Gap junctional communication during human trophoblast differentiation: influence of human chorionic gonadotropin.

Abstract

During pregnancy, the trophoblast develops from the fusion of cytotrophoblastic cells into a syncytiotrophoblast. As the exchange of molecules through gap junctions is considered to play a role in the control of cell and tissue differentiation, the cell to cell diffusion of a fluorescent dye was investigated in human trophoblastic cells differentiating in culture. The fluorescence recovery after photobleaching technique was used to estimate the transfer of 6-carboxyfluorescein from contiguous cellular elements into photobleached cells. Fluorescence recovery follows a slow exponential time course when the cell to cell exchange process is rate limited by the presence of gap junctional channels between contiguous cells, contrasting with a much faster step-like course in the case of fusion of the plasma membranes. In the presence of 10% fetal calf serum, Percoll-purified cytotrophoblastic cells develop into cellular aggregates, then into a syncytium, within 24-48 h after plating. During this in vitro differentiation, fluorescence recoveries after photobleaching with a time course typical for gap junctions were observed between aggregated cytotrophoblastic cells, between cytotrophoblastic cells and syncytiotrophoblasts, and between contiguous syncytiotrophoblasts. The maximum percentage of gap junctional coupling occurs on the fourth day. This fluorescence recovery is attributed to the diffusion of dye through gap junctions, because it can be interrupted by exposure to a known junctional uncoupler (3 mM heptanol). The effects of hCG on this gap junctional communication during trophoblast differentiation were investigated. In the presence of 500 mIU/ml hCG in the culture medium, the percentage of coupled cells was increased at all stages of culture, and the highest proportion of coupled cells was observed after 2 days of culture vs. 4 days in control medium. Moreover, the diffusion rate constant k (the inverse value of the time constant measured on recovery curves) was also significantly increased in the presence of chorionic hormone. It is concluded that during trophoblast differentiation, the development of a cell to cell communication through gap junctions precedes the formation of a morphological syncytium by cell fusion. This gap junctional communication is promoted by hCG. Furthermore, our study confirms the differentiating role and the autocrine action of hCG in the physiology of the trophoblast.

The gap junction protein Cx43 is involved in the bone-targeted metastatic behaviour of human prostate cancer cells

“…The degree of homocellular GJIC between PCa cells was also measured by means of the gap-FRAP (Fluorescence recovery after photobleaching) method [34]. Briefly, cultured PCa cells were loaded with a diffusible fluorescent tracer (7 lg/ml 6-carboxyfluorescein, Sigma) and the fluorescence of investigated cells was dramatically decreased by a laser beam (photobleaching at 405 nm) in order to study the recovery by diffusion from adjacent cells and to measure the diffusion rate constant k.…”

A Positive Feedback Loop between Glial Cells Missing 1 and Human Chorionic Gonadotropin (hCG) Regulates Placental hCGβ Expression and Cell Differentiation

“…A G-protein-coupled receptor has been identified as the receptor for hCG and LH. When hCG binds to its receptor, the coupled G protein(s) activates adenylyl cyclase, leading to an increase in the concentration of intracellular cyclic AMP (cAMP) that activates protein kinase A (PKA) (5,7,8).The expression profile of hCG during pregnancy is physiologically relevant to the rate of placental growth and the degree of syncytiotrophoblast differentiation. Human placenta is composed of villous tissues, of which the outer surface is a multinucleated syncytiotrophoblast (STB) layer overlying mononucleated cytotrophoblasts (CTBs) (9).…”

“…A G-protein-coupled receptor has been identified as the receptor for hCG and LH. When hCG binds to its receptor, the coupled G protein(s) activates adenylyl cyclase, leading to an increase in the concentration of intracellular cyclic AMP (cAMP) that activates protein kinase A (PKA) (5,7,8).…”

Physiological roles of connexins and pannexins in reproductive organs

“…4d-f) [110,[172][173][174][175]. A recent study supports a central role for Cx43 in mediating both GJIC and proteinprotein interactions via the transcription factor Drosophila glial cell missing gene 1 and its downstream target human endogenous retrovirus envelope protein/syncytin-1 with its receptor neutral amino acid transporter B(0) to stimulate trophoblast cell fusion [176].…”