Retinoic acid regulates gap junction intercellular communication in human endometrial stromal cells through modulation of the phosphorylation status of connexin 43

Abstract

Previous studies revealed that gap junction intercellular communication (GJIC) among uterine stromal cells plays critical roles in modulating decidualization, neovasularization, and embryo implantation. Connexin (Cx) proteins are the major component of gap junctions and Cx43 is the most widely expressed connexin in endometrium. Phosphorylation of Cx43 was found to impair gap junction communication in this tissue. Using primary human endometrial stromal cells (ESCs) and a stable high telomerase-expressing ESC transfectant (T-HESC), we found that retinoic acid (RA) altered the phosphorylation status of Cx43 protein such that there was a decrease in the phosphorylated (P1 and P2) species accompanied by an increase in the non-phosphorylated (P0) form. This process is dependent on protein phosphatase 2A (PP2A) activity since selective PP2A inhibitors prevented the ability of RA to dephosphorylate Cx43. Although RA had no effect on total PP2A expression or activity, it significantly increased the intracellular association of Cx43 and PP2A. Inhibition of transcription and protein synthesis by actinomycin D and cycloheximide, respectively, had no effect on the RA-induced changes in the Cx43 phosphorylation pattern. Furthermore, BMS493, a potent antagonist of the classical RA-mediated transcriptional pathway, did not inhibit RA-induced Cx43 dephosphorylation. Our data indicate that RA stimulates physical association of PP2A with Cx43, resulting in the dephosphorylation of Cx43 and, as a consequence, up-regulation of GJIC in ESCs. This process is independent of new mRNA and protein synthesis and suggests a novel mechanism by which aberrant retinoid metabolism can explain certain reproductive disorders manifested by dysfunctional endometrial cell GJIC.

Inhibition of gap junctions relieves the hepatotoxicity of TNF-α

“…All three methods utilized in this study to functionally inhibit GJ function: high-and low-density cell culture, oleamide treatment, and siRNA transfection-mediated inhibition of Cx32 expression demonstrated that inhibiting the function of GJs led to a decrease in the hepatotoxicity of TNF-α. As a corollary, we also attempted to utilize retinoic acid (Wu et al, 2013) to enhance GJ function; however, after retinoic acid treatment, mass cell mortality occurred for unknown reasons. Therefore, the question of whether enhancement of the function of GJs can enhance the cytotoxicity of TNF-α to BRL-3A cells remains uncertain.…”

Influence of gap junction intercellular communication composed of connexin 43 on the antineoplastic effect of adriamycin in breast cancer cells

“…To determine whether GJIC was required for the increase of adriamycin-induced cytotoxicity in cells with GJIC, the breast cancer cells Hs578T and MCF-7 were treated with chemical modulators of GJs prior and during exposure to 6 µM adriamycin [inhibitor, oleamide (22,23) or 18-α-GA (24,25); potentiator, RA (21,26)] at high density (GJ formed) or low density (no GJ formed). Fig.…”

Physiological and pathological implications of retinoid action in the endometrium

“…This disease is characterized by the ectopic localization of endometrial-like tissues in the pelvic cavity, and its pathogenesis involves uncontrollable cell proliferation and is associated with local invasion and distant metastasis. Among the numerous aspects of endometrial behavior regulated by RA are matrix metalloproteinase secretion, gap junctional intracellular communication and the production of various cytokines involved in stromal cell growth, adhesion and differentiation (Wu et al 2013). Interleukin 6 (IL-6), monocyte chemotactic protein 1 (MCP-1), tumor necrosis factor α (TNF-α), vascular endothelial growth factor (VEGF) and connexin43 are all aberrantly expressed in endometriotic lesions (Sawatsri et al 2000, SharpeTimms 2001, Nozaki et al 2006, Sidell et al 2010, Wu et al 2013.…”

“…The expression of these same genes can be modulated by ovarian steroid hormones (i.e., estradiol and progesterone) treatment (Vermot et al 2000, Li & Ong 2003, Rühl et al 2006, Fritzsche et al 2007). The retinoid pathway is known to play vital roles in endometrial development and differentiation (Tanmahasamut & Sidell 2005, Wu et al 2013, Nakajima et al 2016, as well as endometrial neovascularization (Sidell et al 2010) and blastocyst implantation (Han et al 2010, Ma et al 2012.…”

“…Among the numerous aspects of endometrial behavior regulated by RA are matrix metalloproteinase secretion, gap junctional intracellular communication and the production of various cytokines involved in stromal cell growth, adhesion and differentiation (Wu et al 2013). Interleukin 6 (IL-6), monocyte chemotactic protein 1 (MCP-1), tumor necrosis factor α (TNF-α), vascular endothelial growth factor (VEGF) and connexin43 are all aberrantly expressed in endometriotic lesions (Sawatsri et al 2000, SharpeTimms 2001, Nozaki et al 2006, Sidell et al 2010, Wu et al 2013. Thus, many seemingly discordant features of endometriosis, including decreased cell death, increased growth and migration, inflammation and enhanced invasive properties of intraperitoneally seeded endometrial cells, might be accounted for by the dysregulation of RA signaling.…”