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Evidence that MEHP inhibits rat granulosa cell function by a protein kinase C-independent mechanism
1Developmental and Reproductive Toxicology Group, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina.
Abstract:
We have recently shown that mono-(2-ethylhexyl) phthalate (MEHP), the active metabolite of the reproductive toxicant di-(ethylhexyl) phthalate (DEHP), inhibited FSH- but not forskolin-, isoproterenol-, or cholera toxin-stimulated granulosa cell cAMP accumulation in vitro. In addition, MEHP also inhibited FSH-stimulated progesterone production, a cAMP-dependent process. Similar to MEHP, the protein kinase C (PKC) activator, 12-0-tetradecanoyl-phorbol 13-acetate (TPA) has been shown to inhibit rat granulosa cell cAMP accumulation in a FSH-specific manner, and decrease FSH-stimulated progesterone production. Due to the similarity with respect to inhibition of cAMP accumulation, we conducted studies to determine if the inhibitory actions of MEHP on granulosa cell function are mediated via activation of PKC. Treatment of granulosa cells for 48 h with 100 microM MEHP produced no effect on forskolin- or isoproterenol-stimulated progesterone production, indicating that MEHP does not have a post-cyclic AMP site of action with respect to progesterone inhibition. Unlike the FSH-specific effect seen with MEHP, treatment with 10 nM TPA inhibited FSH-, forskolin-, and isoproterenol-stimulated progesterone production. In addition, maximally inhibitory concentrations of TPA and MEHP caused significantly greater inhibition of FSH-stimulated cAMP accumulation than either compound alone. Finally, addition of the progesterone precursor, pregnenolone, reversed the FSH-stimulated progesterone production inhibition by MEHP, but not that by TPA. Taken together, these data indicate that the inhibitory effects of MEHP on granulosa cell function are independent of phorbol ester-sensitive PKC activation.
Insights
Mono-(2-ethylhexyl) phthalate (MEHP) inhibits FSH-stimulated granulosa cell functions, but not through protein kinase C (PKC) activation. This finding is crucial for understanding phthalate toxicity and reproductive health.
Area of Science:
- Reproductive Toxicology
- Endocrinology
- Cellular Biology
Background:
- Di-(ethylhexyl) phthalate (DEHP) is a reproductive toxicant, and its active metabolite, mono-(2-ethylhexyl) phthalate (MEHP), inhibits FSH-stimulated granulosa cell cAMP accumulation and progesterone production.
- Protein kinase C (PKC) activators, like TPA, also inhibit FSH-stimulated cAMP and progesterone production, suggesting a potential shared mechanism.
Purpose of the Study:
- To investigate whether the inhibitory actions of MEHP on granulosa cell function are mediated through the activation of PKC.
- To elucidate the specific mechanisms by which MEHP affects granulosa cell function.
Main Methods:
- Granulosa cells were treated with MEHP and/or TPA.
- Effects on FSH-, forskolin-, and isoproterenol-stimulated cAMP accumulation and progesterone production were measured.
- The role of PKC was assessed by comparing the effects of MEHP and TPA, and by examining the impact of pregnenolone addition.
Main Results:
- MEHP inhibited FSH-stimulated cAMP accumulation and progesterone production, but did not affect forskolin- or isoproterenol-stimulated progesterone production, indicating a pre-cAMP site of action for progesterone inhibition.
- TPA inhibited FSH-, forskolin-, and isoproterenol-stimulated progesterone production.
- Combined treatment with MEHP and TPA resulted in greater inhibition of FSH-stimulated cAMP accumulation than either compound alone.
- Pregnenolone reversed MEHP-induced inhibition of progesterone production, but not TPA-induced inhibition.
Conclusions:
- The inhibitory effects of MEHP on granulosa cell function are independent of phorbol ester-sensitive PKC activation.
- MEHP and TPA exert distinct inhibitory mechanisms on granulosa cell function.
- These findings contribute to understanding the specific pathways involved in phthalate-induced reproductive toxicity.