Interleukin 1β (IL1β) and tumor necrosis factorα (TNFα) partially recapitulate effects of macrophage conditioned medium on cultured endometrial stromal cells

Abstract

OBJECTIVE: To test the hypothesis that IL1β and TNFα mimic the effects of macrophage conditioned medium on gene expression in telomerase-immortalized human endometrial stromal cells (T-HESC) in a continuation of studies designed to elucidate the role of macrophages in the development of endometriosis.DESIGN: Basic research study.MATERIALS AND METHODS: T-HESC cells were treated in culture with vehicle or IL1β (1 ng/ml) ± TNFα (5 ng/ml) (n=3 passages). Total RNA was extracted and gene expression was analyzed by DNA microarray and real time RT-PCR. GeneSpring and GraphPad were used for statistical analysis of microarray and real time RT-PCR data, respectively.RESULTS: DNA microarrays identified differential expression of 259 genes in T-HESC cells in response to IL1β ± TNFα. Approximately 72% of regulated genes matched those differentially expressed in T-HESC cells in response to treatment with macrophage conditioned medium (in press at Reproductive Sciences). Up-regulation of interleukin 8 (IL8), matrix metalloproteinase 3 (MMP3) and tenascin C (TNC) was confirmed by both microarray and real time RT-PCR.CONCLUSION: In previous work we hypothesized that macrophages secrete factors that enhance the development of endometriosis and we reported the effects of macrophage conditioned medium on gene expression in T-HESC cells. One of our goals is to define which cytokines secreted by macrophages are responsible for the gene expression responses of T-HESC cells to macrophage conditioned medium. The current study demonstrates that IL1β and TNFα, cytokines secreted by macrophages, partially recapitulate the effects of macrophage conditioned medium on gene expression in cultured T-HESC cells. Other cytokines secreted by macrophages are expected to complete the recapitulation of gene expression elicited in T-HESC by macrophage conditioned medium. OBJECTIVE: To test the hypothesis that IL1β and TNFα mimic the effects of macrophage conditioned medium on gene expression in telomerase-immortalized human endometrial stromal cells (T-HESC) in a continuation of studies designed to elucidate the role of macrophages in the development of endometriosis. DESIGN: Basic research study. MATERIALS AND METHODS: T-HESC cells were treated in culture with vehicle or IL1β (1 ng/ml) ± TNFα (5 ng/ml) (n=3 passages). Total RNA was extracted and gene expression was analyzed by DNA microarray and real time RT-PCR. GeneSpring and GraphPad were used for statistical analysis of microarray and real time RT-PCR data, respectively. RESULTS: DNA microarrays identified differential expression of 259 genes in T-HESC cells in response to IL1β ± TNFα. Approximately 72% of regulated genes matched those differentially expressed in T-HESC cells in response to treatment with macrophage conditioned medium (in press at Reproductive Sciences). Up-regulation of interleukin 8 (IL8), matrix metalloproteinase 3 (MMP3) and tenascin C (TNC) was confirmed by both microarray and real time RT-PCR. CONCLUSION: In previous work we hypothesized that macrophages secrete factors that enhance the development of endometriosis and we reported the effects of macrophage conditioned medium on gene expression in T-HESC cells. One of our goals is to define which cytokines secreted by macrophages are responsible for the gene expression responses of T-HESC cells to macrophage conditioned medium. The current study demonstrates that IL1β and TNFα, cytokines secreted by macrophages, partially recapitulate the effects of macrophage conditioned medium on gene expression in cultured T-HESC cells. Other cytokines secreted by macrophages are expected to complete the recapitulation of gene expression elicited in T-HESC by macrophage conditioned medium.