Blocking Connexin-43 mediated hemichannel activity protects against early tubular injury in experimental chronic kidney disease

Price, G, Chadjichristos, CE, Kavvadas, P , Tang, SCW, Yiu, WH, Green, CR, Potter, JA, Siamantouras, E, Squires, P and Hills, C (2020) Blocking Connexin-43 mediated hemichannel activity protects against early tubular injury in experimental chronic kidney disease. Cell communication and signalling, 18 (79). ISSN 1478-811X

Full content URL: https://doi.org/10.1186/s12964-020-00558-1

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Blocking Connexin-43 mediated hemichannel activity protects against early tubular injury in experimental chronic kidney disease
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Abstract

Background: Tubulointerstitial fibrosis represents the key underlying pathology of Chronic Kidney Disease (CKD), yet treatment options remain limited. In this study, we investigated the role of connexin43 (Cx43) hemichannel-mediated adenosine triphosphate (ATP) release in purinergic-mediated disassembly of adherens and tight junction complexes in early tubular injury.
Methods: Human primary proximal tubule epithelial cells (hPTECs) and clonal tubular epithelial cells (HK2) were treated with Transforming Growth Factor Beta1 (TGFβ1) ± apyrase, or ATPγS for 48h. For inhibitor studies, cells were co-incubated with Cx43 mimetic Peptide 5, or purinergic receptor antagonists Suramin, A438079 or A804598. Immunoblotting, single-cell force spectroscopy and trans-epithelial electrical resistance assessed protein expression, cell-cell adhesion and paracellular permeability. Carboxyfluorescein uptake and biosensing measured hemichannel activity and real-time ATP release, whilst a heterozygous Cx43+/- mouse model with unilateral ureteral obstruction (UUO) assessed the role of Cx43 in vivo.
Results: Immunohistochemistry of biopsy material from patients with diabetic nephropathy confirmed increased expression of purinergic receptor P2X7. TGFβ1 increased Cx43 mediated hemichannel activity and ATP release in hPTECs and HK2 cells. The cytokine reduced maximum unbinding forces and reduced cell-cell adhesion, which translated to increased paracellular permeability. Changes were reversed when cells were co-incubated with either Peptide 5 or P2-purinoceptor inhibitors. Cx43+/- mice did not exhibit protein changes associated with early tubular injury in a UUO model of fibrosis.
Conclusion: Data suggest that Cx43 mediated ATP release represents an initial trigger in early tubular injury via its actions on the adherens and tight junction complex. Since Cx43 is highly expressed in nephropathy, it represents a novel target for intervention of tubulointerstitial fibrosis in CKD.

Keywords:chronic kidney disease, connexins, hemichannels, ATP, cell adhesion, tubular injury
Subjects:C Biological Sciences > C710 Applied Molecular Biology, Biophysics and Biochemistry
C Biological Sciences > C700 Molecular Biology, Biophysics and Biochemistry
Divisions:College of Science > School of Life Sciences
ID Code:40866
Deposited On:21 May 2020 13:45

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