PROBLEM: Adhesive interaction between trophoblast cells and uterine endometrial basement membrane is one of the critical processes in embryo implantation. This interaction is directly or indirectly regulated by hormones, growth factors, and cytokines. Since tumor necrosis factor-alpha (TNF-alpha) is synthesized by both decidual and trophoblast cells, we hypothesized that TNF-alpha may play a regulatory role in trophoblast cell invasion. To test this hypothesis, we have used in vitro models to determine the effect of TNF-alpha on human trophoblast cell adhesion and motility, two major steps in trophoblast invasion. METHODS: The effect of TNF-alpha on the motility of extended-lifespan first trimester trophoblasts (HTR) and JEG-3 choriocarcinoma cells was tested using the phagokinetic track motility assay. An in vitro adhesion assay was used to determine the effect of TNF-alpha on the adhesion of HTR and JEG-3 cells to laminin, a major basement membrane component. In addition, the effect of TNF-alpha on the surface expression of the laminin receptor beta 1 integrin subunit was examined using flow cytometry. RESULTS: HTR or JEG-3 cells strongly adherent to laminin which was not significantly altered by TNF-alpha treatment. We also measured the effect of TNF-alpha on the surface expression of beta 1 integrin on HTR and JEG-3 cells; no difference was observed between control and treatment groups. Interestingly, the motility of both HTR and choriocarcinoma JEG-3 cells was significantly inhibited by TNF-alpha. CONCLUSIONS: The role of TNF-alpha in human embryo implantation is currently unknown. Our data demonstrate that TNF-alpha does alter trophoblast cell adhesion to laminin, but significantly inhibits trophoblast cell motility in vitro, suggesting that TNF-alpha may play a regulatory role in trophoblast cell invasion.
PROBLEM: Adhesive interaction between trophoblast cells and uterine endometrial basement membrane is one of the critical processes in embryo implantation. This interaction is directly or indirectly regulated by hormones, growth factors, and cytokines. Since tumor necrosis factor-alpha (TNF-alpha) is synthesized by both decidual and trophoblast cells, we hypothesized that TNF-alpha may play a regulatory role in trophoblast cell invasion. To test this hypothesis, we have used in vitro models to determine the effect of TNF-alpha on human trophoblast cell adhesion and motility, two major steps in trophoblast invasion. METHODS: The effect of TNF-alpha on the motility of extended-lifespan first trimester trophoblasts (HTR) and JEG-3 choriocarcinoma cells was tested using the phagokinetic track motility assay. An in vitro adhesion assay was used to determine the effect of TNF-alpha on the adhesion of HTR and JEG-3 cells to laminin, a major basement membrane component. In addition, the effect of TNF-alpha on the surface expression of the laminin receptor beta 1 integrin subunit was examined using flow cytometry. RESULTS:HTR or JEG-3 cells strongly adherent to laminin which was not significantly altered by TNF-alpha treatment. We also measured the effect of TNF-alpha on the surface expression of beta 1 integrin on HTR and JEG-3 cells; no difference was observed between control and treatment groups. Interestingly, the motility of both HTR and choriocarcinoma JEG-3 cells was significantly inhibited by TNF-alpha. CONCLUSIONS: The role of TNF-alpha in human embryo implantation is currently unknown. Our data demonstrate that TNF-alpha does alter trophoblast cell adhesion to laminin, but significantly inhibits trophoblast cell motility in vitro, suggesting that TNF-alpha may play a regulatory role in trophoblast cell invasion.
Authors: Maciej T Nogalski; Gary C T Chan; Emily V Stevenson; Donna K Collins-McMillen; Andrew D Yurochko Journal: J Vis Exp Date: 2012-12-04 Impact factor: 1.355