421. Seminal fluid TGFβ regulates follistatin mRNA expression human Ect1 cervical epithelial cells

Abstract

Introduction of seminal fluid into the female reproductive tract following coitus stimulates a local inflammatory response. Inflammatory leukocyte recruitment is regulated by induction of cytokine and chemokine synthesis in female tract epithelial cells by seminal fluid signalling agents. Affymetrix microarray analysis in immortalised ectocervical epithelial (Ect1) cells identified the potent anti-inflammatory cytokine follistatin (FST) as the most strongly differentially expressed gene, with a ~12-fold increase in mRNA expression induced by seminal fluid. Follistatin has recently been implicated as a key cytokine in early pregnancy by studies in female follistatin null mice, which exhibit infertility as a consequence of failure to resolve the uterine post-mating inflammatory response. The aim of this study was to investigate seminal plasma regulation of follistatin in human Ect1 cervical cells, and to examine the role of the major active seminal fluid constituent, TGFβ, in controlling Ect1 cells follistatin mRNA expression. To confirm Affymetrix findings, qRT–PCR experiments were undertaken in Ect1 cells incubated with 10% pooled human seminal plasma (SP). Primers specific for the tissue bound isoform of follistatin (FST288) as well as both FST288 and the circulating 315 isoforms (FSTall) were used. Ect1 cell incubation with 10%SP elicited 3.8-fold and 4-fold increases in FST288 and FSTall respectively. Incubation of Ect1 cells with TGFβ1, TGFβ2 and TGFβ3 showed differential effects of the three isoforms, with rTGFβ2 inducing FST288 and FSTall, while rTGFβ1 and TGFβ3 exerted little effect.. These results suggest that seminal plasma induces follistatin synthesis after coitus and that TGFβ2 is at least partly responsible for this effect. Follistatin induced by seminal fluid may act to limit the course of inflammation after intercourse, and thereby prevent uncontrolled inflammatory damage. Follistatin induced in the female tissues would be augmented by follistatin delivered from the male, since human seminal plasma also contains a high concentration of this cytokine.