EXPRESSION OF TRANSFORMING GROWTH-FACTOR-BETA (TGF-BETA) ISOFORMS AND TGF-BETA TYPE-II RECEPTOR MESSENGER-RIBONUCLEIC-ACID AND PROTEIN, AND THE EFFECT OF TGF-BETA-S ON ENDOMETRIAL STROMAL CELL-GROWTH AND PROTEIN-DEGRADATION IN-VITRO

Reverse transcription-polymerase chain reaction analysis of total RNA and immunocytochemical observations revealed that human endometrial glandular epithelial and stromal cells in primary culture express messenger RNAs and proteins for transforming growth factor-beta 1 (TGF beta 1), TGF beta 2, and TGF beta 3 as well as TGF beta type II receptor. The epithelial and stromal cells synthesize and secrete into their culture-conditioned medium 2.6 +/- 0.3 and 1.4 +/- 0.2 ng TGF beta 1/10(6) cells, respectively; after transient acidification of the medium, the TGF beta 1 levels were 18.1 +/- 0.4 and 7.8 +/- 0.7 ng/10(6) cells. These cells also contain specific binding sites for [I-125]TGF beta 1, indicated by light microscope autoradiography. TGF beta s at 0.01-10 ng/ml neither stimulated or inhibited subconfluent quiescent stromal cells under serum-free condition nor altered the mitogenic action of 10% fetal bovine serum. However, in the presence of 2% fetal bovine serum, which induced half-maximal stimulation of [H-3]thymidine incorporation, TGF beta 1 and TGF beta 2 at 0.1-0.5 ng/ml and TGF beta 3 at 0.1-2.5 ng/ml significantly stimulated the rate of [H-3]thymidine incorporation into quiescent stromal cells (P < 0.005); they were ineffective at higher concentrations. TGF beta s did not have any effect on cell proliferation, as determined by cell counting; however, at 0.1 ng/ml and higher concentrations, TGF beta s significantly reduced the metabolic activity of stromal cells, as determined by colorimetric 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide assay (P < 0.05). The stimulatory and inhibitory actions of TGF beta s in both assays were reversible using 5-10 mu g/ml TGF beta 1- and TGF beta 2- and 3-6 mu g/ml TGF beta 3-specific neutralizing antibodies. TGF beta 1 at 1 ng/ml had no significant effect on long-lived protein degradation, assayed by incorporation of [C-14]valine into newly synthesized protein by stromal cells, and was similar to the effect of epidermal growth factor or platelet-derived growth factor-BB (10 ng/ml). The data suggest that the TGF beta expression by various endometrial cell types in an autocrine/paracrine manner acts as a negative regulator essential for restraining endometrial growth and transition from proliferation to differentiation stages during the secretory phase after mitogenic stimulation during the proliferative phase of the menstrual cycle.