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Mono-ethylhexyl phthalate stimulates prostaglandin secretion in human placental macrophages and THP-1 cells.

Tetz LM, Aronoff DM, Loch-Caruso R - Reprod. Biol. Endocrinol. (2015)

Bottom Line: Co-treatment of PMs or THP-1 cells with 180 micromolar MEHP and the non-selective COX inhibitor indomethacin reduced MEHP-stimulated PGE2 production.Similarly, co-treatment of PM and THP-1 cells with the COX-2 selective inhibitor NS-398 resulted in a significant decrease in PGE2, suggesting that MEHP-stimulated PGE2 is dependent specifically on increased COX-2 expression.Western blot analysis revealed a significant increase in COX-2 expression in PM and THP-1 cells treated with 180 micromolar MEHP, and no changes in COX-1 expression, supporting the role of COX-2 in MEHP-stimulated PGE2 synthesis.

View Article: PubMed Central - PubMed

Affiliation: Environmental Health Sciences, University of Michigan, Ann Arbor, MI, 48109, USA. lauren.m.tetz@vanderbilt.edu.

ABSTRACT

Background: Diethylhexyl phthalate (DEHP) is widely used as a plasticizer in polyvinyl chloride products. DEHP exposure, which is widespread in the US, increases preterm birth risk; however, the mechanisms driving this relationship are unclear. Because cyclooxygenase-2 (COX-2) dependent prostaglandin synthesis is implicated in preterm birth, we evaluated effects of mono-2-ethylhexyl phthalate (MEHP), the active metabolite of DEHP, on prostaglandin E2 (PGE2) synthesis and COX expression in human placental macrophages (PM). In addition, responses in PM were compared to those in a human macrophage-like cell line, THP-1.

Methods: PM and THP-1 cells were treated for 2, 4, 8, or 24 h with MEHP concentrations ranging from 10 to 180 micromolar. PGE2 concentrations were assessed in culture medium using ELISA, and COX expression was determined by western blot.

Results: Treatment of PM and THP-1 cells with 180 micromolar MEHP for 24 h significantly increased PGE2 release. Co-treatment of PMs or THP-1 cells with 180 micromolar MEHP and the non-selective COX inhibitor indomethacin reduced MEHP-stimulated PGE2 production. Similarly, co-treatment of PM and THP-1 cells with the COX-2 selective inhibitor NS-398 resulted in a significant decrease in PGE2, suggesting that MEHP-stimulated PGE2 is dependent specifically on increased COX-2 expression. Western blot analysis revealed a significant increase in COX-2 expression in PM and THP-1 cells treated with 180 micromolar MEHP, and no changes in COX-1 expression, supporting the role of COX-2 in MEHP-stimulated PGE2 synthesis.

Conclusions: The findings from this study are the first to demonstrate phthalate-stimulated PGE2 synthesis in PM and warrant future studies into COX-2-dependent prostaglandin synthesis as a mechanism of toxicant-associated preterm birth.

No MeSH data available.


Related in: MedlinePlus

MEHP effects on COX-1 and COX-2 protein expression in THP-1 cells. Western blot and densitometry analysis of COX-2 (a and c) and COX-1 (b and d) in THP-1 cells treated for 8 h with DMSO (0.05 % v/v; solvent control), 180 μM MEHP or 500 ng/mL LPS. Western blot analysis was performed on THP-1 cell lysates as described in the “Methods” section. All samples were run alongside COX-1 and COX-2 positive control lysates, seen on the far right of the gel. N = 3–4 experiments. *p < 0.05, compared to solvent controls
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Fig3: MEHP effects on COX-1 and COX-2 protein expression in THP-1 cells. Western blot and densitometry analysis of COX-2 (a and c) and COX-1 (b and d) in THP-1 cells treated for 8 h with DMSO (0.05 % v/v; solvent control), 180 μM MEHP or 500 ng/mL LPS. Western blot analysis was performed on THP-1 cell lysates as described in the “Methods” section. All samples were run alongside COX-1 and COX-2 positive control lysates, seen on the far right of the gel. N = 3–4 experiments. *p < 0.05, compared to solvent controls

Mentions: Western blot densitometric analysis revealed a significant increase in COX-2 protein expression of approximately 2-fold in PMs treated with 180 μM MEHP (Fig. 2a,c) but no significant changes in COX-1 expression (Fig. 2b,c), supporting the role of COX-2 in MEHP-stimulated PGE2 synthesis. MEHP induction of COX-2 was also observed in THP-1 cells (Fig. 3). Furthermore, 180 μM MEHP induced COX-2 expression at levels comparable to those observed with a potent pro-inflammatory stimulus, lipopolysaccharide (LPS; Figs. 2 and 3).Fig. 2


Mono-ethylhexyl phthalate stimulates prostaglandin secretion in human placental macrophages and THP-1 cells.

Tetz LM, Aronoff DM, Loch-Caruso R - Reprod. Biol. Endocrinol. (2015)

MEHP effects on COX-1 and COX-2 protein expression in THP-1 cells. Western blot and densitometry analysis of COX-2 (a and c) and COX-1 (b and d) in THP-1 cells treated for 8 h with DMSO (0.05 % v/v; solvent control), 180 μM MEHP or 500 ng/mL LPS. Western blot analysis was performed on THP-1 cell lysates as described in the “Methods” section. All samples were run alongside COX-1 and COX-2 positive control lysates, seen on the far right of the gel. N = 3–4 experiments. *p < 0.05, compared to solvent controls
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4462084&req=5

Fig3: MEHP effects on COX-1 and COX-2 protein expression in THP-1 cells. Western blot and densitometry analysis of COX-2 (a and c) and COX-1 (b and d) in THP-1 cells treated for 8 h with DMSO (0.05 % v/v; solvent control), 180 μM MEHP or 500 ng/mL LPS. Western blot analysis was performed on THP-1 cell lysates as described in the “Methods” section. All samples were run alongside COX-1 and COX-2 positive control lysates, seen on the far right of the gel. N = 3–4 experiments. *p < 0.05, compared to solvent controls
Mentions: Western blot densitometric analysis revealed a significant increase in COX-2 protein expression of approximately 2-fold in PMs treated with 180 μM MEHP (Fig. 2a,c) but no significant changes in COX-1 expression (Fig. 2b,c), supporting the role of COX-2 in MEHP-stimulated PGE2 synthesis. MEHP induction of COX-2 was also observed in THP-1 cells (Fig. 3). Furthermore, 180 μM MEHP induced COX-2 expression at levels comparable to those observed with a potent pro-inflammatory stimulus, lipopolysaccharide (LPS; Figs. 2 and 3).Fig. 2

Bottom Line: Co-treatment of PMs or THP-1 cells with 180 micromolar MEHP and the non-selective COX inhibitor indomethacin reduced MEHP-stimulated PGE2 production.Similarly, co-treatment of PM and THP-1 cells with the COX-2 selective inhibitor NS-398 resulted in a significant decrease in PGE2, suggesting that MEHP-stimulated PGE2 is dependent specifically on increased COX-2 expression.Western blot analysis revealed a significant increase in COX-2 expression in PM and THP-1 cells treated with 180 micromolar MEHP, and no changes in COX-1 expression, supporting the role of COX-2 in MEHP-stimulated PGE2 synthesis.

View Article: PubMed Central - PubMed

Affiliation: Environmental Health Sciences, University of Michigan, Ann Arbor, MI, 48109, USA. lauren.m.tetz@vanderbilt.edu.

ABSTRACT

Background: Diethylhexyl phthalate (DEHP) is widely used as a plasticizer in polyvinyl chloride products. DEHP exposure, which is widespread in the US, increases preterm birth risk; however, the mechanisms driving this relationship are unclear. Because cyclooxygenase-2 (COX-2) dependent prostaglandin synthesis is implicated in preterm birth, we evaluated effects of mono-2-ethylhexyl phthalate (MEHP), the active metabolite of DEHP, on prostaglandin E2 (PGE2) synthesis and COX expression in human placental macrophages (PM). In addition, responses in PM were compared to those in a human macrophage-like cell line, THP-1.

Methods: PM and THP-1 cells were treated for 2, 4, 8, or 24 h with MEHP concentrations ranging from 10 to 180 micromolar. PGE2 concentrations were assessed in culture medium using ELISA, and COX expression was determined by western blot.

Results: Treatment of PM and THP-1 cells with 180 micromolar MEHP for 24 h significantly increased PGE2 release. Co-treatment of PMs or THP-1 cells with 180 micromolar MEHP and the non-selective COX inhibitor indomethacin reduced MEHP-stimulated PGE2 production. Similarly, co-treatment of PM and THP-1 cells with the COX-2 selective inhibitor NS-398 resulted in a significant decrease in PGE2, suggesting that MEHP-stimulated PGE2 is dependent specifically on increased COX-2 expression. Western blot analysis revealed a significant increase in COX-2 expression in PM and THP-1 cells treated with 180 micromolar MEHP, and no changes in COX-1 expression, supporting the role of COX-2 in MEHP-stimulated PGE2 synthesis.

Conclusions: The findings from this study are the first to demonstrate phthalate-stimulated PGE2 synthesis in PM and warrant future studies into COX-2-dependent prostaglandin synthesis as a mechanism of toxicant-associated preterm birth.

No MeSH data available.


Related in: MedlinePlus