Tom20), reduced mitochondrial import of PKCε, and increased necrotic cell death by ∼70%. Using a rational approach, we designed a 7-amino acid peptide activator of PKCε, derived from an HSP90 homologous sequence located in the C2 domain of PKCε (termed ψεHSP90). Treatment with this peptide (conjugated to the cell permeating TAT protein-derived peptide, TAT47–57) increased PKCε–HSP90 protein–protein interaction, enhanced mitochondrial translocation of PKCε, increased phosphorylation and activity of an intra-mitochondrial PKCε substrate, aldehyde dehydrogenase 2, and reduced cardiac injury in ex vivo and in vivo models of myocardial infarction.
Our results suggest that HSP90-mediated mitochondrial import of PKCε plays an important role in the protection of the myocardium from IR injury.
Protein kinase C epsilon; Mitochondria; Protein–protein interaction; Ischaemia reperfusion; Heat shock protein 90
Numerous conditions promote oxidative stress, leading to the build-up of reactive aldehydes that cause cell damage and contribute to cardiac diseases. Aldehyde dehydrogenases (ALDHs) are important enzymes that eliminate toxic aldehydes by catalysing their oxidation to non-reactive acids. The review will discuss evidence indicating a role for a specific ALDH enzyme, the mitochondrial ALDH2, in combating oxidative stress by reducing the cellular ‘aldehydic load’. Epidemiological studies in humans carrying an inactive ALDH2, genetic models in mice with altered ALDH2 levels, and small molecule activators of ALDH2 all highlight the role of ALDH2 in cardioprotection and suggest a promising new direction in cardiovascular research and the development of new treatments for cardiovascular diseases.
ALDH2; Mitochondria; Ischaemia; Nitroglycerin; Alda-1
Previous studies demonstrate impairment of endothelial-dependent vasodilation after ischemia/reperfusion (I/R). Though we have demonstrated that inhibition of δ protein kinase C (δPKC) at reperfusion reduces myocyte damage and improves cardiac function in a porcine acute myocardial infarction (AMI) model, impact of the selective δPKC inhibitor on epicardial coronary endothelial function remains unknown.
Either δPKC inhibitor (δV1-1, n=5) or saline (n=5) was infused into the left anterior descending artery at the last 1 minute of the 30-minute ischemia by balloon occlusion. In vivo responses to bradykinin (endothelium-dependent vasodilator) or nitroglycerin (endothelium-independent vasodilator) were analyzed at 24 h after I/R using intravascular ultrasound. Vascular responses were calculated as the ratio of vessel area at each time point (30, 60, 90 and 120 seconds after the infusion), divided by values at baseline (before the infusion).
In control pigs, endothelial-dependent vasodilation following bradykinin infusion in infarct-related epicardial coronary artery was impaired, whereas in δPKC inhibitor treated-pigs the endothelial-dependent vasodilation was preserved. Nitroglycerin infusion caused similar vasodilatory responses in the both groups.
This is the first demonstration that a δPKC inhibitor preserves vasodilator capacity in epicardial coronary arteries in an in vivo porcine AMI model. Because endothelial dysfunction correlates with worse outcome in patients with AMI, this preserved endothelial function in epicardial coronary arteries might result in a better clinical outcome.
ultrasonography; angioplasty; myocardial infarction; protein kinases; endothelium
Protein kinase C (PKC) is a family of kinases that are critical in many cellular events. These enzymes are activated by lipid-derived second messengers, are dependent on binding to negatively charged phospholipids and some members also require calcium to attain full activation. The interaction with lipids and calcium activators is mediated by binding to the regulatory domains C1 and C2. In addition, many protein-protein interactions between PKC and other proteins have been described. These include interactions with adaptor proteins, substrates and cytoskeletal elements. Regulation of the interactions between PKC, small molecules and other proteins is essential for signal transduction to occur. Finally, a number of auto-inhibitory intramolecular protein-protein interactions have also been identified in PKC. This chapter focuses on mapping the sites for many of these inter and intramolecular interactions and how this information may be used to generate selective inhibitors and activators of PKC signaling.
The cardioprotective effects of moderate alcohol consumption have been well documented in animal models and in humans. Protection afforded against ischemia and reperfusion injury (I/R) proceeds through an ischemic preconditioning-like mechanism involving the activation of epsilon protein kinase C (εPKC) and is dependent on the time and duration of ethanol treatment. However, the substrates of εPKC and the molecular mechanisms by which the enzyme protects the heart from oxidative damage induced by I/R are not fully described. Using an open-chest model of acute myocardial infarction in vivo, we find that intraperitoneal injection of ethanol (0.5 g/kg) 60 minutes prior to (but not 15 minutes prior to) a 30-minute transient ligation of the left anterior descending coronary artery reduced I/R-mediated injury by 57% (measured as a decrease of creatine phosphokinase release into the blood). Only under cardioprotective conditions, ethanol treatment resulted in the translocation of εPKC to cardiac mitochondria, where the enzyme bound aldehyde dehydrogenase-2 (ALDH2). ALDH2 is an intra-mitochondrial enzyme involved in the detoxification of toxic aldehydes such as 4-hydroxy-2-nonenal (4-HNE) and 4-HNE mediates oxidative damage, at least in part, by covalently modifying and inactivating proteins (by forming 4-HNE adducts). In hearts subjected to I/R after ethanol treatment, the levels of 4-HNE protein adducts were lower and JNK1/2 and ERK1/2 activities were diminished relative to the hearts from rats subjected to I/R in the absence of ethanol. Together, this work provides an insight into the mitochondrial-dependent basis of ethanol-induced and εPKC-mediated protection from cardiac ischemia, in vivo.
Protein-protein interactions sequester enzymes close to their substrates. Protein kinase C (PKC) is one example of a ubiquitous signaling molecule with effects that are dependent upon localization. Short peptides derived from interaction sites between each PKC isozyme and its receptor for activated C kinase act as highly specific inhibitors and have become available as selective drugs in basic research and animal models of human diseases, such as myocardial infarction and hyperglycemia. Whereas the earlier inhibitory peptides are highly specific, we believe that peptides targeting additional interactions between PKC and selective substrates will generate even more selective tools that regulate different functions of individual isozymes. Here, we discuss the methodologies and applications for identifying selective regulators of PKC.
Although heavy alcohol consumption has deleterious effects on heart health, moderate drinking is thought to have cardioprotective effects, reducing the risk of coronary artery disease and improving prognosis after a myocardial infarction. It still is unclear, however, if this effect can be achieved with all types of alcoholic beverages and results from the alcohol itself, from other compounds found in alcoholic beverages, or both. For example, the polyphenolic compound resveratrol, which is found particularly in red wine, can reduce the risk of atherosclerosis; however, it is not clear if the resveratrol levels present in wine are sufficient to achieve this result. Alcohol itself contributes to cardioprotection through several mechanisms. For example, it can improve the cholesterol profile, increasing the levels of “good” cholesterol and reducing the levels of “bad” cholesterol. Alcohol also may contribute to blood clot dissolution and may induce a phenomenon called pre-conditioning, whereby exposure to moderate alcohol levels (like short bouts of blood supply disruption [i.e., ischemia]), and result in reduced damage to the heart tissue after subsequent prolonged ischemia. Finally, the enzyme aldehyde dehydrogenase (ALDH) 2, which is involved in alcohol metabolism, also may contribute to alcohol-related cardioprotection by metabolizing other harmful aldehydes that could damage the heart muscle.
Alcohol consumption; light drinking; moderate drinking; effects and consequences of alcohol and other drug use; beneficial moderate alcohol consumption; risk and protective factors; risk-benefit; cardiovascular system; cardioprotection; wine; red wine; French Paradox; cholesterol; resveratrol; aldehyde dehydrogenase 2
Heart failure (HF) is a chronic syndrome in which pathological cardiac remodeling is an integral part of the disease and mast cell (MC) degranulation-derived mediators have been suggested to play a role in its progression. Protein kinase C (PKC) signaling is a key event in the signal transduction pathway of MC degranulation. We recently found that inhibition of εPKC slows down the progression of hypertension-induced HF in salt-sensitive Dahl rats fed a high-salt diet. We therefore determined whether εPKC inhibition affects MC degranulation in this model. Six week-old male Dahl rats were fed with a high-salt diet to induce systemic hypertension, which resulted in concentric left ventricular hypertrophy at the age of 11 weeks, followed by myocardial dilatation and HF at the age of 17 weeks. We administered εV1-2 an εPKC-selective inhibitor peptide (3 mg/Kg/day), δV1-1, a δPKC-selective inhibitor peptide (3 mg/Kg/day), TAT (negative control; at equimolar concentration; 1.6 mg/Kg/day) or olmesartan (angiotensin receptor blocker [ARB] as a positive control; 3mg/Kg/day) between 11 weeks and 17 weeks. Treatment with εV1-2 attenuated cardiac MC degranulation without affecting MC density, myocardial fibrosis, microvessel patency, vascular thickening and cardiac inflammation in comparison to TAT- or δV1-1-treatment. Treatment with ARB also attenuated MC degranulation and cardiac remodeling, but to a lesser extent when compared to εV1-2. Finally, εV1-2 treatment inhibited MC degranulation in isolated peritoneal MCs. Together, our data suggest that εPKC inhibition attenuates pathological remodeling in hypertension-induced HF, at least in part, by preventing cardiac MC degranulation.
Mast cell degranulation; protein kinase C; PKC-selective inhibitor peptide; cardiac remodeling; heart failure
The response of the myocardium to an ischaemic insult is regulated by two highly homologous protein kinase C (PKC) isozymes, δ and εPKC. Here, we determined the spatial and temporal relationships between these two isozymes in the context of ischaemia/reperfusion (I/R) and ischaemic preconditioning (IPC) to better understand their roles in cardioprotection.
Methods and results
Using an ex vivo rat model of myocardial infarction, we found that short bouts of ischaemia and reperfusion prior to the prolonged ischaemic event (IPC) diminished δPKC translocation by 3.8-fold and increased εPKC accumulation at mitochondria by 16-fold during reperfusion. In addition, total cellular levels of δPKC decreased by 60 ± 2.7% in response to IPC, whereas the levels of εPKC did not significantly change. Prolonged ischaemia induced a 48 ± 11% decline in the ATP-dependent proteasomal activity and increased the accumulation of misfolded proteins during reperfusion by 192 ± 32%; both of these events were completely prevented by IPC. Pharmacological inhibition of the proteasome or selective inhibition of εPKC during IPC restored δPKC levels at the mitochondria while decreasing εPKC levels, resulting in a loss of IPC-induced protection from I/R. Importantly, increased myocardial injury was the result, in part, of restoring a δPKC-mediated I/R pro-apoptotic phenotype by decreasing pro-survival signalling and increasing cytochrome c release into the cytosol.
Taken together, our findings indicate that IPC prevents I/R injury at reperfusion by protecting ATP-dependent 26S proteasomal function. This decreases the accumulation of the pro-apoptotic kinase, δPKC, at cardiac mitochondria, resulting in the accumulation of the pro-survival kinase, εPKC.
Cardioprotection; Ischaemia/reperfusion; Apoptosis; Proteasome; PKC; Ischaemic preconditioning
Two pathways that have been shown to mediate cerebral ischemic damage are the MEK/ERK cascade and the pro-apoptotic δPKC pathway. We investigated the relationship between these pathways in a rat model of focal ischemia by observing and modifying the activation state of each pathway. The ERK1/2 inhibitor, U0126, injected at ischemia onset, attenuated the increase in phosphorylated ERK1/2 (P-ERK1/2) after reperfusion. The δPKC inhibitor, δV1-1, delivered at reperfusion, did not significantly change P-ERK1/2 levels. In contrast, the δPKC activator, ψδRACK, injected at reperfusion, reduced ERK1/2 phosphorylation measured 4 h after reperfusion. Additionally, U0126 pretreatment at ischemia onset reduced infarct size compared with vehicle, but U0126 injected at the onset of reperfusion had no protection. Finally, combination of U0126 injection at ischemia onset plus δV1-1 injection at reperfusion further reduced infarct size, while combination of U0126 delivered at ischemia onset with ψδRACK injected at reperfusion increased infarct size compared with U0126 alone. In conclusion, we find that inhibiting both the MEK/ERK and the δPKC pathways offers greater protection than either alone, indicating they likely act independently.
Cerebral ischemia; MEK/ERK cascade; δPKC; ERK1/2
There is substantial interest in the development of drugs that limit the extent of ischemia-induced cardiac damage caused by myocardial infarction or by certain surgical procedures. Here an unbiased proteomic search identified mitochondrial aldehyde dehydrogenase 2 (ALDH2) as an enzyme whose activation correlates with reduced ischemic heart damage in rodent models. A high-throughput screen yielded a small-molecule activator of ALDH2 (Alda-1) that, when administered to rats prior to an ischemic event, reduced infarct size by 60%, most likely through its inhibitory effect on the formation of cytotoxic aldehydes. In vitro, Alda-1 was a particularly effective activator of ALDH2*2, an inactive mutant form of the enzyme that is found in 40% of East Asian populations. Thus, pharmacologic enhancement of ALDH2 activity may be useful for patients with wildtype or mutant ALDH2 subjected to cardiac ischemia, such as during coronary bypass surgery. (140/140 words)
In response to mild ischemic stress, the brain elicits endogenous survival mechanisms to protect cells against a subsequent lethal ischemic stress, referred to as ischemic tolerance. The molecular signals that mediate this protection are thought to involve the expression and activation of multiple kinases, including protein kinase C (PKC). Here we demonstrate that εPKC mediates cerebral ischemic tolerance in vivo. Systemic delivery of ψεRACK, an εPKC-selective peptide activator, confers neuroprotection against a subsequent cerebral ischemic event when delivered immediately prior to stroke. In addition, activation of εPKC by ψεRACK treatment decreases vascular tone in vivo, as demonstrated by a reduction in microvascular cerebral blood flow. Here we demonstrate the role of acute and transient εPKC in early cerebral tolerance in vivo and suggest that extra-parenchymal mechanisms, such as vasoconstriction, may contribute to the conferred protection.
Ischemia; preconditioning; protein kinase C; cerebral blood flow
Angiogenesis is critical in the progression of prostate cancer. However, the interplay between the proliferation kinetics of tumor endothelial cells (angiogenesis) and tumor cells has not been investigated. Also, protein kinase C (PKC) regulates various aspects of tumor cell growth but its role in prostate cancer has not been investigated in detail. Here, we found that the proliferation rates of endothelial and tumor cells oscillate asynchronously during the growth of human prostate cancer xenografts. Furthermore, our analyses suggest that PKCβII was activated during increased angiogenesis and that PKCβII plays a key role in the proliferation of endothelial cells and tumor cells in human prostate cancer; treatment with a PKCβII-selective inhibitor, βIIV5-3, reduced angiogenesis and tumor cell proliferation. We also find a unique effect of PKCβII inhibition on normalizing pericentrin (a protein regulating cytokinesis), especially in endothelial cells as well as in tumor cells. PKCβII inhibition reduced the level and mislocalization of pericentrin and normalized microtubule organization in the tumor endothelial cells. Although pericentrin has been known to be upregulated in epithelial cells of prostate cancers, its level in tumor endothelium has not been studied in detail. We found that pericentrin is upregulated in human tumor endothelium compared with endothelium adjacent to normal glands in tissues from prostate cancer patients. Our results suggest that a PKCβII inhibitor such as βIIV5-3 may be used to reduce prostate cancer growth by targeting both angiogenesis and tumor cell growth.
Previously we found that neural responses to ethanol and the dopamine D2 receptor (D2) agonist NPA involve both epsilon protein kinase C (εPKC) and cAMP-dependent protein kinase A (PKA). However, little is known about the mechanism underlying ethanol- and D2-mediated activation of εPKC and the relationship to PKA activation. In the present study, we used a new εPKC antibody, 14E6, that selectively recognizes active εPKC when not bound to its anchoring protein εRACK (receptor for activated C-kinase), and PKC isozyme-selective inhibitors and activators, to measure PKC translocation and catalytic activity. We show here that ethanol and NPA activated εPKC and also induced translocation of both εPKC and its anchoring protein, εRACK to a new cytosolic site. The selective εPKC agonist, pseudo-εRACK, activated εPKC but did not cause translocation of the εPKC/εRACK complex to the cytosol. These data suggest a step-wise activation and translocation of εPKC following NPA or ethanol treatment where εPKC first translocates and binds to its RACK and subsequently the εPKC/εRACK complex translocates to a new subcellular site. Direct activation of PKA by Sp-cAMPS, PGE1 or the adenosine A2A receptor is sufficient to cause εPKC translocation to the cytosolic compartment in a process that is dependent on PLC activation and requires PKA activity. These data demonstrate a novel cross-talk mechanism between εPKC and PKA signaling systems. PKA and PKC signaling have been implicated in alcohol rewarding properties in the mesolimbic dopamine system. Cross-talk between PKA and PKC may underlie some of the behaviors associated with alcoholism.
Heart failure (HF) afflicts about 5 million people and causes 300 000 deaths a year in the United States alone. An integral part of the pathogenesis of HF is cardiac remodelling, and the signalling events that regulate it are a subject of intense research. Cardiac remodelling is the sum of responses of the heart to causes of HF, such as ischaemia, myocardial infarction, volume and pressure overload, infection, inflammation, and mechanical injury. These responses, including cardiomyocyte hypertrophy, myocardial fibrosis, and inflammation, involve numerous cellular and structural changes and ultimately result in a progressive decline in cardiac performance. Pharmacological and genetic manipulation of cultured heart cells and animal models of HF and the analysis of cardiac samples from patients with HF are all used to identify the molecular and cellular mechanisms leading to the disease. Protein kinase C (PKC) isozymes, a family of serine–threonine protein kinase enzymes, were found to regulate a number of cardiac responses, including those associated with HF. In this review, we describe the PKC isozymes that play critical roles in specific aspects of cardiac remodelling and dysfunction in HF.
Protein kinase C; Heart failure; Cardiac remodeling; Hypertrophy; Fibrosis and inflammation