Soluble Betaglycan and Tgf-Β Signaling in Human Endometrial and Endometriotic Cells

Abstract

Endometriosis is characterized by presence of endometrial-like tissue outside the uterine cavity. The TGF-β superfamily, consisting of TGF-βs, activins and inhibins, is expressed in the endometrium and is putatively implicated in endometriosis. Betaglycan (BG), a membrane-bound co-receptor and modulator of the TGF-β superfamily ligands, especially of TGF-β2. BG undergoes proteolytic cleavage to release soluble betaglycan (sBG) which often blocks TGF-βs in several physiological and pathological processes. We investigated shedding of betaglycan and its role in TGF-β family signaling in endometrial and endometriotic cell lines. Endometriotic epithelial (12Z) and endometrial stromal (THESC) cells were treated with increasing concentrations of TGF-β1/β2 (1-10 ng/ml), activin A (5-50 ng/ml) or inhibin A (5-50 ng/ml). The level of sBG was evaluated using ELISAs after 24, 48 and 72 hours of stimulation. An ALK-4/5 inhibitor (LY364947, 10 μM) was used to study the signaling pathways. Inhibition of BG shedding was analyzed using tissue inhibitors of metalloproteinases3 (TIMP3, 2.5-10 nM) besides a pan-MMP inhibitor (GM6001, 10 μM). TGF-β1/β2 along with activin A and inhibin A stimulation of 12Z and THESC cells resulted in a significant time- and dose-dependent reduction in BG shedding. Moreover, TGF-β1/2 and activin A-mediated reduction in BG shedding in 12Z cells was found to be TGF-β type I receptor (ALK-5) and activin receptor type-1B (ALK-4)-dependent, respectively. Shedding of BG was significantly attenuated by TIMP3 in a dose- dependent manner and partially (~40%) by the pan-MMP inhibitor, signifying the involvement of matrix metalloproteinases in BG shedding. Collectively, our data suggest involvement of MMPs in shedding of BG and suggest potential involvement of sBG in modulating TGF-β1/β2, activin A and inhibin A signaling in endometriotic and endometrial cells. The signaling mechanisms involved and possible roles in endometriosis merit further investigation.