TLR4 Signaling Is a Major Mediator of the Female Tract Response to Seminal Fluid in Mice1

Abstract

Seminal fluid interacts with epithelial cells lining the female reproductive tract to induce expression of proinflammatory cytokines and chemokines, initiating immune tolerance mechanisms to facilitate pregnancy. TGFB cytokines are key signaling agents in seminal plasma but do not fully account for the female response to seminal fluid. We hypothesized that additional molecular pathways are utilized in seminal fluid signaling. Affymetrix microarray was employed to compare gene expression in the endometrium of mice 8 h after mating with either intact males or seminal fluid deficient (SVX/VAS) males. Bioinformatics analysis revealed TLR4 signaling as a strongly predicted upstream regulator activated by the differentially expressed genes and implicated TGFB signaling as a second key pathway. Quantitative PCR and microbead data confirmed that seminal fluid induces endometrial synthesis of several TLR4-regulated cytokines and chemokines, including CSF3, CXCL1, CXCL2, IL1A, IL6, LIF, and TNF. In primary uterine epithelial cells, CSF3, CXCL1, and CXCL2 were strongly induced by the TLR4 ligand LPS but suppressed by TGFB, while IL1A, TNF, and CSF2 were induced by both ligands. TLR4 was confirmed as essential for the full endometrial cytokine response using mice with a null mutation in Tlr4, where seminal fluid failed to induce endometrial Csf3, Cxcl2, Il6, and Tnf expression. This study provides evidence that TLR4 contributes to seminal fluid modulation of the periconception immune environment. Activation of TLR4 signaling by microbial or endogenous components of seminal fluid is thus implicated as a key element of the female tract response to seminal fluid at the outset of pregnancy in mice.

A role for seminal plasma in modulating pregnancy outcomes in domestic species

“…The importance of toll-like receptor (TLR) 4 signaling has been implicated in SP-induced proinflammatory cytokine expression. In a similar fashion to TLR4 involvement during cumulus oocyte-complex (COC) expansion where endogenous TLR4 ligands induce the expression of proinflammatory mediators important for COC expansion, such as interleukin (IL) 6 (Shimada et al 2008), SP modulates TLR4-induced cytokines like colony-stimulating factor 2 (CSF2) (Shimada et al 2008, Schjenken et al 2015. The importance of SP exposure on embryonic development has also been demonstrated in mice and hamsters (Peitz & Olds-Clarke 1986, O et al 1988.…”

Seminal Fluid Signalling in the Female Reproductive Tract: Implications for Reproductive Success and Offspring Health

“…The molecular basis of the female response to seminal fl uid mostly comes from laboratory studies using the mouse as a model (Robertson 2005 ;Robertson et al 1996 ;Schjenken et al 2015 ). At coitus in the mouse, seminal fl uid is deposited in the uterus and disseminates throughout the female reproductive tract, where it interacts with epithelial cells of the uterine lumen to activate gene expression and synthesis of several proinfl ammatory cytokines and chemokines.…”

“…More recently, several other cytokines and chemokines including interleukin-6 (IL6), tumor necrosis factor (TNF), C-X-C motif chemokine ligand-1 (CXCL1, also known as growth regulated alpha protein, GRO or KC), CXCL2 (macrophage infl ammatory protein-2, MIP2), C-C motif chemokine ligand-3 (CCL3, macrophage infl ammatory protein-1 alpha, MIP1A), and granulocyte colony-stimulating factor (GCSF) have been identifi ed as elevated in the mouse uterus during the immediate post-coital phase (Schjenken et al 2015 ;Sanford et al 1992 ;Robertson et al 1998 ;Pollard et al 1998 ;Johansson et al 2004 ). Seminal plasma is required for much of this response as coitus with males rendered seminal fl uid defi cient, by surgery to remove seminal vesicles and vasectomy, results in either no induction or substantially reduced cytokine induction (Schjenken et al 2015 ). In contrast, delivery of seminal plasma by vasectomised males induces a response comparable to that induced by intact males (Robertson et al 1996 ).…”

The Effect of G-CSF on Infertile Women Undergoing IVF Treatment

“…At the same time, we think that more detailed analysis is needed in order to explore the mechanisms. Schjenken et al and Sharkey et al suggested the levels of G-CSF increased during the interaction of seminal fluid and epithelial cells, further contributing to the conception [4,5]. Meanwhile, as for the patients undergoing assisted reproduction treatment, the better clinical pregnancy outcomes could be observed by adding the G-CSF to embryo culture medium in some extent [6].…”