Beyond Gap Junction Channel Function: the Expression of Cx43 Contributes to Aldosterone-Induced Mesangial Cell Proliferation via the ERK1/2 and PKC Pathways

Abstract

Aims: This study aimed to explore the precise mechanism and signaling pathways of mesangial cell (MC) proliferation from a new point of view considering Connexin 43 (Cx43). Methods: MC proliferation was measured by the incorporation of 3H-thymidine (3H-TdR). Cx43 was over-expressed in MC cells using lipofectamine 2000, and the expression level was tested with reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analyses. The gap junction channel function was explored by Lucifer Yellow scrape loading and dye transfer (SLDT), and the intracellular calcium concentrations ([Ca²⁺]_i) were characterized by confocal microscopy on cells loaded with Fura-3/AM. Results: There was an inverse correlation between Cx43 expression and MC proliferation (P<0.05). SLDT studies revealed that there was no difference in the gap junction channel function between the normal and Aldosterone (Aldo)-stimulated groups (P>0.05). Our data also showed that the mineralcorticoid receptor (MR) antagonist spironolactone, ERK1/2 inhibitor PD98059 and PKC inhibitor GF109203X could attenuate the down-regulation of Cx43 expression in Aldo-induced MC proliferation; however, the PI3K inhibitor LY294002 could block MC proliferation without affecting Cx43 expression at either the mRNA or protein level. In addition, Aldo promoted MC proliferation in parallel with increasing [Ca²⁺]_i (P<0.05), suggesting that the classical PKC pathway might be activated. Conclusions: Our study provides preliminary evidence that Cx43 is an important regulator of Aldo-promoted MC proliferation. Furthermore, reduced Cx43 expression promoted MC proliferation independent of the gap junction channel function, and this process might be mediated through the ERK1/2- and PKC-dependent pathways.

Connexin 43: a New Therapeutic Target Against Chronic Kidney Disease

“…While the podocyte expression of Cx43 increased in animals with GN, it decreased in mesangial cells, showing that Cx43 upregulation is not a universal response to injury in all renal cell populations [64]. In agreement with this study, exposure of cultured mesangial cells to high glucose [65] or aldosterone [66] decreased the expression of Cx43 and induced cytoskeletal rearrangements and proliferation, respectively. The effect of Cx43 on mesangial proliferation, one of the basic histological findings in IgA nephropathy, was attributed to the function of Cx43 hemichannels, since aldosterone had no effect on gap junction formation.…”

“…The effect of Cx43 on mesangial proliferation, one of the basic histological findings in IgA nephropathy, was attributed to the function of Cx43 hemichannels, since aldosterone had no effect on gap junction formation. Transient overexpression of Cx43 abolished the proproliferating effect of aldosterone possibly via Ca +2 /Protein kinase C-related signaling [66]. Treatment of mesangial cells with high glucose, a commonly used in vitro model to mimic diabetic nephropathy, resulted in RhoA/ROCK-mediated cytoskeletal rearrangements, translocation of NF-κB to the nucleus, F-actin accumulation and an enhanced interaction between zonula occludens-1 (ZO-1) and Cx43.…”

Cryptococcus Neoformans Infection and Immune Cell Regulation in Human Monocytes

“…qRT-PCR was performed to verify the data obtained by RNA-seq using SYBR Green (TaKaRa, Japan) using the ABI 7500 SDS system (Applied Biosystems, USA) as previously described [28][29][30]. Primer sequences (5'-3', forward and reverse) used in the present study are as follows: CXCL1, F-TGC CAG CCA CTG TGA TAGAG; R-AGC CCC TTT GTT CTA AGC CAG; CXCL2, F-AGA TCA ATG TGA CGG CAGGG; R-TCT CTG CTC TAA CAC AGA GGGA; CXCL3, F-CGC CCA AAC CGA AGT CATAG; R-GCT CCC CTT GTT CAG TAT CTTTT; CXCL5, F-TCT GCA AGT GTT CGC CATAG; R-TGT CTT CCC TGG GTT CAGAG; CCL1, F-CTC ATT TGC GGA GCA AGA GAT; R-GCC TCT GAA CCC ATC CAA CTG; CCL4, F-GCT TTT CTT ACA CTG CGA GGA; R-CCA GGA TTC ACT GGG ATCAG; CCL7, F-AGA AGG ACC ACC AGT AGCCA; R-ACT AGT CCT ATT CCC TGC CCA; CCR2, F-TAC GGT GCT CCC TGT CAT AAA; R-TAA GAT GAG GAC GAC CAG CAT; IL1B, F-ATG ATG GCT TAT TAC AGT GGCAA; R-GTC GGA GAT TCG TAG CTGGA.…”

Effect of Huaier On the Proliferation of Mesangial Cells in Anti-Thy-1 Nephritis

“…Mesangial cell proliferation plays an important role in progressive glomerular injury that leads to glomerular sclerosis [1]. Although MsPGN is a major cause of end-stage renal failure in many countries, its underlying mechanisms are not fully understood [2, 3]. MsPGN is of two types: IgA nephropathy and non-IgA nephropathy.…”