Stress and Liver Fibrogenesis: Understanding the Role and Regulation of Stress Response Pathways in Hepatic Stellate Cells

Abstract

Stress response pathways are crucial for cells to adapt to physiological and pathologic conditions. Increased transcription and translation in response to stimuli place a strain on the cell, necessitating increased amino acid supply, protein production and folding, and disposal of misfolded proteins. Stress response pathways, such as the unfolded protein response (UPR) and the integrated stress response (ISR), allow cells to adapt to stress and restore homeostasis; however, their role and regulation in pathologic conditions, such as hepatic fibrogenesis, are unclear. Liver injury promotes fibrogenesis through activation of hepatic stellate cells (HSCs), which produce and secrete fibrogenic proteins to promote tissue repair. This process is exacerbated in chronic liver disease, leading to fibrosis and, if unchecked, cirrhosis. Fibrogenic HSCs exhibit activation of both the UPR and ISR, due in part to increased transcriptional and translational demands, and these stress responses play important roles in fibrogenesis. Targeting these pathways to limit fibrogenesis or promote HSC apoptosis is a potential antifibrotic strategy, but it is limited by our lack of mechanistic understanding of how the UPR and ISR regulate HSC activation and fibrogenesis. This article explores the role of the UPR and ISR in the progression of fibrogenesis, and highlights areas that require further investigation to better understand how the UPR and ISR can be targeted to limit hepatic fibrosis progression.

Figure 1

The unfolded protein response (UPR) activates fibrotic genes during hepatic stellate cell (HSC) activation. Endoplasmic reticulum (ER) stress leads to dissociation of the protein chaperone Ig-binding protein (BiP) from the three effector proteins of the UPR [activating transcription factor 6α (ATF6α), protein kinase RNA-like endoplasmic reticulum kinase (PERK), and inositol-requiring enzyme 1α (IRE1α)] and allows them to activate signaling mechanisms aimed at resolving ER stress. A: Dissociated IRE1α oligomerizes, autophosphorylates, and activates signaling cascades through X-box binding protein 1 (XBP1) and apoptosis signal-regulating kinase 1 (ASK1). The endonuclease activity of IRE1α cleaves mRNAs [in a process called regulated IRE1α-dependent decay (RIDD)], including the canonical splicing of XBP1 into the active transcription factor form. XBP1 increases expression of the fibrogenic genes α-smooth muscle actin (α-SMA), collagen I, platelet-derived growth factor receptor-β (PDGFRβ), and transport and Golgi organization protein 1 (TANGO1), as well as the ER stress response mechanisms of ER expansion and autophagy. IRE1α also phosphorylates ASK1, which signals through a phosphorylation cascade involving c-Jun N-terminal kinase (JNK) and the transcription factor CCAAT enhancer-binding protein (c/EBPβ). Like ATF6α, c/EBPβ increases expression of α-SMA, collagen I, and fibronectin, increasing HSC activation and fibrogenesis. B: ATF6α dissociation from BiP reveals Golgi-localization signals to initiate its trafficking to the Golgi, where it is cleaved to release the cytosolic domain. ATF6α localizes to the nucleus, where it acts as a transcription factor. ATF6α increases expression of α-SMA, collagen I, and fibronectin in vitro and in vivo, directly up-regulating HSC activation and fibrogenesis. It also up-regulates thioredoxin domain-containing protein 5 (TXNDC5), which activates JNK and STAT3 to promote fibrogenesis. C: PERK dimerization activates its kinase domain and leads to autophosphorylation. PERK phosphorylates and initiates signaling cascades through the α subunit of eukaryotic translation initiation factor 2 (eIF2α) and heterogeneous nuclear ribonucleoprotein A1 (HNRNPA1). PERK phosphorylation of eIF2α inhibits general translation and promotes preferential translation of stress response genes, including the transcription factor ATF4, whose role in fibrogenesis is unclear. PERK also phosphorylates HNRNPA1, targeting it for degradation and preventing maturation of its downstream target miR-18A, promoting its degradation. miR-18A can target SMAD2, a major protein in fibrogenic transforming growth factor-β signaling, for degradation, so loss of miR-18A increases SMAD2 levels and associated fibrogenic signaling. ∗Shown in vitro. #Shown in vivo. Adapted from UPR Signaling (ATF6, PERK, IRE1) template at BioRender.com (Toronto, ON, Canada; 2023). TIMP1, tissue inhibitor of metalloprotease 1; TRIB3, Tribbles pseudokinase 3.

Figure 2

The potential link between fibrogenesis and integrated stress response (ISR). A: The ISR is initiated by several different stimuli, but regardless of the initiating kinase [heme-regulated inhibitor (HRI), protein kinase R (PKR), general control nonderepressible 2 kinase (GCN2), or protein kinase RNA-like endoplasmic reticulum (ER) kinase (PERK)], ISR signaling begins with phosphorylation of the α subunit of eukaryotic translation initiation factor 2 (eIF2α; P-eIF2α). P-eIF2α limits bulk translation initiation, concomitant with preferential translation of stress response genes, including the transcription factor activating transcription factor 4 (ATF4). ATF4 induces the expression of a diverse set of genes that play critical roles in maintaining cellular homeostasis and responding to stress, including chaperones, oxidation-reduction enzymes, antiviral proteins, amino acid transporters, amino acid biosynthesis enzymes, tRNA synthetase, cytokines, and autophagy genes. ATF4 also up-regulates growth arrest and DNA damage inducible 34 (GADD34), a phosphatase that works in conjunction with another phosphatase, constitutive repressor of eIF2α phosphorylation (CReP), to dephosphorylate eIF2α and restore translation. Other ATF4-regulated genes include glucose transporter type 1 (GLUT1) and genes involved in glycine to serine synthesis [phosphoglycerate dehydrogenase (PHGDH), phosphoserinea aminotransferase 1 (PSAT1), phosphoserine phosphatase (PSPH), and serine hydroxymethyltransferase 2 (SHMT2)]. B: ISR kinases have been shown to have different impacts on fibrogenesis in hepatic stellate cells (HSCs). PERK was up-regulated in the whole liver as well as in HSCs following carbon tetrachloride (CCl₄) treatment and promoted SMAD2 signaling through the heterogeneous nuclear ribonucleoprotein A1–miR-18A pathway. HRI was also up-regulated in the whole liver following CCl₄ treatment, but the impact of HRI signaling in fibrogenesis remains unknown. Histidine starvation induced up-regulation of GCN2 in HSCs, and GCN2 signaling was found to have antifibrotic effects and protect the liver from damage. Lipopolysaccharide (LPS) exposure induced up-regulation of PKR in HSCs, which promoted inflammation and hepatocyte proliferation. Adapted from Integrated Stress Response template at BioRender.com (Toronto, ON, Canada; 2023). BiP, Ig-binding protein; c/EBPβ, CCAAT enhancer-binding protein; dsRNA, double-stranded RNA.