The down-regulation of IL-6-stimulated fibrinogen steady state mRNA and protein levels by human recombinant IL-1 is not PGE2-dependent: Effects of IL-1 receptor antagonist (IL-1RA)

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Abstract
Infections, trauma and inflammatory processes induce a host response with increases in a large group of structurally and functionally diverse plasma proteins. Parental administration of foreign proteins also induce an increase in plasma fibrinogen. Interleukin-6 (IL-6) is a monocyte-derived mediator and has regulatory effects on acute phase protein genes which result in the induction of fibrinogen synthesis in primary hepatocytes, while the addition of interleukin-1 (IL-1) exerts a negative modulating influence on the IL-6-stimulated fibrinogen. In order to understand the mechanisms by which IL-1 inhibits IL-6-stimulated fibrinogen transcription and translation, and since IL-1 is believed to act through PGE2 stimulation, we have studied the influence of PGE2 in IL-6 or IL-1, alone and in combination, on Fg mRNA expression (by Northern blot analysis) and the influence of PGE2, indomethacin, and arachidonic acid on Fg secretion. Moreover, since human recombinant interleukin-1 receptor antagonist (hrIL-1ra) is a strong inhibitor of IL-1 induced IL-1 transcription and translation and has an inhibitory effect on PGE2, we have studied the effects of IL-1ra on the down-regulation of IL-6 stimulated fibrinogen by IL-1, using an Fg ELISA method. Herein we show that using human Hep 3B hepatoma cell cultures, steady-state levels of Fg mRNA is strongly increased in cells treated with IL-6 (10 ng/ml) and IL-6 (10 ng/ml)+PGE2 (5×10−6 M) compared to the control (Nil), while when IL-1 (1 ng/ml) was given in combination with IL-6 10 ng/ml, a strong inhibitory effect was found compared to IL-6 alone. The results were not statistically different from the controls. The addition of PGE2 at 5×10−6 M plus IL-6 10 ng/ml to the cell cultures induced an augmentation of Fg mRNA compared to IL-6 alone. Hep 3B hepatoma cells treated with IL-6, but not IL-1, induced an increase of Fg secretion compared to the control, while cells treated with the combination IL-6+IL-1 produced the same amounts of Fg compared to the control. The addition of PGE2 to IL-6 alone or IL-1+IL-6 did not modify the results. Arachidonic acid also had no effect on the combination IL-1 + IL-6. Similar results were obtained when indomethacin (5–50 μM) was used. When human recombinant IL-1 receptor antagonist (hrIL-1ra) 1–50 μg/ml was added to the Hep 3B hepatoma cell cultures, a dose-response restoration occured on IL-1-inhibited IL-6-stimulated Fg secretion. In these studies we show for the first time that PGE2 is not involved in the inhibition induced by IL-1 on IL-6 gene expression and secretion and that PGE2 (5×10−6 and 5×10−7 M) enhances IL-6-stimulation of Fg expression. Moreover, hrIL-1ra restored the down-regulation produced by IL-1 on IL-6-stimulated Fg translation. These results provide additional biological activities for IL-1, suggesting new and different mechanism(s) of action.

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