Regulation of liver fibrosis by tuning M2 macrophage signaling through IL-4Ralpha

Progression and Regression of liver fibrosis have been linked with the innate immune system, especially macrophages. Depending on stimulation by different cytokines or LPS, macrophages can differentiate roughly into M1 (classical) and M2 (alternative) macrophages. The roles of M1 and M2 in liver fibrosis progression or regression are largely unexplored.

We used the CCl4-model of liver fibrosis progression (6 weeks) and spontaneous regression after omission of the toxin (2 weeks) in C57BL6 mice, and the model of Mdr2KO mice (spontaneous biliary fibrosis progression) to assess the role of M2 and their manipulation in liver fibrosis progression and reversal.

In mice with CCL4-induced liver fibrosis, expression of the M2-inducing, IL4/IL-13 responsive IL-4Ralpha was strongly decreased at week 2 of spontaneous fibrosis reversal. In spontaneously progressive Mdr2 KO mice, expression of the IL-4Ralpha gradually increased until age 6-wk, and decreased thereafter. For functional characterization of M2 macrophages, we applied specific IL-4Ralpha antisense oligonucleotide (ASO) in vitro (murine RAW macrophages) and in vivo (6 weeks CCL4-treated and Mdr2 KO mice). Suppression of IL-4Ralpha in RAW macrophages using the ASO was highly effective (90% at 2 µg/ml). When given to CCL4-treated mice twice weekly at 40 mg/kg i.p. from week-2 to week-4 during the regression phase, the ASO suppressed hepatic expression of IL-4Ralpha by 47%, decreased the M2 markers Arg1 and Mrc1 and increased expression of the M1 markers CCL3, MMP-8 and MMP-9, and of profibrogenic procollagen alpha1(I) RNA. Serum ALT was increased 4.1 fold in ASO-treated mice vs. untreated mice. Mdr2 KO mice that received the ASO from week 6 to week 10 also showed a similar shift from the M2 to the M1 phenotype, and similar regulation of the above genes. Similarly, a higher elevation of ALT in the ASO treated mice indicated mild inflammation due to M2 suppression.

We showed that, somewhat unexpectedly, suppression of M2 macrophage differentiation during fibrosis progression is beneficial, whereas it is detrimental during fibrosis regression. Addressing the IL-4Ralpha with specific inhibitors may be a novel M2 macrophage targeted approach to inhibit liver fibrosis progression.