100: Lipopolysaccharide (LPS) stimulation induces corticotropin releasing hormone (CRH) expression through MyD88 in trophoblast cells

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HORMONE (CRH) EXPRESSION THROUGH MYD88 IN TROPHOBLAST CELLS. OZLEM EQUILS, ANDY UH, CHANTELLE BLUE, GUSTAVO GONZALEZ, CALVIN HOBEL, CedarsSinai Medical Center, Los Angeles, California, Cedars-Sinai Medical Center, Obstetrics and Gynecology, Los Angeles, California, University of California, Los Angeles, California OBJECTIVE: To study the effect of infection on placental CRH expression and the molecular mechanisms involved. STUDY DESIGN: We transiently transfected JEG3 syncytiotrophoblast cell line with CRH-luciferase and beta-galactosidase vectors overnight using Fugene (Roche). The cells were then stimulated with different concentrations of LPS (TLR4 ligand) and Pam3Csk (TLR2 ligand) for various durations. CRH expression was determined by luciferase assay and calorimetric galactosidase assay was performed to correct for transfection efficiency. In order to elucidate the molecular mechanism involved, cells were also transfected with dominant negative (DN) nonsignaling MyD88 and TRIF cDNA; alternatively the cells were transfected with cAMP response element (CRE)-luciferase cDNA. cAMP induces CRH expression in the trophoblasts and was used as the positive control. RESULTS: LPS but not Pam3Csk stimulation led to CRH expression in a dose dependent manner at 24 hours. LPS induced CRH expression was significantly lower than that induced by cAMP. In order to assess whether the effect of LPS to induce CRH expression was secondary to the autocrine effect of LPS-induced cytokines, we stimulated naive JEG3 cells transiently transfected with CRH luciferase cDNA with the supernatant obtained from overnight LPS-treated JEG3 cells and observed that supernatant did not induce CRH expression. Expression of DNMyD88 but not DN-TRIF inhibited the LPS induced CRH expression. cAMP but not LPS stimulation induced CRE expression in JEG3 cells. CONCLUSION: TLR4 but not TLR2 stimulation induces CRH expression in trophoblasts through MyD88 but not CRE. This data suggest that infection with Gram-negative bacteria or exposure to LPS may induce placental CRH expression and play role in the pathogenesis of preterm delivery.