TGFβ1 regulates Scleraxis expression in primary cardiac myofibroblasts by a Smad-independent mechanism.

In cardiac wound healing following myocardial infarction (MI), relatively inactive resident cardiac fibroblasts phenoconvert to hypersynthetic/secretory myofibroblasts that produce large quantities of extracellular matrix (ECM) and fibrillar collagen proteins. Our laboratory and others have identified TGFβ1 as being a persistent stimulus in the chronic and inappropriate wound healing phase that is marked by hypertrophic scarring and eventual stiffening of the entire myocardium, ultimately leading to the pathogenesis of heart failure following MI. Ski is a potent negative regulator of TGFβ/Smad signaling with known antifibrotic effects.

SnoN as a novel negative regulator of TGF-β/Smad signaling: a target for tailoring organ fibrosis.

Remodeling of the extracellular matrix is beneficial during the acute wound healing stage following tissue injury. In the short term, resident fibroblasts and myofibroblasts regulate the matrix remodeling process through production of matricellular protein components that provide structural support to the damaged tissue. This process is largely governed by the transforming growth factor-β1 (TGF-β1) pathway, a critical mediator of the remodeling process.

Autophagy and heart disease: implications for cardiac ischemia-reperfusion damage.

Survival of myocytes and mesenchymal cells in the heart is tightly regulated by a number of adaptive processes that are invoked with the changes that occur within the parenchyma and stroma. Autophagy is implicated in cellular housekeeping duties and maintenance of the integrity of the intracellular milieu by removal of protein aggregates and damaged organelles, whereas under pathophysiological conditions, the chronic up-regulation of autophagy may lead to significant disturbance of homeostatic conditions. Nonetheless, the role of autophagy in heart disease in the context of cardiac ischemia-reperfusion injury is currently unclear.

μ-Calpain-mediated deregulation of cardiac, brain, and kidney NCX1 splice variants.

μ-Calpain is a Ca(2+)-activated protease abundant in mammalian tissues. Here, we examined the effects of μ-calpain on three alternatively spliced variants of NCX1 using the giant, excised patch technique. Membrane patches from Xenopus oocytes expressing either heart (NCX1.

Crystal structures of progressive Ca2+ binding states of the Ca2+ sensor Ca2+ binding domain 1 (CBD1) from the CALX Na+/Ca2+ exchanger reveal incremental conformational transitions.

Na(+)/Ca(2+) exchangers (NCX) constitute a major Ca(2+) export system that facilitates the re-establishment of cytosolic Ca(2+) levels in many tissues. Ca(2+) interactions at its Ca(2+) binding domains (CBD1 and CBD2) are essential for the allosteric regulation of Na(+)/Ca(2+) exchange activity. The structure of the Ca(2+)-bound form of CBD1, the primary Ca(2+) sensor from canine NCX1, but not the Ca(2+)-free form, has been reported, although the molecular mechanism of Ca(2+) regulation remains unclear.

Characterization of zebrafish (Danio rerio) NCX4: a novel NCX with distinct electrophysiological properties.

Members of the Na+/Ca2+ exchanger (NCX) family are important regulators of cytosolic Ca2+ in myriad tissues and are highly conserved across a wide range of species. Three distinct NCX genes and numerous splice variants exist in mammals, many of which have been characterized in a variety of heterologous expression systems. Recently, however, we discovered a fourth NCX gene (NCX4), which is found exclusively in teleost, amphibian, and reptilian genomes.

Frequency-dependent regulation of cardiac Na(+)/Ca(2+) exchanger.

The activity of the cardiac Na(+)/Ca(2+) exchanger (NCX1.1) undergoes continuous modulation during the contraction-relaxation cycle because of the accompanying changes in the electrochemical gradients for Na(+) and Ca(2+). In addition, NCX1.

Inhibitory profile of SEA0400 [2-[4-[(2,5-difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline] assessed on the cardiac Na+-Ca2+ exchanger, NCX1.1.

SEA0400 (2-[4-[(2,5-difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline) has recently been described as a potent and selective inhibitor of Na(+)-Ca(2+) exchange in cardiac, neuronal, and renal preparations. The inhibitory effects of SEA0400 were investigated on the cloned cardiac Na(+)-Ca(2+) exchanger, NCX1.1, expressed in Xenopus laevis oocytes to gain insight into its inhibitory mechanism.

Effects of SEA0400 on mutant NCX1.1 Na+-Ca2+ exchangers with altered ionic regulation.

SEA0400 (SEA) blocks cardiac and neuronal Na+-Ca2+ exchange with the highest affinity of any known inhibitor, yet very little is known about its molecular mechanism of action. Previous data from our lab suggested that SEA stabilizes or modulates the transition of NCX1.1 exchangers into a Na+i-dependent (I1) inactive state.

Inhibition of canine (NCX1.1) and Drosophila (CALX1.1) Na(+)-Ca(2+) exchangers by 7-chloro-3,5-dihydro-5-phenyl-1H-4,1-benzothiazepine-2-one (CGP-37157).

The electrophysiological effects of the benzothiazepine 7-chloro-3,5-dihydro-5-phenyl-1H-4,1-benzothiazepine-2-one (CGP-37157) (CGP) were investigated on the canine (NCX1.1) and Drosophila (CALX1.1) plasmalemmal Na+-Ca2+ exchangers.

Inhibition of Na+/Ca2+ exchange by KB-R7943: transport mode selectivity and antiarrhythmic consequences.

The Na+/Ca2+ exchanger plays a prominent role in regulating intracellular Ca2+ levels in cardiac myocytes and can serve as both a Ca2+ influx and efflux pathway. A novel inhibitor, KB-R7943, has been reported to selectively inhibit the reverse mode (i.e.

Functional role of ionic regulation of Na+/Ca2+ exchange assessed in transgenic mouse hearts.

Na+/Ca2+ exchange is the primary mechanism mediating Ca2+ efflux from cardiac myocytes during diastole and, thus, can prominently influence contractile force. In addition to transporting Na+ and Ca2+, the exchanger is also regulated by these ions. Although structure-function studies have identified protein regions of the exchanger subserving these regulatory processes, their physiological importance is unknown.

Ionic regulatory properties of brain and kidney splice variants of the NCX1 Na(+)-Ca(2+) exchanger.

Ion transport and regulation of Na(+)-Ca(2+) exchange were examined for two alternatively spliced isoforms of the canine cardiac Na(+)-Ca(2+) exchanger, NCX1.1, to assess the role(s) of the mutually exclusive A and B exons. The exchangers examined, NCX1.

Functional differences in ionic regulation between alternatively spliced isoforms of the Na+-Ca2+ exchanger from Drosophila melanogaster.

Ion transport and regulation were studied in two, alternatively spliced isoforms of the Na+-Ca2+ exchanger from Drosophila melanogaster. These exchangers, designated CALX1.1 and CALX1.

Structure-function analysis of CALX1.1, a Na+-Ca2+ exchanger from Drosophila. Mutagenesis of ionic regulatory sites.

Cytoplasmic Na+ and Ca2+ regulate the activity of Na+-Ca2+ exchange proteins, in addition to serving as the transported ions, and protein regions involved in these processes have been identified for the canine cardiac Na+-Ca2+ exchanger, NCX1.1. Although protein regions associated with Na+i- and Ca2+i-dependent regulation are highly conserved among cloned Na+-Ca2+ exchangers, it is unknown whether or not the structure-function relationships characteristic of NCX1.

Effect of binding protein surface charge on palmitate uptake by hepatocyte suspensions.

1. Studies were directed at determining whether hepatocytes, isolated from female Sprague-Dawley rats, facilitate the uptake of protein-bound long-chain fatty acids. We postulated one form of facilitated uptake may occur through an ionic interaction between the protein-ligand complex and the cell surface.

Transport and regulation of the cardiac Na(+)-Ca2+ exchanger, NCX1. Comparison between Ca2+ and Ba2+.

Cardiac muscle fails to relax upon replacement of extracellular Ca2+ with Ba2+. Among the manifold consequences of this intervention, one major possibility is that Na(+)-Ba2+ exchange is inadequate to support normal relaxation. This could occur due to reduced transport rates of Na(+)-Ba2+ exchange and/or by failure of Ba2+ to activate the exchanger molecule at the high affinity regulatory Ca2+ binding site.

Effect of nitric oxide on albumin-palmitate binding.

Bovine serum albumin (albumin) was modified by treatment with nitric oxide (NO) to form S-nitrosoalbumin. Analysis of the reduced sulfhydryl groups showed that more than 99% of the albumin was converted to S-nitrosoalbumin. Using a 1:1 molar ratio of protein:palmitate, the unbound palmitate fraction in the presence of S-nitrosoalbumin was determined to be greater (28%) than in the presence of albumin as determined by heptane: water partitioning.

Coupling of a mutated form of the human beta 2-adrenergic receptor to Gi and Gs. Requirement for multiple cytoplasmic domains in the coupling process.

We constructed five genes encoding mutant human beta 2-adrenergic receptor sequence (beta 2AR) which contained 12-22 amino acid substitutions with corresponding sequence from the human alpha 2AAR in order to assess the receptor domains involved in Gs versus Gi recognition and coupling. Mutant beta 2AR with substitutions in the N (S1)- and C-terminal (S2) portions of the third intracellular loop, the proximal cytoplasmic tail (S3), and two combinations thereof (S2,3 and S1,2,3), were stably expressed in Chinese hamster fibrobasts (CHW-1102), as were the human beta 2AR and alpha 2AAR at comparable receptor levels. All mutant receptors with S2 substitutions (i.

Mutations of the human beta 2-adrenergic receptor that impair coupling to Gs interfere with receptor down-regulation but not sequestration.

The integrity of coupling of the beta 2-adrenergic receptor (beta 2AR) to its guanine nucleotide-binding protein, Gs, and phosphorylation events on the receptor molecule have been proposed to be important determinants in the processes of receptor sequestration and down-regulation. However, little is known about the molecular mechanisms underlying these processes, and the regions of the receptor molecule that specifically subserve sequestration and down-regulation have yet to be delineated. To address these questions, we stably transfected eight mutant beta 2AR genes into Chinese hamster fibroblasts and evaluated the coupling, sequestration, and down-regulation properties of the mutated receptors.

The 5-HT1A receptor: an overview of recent advances.

Progress in the field of neuronal receptor research has accelerated during the last few years due to developments in pharmacology and molecular biology. This is particularly true in the case of the serotonin 5-HT1A receptor. In 1983 the very selective, high affinity 5-HT1A agonist 8-OH-DPAT was developed which allowed the pharmacology and distribution of the 5-HT1A receptor in the central nervous system of the rat and man to be extensively characterized.

Involvement of tyrosine residues located in the carboxyl tail of the human beta 2-adrenergic receptor in agonist-induced down-regulation of the receptor.

Chronic exposure of various cell types to adrenergic agonists leads to a decrease in cell surface beta 2-adrenergic receptor (beta 2AR) number. Sequestration of the receptor away from the cell surface as well as a down-regulation of the total number of cellular receptors are believed to contribute to this agonist-mediated regulation of receptor number. However, the molecular mechanisms underlying these phenomena are not well characterized.

Adrenergic receptors. Models for regulation of signal transduction processes.

Adrenergic receptors are prototypic models for the study of the relations between structure and function of G protein-coupled receptors. Each receptor is encoded by a distinct gene. These receptors are integral membrane proteins with several striking structural features.