Elevated concentration of TNF-alpha induces trophoblast differentiation in mouse blastocyst outgrowths.
Elevated expression of tumour necrosis factor-alpha (TNF-alpha) is associated with adverse pregnancy outcome. This study has examined the expression of TNF-alpha and its receptors (TNF-Rs) by mouse blastocysts and blastocyst outgrowths from day 4 to 9.5 of pregnancy and investigated the effects of elevated TNF-alpha on the inner cell mass (ICM) and trophoblast cells of blastocyst outgrowths. RT-PCR demonstrated TNF-alpha mRNA expression from day 7.5 to 9.5, TNF-R1 from day 6.5 to 9.5 and TNF-R2 from day 5.5 to 7.5 of pregnancy, and in situ hybridisation revealed the trophoblast giant cells (TGCs) of the early placenta as the site of TNF-alpha expression. Day 4 blastocysts were cultured in a physiologically high concentration of TNF-alpha (100 ng/ml) for 72 h to the outgrowth stage and then compared to blastocysts cultured in media alone. TNF-alpha-treated blastocyst outgrowths exhibited a significant reduction in ICM cells (mean +/- SD 23.90+/-10.42 vs 9.37+/-7.45, t-test, P<0.0001) with no significant change in the numbers of trophoblast cells (19.97+/-8.14 vs 21.73+/-7.79, t-test, P=0.39). Within the trophoblast cell population, the TNF-alpha-treated outgrowths exhibited a significant increase in multinucleated cells (14.10+/-5.53 vs 6.37+/-5.80, t-test, P<0.0001) and a corresponding significant decrease in mononucleated cells (5.87+/-3.60 vs 15.37+/-5.87, t-test, P<0.0001). In summary, this study describes the expression of TNF-alpha and its receptors during the peri-implantation period in the mouse. It also reports that elevated TNF-alpha restricts ICM proliferation in the blastocyst and changes the ratio of mononucleated to multinucleated trophoblast cells. These findings suggest a mechanism by which increased expression of TNF-alpha during trophoblast differentiation may be detrimental to pregnancy.
D000818:Animals; D001755:Blastocyst / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects*; D005260:Female; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects*; D017403:In Situ Hybridization; D051379:Mice; D008808:Mice, Inbred CBA; D009924:Organ Culture Techniques; D011247:Pregnancy; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism; D014327:Trophoblasts / Q000166:cytology / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism* / Q000494:pharmacology
D018124:Receptors, Tumor Necrosis Factor; D014409:Tumor Necrosis Factor-alpha