Estrogen modulates the expression of tumor necrosis factor alpha mRNA in phorbol ester-stimulated human monocytic THP-1 cells.
Monokines, including tumor necrosis factor alpha (TNF-alpha), have been implicated in the pathogenesis of several pathologic processes, including atherosclerosis. Because estrogen has been found to offer a certain degree of protection against atherosclerotic progression, we examined the effect of estrogen on the expression of TNF-alpha mRNA in a monocyte-macrophage cell line, THP-1. Cells were exposed to 12-O-tetradecanoylphorbol 13-acetate (TPA, 50 ng/ml) for 48 or 96 h to induce differentiation. Some of the cells were treated with lipopolysaccharide (LPS, 10 micrograms/ml) in the last 3 h and/or ethinyl estradiol (estrogen, 10(-9) M) in the last 20 h. Total cellular RNA was isolated and cDNA synthesized and than coamplified using the polymerase chain reaction (PCR) in the presence of two sets (pairs) of 32P-labeled primers, one for TNF-alpha (product size 325 bp) and the second for the internal control, glyceraldehyde 3-phosphate dehydrogenase (G3PDH; 983 bp). The resultant PCR products were separated by agarose gel electrophoresis, and the ratios of radioactivity incorporated into TNF-alpha PCR products to G3PDH products were used to assess the relative changes in the levels of TNF-alpha mRNA abundance in response to various substances. Treatment with TPA for 48 h induced the expression of TNF-alpha mRNA. Treatment of these TPA-stimulated cells with estrogen caused a 62% decrease in TNF-alpha message abundance (p < 0.01). Similar results were obtained with cells stimulated with TPA for 96 h.(ABSTRACT TRUNCATED AT 250 WORDS)
Shanker
G
G
Department of Comparative Medicine, Bowman Gray School of Medicine, Wake Forest University, Winston-Salem, North Carolina.
Sorci-Thomas
M
M
Adams
M R
MR
D001161:Arteriosclerosis / Q000209:etiology / Q000517:prevention & control; D002460:Cell Line; D004997:Ethinyl Estradiol / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D008264:Macrophages / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D009000:Monocytes / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics* / Q000502:physiology
D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D004997:Ethinyl Estradiol; D013755:Tetradecanoylphorbol Acetate