Hepatic Stellate Cell Activation and Inactivation in NASH-Fibrosis-Roles as Putative Treatment Targets?

Abstract

Hepatic fibrosis is the primary predictor of mortality in patients with non-alcoholic steatohepatitis (NASH). In this process, the activated hepatic stellate cells (HSCs) constitute the principal cells responsible for the deposition of a fibrous extracellular matrix, thereby driving the hepatic scarring. HSC activation, migration, and proliferation are controlled by a complex signaling network involving growth factors, lipotoxicity, inflammation, and cellular stress. Conversely, the clearance of activated HSCs is a prerequisite for the resolution of the extracellular fibrosis. Hence, pathways regulating the fate of the HSCs may represent attractive therapeutic targets for the treatment and prevention of NASH-associated hepatic fibrosis. However, the development of anti-fibrotic drugs for NASH patients has not yet resulted in clinically approved therapeutics, underscoring the complex biology and challenges involved when targeting the intricate cellular signaling mechanisms. This narrative review investigated the mechanisms of activation and inactivation of HSCs with a focus on NASH-associated hepatic fibrosis. Presenting an updated overview, this review highlights key cellular pathways with potential value for the development of future treatment modalities.

Keywords: HSC activation; HSC inactivation; fibrosis; hepatic stellate cells; non-alcoholic fatty liver disease; non-alcoholic steatohepatitis.

Figure 1

A simplified overview of primary drivers of non-alcoholic steatohepatitis (NASH)-induced hepatic fibrosis. The excessive accumulation of triglycerides and free fatty acids increases hepatocellular lipid oxidation generating reactive oxygen species (ROS) and lipotoxicity. This leads to cellular damage and the release of inflammatory cytokines, prompting the activation of resident liver macrophages (Kupffer cells) and the recruitment of circulating immune cells: monocytes and leukocytes. The initial hepatic steatosis then becomes a state of hepatic inflammation and progresses to NASH. The inflammation and sustained lipotoxicity maintain a self-perpetuating vicious circle of increased production of ROS, inflammation, and cell damage, ultimately promoting the activation of hepatic stellate cells (aHSC), which leads to the formation of a fibrogenic extracellular matrix, thus hallmarking the transition to a state of NASH-induced fibrosis.

Figure 2

The hepatic stellate cell phenotypic switch in NASH. In a healthy liver, the hepatic stellate cell (HSC) rests in a quiescent state (qHSC) while residing close to the hepatic sinusoids. qHSCs are considered dormant and non-proliferative, and they are characterized by the cytoplasmatic storage of retinyl esters (vitamin A) in lipid droplets; markers include PPARγ, GFAP, and BAMBI, all expressed in the qHSCs. The accumulation of lipotoxic metabolites, inflammation, and oxidative stress in NASH affects multiple hepatic cell types and leads to the release/activation of several cellular signaling factors, such as growth factors (e.g., increased TGFβ, PDGF, and connective tissue growth factors) and nuclear receptors (e.g., decreased PPARγ and retinoid X receptor activation), thus promoting an HSC phenotypic switch. In this process, qHSCs lose their stored retinyl esters and transdifferentiate into the activated, proliferative, and contractile state (aHSC). aHSCs are characterized by the production of pro-collagens for extracellular matrix deposition and the promotion of HSC activation and fibrogenesis (thus creating a positive feedback loop), as well as the ability to migrate and divide; markers include the expression of αSMA, S100a6, PDGFRβ, and TIMP1. The clearance of aHSCs is necessary for the cessation of matrix deposition, and it can take place through apoptosis or through inactivation. Inactivated HSCs (iHSCs) differentiate towards a more dormant phenotype (e.g., with a decrease of aHSC characteristics and the re-establishment of the cytoplasmic storage of retinyl esters), but they do not completely revert to the qHSC state and have increased sensitivity toward reactivation. aHSC: activated hepatic stellate cell; BAMBI: bone morphogenetic protein and activin membrane bound inhibitor; ECM: extracellular matrix; GFAP: glial fibrillary acidic protein; iHSC: inactivated hepatic stellate cell; PDGFRβ: platelet derived growth factor receptor β; PPARγ: peroxisome proliferator activated receptor γ; qHSC: quiescent hepatic stellate cell; S100a6: S100 calcium-binding protein A6; TGFβ: transforming growth factor beta; TIMP1: tissue inhibitor of metalloproteinase 1; αSMA: alpha smooth muscle actin.

Figure 3

Molecular mechanisms of hepatic stellate cell activation. The activation of hepatic stellate cells involves multiple signaling pathways and receptor systems. 1: TGFβ is one of the most potent fibrogenic factors and is released in response to insults. In HSCs, TGFβ is released through IL-13-dependent induction and via integrin-mediated interactions with extracellular TGFβ stored in a LLC. TGFβ acts through SMAD and non-SMAD pathways to increase collagen synthesis and extracellular matrix deposition. An increased TIMP level inhibits MMP expression and collagen breakdown. 2: PDGF induces RAS-MAPK and PI3K-AKT/PKB signaling that—alongside cytokines and growth factors such as CCL2, CCL5, and CTGF—promotes HSC proliferation and migration. 3: Increased ROS induce ER stress, which (alongside DAMPs) leads to HSC activation. 4: Gut permeability may increase in NASH, and gut-derived and hepatic FC signaling through TLR4 promotes the production of inflammatory cytokines, growth factors, and HSC activation. In addition, TLR4 signaling can indirectly activate HSCs by decreasing the expression of the TGFβ decoy receptor BAMBI, which is also decreased by the inflammatory cytokine IL-1β. 5: In turn, lipotoxic lipid (e.g., palmitic acid) signaling through TLR2 and Hedgehog-derived signaling further contributes to HSC activation. 6: Nuclear receptors also play an important role in HSC activation, being inhibited by RXR, FXR, LXR, PXR, and PPARγ (decreased in activated HSCs). Though all mechanisms of HSC activation remain to be disclosed, this figure illustrates the highly complex cellular signaling patterns involved in NASH-associated HSC activation and the subsequent production of a fibrous extracellular matrix. AKT/PKB: protein kinase B. CTGF: connective tissue growth factor. BAMBI: bone morphogenetic protein and activin membrane-bound inhibitor. CCL: chemokine C-C motif ligand. DAMP: damage-associated molecular patterns. ER: endoplasmic reticulum. FC: free cholesterol. FXR: farnesoid X receptor. HSC: hepatic stellate cell. IL: interleukin. LPS: lipopolysaccharide. LAP: latency-associated protein. LLC: large latent complex. LTBP: latent TGF-β-binding protein. LXR: liver X receptor. MAPK: mitogen-activated protein kinase. MMP: matrix metalloproteinase. NAFLD: non-alcoholic fatty liver disease. PDGF: platelet-derived growth factor. PI3K: phosphoinositide 3-kinase. PPARγ: peroxisome proliferator-activated receptor γ. PXR: pregnane X receptor. ROS: reactive oxygen species. RXR: retinoid X receptor. TIMP: tissue inhibitor of matrix metalloproteinase. TGFβ: tissue growth factor β. TLR: toll-like receptor. SMAD: mothers against decapentaplegic homolog. Arrow heads indicate activation, and transversal lines indicate inhibition.