Stimulation of syncytium formation in vitro in human trophoblast cells by galectin-1.
BACKGROUND
Galectin-1 (gal-1), a member of the mammalian beta-galactoside-binding proteins, exerts biological effects by recognition of glycan ligands, including those involved in cell adhesion and growth regulation. In trophoblast cells, gal-1 binds to cell surface glycoproteins (e.g., Mucin1). It has been demonstrated that gal-1 recognizes appropriate glycotopes on the syncytiotrophoblast and extravillous trophoblast from second trimester human placenta and choriocarcinoma cells BeWo, which reveal two coexisting phenotypes, the cytotrophoblast-like and the syncytiotrophoblast-like phenotype. So the aim of this study was to investigate the effect of gal-1 on syncytium formation in BeWo and human villous trophoblasts (HVT) cells.


METHODS
The effect of gal-1 on syncytium formation was investigated with immunocytochemical and double immunofluorescence stainings, cell-labelling and Real-time RT-PCR. BeWo choriocarcinoma and HVT cells were incubated in vitro for 24 and 48 h in the absence (controls) and presence of gal-1 and forskolin and stained with antibodies against Ki67, beta-catenin, E-cadherin and syncytin. BeWo and HVT cells were incubated with 60 microg/ml gal-1 for 48 h (BeWo) or 96 h (HVT) and cell fusion was detected by fluorescent cell-labelling solution. Finally, BeWo cells were incubated for 1 h or 48 h in the absence and presence of 60 microg/ml gal-1 and Real-time RT-PCR was performed.


RESULTS
We showed with immunocytochemical staining a downregulation of beta-catenin expression in the 24 h BeWo cell culture and with double immunofluorescence staining an inhibition of the beta-catenin and E-cadherin expression in the 48 h BeWo cell culture stimulated with gal-1 or forskolin. The inhibition of E-cadherin was demonstrated on mRNA level in the 1 h BeWo cell culture too. Increased cell fusion was also showed with DiO and DiI fluorescent cell-labelling solution in the 48 h BeWo cell culture. In addition, we demonstrated the downregulation of Ki67 protein expression in the 24 h BeWo cell culture and on mRNA level in the 1 h BeWo cell culture. We also showed the upregulation of syncytin protein and mRNA expression after incubation of the 48 h BeWo cell culture with gal-1 or forskolin. Similar results were obtained with HVT cells: the amount of cell fusion was significantly increased in the gal-1 treated 48 h HVT cell culture in vitro compared to untreated cells as demonstrated with beta-catenin and E-cadherin double immunofluorescence staining. This increase was also shown by fluorescent cell-labelling with DiO and DiI in the 96 h HVT cell culture compared to untreated cells.


CONCLUSIONS
Our data suggest that gal-1 stimulates the syncytium formation in choriocarcinoma cells BeWo and HVT cells in vitro and inhibits the expression of beta-catenin, E-cadherin and in addition Ki67 in BeWo cells. Therefore gal-1 may be a major trigger for the process of trophoblast cell fusion.
Fischer
I
I
Ludwig Maximilians University of Munich, Department of Obstetrics and Gynecology, Maistrasse 11, 80337 Munich, Germany.
Redel
S
S
Hofmann
S
S
Kuhn
C
C
Friese
K
K
Walzel
H
H
Jeschke
U
U
D015820:Cadherins / Q000037:antagonists & inhibitors; D002459:Cell Fusion; D045744:Cell Line, Tumor; D005576:Colforsin / Q000494:pharmacology; D005260:Female; D037483:Galectin 1 / Q000494:pharmacology* / Q000502:physiology; D015686:Gene Products, env / Q000096:biosynthesis; D015726:Giant Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D011257:Pregnancy Proteins / Q000096:biosynthesis; D014327:Trophoblasts / Q000166:cytology*
D015820:Cadherins; D037483:Galectin 1; D015686:Gene Products, env; D011257:Pregnancy Proteins; C405335:syncytin; D005576:Colforsin