Interleukin-33 in the pathogenesis of liver fibrosis: alarming ILC2 and hepatic stellate cells

Interleukin-33 (IL-33), a cytokine of the IL-1 family, has been implicated in the pathogenesis of liver fibrosis for quite some time. Indeed, IL-33 expression is upregulated in both human and murine hepatic fibrosis.^{1, 2} To date, it is believed that IL-33 serves as an ‘alarmin' released by stressed hepatocytes. IL-33 then attracts type 2 innate lymphoid cells (ILC), which trigger the profibrogenic activation of hepatic stellate cells via mediators such as IL-13.^{1, 3} IL-33 signals through a unique IL-1 receptor-related protein that is termed suppressor of tumorigenicity 2 (ST2), also called IL-33R. The binding of IL-33 to ST2 activates nuclear factor-κB and mitogen-activated protein kinases (MAPKs) and drives the production of pro-inflammatory and T helper type 2 (Th2)-associated cytokines.⁴

In the current issue of Cellular & Molecular Immunology, Tan et al. employed a model of cholestatic fibrosis and demonstrated a direct role for IL-33 in the profibrogenic activation of hepatic stellate cells (Figure 1).⁵ First, the authors confirmed that IL-33 is increased in bile duct ligation (BDL)-induced fibrosis in mice and in human cirrhotic liver tissue, which is in line with the current body of literature.^{1, 2, 6} Parenchymal cells (that is, hepatocytes) were the primary cellular source of IL-33. Second, they subjected ST2-deficient mice to surgical BDL and observed reduced hepatic injury, inflammation, and fibrosis. This finding is similar to findings from toxic and dietary fibrosis models that have been studied previously with ST2-deficient mice.^{1, 7} Flow cytometry analysis further revealed that, in the absence of ST2, BDL livers had a lower content of Th1 and Th2 cell infiltrates and reduced neutrophil and macrophage numbers. In addition, the upregulation of IL-33 during BDL was positively correlated with an increase of type 2 ILCs. These data provide further evidence for a link between IL-33 release and the secretion of type 2 inflammatory cytokines, such as IL-4, IL-5, IL-9 and IL-13.⁸

Figure 1.

IL-33 in the pathogenesis of liver fibrosis. Crystal structure analysis has shown that the amino acids 112-270 of interleukin (IL)-33 fold as a typical 12-stranded β-trefoil fold forming a β-barrel.¹⁹ In this issue of Cellular & Molecular Immunology, Tan et al. show that IL-33 expression is upregulated in the livers of mice subjected to bile duct ligation (BDL) and in patients suffering from cirrhosis. IL-33 is released by stressed hepatocytes as an alarmin and attracts innate lymphoid cells type 2 (ILC2) to the liver. Stimulation of cultured hepatic stellate cells (HSC) with IL-33 induces the expression of fibrogenic cytokines (that is, IL-6 and transforming growth factor (TGF)-β) and markers of hepatic fibrogenesis (that is, α-smooth muscle actin (α-SMA), collagen) in activated and transdifferentiated HSC (referred to as myofibroblasts (MFB)).⁵

The relationship between inflammation and IL-33 was further underpinned by the finding that intraperitoneal injections of recombinant IL-33 increased hepatic inflammation in both naive and BDL-injured livers when given 3 days prior to the BDL surgery. However, IL-33 alone failed to enhance the inflammatory response in sham-operated mice, suggesting that IL-33 has the capacity to increase the production of inflammatory cytokines only in injured livers.⁵ In accordance with this role, recombinant IL-33 aggravated hepatic fibrosis in experimental steatohepatitis, but had beneficial effects on hepatic steatosis.⁷

In addition, Tan et al. provide experimental evidence that IL-33/ST2 signaling is critical in activated hepatic stellate cells, the primary matrix-producing cells in the liver. The stimulation of culture-activated hepatic stellate cells with recombinant IL-33 increased the MAPK pathways mediated by ERK, JNK and p38 protein kinases. This resulted in the stimulation of α-smooth muscle actin and collagen expression.⁵ These data suggest a direct profibrogenic role of IL-33 in hepatic stellate cells, which is potentially synergistic with its effects on innate lymphoid cells type 2 (ILC2).¹

Although the current study and complementary fibrosis studies demonstrate overall profibrogenic effects in chronic injury models, studies in acute injury settings sharply contradict this notion. The application of concanavalin A (Con A) induced more hepatic injury in IL-33-deficient mice compared to wild-type controls. Additionally, IL-33-deficient mice given Con A had significantly higher levels of tumor necrosis factor-α and IL-1β and a larger number of natural killer cells infiltrating the liver.⁹ Moreover, the injection of recombinant IL-33 had a protective effect on hepatocytes by simultaneously repressing the expression of proapoptotic genes and activating antiapoptotic genes. IL-33 similarly protected hepatocytes in mice that were subjected to 90 min of partial hepatic ischemia followed by up to 8 h of reperfusion.¹⁰ These data suggest that the release of IL-33 by injured hepatocytes is a protective mechanism in acute (and massive) liver damage, but that it aggravates tissue fibrosis in chronic injury. These opposing functions need to be considered when developing new therapeutics intended to target this pathway.¹¹

ST2, the IL-33 receptor, is highly conserved across species and is produced in two isoforms from alternative 3' processing of mRNAs.^{12, 13} The membrane-bound isoform (ST2L or IL1RL1-b) induces immune responses after binding to IL-33, while the soluble isoforms (soluble ST2 (sST2) or IL1RL1-a) act as a decoy receptor for IL-33.¹⁴ Tan et al. used a knockout mouse in which the targeting vector deleted the majority of exons 4 and 5 of the ST2 gene. These exons are common to both the membrane-bound and soluble forms of the protein.¹⁵ It is speculated that each receptor variant has a different role in the pathogenesis of various liver diseases. Moreover, sST2 could potentially serve as a circulating biomarker to reflect IL-33 activation and fibrosis in patients with liver diseases. In fact, sST2 serum levels differ between hepatitis B virus (HBV) infected patients with mild and severe fibrosis.¹⁶ Similarly, the plasma levels of sST2 were found to be associated with disease severity and mortality in patients with HBV-related acute-on-chronic liver failure. Elevated concentrations of this decoy receptor might indicate adverse prognosis in hepatocellular carcinoma.^{17, 18}

Since its discovery in 2005,⁴ it has become evident that IL-33 is a versatile cytokine that influences the formation and severity of liver disease. The study by Tan et al. emphasizes the crucial role of IL-33 in the formation of liver fibrosis. This cytokine ‘alarms' not only ILC2 but also the collagen-producing hepatic stellate cells during chronic injury. These data strongly support ongoing efforts to target the IL-33/ST2 signaling pathway in the development of new therapeutics for patients with liver diseases.