Corticotropin-releasing hormone increases the expression of the prostaglandin E(2) receptor subtype EP1 in amnion WISH cells.

The purpose of this study was to investigate the effect of corticotropin-releasing hormone (CRH) on the expression of the prostaglandin (PG) E(2) EP1 receptor subtype and PGE(2) production in amnion WISH cells (AWC). AWC cultures were incubated with CRH. Culture fluid was collected for PGE(2) measurement, and the cells were collected and analyzed for EP1 protein and mRNA. Immunohistochemical localization of the EP1 receptor was also performed. Incubation of AWC with CRH resulted in a dose-dependent increase (r = 0.97) in the level of EP1 receptor protein (P < 0.001). Coincubation of AWC with CRH and indomethacin resulted in the decreased production of PGE(2) while having no effect on EP1 receptor expression. A significant but not dose-dependent increase in EP1 mRNA expression was also observed (P < 0.01). Immunohistochemical evaluation verified cell membrane localization of the receptor in both stimulated and unstimulated cells and confirmed the increased expression of EP1 receptor in response to CRH. Incubation of AWC with CRH also resulted in increased culture fluid PGE(2) levels (P < 0.01). These results suggest that the role CRH plays in the initiation of labor may also involve the promotion of elevated PGE(2) levels and increased expression of the EP1 receptor in amnion.

Modulation of the prostaglandin E receptor: a possible mechanism for infection-induced preterm labor.

OBJECTIVE: To evaluate the modulatory effects of interleukin (IL)-1beta and prostaglandin (PG)E2 on the PGE2 receptor subtype EP1 in amnion cell cultures. METHODS: Amnion cell cultures were incubated in increasing concentrations of (IL)-1beta or PGE2. Cultures were also incubated in high concentrations of IL-1beta and PGE2 in combination. Changes in EP1 receptor levels were evaluated by western and northern blot analysis. Culture fluid PGE2 levels were measured by enzyme-linked immunosorbent assay. RESULTS: EP1 receptor protein levels decreased with increasing levels of PGE2 (r = -0.82, P < .05). EP1 receptor protein (r = 0.95, P < .05), EP1 mRNA (r = 0.95, P < .01), and culture fluid PGE2 levels (P < .01) were all increased after IL-1beta administration. EP1 receptor levels also increased approximately fourfold in response to IL-1beta incubation even in the presence of high agonist (PGE2) concentrations (P < .01). CONCLUSION: The results of this study show that IL-1beta might be involved in infection-induced preterm labor by interfering with the normal regulation of EP1 receptor levels and with the promotion of increased PGE2 production in amnion tissue.

The induction of cyclooxygenase-2 by interleukin-4 in human umbilical vein endothelial cells: a possible role in regulation of fetal vascular tone.

OBJECTIVE: We sought to determine a possible role for interleukin-4 in the control of umbilical cord blood flow by evaluating its effect on cyclooxygenase-2 production of a vasoactive prostaglandin. STUDY DESIGN: Human umbilical vein endothelial cells in culture were incubated for 16 hours in media containing interleukin-4 in concentrations from 5 to 100 ng/ml. Prostaglandin E2 concentrations in the culture media were measured using a monoclonal enzyme-immunoassay. Concentrations of cyclooxygenase-1 and cyclooxygenase-2 were determined by Western blot analysis on cell homogenates. Statistical comparisons between prostaglandin E2, cyclooxygenase-1, and cyclooxygenase-2 concentrations for each interleukin-4 concentration were performed using a one way analysis of variance. RESULTS: Incubation of human umbilical vein endothelial cells in media containing interleukin-4 resulted in a significant increase in both prostaglandin E2 and cyclooxygenase-2 for interleukin-4 concentrations greater than 50 ng/ml (p < 0.05). Cyclooxygenase-1 levels were not affected. CONCLUSIONS: We suggest that interleukin-4 may have a role in the regulation of umbilical blood flow mediated through the induction of cyclooxygenase-2.

Tumor necrosis factor-alpha upregulates the prostaglandin E2 EP1 receptor subtype and the cyclooxygenase-2 isoform in cultured amnion WISH cells.

Recent studies have demonstrated a strong correlation between infection and preterm labor. Preterm delivery is also associated with high levels of cytokines and prostaglandins in amniotic fluid. The purpose of this study was to investigate the effect of tumor necrosis factor-alpha (TNF-alpha) on the levels of cyclooxygenase, prostaglandin E2 production (PGE2), and expression of the PGE2 receptor subtype EP1 in amnion WISH cell culture. Amnion WISH cell cultures were incubated in increasing concentrations of TNF-alpha (0-50 ng/ml). Changes in cyclooxygenase and EP1 receptor proteins were evaluated by Western blot analysis. Changes in EP1 mRNA were evaluated by Northern blot, and culture fluid concentrations of PGE2 were estimated by enzyme immunoassay (EIA). EP1 protein (p<0.01), EP1 mRNA (p<0.05), cyclooxygenase-2 (COX-2) protein (p<0.001), and PGE2 concentrations (p<0.01) all increased with increasing concentrations of TNF-alpha. Changes in COX-1 protein were not observed following TNF-alpha-incubation. The results suggest that TNF-alpha may play a role in infection-induced preterm labor by its pleiotropic ability to simultaneously stimulate COX-2 activity, PGE2 concentrations, and PGE2 EP1 receptor levels in human amnion.

The effect of interleukin-1 beta and interleukin-4 on the expression of prostaglandin receptors EP1 and EP3 in amnion WISH cells.

PROBLEM: Although prostaglandin E2 (PGE2) is believed to modulate biochemical and immunological events leading to parturition, the role of prostaglandin E receptors during labor has not been investigated. METHOD OF STUDY: Amnion WISH cells were incubated in media containing increasing concentrations of either interleukin-1 beta (IL-1 beta) or IL-4. Increased EP1 and EP3 protein expression was determined by Western blot analysis with peptide-specific antibodies. Concomitant measurements of culture media PGE2 were made by an enzyme immunoassay. RESULTS: Incubation of WISH cells with IL-1 beta or IL-4 caused a two- to three-fold increase in EP1 protein levels. IL-1 beta and IL-4 also caused six- and two-fold increases, respectively, in culture fluid PGE2 concentrations. IL-1 beta or IL-4 had no effect on EP3 protein levels. CONCLUSIONS: Based on these results, it is proposed that IL-1 beta and IL-4 may be involved in the initiation and promotion of labor by inducing EP1 levels and PGE2 production in amnion.

The induction of cyclooxygenase-2 (COX-2) in intact human amnion tissue by interleukin-4.

Infection is a major cause of preterm labor. Amniotic fluid from women in preterm labor associated with intrauterine infection contains increased concentrations of cytokines. The mechanism underlying this association may be a cytokine-mediated stimulation of amnion cell prostaglandin production. The biosynthesis of prostaglandins from arachidonic acid is regulated by the enzyme cyclooxygenase which exists in two forms; the constitutive form (COX-1) and the other mitogen inducible (COX-2). The purpose of this study was to evaluate the effect of the cytokine interleukin-4 (IL-4) on cyclooxygenase activity and PGE2 production in amnion. Amnion tissue was taken at caesarean section from term women not in labor and immediately incubated for 2 hours in media containing concentrations of IL-4 ranging from 1 to 100 ng/ml. An increase in both COX-2 enzyme and prostaglandin E2 (PGE2) production was observed for all concentrations of IL-4 greater than 25 ng/ml (P < 0.05, n = 8). No change in COX-1 was observed. Our data suggest that the cytokine IL-4 may be involved in the pathogenesis of premature labor by inducing COX-2 in amnion tissue resulting in increased production of PGE2 and subsequent myometrial activity.