Chronic antagonism of nuclear factor-κb activity in cytotrophoblasts by dexamethasone: A potential mechanism for antiinflammatory action of glucocorticoids in human placenta

Circulating glucocorticoids are present in increasing quantities as human gestation progresses, peaking during labor whether it occurs before or at term. Although the precise role of glucocorticoids in pregnancy is not well defined, it is clear that glucocorticoids suppress inflammation in many cell types by antagonizing the acute stimulatory actions of members of the Rel/nuclear factor-κB (NF-κB) family on cytokine gene expression. In the present study we tested the hypothesis that during pregnancy, glucocorticoids chronically suppress inflammation in the human placenta. Cytotrophoblasts obtained from human term placentas were maintained for 48 h in culture medium supplemented with 10% charcoal-stripped calf serum with and without 100 nmol/L dexamethasone (DEX). Enzyme-linked immunosorbent assay studies revealed that cytotrophoblasts constitutively express interleukin-8 (IL-8), a known mediator of placental inflammation, between 24-96 h of culture. A 48-h treatment of cytotrophoblasts with 100 nmol/L DEX significantly reduced the production of IL-8 to 24 ± 1% of control levels (P < 0.01). DEX and cortisol mediated a dose-dependent inhibition of IL-8 expression, with ED₅₀ values of 5 and 50 nmol/L, respectively. DEX treatment also significantly reduced levels of IL-6 and tumor necrosis factor-α in culture medium, suggesting that glucocorticoids coordinately reduce cytokine levels in cytotrophoblasts. As cytokine expression is regulated by NF-κB and activator protein-1 (AP-1) transcription factors, electrophoretic mobility shift assays (n = 4) were used to determine whether DEX treatment altered the binding of nuclear proteins from cytotrophoblasts to labeled oligonucleotides corresponding to the κB and AP-1 response elements. We observed that a 48-h treatment of cytotrophoblasts with 100 nmol/L DEX markedly reduced binding of nuclear extracts from cytotrophoblasts to the κB response element. DEX treatment promoted a relatively smaller reduction of binding to the AP-1 response element. Northern blotting experiments revealed that DEX treatment did not alter the level of IκB, p50, or p65 messenger ribonucleic acid, suggesting that the antiinflammatory action of glucocorticoid in cytotrophoblasts did not directly involve alterations in the level of NF-κB proteins. Our results demonstrate a novel chronic suppressive action of glucocorticoid on cytokine production and nuclear binding of NF-κB and AP-1 proteins in cytotrophoblasts, providing a potential mechanism through which glucocorticoids may suppress inflammation at maternal-fetal interfaces across gestation.