Expression of Functional Aromatic Hydrocarbon Receptor and Aromatic Hydrocarbon Nuclear Translocator Proteins in Murine Bone Marrow Stromal Cells

Abstract

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) acting through the aromatic hydrocarbon receptor (AhR) and its dimerization partner, the AhR nuclear translocator protein (arnt), elicits numerous toxicological effects including immunosuppression and thymic atrophy. Previous work has shown that TCDD alters bone marrow prothymocyte populations. These effects could be mediated at the lymphocyte level directly and/or through effects on bone marrow stromal cells, a population important in the support of lymphopoiesis. The purpose of this study was to characterize AhR and arnt expression in three murine bone marrow stromal cell lines (S17, M2-10B4, and BMS2) and in primary stromal cell cultures. Immunoblot analysis detected AhR protein in M2-10B4 and BMS2 cells. AhR protein was also detected in the primary cultures. Arnt protein could be detected in all cell cultures. Electrophoretic mobility shift assays detected TCDD-dependent dioxin-responsive element (DRE) binding in all three cell lines. DNA binding was sequence-specific and dependent on AhR, as demonstrated by the addition of unlabeled DRE DNA or of anti-AhR antibody. Results obtained with the primary cultures paralleled those seen with the stromal cell lines. The ED₅₀for induction of TCDD-dependent DRE binding in M2-10B4 cells was 0.21 nM. TCDD treatment did not induce stromal P4501A1 mRNA expression but did increase P4501B1 mRNA levels in all three cell lines and in the primary cultures. These results indicate that murine bone marrow stromal cells express AhR and arnt proteins. Furthermore, these proteins are functional in terms of their DRE-binding ability and potential to regulate mRNA levels in a gene-specific fashion.