Interleukin-1 regulation of corticotropin-releasing factor (CRF), glucocorticoid receptor, c-fos and c-jun messenger RNA in the NPLC-KC cell line

Abstract

The NPLC-KC human hepatoma cell line expresses corticotropin-releasing factor (CRF) and it has been demonstrated that CRF secretion and synthesis in this cell line increases in response to activators of the protein kinase A (PKA) and C (PKC) pathways as well as interleukin-1 (IL1). CRF expression with all three agents can be inhibited with the synthetic steroid-dexamethasone (DEX). In this report, we have examined the effect of IL1 ( β form) in the presence and absence of DEX on CRF mRNA (mRNA) expression as well as the expression of human glucocorticoid receptor (GR) mRNA and the mRNA of the proto-oncogenes (c-jun and c-fos) that have been implicated in CRF regulation. NPLC-KC cells were incubated with picomolar concentrations of IL1. Following this total RNA was extracted from the cells and Northern Blots were probed with 32P-labelled human DNA probes for the CRF, GR, c-jun and c-fos genes. Levels of mRNA expression were measured using a PhosphoImager and were normalized to mRNA levels of control probe glyceraldehyde-3-phosphate dehydrogenase (GAPD). CRF mRNA was significantly increased with IL1 treatment in a time and concentration dependent manner. CRF mRNA expression increased with increasing concentrations of IL1 over the range of 1–100 pM; expression of CRF mRNA also peaked after 24 h of 100 pM IL1 treatment and reached a level of expression approximately seven times higher than control. This pattern of expression was significantly inhibited in the presence of 100 nM DEX. Levels of the GR, c-fos and c-jun mRNAs were also significantly increased in the presence of IL1 and inhibited when DEX was co-incubated with IL1. The results reveal that IL1 stimulation of CRF mRNA expression by IL1 in the NPLC-KC cell line is accompanied by activation of GR mRNA as well as the mRNA of the immediate early genes—c-fos and c-jun. The results also demonstrate that this cell line may serve as a model system for the molecular mechanisms by which IL1 regulates CRF in central nervous system (CNS) neurons.