Modulating OPG and TGF-β1 mRNA expression via bioelectrical stimulation.

Background: Bone remodeling is a lifelong process that ranges from orthodontic tooth movement/alignment to bone damage/healing, to overall bone health. Osteoprotegerin (OPG) and transforming growth factor β1 (TGF-β1) are secreted by osteoblasts and participate in bone remodeling. OPG promotes bone remineralization and stabilization prominent in post-mechanical repositioning of the teeth in the dental alveolus.

Distinct moieties underlie biphasic H gating of connexin43 channels, producing a pH optimum for intercellular communication.

Most mammalian cells can intercommunicate via connexin-assembled, gap-junctional channels. To regulate signal transmission, connexin (Cx) channel permeability must respond dynamically to physiological and pathophysiological stimuli. One key stimulus is intracellular pH (pH), which is modulated by a tissue's metabolic and perfusion status.

Mono-Heteromeric Configurations of Gap Junction Channels Formed by Connexin43 and Connexin45 Reduce Unitary Conductance and Determine both Voltage Gating and Metabolic Flux Asymmetry.

In cardiac tissues, the expression of multiple connexins (Cx40, Cx43, Cx45, and Cx30.2) is a requirement for proper development and function. Gap junctions formed by these connexins have distinct permeability and gating mechanisms.

Modulation of Asymmetric Flux in Heterotypic Gap Junctions by Pore Shape, Particle Size and Charge.

Gap junction channels play a vital role in intercellular communication by connecting cytoplasm of adjoined cells through arrays of channel-pores formed at the common membrane junction. Their structure and properties vary depending on the connexin isoform(s) involved in forming the full gap junction channel. Lack of information on the molecular structure of gap junction channels has limited the development of computational tools for single channel studies.

Computational simulations of asymmetric fluxes of large molecules through gap junction channel pores.

Gap junction channels are formed out of connexin isoforms, which enable molecule and ion selective diffusion amongst neighboring cells. HeLa cells expressing distinct connexins (Cx) allow the formation of heterotypic channels, where we observed a molecular charge-independent preferential flux of large fluorescent molecules in the Cx45 to Cx43 direction. We hypothesize that the pore's shape is a significant factor along-side charge and transjunctional voltages for this asymmetric flux.

Evaluation of Multiple Biological Therapies for Ischemic Cardiac Disease.

The development of cell- and gene-based strategies for regenerative medicine offers a therapeutic option for the repair and potential regeneration of damaged cardiac tissue post-myocardial infarction (MI). Human umbilical cord subepithelial cell-derived stem cells (hUC-SECs), human bone marrow-derived mesenchymal stem cells (hBM-MSCs), and human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), all derived from human tissue, have been shown to have in vitro and in vivo therapeutic potential. Additionally, S100a1, VEGF165, and stromal-derived factor-1α (SDF-1α) genes all have the potential to improve cardiac function and/or effect adverse remodeling.

Novel xeno-free human heart matrix-derived three-dimensional scaffolds.

Rationale: Myocardial infarction (MI) results in damaged heart tissue which can progress to severely reduce cardiac function, leading to death. Recent studies have injected dissociated, suspended cardiac cells into coronary arteries to restore function with limited results attributed to poor cell retention and cell death. Extracellular matrix (ECM) injected into damaged cardiac tissue sites show some promising effects.

Functional and pharmacological analysis of cardiomyocytes differentiated from human peripheral blood mononuclear-derived pluripotent stem cells.

Advances in induced pluripotent stem cell (iPSC) technology have set the stage for routine derivation of patient- and disease-specific human iPSC-cardiomyocyte (CM) models for preclinical drug screening and personalized medicine approaches. Peripheral blood mononuclear cells (PBMCs) are an advantageous source of somatic cells because they are easily obtained and readily amenable to transduction. Here, we report that the electrophysiological properties and pharmacological responses of PBMC-derived iPSC CM are generally similar to those of iPSC CM derived from other somatic cells, using patch-clamp, calcium transient, and multielectrode array (MEA) analyses.

Absence of glucose transporter 4 diminishes electrical activity of mouse hearts during hypoxia.

Insulin resistance, which characterizes type 2 diabetes, is associated with reduced translocation of glucose transporter 4 (GLUT4) to the plasma membrane following insulin stimulation, and diabetic patients with insulin resistance show a higher incidence of ischaemia, arrhythmias and sudden cardiac death. The aim of this study was to examine whether GLUT4 deficiency leads to more severe alterations in cardiac electrical activity during cardiac stress due to hypoxia. To fulfil this aim, we compared cardiac electrical activity from cardiac-selective GLUT4-ablated (G4H-/-) mouse hearts and corresponding control (CTL) littermates.

The voltage-sensitive dye di-4-ANEPPS slows conduction velocity in isolated guinea pig hearts.

Background: Voltage-sensitive dyes are important tools for mapping electrical activity in the heart. However, little is known about the effects of voltage-sensitive dyes on cardiac electrophysiology.

Objective: To test the hypothesis that the voltage-sensitive dye di-4-ANEPPS modulates cardiac impulse propagation.

The maximal downstroke of epicardial potentials as an index of electrical activity in mouse hearts.

The maximal upstroke of transmembrane voltage (dV(m)/dt(max)) has been used as an indirect measure of sodium current I(Na) upon activation in cardiac myocytes. However, sodium influx generates not only the upstroke of V(m), but also the downstroke of the extracellular potentials V(e) including epicardial surface potentials V(es). The purpose of this study was to evaluate the magnitude of the maximal downstroke of V(es) (|dV(es)/dt (min)|) as a global index of electrical activation, based on the relationship of dV(m)/dt(max) to I(Na).

The connexin43 C-terminal region mediates neuroprotection during stroke.

Connexin43 plays an important role in neuroprotection in experimental stroke models; reducing the expression of this gap junction protein in astrocytes enhances injury upon middle cerebral artery occlusion (MCAO). Because the C-terminal region of connexin43 isimportant for channel activity, we carried out MCAO stroke experiments in mice expressing a truncated form of connexin43 (Cx43DeltaCT mice). Brain sections were analyzed for infarct volume, astrogliosis, and inflammatory cell invasion 4 days after MCAO.

Electrophysiological modeling of fibroblasts and their interaction with myocytes.

Experimental studies have shown that cardiac fibroblasts are electrically inexcitable, but can contribute to electrophysiology of myocardium in various manners. The aim of this computational study was to give insights in the electrophysiological role of fibroblasts and their interaction with myocytes. We developed a mathematical model of fibroblasts based on data from whole-cell patch clamp and polymerase chain reaction (PCR) studies.

Gap junction channel gating modulated through protein phosphorylation.

As a ubiquitous post-translation modification process, protein phosphorylation has proven to be a key mechanism in regulating the function of several membrane proteins, including transporters and channels. Connexins, pannexins, and innexins are protein families that form gap junction channels essential for intercellular communication. Connexins have been intensely studied, and most of their isoforms are known to be phosphorylated by protein kinases that lead to modifications in tyrosine, serine, and threonine residues, which have been reported to affect, in one way or another, intercellular communication.

SRC utilizes Cas to block gap junctional communication mediated by connexin43.

The Src tyrosine kinase phosphorylates Cas (Crk-associated substrate) to confer anchorage independence and invasive growth potential to transformed cells. Gap junctional communication is often lower between aggressive tumor cells compared with normal or benign precursors. The gap junction protein connexin43 (Cx43) is a tumor suppressor that can inhibit tumor cell growth.

Transfection of mammalian cells with connexins and measurement of voltage sensitivity of their gap junctions.

Vertebrate gap junction channels are formed by a family of more than 20 connexin proteins. These gap junction proteins are expressed with overlapping cellular and tissue specificity, and coding region mutations can cause human hereditary diseases. Here we present a summary of what has been learned from voltage clamp studies performed on cell pairs either endogenously expressing gap junctions or in which connexins are exogenously expressed.

Connexin phosphorylation as a regulatory event linked to channel gating.

The main proteins required for functional gap junction channels are known as connexins and most of their isoforms indicate that they can become phosphorylated. Connexin phosphorylation has been reported to participate in modifying junctional communication and the mechanisms involved apparently depend on which kinase becomes involved. Although multiple reports have suggested a strong influence of phosphorylation on channel gating, not enough physiological studies have been performed to determine precisely the gating mechanisms implicated.

Biophysical evidence that connexin-36 forms functional gap junction channels between pancreatic mouse beta-cells.

Connexin-36 (Cx36) is the only gap junction protein that has been unambiguously identified in rodent pancreatic beta-cells. However, properties of gap junction channel unitary currents between beta-cells remain unrevealed. To address whether Cx36 forms functional channels in beta-cells, we characterized biophysical properties of macro- and microscopic junctional currents recorded from dual whole cell voltage clamp isolated pairs of dispersed mouse beta-cells.

Biophysical properties of homomeric and heteromultimeric channels formed by cardiac connexins.

Substantial advances have been made in characterizing the biophysical properties of channels formed exclusively by connexin isoforms expressed mainly in the heart, e.g., Cx43, Cx45 or Cx40.

A carboxyl terminal domain of connexin43 is critical for gap junction plaque formation but not for homo- or hetero-oligomerization.

We have initiated a series of experiments to analyze the biosynthesis and oligomerization of Cx43 in cells containing other connexins through the expression of site-directed mutants and chimeric connexin polypeptides. Here we report studies concerning a mutant of Cx43 (Cx43tr) that has been truncated after amino acid 251 to remove most of the Cx43 carboxy-terminal region. In stably transfected HeLa cells, full length Cx43 localized primarily to appositional membranes while much more Cx43tr was observed in the cytoplasm.

Gap junctions between cells expressing connexin 43 or 32 show inverse permselectivity to adenosine and ATP.

Gap junctions, composed of proteins from the connexin family, are the only channels that directly connect the cytoplasm of adjacent cells to allow for the intercellular transfer of small hydrophilic molecules. Gap junctional communication is essential for proper development and health in animals and humans. Whereas the study of biological molecules that pass through gap junctions is extremely important, the identification of endogenous transjunctional metabolites is challenging.

Connexin43 and connexin45 form heteromeric gap junction channels in which individual components determine permeability and regulation.

Two gap junction proteins, connexin43 (Cx43) and connexin45 (Cx45), are coexpressed in many cardiac and other cells. Homomeric channels formed by these proteins differ in unitary conductance, permeability, and regulation. We sought to determine the ability of Cx43 and Cx45 to oligomerize with each other to form heteromeric gap junction channels and to determine the functional and regulatory properties of these heteromeric channels.

Role of the carboxyl terminal of connexin43 in transjunctional fast voltage gating.

Previous studies show that chemical regulation of connexin43 (Cx43) gap junction channels depends on the integrity of the carboxyl terminal (CT) domain. Experiments using Xenopus oocytes show that truncation of the CT domain alters the time course for current inactivation; however, correlation with the behavior of single Cx43 channels has been lacking. Furthermore, whereas chemical gating is associated with a "ball-and-chain" mechanism, there is no evidence whether transjunctional voltage regulation for Cx43 follows a similar model.