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Review
. 2020 Jan;16(1):11-31.
doi: 10.1038/s41584-019-0324-5. Epub 2019 Dec 2.

Evasion of apoptosis by myofibroblasts: a hallmark of fibrotic diseases

Boris Hinz  1 , David Lagares  2   3   4
Affiliations
Review

Evasion of apoptosis by myofibroblasts: a hallmark of fibrotic diseases

Boris Hinz et al. Nat Rev Rheumatol. 2020 Jan.

Abstract

Organ fibrosis is a lethal outcome of autoimmune rheumatic diseases such as systemic sclerosis. Myofibroblasts are scar-forming cells that are ultimately responsible for the excessive synthesis, deposition and remodelling of extracellular matrix proteins in fibrosis. Advances have been made in our understanding of the mechanisms that keep myofibroblasts in an activated state and control myofibroblast functions. However, the mechanisms that help myofibroblasts to persist in fibrotic tissues remain poorly understood. Myofibroblasts evade apoptosis by activating molecular mechanisms in response to pro-survival biomechanical and growth factor signals from the fibrotic microenvironment, which can ultimately lead to the acquisition of a senescent phenotype. Growing evidence suggests that myofibroblasts and senescent myofibroblasts, rather than being resistant to apoptosis, are actually primed for apoptosis owing to concomitant activation of cell death signalling pathways; these cells are poised to apoptose when survival pathways are inhibited. This knowledge of apoptotic priming has paved the way for new therapies that trigger apoptosis in myofibroblasts by blocking pro-survival mechanisms, target senescent myofibroblast for apoptosis or promote the reprogramming of myofibroblasts into scar-resolving cells. These novel strategies are not only poised to prevent progressive tissue scarring, but also have the potential to reverse established fibrosis and to regenerate chronically injured tissues.

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Conflict of interest statement

Competing interests

D.L. declares that he has received research funding from Boehringer Ingelheim, Indalo Therapeutics and Unity Biotechnology. D.L. also has a financial interest in Mediar Therpeutics, which is developing treatments for organ fibrosis. D.L.’s interests were reviewed and are managed by MGH and Partners HealthCare in accordance with their conflict of interest policies. B.H. declares no competing interests.

Figures

Fig. 1 |
Fig. 1 |. Origin, functions and fate of myofibroblasts during tissue repair and fibrosis.
a | Activated myofibroblasts are important orchestrators of wound healing. During tissue injury, fibroblasts can differentiate into myofibroblasts. In the skin, two dermal fibroblast populations (reticular and papillary), which are derived from a common embryonic origin, have distinct functions during tissue repair. Whereas papillary fibroblasts do not express α-smooth muscle actin (α-SMA) and are involved in the regeneration of skin structures such as hair follicles, reticular fibroblasts differentiate into collagen-producing α-SMA+ myofibroblasts by the action of biomechanical (extracellular matrix (ECM) stiffness) and biochemical factors such as transforming growth factor-β1 (TGFβ1) and connective tissue growth factor (CTGF). In specific conditions and experimental models, myofibroblasts can also arise from epithelial and endothelial cells via epithelial-to-mesenchymal transition or endothelial-to-mesenchymal transition, respectively, and from mesenchymal stem cells, pericytes and pre-adipocytes or adipocytes. Myofibroblasts are characterized by increased synthesis of ECM proteins and by the expression of α-SMA, which confers a contractile phenotype that promotes wound closure. Activated myofibroblasts also produce anti-inflammatory cytokines such as TGFβ1 and IL-10, thereby shifting macrophages towards an ECM-degrading phenotype. During the resolution of wound healing, myofibroblasts can follow several different cell fates. Myofibroblasts can die via apoptosis, a process induced by ECM softening (stress release) or soluble pro-apoptotic factors such as IL-1β, fibroblast growth factor 1 (FGF1) and prostaglandin E2 (PGE2). Alternatively, they can deactivate, become scar-resolving fibroblasts or become senescent via the action of CCN family member 1 (CCN1); these cell types all participate in ECM degradation in partnership with pro-resolving macrophages. b | Myofibroblasts and senescent myofibroblasts can also escape apoptosis and continue to remodel tissue beyond repair, which results in pathological scarring and the development of fibrotic disease. LOXs, lysyl oxidases; MMPs, matrix metalloproteinases; TIMPs, tissue inhibitors of metalloproteinases.
Fig. 2 |
Fig. 2 |. Intrinsic and extrinsic apoptosis pathways.
Apoptosis is controlled by two distinct yet connected pathways. The intrinsic pathway is triggered by intracellular death stimuli, such as DNA damage, radiation, nutrient deprivation, oxidative stress or oncogene activation, which induce apoptosis by promoting mitochondrial outer membrane permeabilization (MOMP) and cytochrome c (cyt c)-dependent activation of caspase-9, which then activates caspase-3 and caspase-7. This pathway of apoptosis is tightly controlled by the BCL-2 family of proteins, which are classified on the basis of their structural homology and function into sensitizers, pro-survival proteins, activators and effectors. MOMP is controlled by homo-oligomerization of the effector proteins BCL-associated X protein (BAX) and BCL-2 homologous antagonist/killer (BAK), which are activated by the activator proteins BCL-2-like protein 11 (BCL2L11; also known as BIM), p53-upregulated modulator of apoptosis (PUMA) and BH3-interacting domain death agonist (BID). Pro-survival proteins such as BCL-2, BCL-XL, BCL-W, induced myeloid leukaemia cell differentiation protein MCL1 (MCL1) and BCL-2-related protein A1 (BCL2A1; also known as BFL1) can bind and sequester both activators and effectors, thereby preventing their interaction and the induction of MOMP. Sensitizer proteins (such as BCL-2-associated death promoter (BAD), BCL-2 interaction killer (BIK), BCL-2 modifying factor (BMF), activator of apoptosis harakiri (HRK) and phorbol-12-myristate-13-acetate-induced protein 1 (PMAIP1; also known as NOXA) are a distinct group of proteins that promote apoptosis by binding and blocking pro-survival proteins, thereby releasing formerly bound activator and effector proteins. The extrinsic pathway is initiated by extracellular ligands such as FAS ligand (FASL), TNF and TNF ligand superfamily member 10 (TNFSF10; also known as TRAIL), which bind to the cell-surface receptors FAS, TNF receptors (TNFRs) and death receptor 4 (DR4) and DR5, respectively. The signal is transmitted via FAS-associated death domain (FADD), which triggers the activation of caspase-8 and caspase-10, ultimately initiating apoptosis through cleavage and activation of pro-caspase-3 and pro-caspase-7. BH3, BCL-2 homology domain 3; XIAP, X-linked inhibitor of apoptosis protein.
Fig. 3 |
Fig. 3 |. Mitochondrial priming of myofibroblasts.
Mitochondrial priming refers to the proximity of mitochondria to the apoptosis threshold and is determined by the relative expression of pro-apoptotic (effectors, activators and sensitizers) and pro-survival members of the BCL-2 family of proteins. ‘Unprimed’ fibroblasts express low amounts of activators and/or effectors, leading to an apoptosis-resistant phenotype. Conversely, high expression of sensitizers, activators and/or effectors results in high mitochondrial priming, ultimately triggering cytochrome-c-mediated mitochondrial outer membrane permeabilization and apoptosis. However, myofibroblasts can survive with high mitochondrial priming if a pro-survival mechanism is activated. In this cellular state, known as ‘primed for death’, cells are poised to die and become dependent on one or more pro-survival proteins to sequester pro-apoptotic proteins and ensure survival. BCL-2 homology domain 3 (BH3) mimetic drugs can trigger the intrinsic pathway of apoptosis in primed-for-death myofibroblasts by binding to pro-survival proteins, resulting in the release of activator proteins, which can then bind to and activate effector proteins.
Fig. 4 |
Fig. 4 |. Molecular control of myofibroblast activation and survival.
a | Myofibroblast activation is jointly promoted by biomechanical and biochemical cues. Biomechanical signalling induced by extracellular matrix (ECM) stiffness activates mechanotransduction pathways that directly control α-smooth muscle actin (α-SMA) transcription. These pathways involve β1 integrin, focal adhesion kinase (FAK) and RHO-associated protein kinase (ROCK), together with increasing traction forces in fibroblasts. Increased actomyosin activity causes the nuclear translocation of myocardin-related transcription factor (MRTF), as well as the transcriptional co-activators yes-associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ), which regulate α-SMA expression by binding and activating other transcription factors, such as serum response factor (SRF), TEA domain family member (TEAD), T cell factor/lymphoid enhancer-binding factor (TCF/LEF) and β-catenin. ECM stiffness and mechanical forces also regulate force-dependent activation of latent transforming growth factor-β1 (TGFβ1) by increasing resistance to traction forces generated by fibroblasts. In this mechanism, extracellular latent TGFβ1 (TGFβ1 with its latency-associated peptide (L AP)) is released from latent TGFβ1 binding protein stores when αv integrins respond to mechanical pulling forces. Once activated, TGFβ1 binds to TGFβ receptors and promotes canonical mothers against decapentaplegic homolog 3 (SMAD3) activation. Activated SMAD3 binds to SMAD4 and translocates to the nucleus, where it binds to SMAD-binding elements in the promoters of fibrogenic genes, such as ACTA2 (encoding α-SMA). Together, myofibroblast activation is controlled by both the TGFβ–SMAD pathway, as well as biomechanical pathways such as integrin–FAK–ROCK–MRTF–YAP–TAZ signalling. b | Mitochondria in activated myofibroblasts contain large amounts of pro-apoptotic factors, which force these cells to activate pro-survival mechanisms to ensure survival. The intrinsic pathway of apoptosis is directly inhibited by TGFβ1 via activation of ABL signalling, which increases the amount of the pro-survival proteins BCL-2 and BCL-XL. TGFβ1 also blocks the intrinsic pathway by inhibiting the pro-apoptotic protein BCL-2-associated death promoter (BAD) via the FAK–PI3K–AKT signalling pathway, and TGFβ1-mediated FAK signalling blocks the extrinsic pathway of apoptosis by inhibiting sphingomyelinase (ASMase), an enzyme that controls FAS-mediated apoptosis by generating ceramide from sphingomyelin. Biomechanical signalling similarly inhibits the intrinsic pathway via the biomechanically regulated TGFβ–FAK–YAP1–TAZ–BCL-XL and TGFβ–ROCK–MRTF–BCL-2 pathways. Moreover, ECM stiffness induces expression of the microRNAs miR-21 and miR-29a, which promote the survival of myofibroblasts by increasing the expression of pro-survival BCL-2 proteins. Both ECM stiffness and TGFβ1 can also block apoptosis by increasing amounts of X-linked inhibitor of apoptosis protein (XIAP), a direct caspase inhibitor. AKT, pro-survival protein kinase B; BAK, BCL-2 homologous antagonist/killer; BAX, BCL-associated X protein; BID, BH3-interacting domain death agonist; BIM, BCL-2-interacting mediator of cell death (also known as BCL2L11); PI3K, phosphoinositide 3-kinase; TGFβRII, transforming growth factor-β receptor II.
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