Progesterone and its metabolites 5-dihydroprogesterone and 5-3-tetrahydroprogesterone decrease LPS-induced NO release in the murine microglial cell line, BV-2.

Abstract

OBJECTIVE: Microglial cells, important immunosurveillance cells in the nervous system, respond to pathogens with an increase in inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) release. Because excessive release of NO may be neurotoxic, tight regulation of nitrosative stress is required. DESIGN AND SETTING: We cultured the murine microglial cell-line, BV-2, with bacterial lipopolysaccharide (LPS) to induce NO synthesis and quantified the impact of progesterone and its metabolites, 5alpha-dihydroprogesterone (DHP) and 5alpha-3alpha-tetrahydroprogesterone (THP), on NO release. RESULTS: Our in vitro experiments showed that in BV-2 cells LPS-induced NO release is suppressed by progesterone and THP. Both neurosteroids decreased NO release by about 40% when used at a concentration of 10 microM. NO release was less sensitive to DHP. This neurosteroid decreased NO release only by 20% when used at a concentration of 10 microM. NO release was sensitive to N(G)-methyl-L-arginine (L-NMMA), a completive inhibitor of NOS, indicating that LPS-induced NO release is mediated by NOS activity. Trypan blue exclusion experiments showed that the ratio of viable to dead cells did not vary using different concentrations of progesterone. Furthermore, progesterone did not increase apoptosis or necrosis when estimated by the distribution of DAPI-labelled condensed chromatin. CONCLUSION: These experiments indicate that the decline in NO release is mainly due to modulation of NOS activity or expression. Because progesterone, DHP, and THP are synthesized in astrocytes and oligodendrocytes, these neurosteroids may locally suppress an immune response.