Connexin 43 Hemichannels Regulate the Expression of Wound Healing-Associated Genes in Human Gingival Fibroblasts

Tarzemany, Rana; Jiang, Guoqiao; Jiang, Jean X.; Larjava, Hannu; Häkkinen, Lari

doi:10.1038/s41598-017-12672-1

Cited by 46 publications

(56 citation statements)

References 79 publications

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“…After stimulation, AS HBFs exhibited different expression patterns of some proteins compared to NA HBFs. The most important and notable differences are amplified levels of α-SMA and connexin (Cx) 43 (protein involved in the intercellular transfer of small metabolites and ions via hexameric channels termed gap junctions) [ 236 , 237 ] in response to TGF-β administration in AS HBFs compared to NA HBFs (Fig. 1 —upregulation of α-SMA and Cx43) [ 46 , 223 , 234 , 238 ].…”

Section: Fibroblast Featuresmentioning

confidence: 99%

Fibroblast-to-myofibroblast transition in bronchial asthma

Michalik

Wójcik-Pszczoła

Paw

et al. 2018

Cell. Mol. Life Sci.

122

118

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Bronchial asthma is a chronic inflammatory disease in which bronchial wall remodelling plays a significant role. This phenomenon is related to enhanced proliferation of airway smooth muscle cells, elevated extracellular matrix protein secretion and an increased number of myofibroblasts. Phenotypic fibroblast-to-myofibroblast transition represents one of the primary mechanisms by which myofibroblasts arise in fibrotic lung tissue. Fibroblast-to-myofibroblast transition requires a combination of several types of factors, the most important of which are divided into humoural and mechanical factors, as well as certain extracellular matrix proteins. Despite intensive research on the nature of this process, its underlying mechanisms during bronchial airway wall remodelling in asthma are not yet fully clarified. This review focuses on what is known about the nature of fibroblast-to-myofibroblast transition in asthma. We aim to consider possible mechanisms and conditions that may play an important role in fibroblast-to-myofibroblast transition but have not yet been discussed in this context. Recent studies have shown that some inherent and previously undescribed features of fibroblasts can also play a significant role in fibroblast-to-myofibroblast transition. Differences observed between asthmatic and non-asthmatic bronchial fibroblasts (e.g., response to transforming growth factor β, cell shape, elasticity, and protein expression profile) may have a crucial influence on this phenomenon. An accurate understanding and recognition of all factors affecting fibroblast-to-myofibroblast transition might provide an opportunity to discover efficient methods of counteracting this phenomenon.

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Section: Fibroblast Featuresmentioning

confidence: 99%

Fibroblast-to-myofibroblast transition in bronchial asthma

Michalik

Wójcik-Pszczoła

Paw

et al. 2018

Cell. Mol. Life Sci.

122

118

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“…Moreover, in our SL/DT assay protocol, the cells are washed with CaMg-PBS prior the loading of dyes dissolved also in CaMg-PBS, which should inhibit hemichannel activity. It has been demonstrated that the same in vitro model can be used for evaluation of both hemichannel activity by dye uptake assay, and GJIC by SL-DT assay [42][43][44] . Interestingly, some chemicals can regulate hemichannels and gap junction channels differently.…”

Section: Discussionmentioning

confidence: 99%

Improved multiparametric scrape loading-dye transfer assay for a simultaneous high-throughput analysis of gap junctional intercellular communication, cell density and viability

Dydowiczová

Brózman

Babica

et al. 2020

Sci Rep

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Gap junctional intercellular communication (GJic) is a vital cellular process required for maintenance of tissue homeostasis. In vitro assessment of GJic represents valuable phenotypic endpoint that could be effectively utilized as an integral component in modern toxicity testing, drug screening or biomedical in vitro research. However, currently available methods for quantifying GJic with higher-throughputs typically require specialized equipment, proprietary software and/or genetically engineered cell models. to overcome these limitations, we present here an innovative adaptation of traditional, fluorescence microscopy-based scrape loading-dye transfer (SL-DT) assay, which has been optimized to simultaneously evaluate GJic, cell density and viability. this multiparametric method was demonstrated to be suitable for various multiwell microplate formats, which facilitates an automatized image acquisition. The assay workflow is further assisted by an open source-based software tools for batch image processing, analysis and evaluation of GJic, cell density and viability. our results suggest that this approach provides a simple, fast, versatile and cost effective way for in vitro high-throughput assessment of GJic and other related phenotypic cellular events, which could be included into in vitro screening and assessment of pharmacologically and toxicologically relevant compounds. Gap junctional intercellular communication (GJIC) allows an exchange of low molecular weight molecules (<1.2 kDa) between adjacent cells in both vertebrates and invertebrates 1. GJIC is mediated through intercellular channels build from connexin proteins in vertebrates and from innexin proteins in invertebrates 1,2. GJIC plays a central role in coordinating cell-to-cell communication and integration of signal transduction pathways controlling gene expression and cell behaviour in order to receive coordinated and collective responses from the cells across a tissue of multicellular organism 3,4. Dysregulation of GJIC and GJIC-dependent signal integration, for example by drugs or environmental contaminants, can disrupt the normal homeostatic control of a cell behaviour and lead to numerous adverse outcomes such as cardiovascular 5 , pulmonary 6 or reproductive system 7,8 diseases and cancer 4,9,10. On the other hand, timed and targeted upregulation or downregulation of GJIC, connexin channel or hemichannel activity, induced by pharmacological agents, is currently widely discussed as potential therapeutic approach for treatment of various diseases 10-12. Accordingly, GJIC can be effectively utilized in the modern toxicological and pharmacological research, and GJIC analysis could be a valuable component of any biological study. However, in vitro assessment of GJIC has been used relatively less frequently in the modern research in comparison to other and more traditional phenotypic assays, such as evaluation of cellular metabolic activity, cell proliferation, cell cycle, autophagy, apoptosis, cell migration, cytoskeletal rearangements 13. This m...

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“…Recent studies using Gap27 have also shown that this peptide is effective in improving rabbit corneal wound healing [ 39 ] and in primary human gingival fibroblasts, isolated from donors aged 26–48 years of age [ 29 ]. A further peptide TAT-Gap19, has also recently been reported to enhance scrape wound closure of human gingival fibroblasts [ 40 ]. TAT-Gap19, targeted to the intracellular loop of Cx43, was designed as a cell permeant peptide and has been extensively used to characterise interactions between the intracellular loop of Cx43 and the carboxy terminal tail [ 41 , 42 ].…”

Section: Discussionmentioning

confidence: 99%

“…This re-enforces our previous findings where irradiated juvenile fibroblasts were used and Gap27 effectively enhanced wound closure [ 22 ]. Hence in JFF cells and keratinocytes proliferation factors are not involved in the enhanced cell migration response, suggesting hemichannel signalling plays an important role in co-ordinating cellular responses [ 30 , 40 ].…”

Section: Discussionmentioning

confidence: 99%

“…Modifying Cx43 expression or function by antisense oligonucleotides and peptide αCTI have also been associated with alterations in ECM deposition [ 20 , 33 ]. Further studies by Tarzemany et al systematically reviewed the impact of Gap27 and more recently TAT-Gap19 on gene expression and cell signalling pathways involved in wound closure events in human adult gingival fibroblasts [ 29 , 40 ]. Both peptides effectively enhanced wound closure rates and gene array analysis after 24 h indicated changes in expression of a panel of genes related to ECM deposition including a number of MMP proteins and CTGF, in agreement with our previous studies on JFF cells [ 23 ].…”

Section: Discussionmentioning

confidence: 99%

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The Connexin Mimetic Peptide Gap27 and Cx43-Knockdown Reveal Differential Roles for Connexin43 in Wound Closure Events in Skin Model Systems

Faniku

O'Shaughnessy

Lorraine

et al. 2018

IJMS

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In the epidermis, remodelling of Connexin43 is a key event in wound closure. However, controversy between the role of connexin channel and non-channel functions exist. We compared the impact of SiRNA targeted to Connexin43 and the connexin mimetic peptide Gap27 on scrape wound closure rates and hemichannel signalling in adult keratinocytes (AK) and fibroblasts sourced from juvenile foreskin (JFF), human neonatal fibroblasts (HNDF) and adult dermal tissue (ADF). The impact of these agents, following 24 h exposure, on GJA1 (encoding Connexin43), Ki67 and TGF-β1 gene expression, and Connexin43 and pSmad3 protein expression levels, were examined by qPCR and Western Blot respectively. In all cell types Gap27 (100 nM–100 μM) attenuated hemichannel activity. In AK and JFF cells, Gap27 (100 nM–100 μM) enhanced scrape wound closure rates by ~50% but did not influence movement in HNDF or ADF cells. In both JF and AK cells, exposure to Gap27 for 24 h reduced the level of Cx43 protein expression but did not affect the level in ADF and HNDF cells. Connexin43-SiRNA enhanced scrape wound closure in all the cell types under investigation. In HDNF and ADF, Connexin43-SiRNA enhanced cell proliferation rates, with enhanced proliferation also observed following exposure of HDNF to Gap27. By contrast, in JFF and AK cells no changes in proliferation occurred. In JFF cells, Connexin43-SiRNA enhanced TGF-β1 levels and in JFF and ADF cells both Connexin43-SiRNA and Gap27 enhanced pSmad3 protein expression levels. We conclude that Connexin43 signalling plays an important role in cell migration in keratinocytes and foreskin derived fibroblasts, however, different pathways are evoked and in dermal derived adult and neonatal fibroblasts, inhibition of Connexin43 signalling plays a more significant role in regulating cell proliferation than cell migration.

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Connexin 43 Hemichannels Regulate the Expression of Wound Healing-Associated Genes in Human Gingival Fibroblasts

Cited by 46 publications

References 79 publications

Fibroblast-to-myofibroblast transition in bronchial asthma

Fibroblast-to-myofibroblast transition in bronchial asthma

Improved multiparametric scrape loading-dye transfer assay for a simultaneous high-throughput analysis of gap junctional intercellular communication, cell density and viability

The Connexin Mimetic Peptide Gap27 and Cx43-Knockdown Reveal Differential Roles for Connexin43 in Wound Closure Events in Skin Model Systems

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