Interleukin-15 expression is induced by TGF-beta in hepatocytes: A new role for IL-15 in liver disease?

Liver fibrogenesis is a complex process involving the participation of many meidators. TGF-beta, a key pro-fibrogenic cytokine, drives the fibrogenic process in part by activation of hepatic stellate cells. We recently described a new role for hepatocytes in the process of fibrogenesis, via TGF-beta-induced expression and secretion of CTGF, another potent profibrogenic cytokine, highlighting the importance of hepatocytes in the fibrotic process. Interleukin-15, a potent activator of NK cells, has emerged as an important new player in several models of liver damage, but its precise role has not yet been determined. Microarray analysis of primary mouse hepatocytes stimulated with TGF-beta showed that IL-15 is one of the strongest induced targets 24h after stimulation. Real Time PCR confirmed that TGFbeta induces IL-15 mRNA up to 80 fold in primary hepatocytes, and up to 10 fold in AML12 cells. By siRNA we determined that Smad3 but not Smad2 is partially necessary for IL-15 mRNA induction by TGF-beta. Besides the canonical Smad2/3 pathway, TGF-beta activates the MAPK ERK and JNK, and the PI-3K-Akt pathway. Using chemical inhibitors we determined that although ALK5 activity is essential, ERK and JNK pathways cooperate in this induction, whereas p38 and Akt are not relevant. IL-15 is expressed as two spliced isoforms, differing in the length of the signal peptide. By PCR we observed TGF-beta induction of both splice variants of IL-15. In silico analysis of a -1,8kb region of mouse IL-15 promoter indicated binding sites for Smad3, Fast-1 and AP-1 highlighting the potential cooperation between Smad3 and AP-1 transcriptional complexes. By direct and two-site ELISA we detected IL-15 and IL-15-IL-15Ralpha complexes in cell pellets but not in supernatants, indicating that IL-15 and IL-15/IL-15R complex are not secreted. Immunofluorescence revealed that IL-15 is present in the cytosol of the cells, which might indicate that another mechanism might lead to its secretion.

Our results indicate that a new potent cytokine is induced in hepatocytes by TGFbeta. We are currently investigating the biological significance of this finding.