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Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression

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A special issue of International Journal of Molecular Sciences (ISSN 1422-0067). This special issue belongs to the section "Molecular Pathology, Diagnostics, and Therapeutics".

Deadline for manuscript submissions: closed (30 September 2021) | Viewed by 30113

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Special Issue Editors

Prof. Dr. Paul Squires

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Guest Editor

Joseph Banks Laboratories, School of Life, Sciences University of Lincoln, Lincoln LN6 7DL, UK
Interests: diabetes; cell-to-cell communication; connexins
Special Issues, Collections and Topics in MDPI journals

Dr. Claire Elizabeth Hills

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Guest Editor

School of Life Sciences, University of Lincoln, Lincoln, UK
Interests: diabetes; cell-to-cell communication; connexins
Special Issues, Collections and Topics in MDPI journals

Special Issue Information

Dear Colleagues,

Cell contact, coupling, and communication are essential in maintaining normal physiological activity. Recent studies suggest that altered connexin expression and function are linked to remodeling of the extracellular matrix, integrin activation, and downstream purinergic signaling. As a consequence, changes in hemichannel and/or gap-junction connexin-mediated cell-to-cell communication are associated with inflammation and fibrosis, and have been linked to multiple diseases, including conditions of the heart, lung, liver, eye, and kidneys. In light of this, the potential impact of connexin-targeted therapeutics attracts considerable attention. In the current Special Issue, we will collectively highlight recent advances in the field of connexin biology, the link to disease development and progression, and the future therapeutic potential of connexin mimetics.

Prof. Dr. Paul Squires
Guest Editor

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Please visit the Instructions for Authors page before submitting a manuscript. There is an Article Processing Charge (APC) for publication in this open access journal. For details about the APC please see here. Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

connexins
hemichannels
gap junctions
ATP
fibrosis
inflammation
purinergic
connexin mimetics
extracellular matrix

Published Papers (11 papers)

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Research

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16 pages, 3061 KiB

Open AccessArticle

Interrogation of Carboxy-Terminus Localized GJA1 Variants Associated with Erythrokeratodermia Variabilis et Progressiva

by Sergiu A. Lucaciu, Qing Shao, Rhett Figliuzzi, Kevin Barr, Donglin Bai and Dale W. Laird

Int. J. Mol. Sci. 2022, 23(1), 486; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms23010486 - 01 Jan 2022

Cited by 4 | Viewed by 1787

Abstract

Although inherited GJA1 (encoding Cx43) gene mutations most often lead to oculodentodigital dysplasia and related disorders, four variants have been linked to erythrokeratodermia variabilis et progressiva (EKVP), a skin disorder characterized by erythematous and hyperkeratotic lesions. While two autosomal-dominant EKVP-linked GJA1 mutations have been shown to lead to augmented hemichannels, the consequence(s) of keratinocytes harboring a de novo P283L variant alone or in combination with a de novo T290N variant remain unknown. Interestingly, these variants reside within or adjacent to a carboxy terminus polypeptide motif that has been shown to be important in regulating the internalization and degradation of Cx43. Cx43-rich rat epidermal keratinocytes (REKs) or Cx43-ablated REKs engineered to express fluorescent protein-tagged P283L and/or T290N variants formed prototypical gap junctions at cell–cell interfaces similar to wildtype Cx43. Dye coupling and dye uptake studies further revealed that each variant or a combination of both variants formed functional gap junction channels, with no evidence of augmented hemichannel function or induction of cell death. Tracking the fate of EKVP-associated variants in the presence of the protein secretion blocker brefeldin A, or an inhibitor of protein synthesis cycloheximide, revealed that P283L or the combination of P283L and T290N variants either significantly extended Cx43 residency on the cell surface of keratinocytes or delayed its degradation. However, caution is needed in concluding that this modest change in the Cx43 life cycle is sufficient to cause EKVP, or whether an additional underlying mechanism or another unidentified gene mutation is contributing to the pathogenesis found in patients. This question will be resolved if further patients are identified where whole exome sequencing reveals a Cx43 P283L variant alone or, in combination with a T290N variant, co-segregates with EKVP across several family generations. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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11 pages, 1436 KiB

Open AccessArticle

Diazoxide Needs Mitochondrial Connexin43 to Exert Its Cytoprotective Effect in a Cellular Model of CoCl₂-Induced Hypoxia

by Michela Pecoraro, Stefania Marzocco and Ada Popolo

Int. J. Mol. Sci. 2021, 22(21), 11599; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms222111599 - 27 Oct 2021

Cited by 8 | Viewed by 1772

Abstract

Hypoxia is the leading cause of death in cardiomyocytes. Cells respond to oxygen deprivation by activating cytoprotective programs, such as mitochondrial connexin43 (mCx43) overexpression and the opening of mitochondrial K_ATP channels, aimed to reduce mitochondrial dysfunction. In this study we used an in vitro model of CoCl₂-induced hypoxia to demonstrate that mCx43 and K_ATP channels cooperate to induce cytoprotection. CoCl₂ administration induces apoptosis in H9c2 cells by increasing mitochondrial ROS production, intracellular and mitochondrial calcium overload and by inducing mitochondrial membrane depolarization. Diazoxide, an opener of K_ATP channels, reduces all these deleterious effects of CoCl₂ only in the presence of mCx43. In fact, our results demonstrate that in the presence of radicicol, an inhibitor of Cx43 translocation to mitochondria, the cytoprotective effects of diazoxide disappear. In conclusion, these data confirm that there exists a close functional link between mCx43 and K_ATP channels. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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17 pages, 3458 KiB

Open AccessArticle

Dysregulation of Connexin Expression Plays a Pivotal Role in Psoriasis

by Erin M. O’Shaughnessy, William Duffy, Laura Garcia-Vega, Keith Hussey, A. David Burden, Mozheh Zamiri and Patricia E. Martin

Int. J. Mol. Sci. 2021, 22(11), 6060; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22116060 - 04 Jun 2021

Cited by 4 | Viewed by 2667

Abstract

Background: Psoriasis, a chronic inflammatory disease affecting 2–3% of the population, is characterised by epidermal hyperplasia, a sustained pro-inflammatory immune response and is primarily a T-cell driven disease. Previous work determined that Connexin26 is upregulated in psoriatic tissue. This study extends these findings. Methods: Biopsies spanning psoriatic plaque (PP) and non-involved tissue (PN) were compared to normal controls (NN). RNA was isolated and subject to real-time PCR to determine gene expression profiles, including GJB2/CX26, GJB6/CX30 and GJA1/CX43. Protein expression was assessed by immunohistochemistry. Keratinocytes and fibroblasts were isolated and used in 3D organotypic models. The pro-inflammatory status of fibroblasts and 3D cultures was assessed via ELISA and RnD cytokine arrays in the presence or absence of the connexin channel blocker Gap27. Results: Connexin26 expression is dramatically enhanced at both transcriptional and translational level in PP and PN tissue compared to NN (>100x). In contrast, CX43 gene expression is not affected, but the protein is post-translationally modified and accumulates in psoriatic tissue. Fibroblasts isolated from psoriatic patients had a higher inflammatory index than normal fibroblasts and drove normal keratinocytes to adopt a “psoriatic phenotype” in a 3D-organotypic model. Exposure of normal fibroblasts to the pro-inflammatory mediator peptidoglycan, isolated from Staphylococcus aureus enhanced cytokine release, an event protected by Gap27. Conclusion: dysregulation of the connexin26:43 expression profile in psoriatic tissue contributes to an imbalance of cellular events. Inhibition of connexin signalling reduces pro-inflammatory events and may hold therapeutic benefit. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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17 pages, 6490 KiB

Open AccessArticle

Connexin 43 and Connexin 26 Involvement in the Ponatinib-Induced Cardiomyopathy: Sex-Related Differences in a Murine Model

by Rosalinda Madonna, Stefania Moscato, Enza Polizzi, Damiana Pieragostino, Maria Concetta Cufaro, Piero Del Boccio, Francesco Bianchi, Raffaele De Caterina and Letizia Mattii

Int. J. Mol. Sci. 2021, 22(11), 5815; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22115815 - 28 May 2021

Cited by 14 | Viewed by 2242

Abstract

Cardiac connexins (Cxs) are proteins responsible for proper heart function. They form gap junctions that mediate electrical and chemical signalling throughout the cardiac system, and thus enable a synchronized contraction. Connexins can also individually participate in many signal transduction pathways, interacting with intracellular proteins at various cellular compartments. Altered connexin expression and localization have been described in diseased myocardium and the aim of this study is to assess the involvement of Cx43, Cx26, and some related molecules in ponatinib-induced cardiac toxicity. Ponatinib is a new multi-tyrosine kinase inhibitor that has been successfully used against human malignancies, but its cardiotoxicity remains worrisome. Therefore, understanding its signaling mechanism is important to adopt potential anti cardiac damage strategies. Our experiments were performed on hearts from male and female mice treated with ponatinib and with ponatinib plus siRNA-Notch1 by using immunofluorescence, Western blotting, and proteomic analyses. The altered cardiac function and the change in Cxs expression observed in mice after ponatinib treatment, were results dependent on the Notch1 pathway and sex. Females showed a lower susceptibility to ponatinib than males. The downmodulation of cardiac Cx43, Cx26 and miR-122, high pS368-Cx43 phosphorylation, cell viability and survival activation could represent some of the female adaptative/compensatory reactions to ponatinib cardiotoxicity. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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19 pages, 27447 KiB

Open AccessArticle

Collagen I Modifies Connexin-43 Hemichannel Activity via Integrin α2β1 Binding in TGFβ1-Evoked Renal Tubular Epithelial Cells

by Joe A. Potter, Gareth W. Price, Chelsy L. Cliff, Colin R. Green, Paul E. Squires and Claire E. Hills

Int. J. Mol. Sci. 2021, 22(7), 3644; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22073644 - 31 Mar 2021

Cited by 11 | Viewed by 3830

Abstract

Chronic Kidney Disease (CKD) is associated with sustained inflammation and progressive fibrosis, changes that have been linked to altered connexin hemichannel-mediated release of adenosine triphosphate (ATP). Kidney fibrosis develops in response to increased deposition of extracellular matrix (ECM), and up-regulation of collagen I is an early marker of renal disease. With ECM remodeling known to promote a loss of epithelial stability, in the current study we used a clonal human kidney (HK2) model of proximal tubular epithelial cells to determine if collagen I modulates changes in cell function, via connexin-43 (Cx43) hemichannel ATP release. HK2 cells were cultured on collagen I and treated with the beta 1 isoform of the pro-fibrotic cytokine transforming growth factor (TGFβ1) ± the Cx43 mimetic Peptide 5 and/or an anti-integrin α2β1 neutralizing antibody. Phase microscopy and immunocytochemistry observed changes in cell morphology and cytoskeletal reorganization, whilst immunoblotting and ELISA identified changes in protein expression and secretion. Carboxyfluorescein dye uptake and biosensing measured hemichannel activity and ATP release. A Cytoselect extracellular matrix adhesion assay assessed changes in cell-substrate interactions. Collagen I and TGFβ1 synergistically evoked increased hemichannel activity and ATP release. This was paralleled by changes to markers of tubular injury, partly mediated by integrin α2β1/integrin-like kinase signaling. The co-incubation of the hemichannel blocker Peptide 5, reduced collagen I/TGFβ1 induced alterations and inhibited a positive feedforward loop between Cx43/ATP release/collagen I. This study highlights a role for collagen I in regulating connexin-mediated hemichannel activity through integrin α2β1 signaling, ahead of establishing Peptide 5 as a potential intervention. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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21 pages, 6038 KiB

Open AccessArticle

Danegaptide Prevents TGFβ1-Induced Damage in Human Proximal Tubule Epithelial Cells of the Kidney

by Paul E. Squires, Gareth W. Price, Ulrik Mouritzen, Joe A. Potter, Bethany M. Williams and Claire E. Hills

Int. J. Mol. Sci. 2021, 22(6), 2809; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22062809 - 10 Mar 2021

Cited by 6 | Viewed by 3261

Abstract

Chronic kidney disease (CKD) is a global health problem associated with a number of comorbidities. Recent evidence implicates increased hemichannel-mediated release of adenosine triphosphate (ATP) in the progression of tubulointerstitial fibrosis, the main underlying pathology of CKD. Here, we evaluate the effect of danegaptide on blocking hemichannel-mediated changes in the expression and function of proteins associated with disease progression in tubular epithelial kidney cells. Primary human proximal tubule epithelial cells (hPTECs) were treated with the beta1 isoform of the pro-fibrotic cytokine transforming growth factor (TGFβ1) ± danegaptide. qRT-PCR and immunoblotting confirmed mRNA and protein expression, whilst a cytokine antibody array assessed the expression/secretion of proinflammatory and profibrotic cytokines. Carboxyfluorescein dye uptake and ATP biosensing measured hemichannel activity and ATP release, whilst transepithelial electrical resistance was used to assess paracellular permeability. Danegaptide negated carboxyfluorescein dye uptake and ATP release and protected against protein changes associated with tubular injury. Blocking Cx43-mediated ATP release was paralleled by partial restoration of the expression of cell cycle inhibitors, adherens and tight junction proteins and decreased paracellular permeability. Furthermore, danegaptide inhibited TGFβ1-induced changes in the expression and secretion of key adipokines, cytokines, chemokines, growth factors and interleukins. The data suggest that as a gap junction modulator and hemichannel blocker, danegaptide has potential in the future treatment of CKD. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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19 pages, 3277 KiB

Open AccessArticle

Differential Action of Connexin Hemichannel and Pannexin Channel Therapeutics for Potential Treatment of Retinal Diseases

by Mohd N. Mat Nor, Ilva D. Rupenthal, Colin R. Green and Monica L. Acosta

Int. J. Mol. Sci. 2021, 22(4), 1755; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22041755 - 10 Feb 2021

Cited by 12 | Viewed by 2182

Abstract

Dysregulation of retinal function in the early stages of light-induced retinal degeneration involves pannexins and connexins. These two types of proteins may contribute to channels that release ATP, leading to activation of the inflammasome pathway, spread of inflammation and retinal dysfunction. However, the effect of pannexin channel block alone or block of both pannexin channels and connexin hemichannels in parallel on retinal activity in vivo is unknown. In this study, the pannexin channel blocker probenecid and the connexin hemichannel blocker tonabersat were used in the light-damaged rat retina. Retinal function was evaluated using electroretinography (ERG), retinal structure was analyzed using optical coherence tomography (OCT) imaging and the tissue response to light-induced injury was assessed immunohistochemically with antibodies against glial fibrillary acidic protein (GFAP), Ionized calcium binding adaptor molecule 1 (Iba-1) and Connexin43 (Cx43). Probenecid did not further enhance the therapeutic effect of connexin hemichannel block in this model, but on its own improved activity of certain inner retina neurons. The therapeutic benefit of blocking connexin hemichannels was further evaluated by comparing these data against results from our previously published studies that also used the light-damaged rat retina model. The analysis showed that treatment with tonabersat alone was better than probenecid alone at restoring retinal function in the light-damaged retina model. The results assist in the interpretation of the differential action of connexin hemichannel and pannexin channel therapeutics for potential treatment of retinal diseases. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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Review

Jump to: Research

11 pages, 834 KiB

Open AccessReview

New Insights into Pulmonary Hypertension: A Role for Connexin-Mediated Signalling

by Myo Htet, Jane. E. Nally, Patricia. E. Martin and Yvonne Dempsie

Int. J. Mol. Sci. 2022, 23(1), 379; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms23010379 - 29 Dec 2021

Cited by 5 | Viewed by 1822

Abstract

Pulmonary hypertension is a serious clinical condition characterised by increased pulmonary arterial pressure. This can lead to right ventricular failure which can be fatal. Connexins are gap junction-forming membrane proteins which serve to exchange small molecules of less than 1 kD between cells. Connexins can also form hemi-channels connecting the intracellular and extracellular environments. Hemi-channels can mediate adenosine triphosphate release and are involved in autocrine and paracrine signalling. Recently, our group and others have identified evidence that connexin-mediated signalling may be involved in the pathogenesis of pulmonary hypertension. In this review, we discuss the evidence that dysregulated connexin-mediated signalling is associated with pulmonary hypertension. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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22 pages, 1333 KiB

Open AccessReview

Mechanisms of Connexin Regulating Peptides

by D. Ryan King, Meghan W. Sedovy, Xinyan Leng, Jianxiang Xue, Samy Lamouille, Michael Koval, Brant E. Isakson and Scott R. Johnstone

Int. J. Mol. Sci. 2021, 22(19), 10186; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms221910186 - 22 Sep 2021

Cited by 13 | Viewed by 3840

Abstract

Gap junctions (GJ) and connexins play integral roles in cellular physiology and have been found to be involved in multiple pathophysiological states from cancer to cardiovascular disease. Studies over the last 60 years have demonstrated the utility of altering GJ signaling pathways in experimental models, which has led to them being attractive targets for therapeutic intervention. A number of different mechanisms have been proposed to regulate GJ signaling, including channel blocking, enhancing channel open state, and disrupting protein-protein interactions. The primary mechanism for this has been through the design of numerous peptides as therapeutics, that are either currently in early development or are in various stages of clinical trials. Despite over 25 years of research into connexin targeting peptides, the overall mechanisms of action are still poorly understood. In this overview, we discuss published connexin targeting peptides, their reported mechanisms of action, and the potential for these molecules in the treatment of disease. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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10 pages, 3493 KiB

Open AccessReview

CO₂-Sensitive Connexin Hemichannels in Neurons and Glia: Three Different Modes of Signalling?

by Emily Hill, Nicholas Dale and Mark J. Wall

Int. J. Mol. Sci. 2021, 22(14), 7254; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22147254 - 06 Jul 2021

Cited by 2 | Viewed by 2204

Abstract

Connexins can assemble into either gap junctions (between two cells) or hemichannels (from one cell to the extracellular space) and mediate cell-to-cell signalling. A subset of connexins (Cx26, Cx30, Cx32) are directly sensitive to CO₂ and fluctuations in the level within a physiological range affect their open probability, and thus, change cell conductance. These connexins are primarily found on astrocytes or oligodendrocytes, where increased CO₂ leads to ATP release, which acts on P2X and P2Y receptors of neighbouring neurons and changes excitability. CO₂-sensitive hemichannels are also found on developing cortical neurons, where they play a role in producing spontaneous neuronal activity. It is plausible that the transient opening of hemichannels allows cation influx, leading to depolarisation. Recently, we have shown that dopaminergic neurons in the substantia nigra and GABAergic neurons in the VTA also express Cx26 hemichannels. An increase in the level of CO₂ results in hemichannel opening, increasing whole-cell conductance, and decreasing neuronal excitability. We found that the expression of Cx26 in the dopaminergic neurons in the substantia nigra at P7-10 is transferred to glial cells by P17-21, displaying a shift from being inhibitory (to neuronal activity) in young mice, to potentially excitatory (via ATP release). Thus, Cx26 hemichannels could have three modes of signalling (release of ATP, excitatory flickering open and shut and inhibitory shunting) depending on where they are expressed (neurons or glia) and the stage of development. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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25 pages, 1507 KiB

Open AccessReview

Connexins—Therapeutic Targets in Cancers

by Magdalena Nalewajska, Małgorzata Marchelek-Myśliwiec, Martyna Opara-Bajerowicz, Violetta Dziedziejko and Andrzej Pawlik

Int. J. Mol. Sci. 2020, 21(23), 9119; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms21239119 - 30 Nov 2020

Cited by 16 | Viewed by 3131

Abstract

Connexins (Cx) are members of a protein family that forms intercellular channels localised in gap junction (GJ) plaques and single transmembrane channels called hemichannels. They participate in intercellular communication or communication between the intracellular and extracellular environments. Connexins affect cell homeostasis, growth and differentiation by enabling the exchange of metabolites or by interfering with various signalling pathways. Alterations in the functionality and the expression of connexins have been linked to the occurrence of many diseases. Connexins have been already linked to cancers, cardiac and brain disorders, chronic lung and kidney conditions and wound healing processes. Connexins have been shown either to suppress cancer tumour growth or to increase tumorigenicity by promoting cancer cell growth, migration and invasiveness. A better understanding of the complexity of cancer biology related to connexins and intercellular communication could result in the design of novel therapeutic strategies. The modulation of connexin expression may be an effective therapeutic approach in some types of cancers. Therefore, one important challenge is the search for mechanisms and new drugs, selectively modulating the expression of various connexin isoforms. We performed a systematic literature search up to February 2020 in the electronic databases PubMed and EMBASE. Our search terms were as follows: connexins, hemichannels, cancer and cancer treatment. This review aims to provide information about the role of connexins and gap junctions in cancer, as well as to discuss possible therapeutic options that are currently being studied. Full article

(This article belongs to the Special Issue Therapeutic Potential of Targeting Connexins in Managing Disease Onset and Progression)

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