| It has become increasingly evident that pituitary cytokines act as mediators of hormone production and pituitary tumor pathogenesis. Cytokines such as interleukin 1, 2 and 6 are detectable both in normal pituitary tissue and in human pituitary tumors, they may form an intra-pituitary signal network which synergize hypothalamic or peripheral hormones to modulate pituitary trophic hormone secretion. But till now, the molecular regulation mechanisms of cytokines controlling pituitary function is remain undefined. In this regard, we have used AtT20 cells (mouse pituitary ACTH secreting tumor cell line) as a model system to characterize the IL1β-mediated ACTH secretion, pro-opiomelanocortin(POMC) mRNA level, especially to measure IL1β-mediated changes in POMC gene promoter activity, in order to explore the effect of cytokines on POMC gene expression regulation in pituitary tumors, and also provide molecular evidence of Immune-Neuroendocrinology interactions.In the present study, we first examined the human recombinant (rh) IL1β-mediated ir-ACTH secretion by radiommunoassay(RIA) and POMC mRNA levels by RT-PCR and Southern blot analysis. The results showed that, rhIL1β (0.1pmol/L~1000pmol/L) dose-dependent increased ir-ACTH secretion from 1 hour to 12 hours and POMC mRNA level accompanied elevation 3hours after rhIL1β(1000pmol/L) treatment in AtT20 cells. Therefore, IL1β was found to enhance POMC gene expression mainly in the transcriptional level.In order to further investigate IL1β-mediated changes in POMC gene promoter activity and to determine the IL1β-induced POMC cis-elements, we need to construct POMC promoter-luciferase fusion reporter plasmids as the first step. By using the proper single restriction enzyme site, a rPOMC 5' promoter proximal fragment (-480/+63bp) from JA300 plasmid was inserted into luciferase reporter vector pGL2-Basic, constructed pGL2-POMC480(rPOMC-480/+63bp) recombinant plasmids, as well as progressively...
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