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The article discusses a study which examined the efficacy and safety of perioperative treatment with methylprednisolone on the development of lung injury after pulmonary thromboendarterectomy. Considered as the common complication of PTE, lung injury is characterized by edema resulting to hypoxemia and radiographic opacities in areas of the lungs that have been reperfused. Based on the results, it was concluded that the use of methylprednisolone does not reduce the incidence of lung injury.

Myocardial ischemia/reperfusion injury is a major cause of morbidity and mortality. The molecular signaling pathways involved in cardiac protection from myocardial ischemia/reperfusion injury are complex. An emerging idea in signal transduction suggests the existence of spatially organized complexes of signaling molecules in lipid-rich microdomains of the plasma membrane known as caveolae. Caveolins-proteins abundant in caveolae-provide a scaffold to organize, traffic, and regulate signaling molecules. Numerous signaling molecules involved in cardiac protection are known to exist within caveolae or interact directly with caveolins. Over the last 4 years, our laboratories have explored the hypothesis that caveolae are vitally important to cardiac protection from myocardial ischemia/reperfusion injury. We have provided evidence that (1) caveolae and the caveolin isoforms 1 and 3 are essential for cardiac protection from myocardial ischemia/reperfusion injury, (2) stimuli that produce preconditioning of cardiac myocytes, including brief periods of ischemia/reperfusion and exposure to volatile anesthetics, alter the number of membrane caveolae, and (3) cardiac myocyte-specific overexpression of caveolin-3 can produce innate cardiac protection from myocardial ischemia/reperfusion injury. The work demonstrates that caveolae and caveolins are critical elements of signaling pathways involved in cardiac protection and suggests that caveolins are unique targets for therapy in patients at risk of myocardial ischemia. [ABSTRACT FROM AUTHOR]

Scar formation occurs during the late stages of the inflammatory response but, when excessive, produces fibrosis that can lead to functional and structural damage of tissues. Here, we show that the profibrogenic agonist, transforming growth factor β1, transcriptionally decreases expression of Exchange protein activated by cAMP 1 (Epac1) in fibroblasts/fibroblast-like cells from multiple tissues (i.e., cardiac, lung, and skin fibroblasts and hepatic stellate cells). Overexpression of Epac1 inhibits transforming growth factor β1-induced collagen synthesis, indicating that a decrease of Epac1 expression appears to be necessary for the fibrogenic phenotype, an idea supported by evidence that Epac1 expression in cardiac fibroblasts is inhibited after myocardial infarction. Epac and protein kinase A, a second mediator of cAMP action, have opposite effects on migration but both inhibit synthesis of collagen and DNA by fibroblasts. Epac is preferentially activated by low concentrations of cAMP and stimulates migration via the small G protein Rap1 but inhibits collagen synthesis in a Rap1-independent manner. The regulation of Epac expression and activation thus appear to be critical for the integration of pro- and anti-fibrotic signals and for the regulation of fibroblast function. [ABSTRACT FROM AUTHOR]

N-Methyl-D-aspartate (NMDA) receptor (NMDAR) activation and downstream signaling are important for neuronal function. Activation of prosurvival Src family kinases and extracellular signal-regulated kinase (ERK) 1/2 is initiated by NMDAR activation, but the cellular organization of these kinases in relation to NMDARs is not entirely clear. We hypothesized that caveolin-1 scaffolds and coordinates protein complexes involved in NMDAR signaling and that this organization is necessary for neuronal preconditioning, whereby NMDAR activation protects neurons from subsequent ischemic cell death. We found that sublethal ischemia (SLI) or preconditioning via NMDA treatment of primary cortical neurons from neonatal rats or mice increases expression of phosphorylated (P) caveolin-1, P-Src, and P-ERK1/2. The NMDAR antagonist, MK801, or the Src inhibitor, PP2, attenuated SLI-induced preconditioning. NMDAR2B distributed to buoyant fractions and heavy fractions, partially colocalized with caveolin-1 and the membrane raft marker, cholera toxin B. Cultures of primary neurons treated with caveolin-1 small interfering RNA or from caveolin-1-/- mice lacked the NMDA-mediated increase in P-Src and PERK, as well as SLI- and NMDA-induced preconditioning. Adenovirally mediated expression of caveolin-1 in neurons from caveolin-1-/- mice restored NMDA-mediated enhancement of P-Src and P-ERK1/2, redistributed NMDAR2B to buoyant fractions, and enhanced NMDAR2B localization to membrane rafts. We conclude that caveolin-1, perhaps via its ability to scaffold key signaling components, is essential for NMDAR localization to neuronal membrane rafts, NMDAR/Src tyrosine kinase family/ERK signaling, and protection of neurons from ischemic injury and cell death. [ABSTRACT FROM AUTHOR]

Myocardial infarction (MI) results in left ventricular remodeling (e.g., ventricular hypertrophy, dilatation, and fibrosis). Fibrosis contributes to increased myocardial stiffening, impaired ventricular filling and function, and reduced cardiac output. Adenylyl cyclase (AC) expression and activity are reduced in animal models of heart failure. Stimulation of AC can inhibit extracellular matrix production in isolated cardiac fibroblasts; however, a role for reduced AC expression and activity in fibrosis associated with cardiac remodeling after chronic MI has never been determined. We tested the hypothesis that AC expression and activity are reduced in cardiac fibroblasts after chronic (18 wk) MI. Rats underwent coronary artery ligation or sham surgery (control), and echocardiography was used to assess left ventricular remodeling 1, 3, 5, 7, 10, 12, and 18 wk after surgery. Cardiac fibroblasts were isolated from the noninfarcted myocardium and compared for differences in AC activity and collagen synthesis. End-diastolic dimension was increased [control: 0.76 ± 0.02 cm and Ml: 1.0 ± 0.02 cm (means ± SE), P < 0.001] and fractional shortening was decreased (control: 44 ± 2% and MI: 17 ± 2%, P < 0.001) in MI compared with control rats. Basal and forskolin-stimulated cAMP production were decreased by 90% and 93%, respectively, and AC5/6 expression was decreased 39% in fibroblasts isolated from MI rats compared with sham controls. Serum-stimulated collagen production was increased twofold and forskolin-mediated inhibition of collagen synthesis was reduced in fibroblasts from MI rats compared with controls. Our data demonstrate that AC expression and activity are reduced and collagen production is increased in cardiac fibroblasts of rats after MI. [ABSTRACT FROM AUTHOR]

Delayed cardiac protection mediated by 12-1ipoxygenase (12-LO) expression and activity has been linked to opioid receptor stimulation. The role of 12-LO in volatile anesthetic-induced delayed cardiac protection has not been determined. We tested the hypothesis that expression and activity of 12-LO mediate delayed cardiac protection induced by isoflurane in the mouse heart in vivo. Mice were pretreated with 1.4% isoflurane for 30 min and allowed to recover for 1, 12, or 24 h. Immunoblot analysis showed isofturane significantly enhanced 12-LO protein expression at 12 and 24 h after isoflurane exposure, and this induction of 12-LO was confirmed by immunohistochemistry of whole heart sections at 24 h. The induced protein expression appeared to be localized to intercalated disc regions adjoining adjacent cardiac myocytes. Additionally, mice ± isoflurane (24 h previously) were subjected to 30 min coronary artery occlusion followed by 2 h of reperfusion in the presence and absence of a 12-LO inhibitor. Isoflurane reduced infarct size (27.1 ± 2.2% of the area at risk; n = 8) compared with the control group (44.6 ± 3.6%, n = 8). Baicalein (3 mg/kg), a selective 12-LO inhibitor, blocked the delayed protective effects of isoflurane pretreatment on infarct size (40.6 ± 3.6%, n = 8). These data suggest that 12-LO is an important mediator of isoflurane-induced delayed preconditioning. [ABSTRACT FROM AUTHOR]

We tested the hypothesis that deletion of adenylyl cyclase type V (ACV) would be associated with decreased left ventricular (LV) contractile function and responsiveness to β–adrenergic receptor (βAR) stimulation. Absence of cardiac ACV expression was confirmed by RT–PCR and immunoblotting in ACV–deleted mice (ACV -/-). Compared to sibling mice with normal amounts of ACV (CON), basal and water–soluble forskolin derivative NKH477–stimulated cAMP production was reduced in both LV homogenates and in isolated cardiac myocytes. Basal LV +dP/dt (isolated perfused hearts) was increased (CON: 3649 ± 247 mmHg/s; ACV -/-: CON: –6.83 ± 0.14 M; ACV: –5.99 ± 0.15 M; p = 0.0007, n = 10). The initial rates of ATP–dependent sarcoplasmic reticulum calcium uptake, assessed in LV homogenates, showed that AC50: CON: –6.83 ± 0.14 M; ACV -/-: CON: –5.94 ± 0.03 M; ACV: –6.09 ± 0.02 M; p = 0.001, n = 8). AC2+ (log EC50: CON: –5.94 ± 0.03 M; ACV -/-: –6.09 ± 0.02 M; p = 0.001, n = 8). ACV deletion is also associated with increased phospholamban phosphorylation, decreased type 1 protein phosphatase catalytic subunit content and activity, and reduced cardiac Gαs protein content. In conclusion, ACV deletion has a favorable effect on basal LV function despite reduced cAMP levels. Increased SERCA2a affinity for Ca2+ and increased phospholamban phosphorylation are contributing factors. However, ACV deletion is associated with reduced LV contractile responsiveness to βAR stimulation, an effect that is associated with reduced Gαs protein content and reduced cAMP generating capacity in cardiac myocytes. [ABSTRACT FROM AUTHOR]

Transformation of fibroblasts to myofibroblasts, characterized by expression of α-smooth muscle actin (α-SMA) and production of extracellular matrix (ECM) components, is a key event in connective tissue remodeling. Approaches to inhibit this transformation are needed in tissues, such as the heart, where excessive ECM production by cardiac fibroblasts (CFs) causes fibrosis, myocardial stiffening, and cardiac dysfunction. We tested whether adenylyl cyclase (AC) activation (increased CAMP levels) modulates the transformation of adult rat CF to myofibroblasts, as assessed by immunofluorescent microscopy, immunoblotting, and collagen synthesis. A 24-h incubation of CF with TGF-β or angiotensin II increased α-SMA expression, which was inhibited by the AC agonist forskolin and a CAMP analog that activates protein kinase A. Treatment with forskolin blunted serum-, TGF-β-, and angiotensin II-stimulated collagen synthesis. CFs engineered to overexpress type 6 AC had enhanced forskolin-promoted cAMP formation, greater inhibition by forskolin of TGF-β-stimulated α-SMA expression, and a decrease in the EC50 of forskolin to reduce serum-stimulated collagen synthesis. The AC stimulatory agonist adrenomedullin inhibited collagen synthesis in CF that overexpressed AC6 but not in controls. Thus, AC stimulation blunts collagen synthesis and, in parallel, the transformation of adult rat CF to myofibroblasts. AC overexpression enhances these effects, "uncovering" an inhibition by adrenomedullin. These findings implicate cAMP as an inhibitor of ECM formation by means of blockade of the transformation of CF to myofibroblasts and suggest that increasing AC expression, thereby enhancing CAMP generation through stimulation of receptors expressed on CF, could provide a means to attenuate and prevent cardiac fibrosis and its sequelae. [ABSTRACT FROM AUTHOR]

We performed indirect intracoronary delivery of adenovirus vectors in mice and explored techniques including hypothermia and pharmacological means to increase cardiac gene transfer. Mice were maintained in a normothermic state or cooled to 25δC. The aorta or both the pulmonary artery and aorta were clamped while a needle was advanced into the left ventricular cavity to deliver adenovirus vectors encoding enhanced green fluorescent protein (EGFP) or murine adenylyl cyclase type VI (ACVI )with saline, sodium nitroprusside, acetylcholine, or serotonin. Clamping was maintained for 30 s (normothermia) or 2 min (25°C) after adenovirus administration. Mice were killed 7 or 21 days later, and hearts were examined for EGFP expression. Compared with clamping the aorta alone and with no cooling, gene transfer was increased as follows: 1) 1.3-fold with hypothermia to extend dwell time; 2) 4.5-fold by clamping the aorta and the pulmonary artery; 3) 11.4-fold with nitroprusside administration; 4) 11.8-fold with serotonin addition, and 5) 14.3-fold with acetylcholine delivery. Gene expression remained substantial at 21 days, and no significant inflammatory response was seen. Efficacy of the method was tested by performing gene transfer of adenovirus encoding ACVI. Fourteen days after gene transfer, hearts isolated from mice that received adenovirus encoding ACVI showed increased contractile function. Indirect intracoronary delivery of adenovirus vectors in mice is associated with efficient cardiac gene transfer and increased left ventricular function after ACVI gene transfer. [ABSTRACT FROM AUTHOR]

Objective: We have previously shown that cardiac-directed expression of adenylycyclase type VI (ACVI) increases heart function in transgenic mice, and improves heart function and survival in murine cardiomyopathy. However, a potential problem of crossbreeding paradigms that use lines with two constitutively active transgenes is that results can be obfuscated by interactions between transgenes during growth and development. Methods: To develop a model that could be used subsequently to address this generic problem, transgenic mice with tetracycline (tet)-regulated cardiac-specific expression of ACVI were generated. In this transgenic strain, the expression of a tet-controlled transactivator (tTA) was under control of the rat α-myosin heavy chain promoter. Expression of the ACVI gene was driven by a tet-response element (TRE) and a minimal CMV promoter. Results: Homogenates of hearts showed no change in ACVI protein content during tet suppression (doxycycline), confirming successful suppression of transgene expression. Removal of tet suppression for 10 days was associated with a 10-fold increase in cardiac ACVI protein content. A similar increase in mRNA was observed (Northern blot analysis). The estimated half-life of newly synthesized cardiac ACVI protein was 2–3 days. Isolated cardiac myocytes from animals that had tet-suppression removed for 10 days showed increased cAMP production in response to forskolin stimulation (Transgene Off: 15±6 fmol/μg; Transgene On: 39±14 fmol/μg; n=5 each group; P=0.004) and also to isoproterenol stimulation (Transgene Off: 20±5 fmol/μg; Transgene On: 31±12 fmol/μg; n=5 each group; P=0.035) and hearts isolated from these animals showed marked increased left ventricular peak dP/dt in response to dobutamine stimulation (P=0.009) indicating that inducible cardiac ACVI is functionally coupled and recruitable. Conclusion: We have generated transgenic mice with controlled cardiac-specific expression of ACVI, provided detailed information regarding the kinetics of transgene expression and suppression and estimated the half-life of cardiac ACVI protein to be 2–3 days. Finally, we have shown, for the first time, that controlled cardiac-directed expression of a transgene can increase cardiac myocyte cAMP generation and left ventricular contractile function. [Copyright &y& Elsevier]

Inhaling aerosols from electronic nicotine delivery systems, such as e-cigarettes (e-cigs), may pose health risks beyond those caused by nicotine intake. Exposure to e-cig aerosols can lead to the release of exosomes and metabolites into the bloodstream, potentially affecting mitochondrial physiology across the body, leading to chronic inflammatory diseases. In this study we assessed the effects of e-cig use by young healthy human subjects on the circulating exosome profile and markers of cell stress, and also defined the effects of e-cig user plasma on mitochondrial function in endothelial cells (EA. Hy 926) and epithelial cells (A549) via adoptive transfer. E-cig users had altered plasma exosome profiles, with significantly increased levels of cell free mitochondrial DNA (mtDNA), protein carbonyls, and 4-HNE relative to non-users. Plasma from e-cig users decreased maximal mitochondrial respiration and spare capacity of cells, while also increasing metabolic stress, as evidenced by changes in mitochondrial phenotype from basal to stressed in both endothelial and epithelial cells, which was corroborated by electron microscopy demonstrating structural changes in mitochondria. Mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) levels significantly increased in e-cig plasma-subjected cells. Overall, we identified alterations in plasma exosome profiles and increased markers of mitochondrial stress in e-cig users and evidence that circulating factors within plasma from e-cig users drives metabolic stress in endothelial and epithelial cells. Our results imply that e-cig use adversely affects mitochondrial function, leading to stress and potentially chronic inflammation across the body. Inhalation of e-cigarette aerosols alters circulating nanoparticles in e-cigarette users. Factors within e-cig user plasma change metabolism within endothelial and epithelial cells, which may be primarily due to oxidative stress. [Display omitted] • E-cig use induces stress both in human subjects and cell culture models. • E-cig users show significantly higher plasma mitochondrial DNA than non-users. • E-cig plasma exposure alters respiration, stressing epithelial and endothelial cells. [ABSTRACT FROM AUTHOR]

Disruptions in normal attachment, signaled by intense separation distress, can be conceptualized as a central risk factor underlying the development of anorexia and bulimia nervosa. In a preliminary study of the hypothesis, Hansburg's Separation Anxiety Test, derived from Bowlby's attachment theory, was administered to eating- disordered inpatients. These patients evidenced significantly more severe separation and attachment difficulties than is normal in adolescence and in adults under- going developmentally based relationship crises. They appear to make no cognitive distinction between brief, everyday leavetakings and more permanent breaks. The implications of these findings, and of Bowlby's attachment theory, for the understanding and treatment of eating disorders is discussed. [ABSTRACT FROM AUTHOR]

A postulated capability of riboflavin and flavin mononucleotide to protect DNA from damage by far-ultraviolet irradiation was investigated in purified reaction mixtures and in cultured mammalian cells. Purified calf thymus DNA, admixed with flavin at approximately physiological ratios, was protected from undergoing irradiation-induced changes in spectral absorbance and viscosity compatible with those accompanying pyrimidine dimerization. Irradiation-induced changes in unprotected DNA were reversed by a subsequent treatment of the DNA with flavin in the presence of visible light. There was no apparent interaction between flavin and non-irradiated DNA. These results suggest that photoexcited flavin interacts directly with the ultraviolet photoproduct in DNA and does not function simply as an energy sink to dissipate photons absorbed by DNA. Cultured embryonic hamster cells were protected from ultraviolet-induced inhibition of growth by a preliminary exposure to relatively large amounts of flavin. This effect was not due to a vitamin- like promotion of cell growth by the flavin. The observation that flavin concentrated transiently in the cytoplasm of exponentially growing hamster cells during a 6 hr incubation period suggested that a shielding action may cooperate with the protective photochemical mechanism in vivo. It was concluded that flavins may function physiologically to protect the genetic material of tissues exposed to actinic radiations, and that when flavin levels in these tissues are inadequate, either because of nutritional deficiency or excessive intensity of irradiation, the damage which results may provide one pathogenetic avenue for the characteristic integumentary lesions of riboflavin deficiency disease. [ABSTRACT FROM AUTHOR]

The noble gas helium (He) induces cardioprotection in vivo through unknown molecular mechanisms. He can interact with and modify cellular membranes. Caveolae are cholesterol and sphingolipid-enriched invaginations of the plasma-membrane-containing caveolin (Cav) proteins that are critical in protection of the heart. Mice (C57BL/6J) inhaled either He gas or adjusted room air. Functional measurements were performed in the isolated Langendorff perfused heart at 24 h post He inhalation. Electron paramagnetic resonance spectrometry (EPR) of samples was carried out at 24 h post He inhalation. Immunoblotting was used to detect Cav-1/3 expression in whole-heart tissue, exosomes isolated from platelet free plasma (PFP) and membrane fractions. Additionally, transmission electron microscopy analysis of cardiac tissue and serum function and metabolomic analysis were performed. In contrast to cardioprotection observed in in vivo models, the isolated Langendorff perfused heart revealed no protection after He inhalation. However, levels of Cav-1/3 were reduced 24 h after He inhalation in whole-heart tissue, and Cav-3 was increased in exosomes from PFP. Addition of serum to muscle cells in culture or naïve ventricular tissue increased mitochondrial metabolism without increasing reactive oxygen species generation. Primary and lipid metabolites determined potential changes in ceramide by He exposure. In addition to direct effects on myocardium, He likely induces the release of secreted membrane factors enriched in caveolae. Our results suggest a critical role for such circulating factors in He-induced organ protection. [ABSTRACT FROM AUTHOR]