Protein Kinase A Is a Master Regulator of Physiological and Pathological Cardiac Hypertrophy.

BACKGROUND: The sympathoadrenergic system and its major effector PKA (protein kinase A) are activated to maintain cardiac output coping with physiological or pathological stressors. If and how PKA plays a role in physiological cardiac hypertrophy (PhCH) and pathological CH (PaCH) are not clear. METHODS: Transgenic mouse models expressing the PKA inhibition domain (PKAi) of PKA inhibition peptide alpha (PKIalpha)-green fluorescence protein (GFP) fusion protein (PKAi-GFP) in a cardiac-specific and inducible manner (cPKAi) were used to determine the roles of PKA in physiological CH during postnatal growth or induced by swimming, and in PaCH induced by transaortic constriction (TAC) or augmented Ca2+ influx. Kinase profiling was used to determine cPKAi specificity. Echocardiography was used to determine cardiac morphology and function. Western blotting and immunostaining were used to measure protein abundance and phosphorylation. Protein synthesis was assessed by puromycin incorporation and protein degradation by measuring protein ubiquitination and proteasome activity. Neonatal rat cardiomyocytes (NRCMs) infected with AdGFP (GFP adenovirus) or AdPKAi-GFP (PKAi-GFP adenovirus) were used to determine the effects and mechanisms of cPKAi on myocyte hypertrophy. rAAV9.PKAi-GFP was used to treat TAC mice. RESULTS: (1) cPKAi delayed postnatal cardiac growth and blunted exercise-induced PhCH; (2) PKA was activated in hearts after TAC due to activated sympathoadrenergic system, the loss of endogenous PKIα (PKA inhibition peptide α), and the stimulation by noncanonical PKA activators; (3) cPKAi ameliorated PaCH induced by TAC and increased Ca2+ influxes and blunted neonatal rat cardiomyocyte hypertrophy by isoproterenol and phenylephrine; (4) cPKAi prevented TAC-induced protein synthesis by inhibiting mTOR (mammalian target of rapamycin) signaling through reducing Akt (protein kinase B) activity, but enhancing inhibitory GSK-3α (glycogen synthase kinase-3α) and GSK-3ß signals; (5) cPKAi reduced protein degradation by the ubiquitin-proteasome system via decreasing RPN6 phosphorylation; (6) cPKAi increased the expression of antihypertrophic atrial natriuretic peptide (ANP); (7) cPKAi ameliorated established PaCH and improved animal survival. CONCLUSIONS: Cardiomyocyte PKA is a master regulator of PhCH and PaCH through regulating protein synthesis and degradation. cPKAi can be a novel approach to treat PaCH.

Cardiomyocyte PKA Ablation Enhances Basal Contractility While Eliminates Cardiac ß-Adrenergic Response Without Adverse Effects on the Heart.

RATIONALE: PKA (Protein Kinase A) is a major mediator of ß-AR (ß-adrenergic) regulation of cardiac function, but other mediators have also been suggested. Reduced PKA basal activity and activation are linked to cardiac diseases. However, how complete loss of PKA activity impacts on cardiac physiology and if it causes cardiac dysfunction have never been determined. OBJECTIVES: We set to determine how the heart adapts to the loss of cardiomyocyte PKA activity and if it elicits cardiac abnormalities. METHODS AND RESULTS: (1) Cardiac PKA activity was almost completely inhibited by expressing a PKA inhibitor peptide in cardiomyocytes (cPKAi) in mice; (2) cPKAi reduced basal phosphorylation of 2 myofilament proteins (TnI [troponin I] and cardiac myosin binding protein C), and one longitudinal SR (sarcoplasmic reticulum) protein (PLB [phospholamban]) but not of the sarcolemmal proteins (Cav1.2 α1c and PLM [phospholemman]), dyadic protein RyR2, and nuclear protein CREB (cAMP response element binding protein) at their PKA phosphorylation sites; (3) cPKAi increased the expression of CaMKII (Ca2+/calmodulin-dependent kinase II), the Cav1.2 ß subunits and current, but decreased CaMKII phosphorylation and CaMKII-mediated phosphorylation of PLB and RyR2; (4) These changes resulted in significantly enhanced myofilament Ca2+ sensitivity, prolonged contraction, slowed relaxation but increased myocyte Ca2+ transient and contraction amplitudes; (5) Isoproterenol-induced PKA and CaMKII activation and their phosphorylation of proteins were prevented by cPKAi; (6) cPKAi abolished the increases of heart rate, and cardiac and myocyte contractility by a ß-AR agonist (isoproterenol), showing an important role of PKA and a minimal role of PKA-independent ß-AR signaling in acute cardiac regulation; (7) cPKAi mice have partial exercise capability probably by enhancing vascular constriction and ventricular filling during ß-AR stimulation; and (8) cPKAi mice did not show any cardiac functional or structural abnormalities during the 1-year study period. CONCLUSIONS: PKA activity suppression induces a unique Ca2+ handling phenotype, eliminates ß-AR regulation of heart rates and cardiac contractility but does not cause cardiac abnormalities.

GNAI1 and GNAI3 Reduce Colitis-Associated Tumorigenesis in Mice by Blocking IL6 Signaling and Down-regulating Expression of GNAI2.

BACKGROUND & AIMS: Interleukin 6 (IL6) and tumor necrosis factor contribute to the development of colitis-associated cancer (CAC). We investigated these signaling pathways and the involvement of G protein subunit alpha i1 (GNAI1), GNAI2, and GNAI3 in the development of CAC in mice and humans. METHODS: B6;129 wild-type (control) or mice with disruption of Gnai1, Gnai2, and/or Gnai3 or conditional disruption of Gnai2 in CD11c+ or epithelial cells were given dextran sulfate sodium (DSS) to induce colitis followed by azoxymethane (AOM) to induce carcinogenesis; some mice were given an antibody against IL6. Feces were collected from mice, and the compositions of microbiomes were analyzed by polymerase chain reactions. Dendritic cells (DCs) and myeloid-derived suppressor cells (MDSCs) isolated from spleen and colon tissues were analyzed by flow cytometry. We performed immunoprecipitation and immunoblot analyses of colon tumor tissues, MDSCs, and mouse embryonic fibroblasts to study the expression levels of GNAI1, GNAI2, and GNAI3 and the interactions of GNAI1 and GNAI3 with proteins in the IL6 signaling pathway. We analyzed the expression of Gnai2 messenger RNA by CD11c+ cells in the colonic lamina propria by PrimeFlow, expression of IL6 in DCs by flow cytometry, and secretion of cytokines in sera and colon tissues by enzyme-linked immunosorbent assay. We obtained colon tumor and matched nontumor tissues from 83 patients with colorectal cancer having surgery in China and 35 patients with CAC in the United States. Mouse and human colon tissues were analyzed by histology, immunoblot, immunohistochemistry, and/or RNA-sequencing analyses. RESULTS: GNAI1 and GNAI3 (GNAI1;3) double-knockout (DKO) mice developed more severe colitis after administration of DSS and significantly more colonic tumors than control mice after administration of AOM plus DSS. Development of increased tumors in DKO mice was not associated with changes in fecal microbiomes but was associated with activation of nuclear factor (NF) κB and signal transducer and activator of transcription (STAT) 3; increased levels of GNAI2, nitric oxide synthase 2, and IL6; increased numbers of CD4+ DCs and MDSCs; and decreased numbers of CD8+ DCs. IL6 was mainly produced by CD4+/CD11b+, but not CD8+, DCs in DKO mice. Injection of DKO mice with a blocking antibody against IL6 reduced the expansion of MDSCs and the number of tumors that developed after CAC induction. Incubation of MDSCs or mouse embryonic fibroblasts with IL6 induced activation of either NF-κB by a JAK2-TRAF6-TAK1-CHUK/IKKB signaling pathway or STAT3 by JAK2. This activation resulted in expression of GNAI2, IL6 signal transducer (IL6ST, also called GP130) and nitric oxide synthase 2, and expansion of MDSCs; the expression levels of these proteins and expansion of MDSCs were further increased by the absence of GNAI1;3 in cells and mice. Conditional disruption of Gnai2 in CD11c+ cells of DKO mice prevented activation of NF-κB and STAT3 and changes in numbers of DCs and MDSCs. Colon tumor tissues from patients with CAC had reduced levels of GNAI1 and GNAI3 and increased levels of GNAI2 compared with normal tissues. Further analysis of a public human colorectal tumor DNA microarray database (GSE39582) showed that low Gani1 and Gnai3 messenger RNA expression and high Gnai2 messenger RNA expression were significantly associated with decreased relapse-free survival. CONCLUSIONS: GNAI1;3 suppresses DSS-plus-AOM-induced colon tumor development in mice, whereas expression of GNAI2 in CD11c+ cells and IL6 in CD4+/CD11b+ DCs appears to promote these effects. Strategies to induce GNAI1;3, or block GNAI2 and IL6, might be developed for the prevention or therapy of CAC in patients.

HIF1 mediates a switch in pyruvate kinase isoforms after myocardial infarction.

Alternative splicing of RNA is an underexplored area of transcriptional response. We expect that early changes in alternatively spliced genes may be important for responses to cardiac injury. Hypoxia inducible factor 1 (HIF1) is a key transcription factor that rapidly responds to loss of oxygen through alteration of metabolism and angiogenesis. The goal of this study was to investigate the transcriptional response after myocardial infarction (MI) and to identify novel, hypoxia-driven changes, including alternative splicing. After ligation of the left anterior descending artery in mice, we observed an abrupt loss of cardiac contractility and upregulation of hypoxic signaling. We then performed RNA sequencing on ischemic heart tissue 1 and 3 days after infarct to assess early transcriptional changes and identified 89 transcripts with altered splicing. Of particular interest was the switch in Pkm isoform expression (pyruvate kinase, muscle). The usually predominant Pkm1 isoform was less abundant in ischemic hearts, while Pkm2 and associated splicing factors (hnRNPA1, hnRNPA2B1, Ptbp1) rapidly increased. Despite increased Pkm2 expression, total pyruvate kinase activity remained reduced in ischemic myocardial tissue. We also demonstrated HIF1 binding to PKM by chromatin immunoprecipitation, indicating a direct role for HIF1 in mediating this isoform switch. Our study provides a new, detailed characterization of the early transcriptome after MI. From this analysis, we identified an HIF1-mediated alternative splicing event in the PKM gene. Pkm1 and Pkm2 play distinct roles in glycolytic metabolism and the upregulation of Pkm2 is likely to have important consequences for ATP synthesis in infarcted cardiac muscle.

Increasing T-type calcium channel activity by ß-adrenergic stimulation contributes to ß-adrenergic regulation of heart rates.

KEY POINTS: Cav3.1 T-type Ca2+ channel current (ICa-T ) contributes to heart rate genesis but is not known to contribute to heart rate regulation by the sympathetic/ß-adrenergic system (SAS). We show that the loss of Cav3.1 makes the beating rates of the heart in vivo and perfused hearts ex vivo, as well as sinoatrial node cells, less sensitive to ß-adrenergic stimulation; it also renders less conduction acceleration through the atrioventricular node by ß-adrenergic stimulation. Increasing Cav3.1 in cardiomyocytes has the opposite effects. ICa-T in sinoatrial nodal cells can be upregulated by ß-adrenergic stimulation. The results of the present study add a new contribution to heart rate regulation by the SAS system and provide potential new mechanisms for the dysregulation of heart rate and conduction by the SAS in the heart. T-type Ca2+ channel can be a target for heart disease treatments that aim to slow down the heart rate ABSTRACT: Cav3.1 (α1G ) T-type Ca2+ channel (TTCC) is expressed in mouse sinoatrial node cells (SANCs) and atrioventricular (AV) nodal cells and contributes to heart rate (HR) genesis and AV conduction. However, its role in HR regulation and AV conduction acceleration by the ß-adrenergic system (SAS) is unclear. In the present study, L- (ICa-L ) and T-type (ICa-T ) Ca2+ currents were recorded in SANCs from Cav3.1 transgenic (TG) and knockout (KO), and control mice. ICa-T was absent in KO SANCs but enhanced in TG SANCs. In anaesthetized animals, different doses of isoproterenol (ISO) were infused via the jugular vein and the HR was recorded. The EC50 of the HR response to ISO was lower in TG mice but higher in KO mice, and the maximal percentage of HR increase by ISO was greater in TG mice but less in KO mice. In Langendorff-perfused hearts, ISO increased HR and shortened PR intervals to a greater extent in TG but to a less extent in KO hearts. KO SANCs had significantly slower spontaneous beating rates than control SANCs before and after ISO; TG SANCs had similar basal beating rates as control SANCs probably as a result of decreased ICa-L but a greater response to ISO than control SANCs. ICa-T in SANCs was significantly increased by ISO. ICa-T upregulation by ß-adrenergic stimulation contributes to HR and conduction regulation by the SAS. TTCC can be a target for slowing the HR.

The TGF-ß pathway mediates doxorubicin effects on cardiac endothelial cells.

Elevated ALK4/5 ligands including TGF-ß and activins have been linked to cardiovascular remodeling and heart failure. Doxorubicin (Dox) is commonly used as a model of cardiomyopathy, a condition that often precedes cardiovascular remodeling and heart failure. In 7-8-week-old C57Bl/6 male mice treated with Dox we found decreased capillary density, increased levels of ALK4/5 ligand and Smad2/3 transcripts, and increased expression of Smad2/3 transcriptional targets. Human cardiac microvascular endothelial cells (HCMVEC) treated with Dox also showed increased levels of ALK4/5 ligands, Smad2/3 transcriptional targets, a decrease in proliferation and suppression of vascular network formation in a HCMVEC and human cardiac fibroblasts co-culture assay. Our hypothesis is that the deleterious effects of Dox on endothelial cells are mediated in part by the activation of the TGF-ß pathway. We used the inhibitor of ALK4/5 kinases SB431542 (SB) in concert with Dox to ascertain the role of TGF-ß pathway activation in doxorubicin induced endothelial cell defects. SB prevented the suppression of HCMVEC proliferation in the presence of TGF-ß2 and activin A, and alleviated the inhibition of HCMVEC proliferation by Dox. SB also prevented the suppression of vascular network formation in co-cultures of HCMVEC and human cardiac fibroblasts treated with Dox. Our results show that the inhibition of the TGF-ß pathway alleviates the detrimental effects of Dox on endothelial cells in vitro.

Persistent increases in Ca(2+) influx through Cav1.2 shortens action potential and causes Ca(2+) overload-induced afterdepolarizations and arrhythmias.

Persistent elevation of Ca(2+) influx due to prolongation of the action potential (AP), chronic activation of the ß-adrenergic system and molecular remodeling occurs in stressed and diseased hearts. Increases in Ca(2+) influx are usually linked to prolonged myocyte action potentials and arrhythmias. However, the contribution of chronic enhancement of Cav1.2 activity on cardiac electrical remodeling and arrhythmogenicity has not been completely defined and is the subject of this study. Chronically increased Cav1.2 activity was produced with a cardiac specific, inducible double transgenic (DTG) mouse system overexpressing the ß2a subunit of Cav (Cavß2a). DTG myocytes had increased L-type Ca(2+) current (ICa-L), myocyte shortening, and Ca(2+) transients. DTG mice had enhanced cardiac performance, but died suddenly and prematurely. Telemetric electrocardiograms revealed shortened QT intervals in DTG mice. The action potential duration (APD) was shortened in DTG myocytes due to significant increases of potassium currents and channel abundance. However, shortened AP in DTG myocytes did not fully limit excess Ca(2+) influx and increased the peak and tail ICa-L. Enhanced ICa promoted sarcoplasmic reticulum (SR) Ca(2+) overload, diastolic Ca(2+) sparks and waves, and increased NCX activity, causing increased occurrence of early and delayed afterdepolarizations (EADs and DADs) that may contribute to premature ventricular beats and ventricular tachycardia. AV blocks that could be related to fibrosis of the AV node were also observed. Our study suggests that increasing ICa-L does not necessarily result in AP prolongation but causes SR Ca(2+) overload and fibrosis of AV node and myocardium to induce cellular arrhythmogenicity, arrhythmias, and conduction abnormalities.

Daytime somnolence as an early sign of cognitive decline in a community-based study of older people.

OBJECTIVE: This study aimed to examine the association between self-reported sleep problems and cognitive decline in community-dwelling older people. We hypothesized that daytime somnolence predicts subsequent cognitive decline. METHODS: This is a longitudinal study in a 3.2-year follow-up, with 18-month intervals. The setting is the Washington Heights-Inwood Community Aging Project. There were 1098 participants, who were over 65 years old and recruited from the community. Sleep problems were estimated using five sleep categories derived from the RAND Medical Outcome Study Sleep Scale: sleep disturbance, snoring, awaken short of breath/with a headache, sleep adequacy, and daytime somnolence. Four distinct cognitive composite scores were calculated: memory, language, speed of processing, and executive functioning. We used generalized estimating equations analyses with cognitive scores as the outcome, and time, sleep categories and their interactions as the main predictors. Models were initially unadjusted and then adjusted for age, gender, education, ethnicity, depression, and apolipoprotein E-ε4 genotype. RESULTS: Increased daytime somnolence (including feeling drowsy/sleepy, having trouble staying awake, and taking naps during the day) was linked to slower speed of processing both cross-sectionally (B = -0.143, p = 0.047) and longitudinally (B = -0.003, p = 0.027). After excluding the demented participants at baseline, the results remained significant (B = -0.003, p = 0.021). CONCLUSIONS: Our findings suggest that daytime somnolence may be an early sign of cognitive decline in the older population.

Cardiotoxic and cardioprotective features of chronic ß-adrenergic signaling.

RATIONALE: In the failing heart, persistent ß-adrenergic receptor activation is thought to induce myocyte death by protein kinase A (PKA)-dependent and PKA-independent activation of calcium/calmodulin-dependent kinase II. ß-adrenergic signaling pathways also are capable of activating cardioprotective mechanisms. OBJECTIVE: This study used a novel PKA inhibitor peptide to inhibit PKA activity to test the hypothesis that ß-adrenergic receptor signaling causes cell death through PKA-dependent pathways and cardioprotection through PKA-independent pathways. METHODS AND RESULTS: In PKA inhibitor peptide transgenic mice, chronic isoproterenol failed to induce cardiac hypertrophy, fibrosis, and myocyte apoptosis, and decreased cardiac function. In cultured adult feline ventricular myocytes, PKA inhibition protected myocytes from death induced by ß1-adrenergic receptor agonists by preventing cytosolic and sarcoplasmic reticulum Ca(2+) overload and calcium/calmodulin-dependent kinase II activation. PKA inhibition revealed a cardioprotective role of ß-adrenergic signaling via cAMP/exchange protein directly activated by cAMP/Rap1/Rac/extracellular signal-regulated kinase pathway. Selective PKA inhibition causes protection in the heart after myocardial infarction that was superior to ß-blocker therapy. CONCLUSIONS: These results suggest that selective block of PKA could be a novel heart failure therapy.

Cardiac G-protein-coupled receptor kinase 2 ablation induces a novel Ca2+ handling phenotype resistant to adverse alterations and remodeling after myocardial infarction.

BACKGROUND: G-protein-coupled receptor kinase 2 (GRK2) is a primary regulator of ß-adrenergic signaling in the heart. G-protein-coupled receptor kinase 2 ablation impedes heart failure development, but elucidation of the cellular mechanisms has not been achieved, and such elucidation is the aim of this study. METHODS AND RESULTS: Myocyte contractility, Ca(2+) handling and excitation-contraction coupling were studied in isolated cardiomyocytes from wild-type and GRK2 knockout (GRK2KO) mice without (sham) or with myocardial infarction (MI). In cardiac myocytes isolated from unstressed wild-type and GRK2KO hearts, myocyte contractions and Ca(2+) transients were similar, but GRK2KO myocytes had lower sarcoplasmic reticulum (SR) Ca(2+) content because of increased sodium-Ca(2+) exchanger activity and inhibited SR Ca(2+) ATPase by local protein kinase A-mediated activation of phosphodiesterase 4 resulting in hypophosphorylated phospholamban. This Ca(2+) handling phenotype is explained by a higher fractional SR Ca(2+) release induced by increased L-type Ca(2+) channel currents. After ß-adrenergic stimulation, GRK2KO myocytes revealed significant increases in contractility and Ca(2+) transients, which were not mediated through cardiac L-type Ca(2+) channels but through an increased SR Ca(2+). Interestingly, post-MI GRK2KO mice showed better cardiac function than post-MI control mice, which is explained by an improved Ca(2+) handling phenotype. The SR Ca(2+) content was better maintained in post-MI GRK2KO myocytes than in post-MI control myocytes because of better-maintained L-type Ca(2+) channel current density and no increase in sodium-Ca(2+) exchanger in GRK2KO myocytes. An L-type Ca(2+) channel blocker, verapamil, reversed some beneficial effects of GRK2KO. CONCLUSIONS: These data argue for novel differential regulation of L-type Ca(2+) channel currents and SR load by GRK2. G-protein-coupled receptor kinase 2 ablation represents a novel beneficial Ca(2+) handling phenotype resisting adverse remodeling after MI.

Increasing cardiac contractility after myocardial infarction exacerbates cardiac injury and pump dysfunction.

RATIONALE: Myocardial infarction (MI) leads to heart failure (HF) and premature death. The respective roles of myocyte death and depressed myocyte contractility in the induction of HF after MI have not been clearly defined and are the focus of this study. OBJECTIVES: We developed a mouse model in which we could prevent depressed myocyte contractility after MI and used it to test the idea that preventing depression of myocyte Ca(2+)-handling defects could avert post-MI cardiac pump dysfunction. METHODS AND RESULTS: MI was produced in mice with inducible, cardiac-specific expression of the ß2a subunit of the L-type Ca(2+) channel. Myocyte and cardiac function were compared in control and ß2a animals before and after MI. ß2a myocytes had increased Ca(2+) current; sarcoplasmic reticulum Ca(2+) load, contraction and Ca(2+) transients (versus controls), and ß2a hearts had increased performance before MI. After MI, cardiac function decreased. However, ventricular dilation, myocyte hypertrophy and death, and depressed cardiac pump function were greater in ß2a versus control hearts after MI. ß2a animals also had poorer survival after MI. Myocytes isolated from ß2a hearts after MI did not develop depressed Ca(2+) handling, and Ca(2+) current, contractions, and Ca(2+) transients were still above control levels (before MI). CONCLUSIONS: Maintaining myocyte contractility after MI, by increasing Ca(2+) influx, depresses rather than improves cardiac pump function after MI by reducing myocyte number.

The activin receptor ligand trap ActRIIB:ALK4-Fc ameliorates cardiomyopathy induced by neuromuscular disease and diabetes.

Cardiomyopathies are ascribed to a variety of etiologies, present with diverse clinical phenotypes, and lack disease-modifying treatments. Mounting evidence implicates dysregulated activin receptor signaling in heart disease and highlights inhibition of this pathway as a potential therapeutic target. Here, we explored the effects of activin ligand inhibition using ActRIIB:ALK4-Fc, a heterodimeric receptor fusion protein, in two mechanistically distinct murine models of cardiomyopathy. Treatment with ActRIIB:ALK4-Fc significantly improved systolic or diastolic function in cardiomyopathy induced by neuromuscular disease or diabetes mellitus. Moreover, ActRIIB:ALK4-Fc corrected Ca2+ handling protein expression in diseased heart tissues, suggesting that activin signaling inhibition could alleviate cardiomyopathies in part by rebalancing aberrant intracellular Ca2+ homeostasis-a common underlying pathomechanism in diverse heart diseases.

Calcium influx through Cav1.2 is a proximal signal for pathological cardiomyocyte hypertrophy.

Pathological cardiac hypertrophy (PCH) is associated with the development of arrhythmia and congestive heart failure. While calcium (Ca(2+)) is implicated in hypertrophic signaling pathways, the specific role of Ca(2+) influx through the L-type Ca(2+) channel (I(Ca-L)) has been controversial and is the topic of this study. To determine if and how sustained increases in I(Ca-L) induce PCH, transgenic mouse models with low (LE) and high (HE) expression levels of the ß2a subunit of Ca(2+) channels (ß2a) and in cultured adult feline (AF) and neonatal rat (NR) ventricular myocytes (VMs) infected with an adenovirus containing a ß2a-GFP were used. In vivo, ß2a LE and HE mice had increased heart weight to body weight ratio, posterior wall and interventricular septal thickness, tissue fibrosis, myocyte volume, and cross-sectional area and the expression of PCH markers in a time- and dose-dependent manner. PCH was associated with a hypercontractile phenotype including enhanced I(Ca-L), fractional shortening, peak Ca(2+) transient, at the myocyte level, greater ejection fraction, and fractional shortening at the organ level. In addition, LE mice had an exaggerated hypertrophic response to transverse aortic constriction. In vitro overexpression of ß2a in cultured AFVMs increased I(Ca-L), cell volume, protein synthesis, NFAT, and HDAC translocations and in NRVMs increased surface area. These effects were abolished by the blockade of I(Ca-L), intracellular Ca(2+), calcineurin, CaMKII, and SERCA. In conclusion, increasing I(Ca-L) is sufficient to induce PCH through the calcineurin/NFAT and CaMKII/HDAC pathways. Both cytosolic and SR/ER-nuclear envelop Ca(2+) pools were shown to be involved.

Importance of symptomatic cerebral infarcts on cognitive performance in patients with Alzheimer's disease.

The coexistence of cerebral infarcts and Alzheimer's disease (AD) is common, but the influence of symptomatic cerebral infarcts on cognition is uncertain in AD. We hypothesize that symptomatic cerebral infarcts may provide an additive cognitive factor contributing to dementia in the AD population. We studied 1,001 clinically probable or possible AD patients in the Alzheimer Disease Research Center (ADRC) database. Linear regression was used to evaluate for an association between symptomatic cerebral infarcts and memory, language, executive function, abstract reasoning, and visuospatial performance, separately. Models were adjusted for covariates including age, gender, education, ethnicity, hypertension, diabetes mellitus, heart disease, clinical dementia rating, the presence of silent cerebral infarcts, and multiplicity or location of infarcts. Clinical history of stroke was present in 107 patients, radiological infarcts in 308 patients, and 68 patients with both were considered to have symptomatic infarcts. Adjusting for all covariates, AD patients with symptomatic infarcts had more impairment of executive function (P < 0.05). The influence of cerebral infarcts is neither general nor diffuse, and the presence of clinical history may have a more important influence on executive performance in AD.

Enhanced basal contractility but reduced excitation-contraction coupling efficiency and beta-adrenergic reserve of hearts with increased Cav1.2 activity.

Cardiac remodeling during heart failure development induces a significant increase in the activity of the L-type Ca(2+) channel (Cav1.2). However, the effects of enhanced Cav1.2 activity on myocyte excitation-contraction (E-C) coupling, cardiac contractility, and its regulation by the beta-adrenergic system are not clear. To recapitulate the increased Cav1.2 activity, a double transgenic (DTG) mouse model overexpressing the Cavbeta2a subunit in a cardiac-specific and inducible manner was established. We studied cardiac (in vivo) and myocyte (in vitro) contractility at baseline and upon beta-adrenergic stimulation. E-C coupling efficiency was evaluated in isolated myocytes as well. The following results were found: 1) in DTG myocytes, L-type Ca(2+) current (I(Ca,L)) density, myocyte fractional shortening (FS), peak Ca(2+) transients, and sarcoplasmic reticulum (SR) Ca(2+) content (caffeine-induced Ca(2+) transient peak) were significantly increased (by 100.8%, 48.8%, 49.8%, and 46.8%, respectively); and 2) cardiac contractility evaluated with echocardiography [ejection fraction (EF) and (FS)] and invasive intra-left ventricular pressure (maximum dP/dt and -dP/dt) measurements were significantly greater in DTG mice than in control mice. However, 1) the cardiac contractility (EF, FS, dP/dt, and -dP/dt)-enhancing effect of the beta-adrenergic agonist isoproterenol (2 microg/g body wt ip) was significantly reduced in DTG mice, which could be attributed to the loss of beta-adrenergic stimulation on contraction, Ca(2+) transients, I(Ca,L), and SR Ca(2+) content in DTG myocytes; and 2) E-C couplng efficiency was significantly lower in DTG myocytes. In conclusion, increasing Cav1.2 activity by promoting its high-activity mode enhances cardiac contractility but decreases E-C coupling efficiency and the adrenergic reserve of the heart.

Alpha B-crystallin suppresses pressure overload cardiac hypertrophy.

AlphaB-crystallin (CryAB) is the most abundant small heat shock protein (HSP) constitutively expressed in cardiomyocytes. Gain- and loss-of-function studies demonstrated that CryAB can protect against myocardial ischemia/reperfusion injury. However, the role of CryAB or any HSPs in cardiac responses to mechanical overload is unknown. This study addresses this issue. Nontransgenic mice and mice with cardiomyocyte-restricted transgenic overexpression of CryAB or with germ-line ablation of the CryAB/HSPB2 genes were subjected to transverse aortic constriction or sham surgery. Two weeks later, cardiac responses were analyzed by fetal gene expression profiling, cardiac function analyses, and morphometry. Comparison among the 3 sham surgery groups reveals that CryAB overexpression is benign, whereas the knockout is detrimental to the heart as reflected by cardiac hypertrophy and malfunction at 10 weeks of age. Compared to nontransgenic mice, transgenic mouse hearts showed significantly reduced NFAT transactivation and attenuated cardiac hypertrophic responses to transverse aortic constriction but unchanged cardiac function, whereas NFAT transactivation was significantly increased in cardiac and skeletal muscle of the knockout mice at baseline, and they developed cardiac insufficiency at 2 weeks after transverse aortic constriction. CryAB overexpression in cultured neonatal rat cardiomyocytes significantly attenuated adrenergic stimulation-induced NFAT transactivation and hypertrophic growth. We conclude that CryAB suppresses cardiac hypertrophic responses likely through attenuating NFAT signaling and that CryAB and/or HSPB2 are essential for normal cardiac function.

Enhanced dissipation of xenobiotic agrochemicals harnessing soil microbiome in the tillage-reduced rice-dominated agroecosystem.

The ubiquitous contamination generating from the frequent input of agrochemicals is a major hurdle affecting the ecological sustainability of agroecosystems. Here, we investigated the dissipation of multiple pesticides in the subtropical rice-dominated landscapes under tillage intensity management, and unveiled the vital role of soil microbiome in promoting xenobiotic degradation. Three commonly used pesticides (including herbicide butachlor, insecticide clothianidin and fungicide tricyclazole) showed rapid dissipation dynamics in the field where the reduction of tillage intensity with straw incorporation was conducted. In response to pesticide exposure, soil microbial communities adapted quickly with slight changes in community composition and diversity. Meanwhile, the microbial xenobiotic degradation-related functions were stimulated, which possibly related to the increased organic carbon and nitrogen in soil. Importantly, these shifts and effects on microbial communities and functions gradually declined after a length of rice growing seasons, suggesting the flexibility of soil microbiome in tackling with long-term xenobiotic disturbance to maintain a diverse and vibrant soil biota. Overall, our study that displayed the enhanced agrochemical dissipation which benefited markedly from the interaction of tillage management and soil microbial functioning, provides important basis and insights for facilitating green, balanced and sustainable agriculture.

Plasma homocysteine and risk of mild cognitive impairment.

BACKGROUND AND OBJECTIVE: There are conflicting data relating homocysteine levels to the risk of Alzheimer's disease (AD). We sought to explore whether fasting plasma homocysteine is associated with the risk of mild cognitive impairment (MCI), an intermediate stage to dementia. METHODS: Fasting levels of plasma homocysteine were obtained from 678 elderly subjects chosen at random from a cohort of Medicare recipients. There were longitudinal data in 516 subjects without MCI or dementia at baseline who were followed for 2,705 person-years. The relation of plasma homocysteine with prevalent and incident all-cause MCI, amnestic MCI and non-amnestic MCI was assessed using logistic and Cox proportional hazards regression analyses. RESULTS: There were 162 cases of prevalent MCI and 132 cases of incident MCI in 5.2 years of follow-up. There was no association between plasma homocysteine and prevalence of MCI or amnestic or non-amnestic MCI in the cross-sectional analyses. There was no association between higher homocysteine levels and a lower risk of all-cause MCI. Consistent with the cross-sectional analyses, there was no specific association with the amnestic or non-amnestic subtype of MCI in crude or adjusted models. CONCLUSION: Plasma homocysteine levels measured at baseline were not related to MCI or its subtypes in an elderly multiethnic cohort.

Plasma lipid levels in the elderly are not associated with the risk of mild cognitive impairment.

BACKGROUND: There are conflicting data relating plasma lipids to the risk of Alzheimer's disease (AD). We explored the association of plasma lipids to mild cognitive impairment (MCI), a transitional stage between normal cognition and dementia, in a prospective community-based cohort study among randomly sampled Medicare recipients > or =65 years. Baseline data were collected from 1992 to 1994, follow-up data were collected at 18-month intervals. METHODS: Multivariate proportional hazards regression was used to relate plasma lipid levels to incident total MCI, amnestic MCI and nonamnestic MCI in 854 persons without MCI or dementia at baseline. RESULTS: There were 324 cases of incident MCI, 153 cases of amnestic MCI and 171 cases of nonamnestic MCI during 4,189 person-years of follow-up. Higher levels of total cholesterol and LDL were associated with a decreased risk of total MCI in models adjusting for age and sex. However, these associations were attenuated after adjusting for ethnicity, education, APOEepsilon4 and vascular risk factors. There was no association between lipids and the risk of amnestic or nonamnestic MCI, and there was no effect of lipid-lowering treatment on MCI risk. CONCLUSIONS: Plasma lipid levels or lipid-lowering treatment in the elderly are not associated with the risk of MCI.

Relation of plasma lipids to all-cause mortality in Caucasian, African-American and Hispanic elders.

OBJECTIVES: to investigate the relation of plasma lipids to all-cause mortality in a multi-ethnic cohort of non-demented elderly. SETTING: community-based sample of Medicare recipients, 65 years and older, residing in Northern Manhattan. PARTICIPANTS: about two thousand five hundred and fifty-six non-demented elderly, 65-103 years. Among participants, 66.1% were women, 27.6% were White/non-Hispanic, 31.2% were African-American and 41.2% were Hispanic. METHODS: a standardised assessment, including functional ability, medical history, physical and neurological examination and a neuropsychological battery was conducted. Vital status was ascertained through the National Death Index (NDI). We used survival analyses stratified by race and ethnicity to examine the relation of plasma lipids to subsequent all-cause mortality. RESULTS: hispanics had the best overall survival, followed by African-Americans and Whites. Whites and African-Americans in the lowest quartiles of total cholesterol, non-HDL cholesterol and low-density lipoprotein cholesterol (LDL cholesterol) were approximately twice as likely to die as those in the highest quartile (White HR: 2.2, for lowest total cholesterol quartile; HR: 2.3, for lowest non-HDL cholesterol quartile; and HR: 1.8, for lowest LDL cholesterol quartile. African-American HR: 1.9, for lowest total cholesterol, HR: 2.0, for lowest non-HDL cholesterol and HR: 1.9, for lowest LDL cholesterol). In contrast, plasma lipid levels were not related to mortality risk among Hispanics. CONCLUSIONS: hispanic ethnicity modifies the associations between lipid levels and all-cause mortality in the elderly.