Monocytes are regarded as precursor cells of the mononuclear phagocytic system, with macrophages being one of the key members of this cellular system. Within the macrophage population, there exist various subpopulations of macrophages, each with its characteristics and functions.

Macrophages have highly diverse roles in maintaining the body’s integrity, including direct participation in pathogen elimination and tissue repair during aseptic inflammatory conditions. Their functions vary across different tissues, playing crucial roles in tissue development, immune response to pathogens, surveillance and monitoring of tissue changes, and maintenance of tissue homeostasis.

In 1971, Gabbiani and his colleagues discovered and characterized myofibroblasts, which are fibroblasts modified to exhibit active contraction in rat wound granulation tissue. This was the first time it had been shown that myofibroblasts promote dermal wound contraction (101). Myofibroblasts are a subset of activated fibroblasts that express molecular markers such as α-SMA and the fibronectin (FN) splice variant extracellular domain (ED)-A FN (102). Hyperactive myofibroblasts, marked by the expression of α-SMA, are primarily responsible for the production of pathogenic collagen tissue fibrosis (7, 103). One of the defining features of myofibroblasts is the development of in vivo stress fibers and contractile force (104). They exhibit morphological and structural characteristics similar to smooth muscle cells, including a flat and irregular morphology, developed cell-ECM interactions, and intercellular space junctions (105). The activation of myofibroblasts is crucial for physiological and pathological tissue repair. Myofibroblasts are the main ECM secretory cells in wound healing and fibrosis and are mainly responsible for the contractility of scar tissue when it matures (106). Myofibroblasts combine the ECM synthesis characteristics of fibroblasts with the cytoskeletal characteristics of contractile smooth muscle cells, regulating connective tissue remodeling (107).

Defining characteristics of myofibroblasts include abundant rough endoplasmic reticulum, moderately developed peripheral myofilaments with focal density, fibronectin, and α-SMA immunostaining (108). In wound granulation tissue, myofibroblasts coexist with prominent endoplasmic reticulum and contractile microfilaments (109). The transformation of myofibroblasts is triggered by integrating neurohumoral, cytokine, growth factor, and mechanical signals from the extracellular environment (110). Myofibroblast differentiation is a critical event for wound healing, tissue repair, and chronic fibrosis (104, 107, 111). At least three local events are required for the differentiation of α-SMA-positive myofibroblasts: accumulation of biologically active TGF-β1, the presence of specialized ECM proteins like ED-A splice variants of fibronectin, and high extracellular stress are caused by the mechanical properties of ECM and cellular remodeling activity (104). The mechanical resistance of the ECM, combined with the action of fibrotic TGF-β1, is the primary stimulus for the differentiation and persistence of myofibroblasts (104).

Myofibroblasts can originate from various sources, including epithelial-mesenchymal transition (EMT) (7), endothelial-mesenchymal transition (112, 113), resident fibroblast or pericyte proliferation (114), and the newly discovered phenomenon of MMT (115). Experimental evidence demonstrates that about 50% of myofibroblast accumulation comes from local proliferation of resident tissue fibroblasts, while approximately 35% comes from bone marrow-derived cells (116). Bone marrow transplantation studies have demonstrated the ability of bone marrow-derived cells to populate distal tissue sites (115, 117, 118).

Myofibroblasts pose hazards in various ways. They are the primary cells responsible for collagen production in tissue fibrosis, and their contraction and ECM remodeling activity play a crucial role in fibrotic diseases (119–121). The fate of myofibroblasts in injured tissues, regardless of their origin, may ultimately determine whether healing occurs normally or progress to end-stage fibrosis (107). Persistent myofibroblast activity leads to progressive tissue fibrosis and distortion of the typical tissue architecture, resulting in organ failure and, ultimately, death (89). While the high contractile force generated by myofibroblasts is beneficial for physiological tissue remodeling, excessive force can be detrimental to tissue function, as seen in hypertrophic scars, fibrotic diseases, and stromal reactions to tumors (111).

Myofibroblasts are also critical components of the matrix reaction around hepatocellular carcinoma, contributing to the extracellular matrix component (122, 123). Activated hepatic stellate cells, portal vein fibroblasts, and bone marrow-derived myofibroblasts have been identified as central collagen-producing cells in the damaged liver (91). They play significant roles in renal fibrosis and are implicated in its pathogenesis (124). Additionally, myofibroblasts contribute to chronic cardiac fibrosis (110). Experimental and clinical observations suggest that myofibroblasts produce pro-invasive signals that may be associated with cancer progression and pain (125). Myofibroblasts present in the matrix reaction of epithelial tumors may contribute to the progression of cancer invasion (126, 127).

PF is a chronic and progressive irreversible pulmonary interstitial disease that poses a significant public threat health (128). It is a characteristic feature of a large class of interstitial lung diseases (ILD) (129, 130). Symptoms of PF typically include shortness of breath, unproductive cough, weight loss, and fatigue due to hypoxia (131). It is characterized by thickened fibrotic alveolar walls leading to impaired gas transfer, restricted ventilatory patterns, and, as a result, respiratory failure (132, 133).

Pre-existing inflammation is a key factor in PF development. Acute lung injury (ALI) and its more severe manifestation, acute respiratory distress syndrome (ARDS), are specific forms of lung inflammation characterized by diffuse alteration of the alveoli, non-cardiogenic lung edema, and local and systemic inflammation (134–137). Inflammatory cascades contribute to the pathogenesis of ALI, resulting in increased permeability of lung capillary vessels and diffuse alveolar damage (138–140). The pathomorphological changes in the lungs during ALI/ARDS include neutrophilic inflammatory infiltration, diffuse alveolar damage, alveolar and interstitial edema, hyalin membrane formation in the exudative phase, and ECM deposition in the proliferative phase (139, 141, 142).

PF is a heterogeneous disease characterized by a distinct pattern of tissue pathology and comprises a large number of chronic respiratory pathologies accompanied by connective tissue growth in various lung compartments, among which interstitial lung disease (ILD) and idiopathic PF (IPF) are the most severe and irreversible ones with progressive fibrosing of the lung parenchyma (130, 143–145). IPF, specifically, is a significant type of pulmonary fibrosis, predominantly affecting the elderly, with high mortality and poor prognosis (146). It can cause dyspnea, cough, impaired lung function, and death (147–149). The prevalence of IPF is around 10 cases per 100,000 population, while ILDs have a prevalence of 19.4 cases per 100,000 population (150, 151). In 2014, two drugs, pirfenidone and nintedanib, were approved by the FDA for the treatment of PF (152). However, effective therapeutic options for PF are still lacking, and current treatments only delay disease progression without providing a complete cure. Moreover, these drugs have undesirable side effects, such as gastric and intestinal bleeding and severe diarrhea. Lung transplantation is the last resort for patients, offering some extension of lifespan, but it is not accessible to most individuals. Therefore, studying the molecular mechanisms underlying the transition from acute lung inflammation to PF and identifying new molecular markers and promising therapeutic targets for preventing PF development remain important objectives.

Macrophages, as innate immune cells with antibacterial and phagocytic activity, play a significant role in PF. They are the most abundant immune cell population, accounting for about 70% (153). They are widely distributed in the lung and alveolar tissue and are involved in almost all the physiological and pathological processes of the lung (154). They are the host lung defense, indispensable paramount sentry (155, 156), and also play a vital role in the pathogenesis of PF. Macrophage infiltration is observed in PF (157). Macrophages are involved in all stages of lung injury and repair and can both promote and inhibit fibrosis. They play an essential role in the removal of lung pathogens clearance and maintaining homeostasis (157, 158). The pathogenic role of macrophages in PF has been investigated in multiple studies, involving reactive oxygen species generation (159–161), stimulation of proteinase-activated receptors (162, 163), and secretion of pro-fibrotic cytokines (164, 165).

There are three main types of pulmonary macrophages: alveolar macrophages (AM), interstitial macrophages (IM), and bronchial macrophages (BM), with AM accounting for more than 90% (166). Different subtypes of macrophages play distinct roles in lung injury, repair, and fibrosis (167). Single-cell sequencing of lung tissue from patients with PF have confirmed that alveolar macrophages play an essential role in PF (168–170). Alveolar macrophages are the first cells to come into contact with external pathogens and irritants, initiating and later resolving lung immune responses. Additionally, macrophages have other organ-specific functions, such as surfactant utilization and absorption of apoptosing and destroying cells (171–174). Monocyte-derived macrophages are key drivers of PF and supplement alveolar macrophages that are lost immediately upon injury (175, 176).

The effect of macrophages on PF is mainly related to their polarization, which occurs during the repeated damage and abnormal repair of alveolar epithelial cells (177, 178). Epithelial apoptosis is a critical component of fibrotic disease in many organs, including the lung (179, 180). Down-regulating the pro-fibrosis activity of alveolar macrophages or depleting this group of cells can effectively treat experimental PF (181–183). Macrophages can polarize into either a pro-inflammatory M1 phenotype or an alternatively activated M2 phenotype, depending on the microenvironment in which they reside (184). In response to lung injury, macrophages undergo a transition into pro-inflammatory M1 phenotypes and begin to secrete pro-inflammatory cytokines (TNF-α, IL-6, IL-1) and chemokines (IL-8, CCL7, CCL2), which leads to the increased chemotaxis and progressive enrichment of alveolar spaces by monocytes and neutrophils (185), which aggravate the pulmonary inflammatory response. On the other hand, M2 polarization releases various cytokines, such as TGF-β1 and IL-10, promoting the generation of myofibroblasts and the deposition of extracellular matrix, ultimately leading to PF.

During tissue damage and early inflammation stages, the activation of M1 macrophages promotes inflammation through extracellular matrix-degrading MMP and pro-inflammatory cytokines. An active cytokine environment, including Th1 cytokines, IL2, IFN-γ, and TNF-α, drives M1 macrophage activation. In contrast, other types of interstitial lung diseases (ILDs), including PF, often have a higher proportion of anti-inflammatory M2 macrophages (186) ( Figure 2 ).

In the progression of PF, M1, and M2, macrophages are recruited to the site of the lung tissue injury site to regulate the fibrotic process after basement membrane destruction. M1 macrophages play a crucial role in matrix degradation by directly and indirectly producing MMP and various anti-fibrotic cytokines, essential for ECM remodeling and help reduce the pathological fibrous proliferation observed in late ALI (187). In contrast, M2 macrophages promote fibrous proliferation and ECM deposition in lung tissue (188, 189). Therefore, the degree of PF depends on the balance between M1 and M2 macrophages in the local microenvironment of lung tissue injury. Studies have shown that macrophages, predominantly M2 macrophages, contribute to the pathogenesis of PF (155, 190). M2 macrophages are the primary source of TGF-β1 and platelet-derived growth factors that induce fibroblast differentiation into myofibroblasts, initiating PF (191). Macrophage subsets may regulate fibrosis by differentiating into myofibroblasts, acting as sources of cytokines and growth factors with fibrotic properties, and secreting proteases involved in matrix remodeling (192). Therefore, the number and phenotype of macrophages are considered essential for the pathological process of PF (193, 194). While macrophages are essential for lung defense, they can also lead to tissue damage (195). Different subtypes of macrophages play distinct roles in lung injury, repair, and fibrosis (196).

The main morphological characteristics of PF, such as ECM deposition and remodeling of lung architecture, are consequences of a disbalance between two physiological processes in the lungs: (1) proliferation/apoptosis of fibroblasts and myofibroblasts; (2) synthesis/degradation of ECM components (197). These processes are closely interconnected, and the disruption of fibroblast and myofibroblast functioning is the primary driver behind the imbalance of ECM homeostasis and the development of PF. The fibroblastic phenotype present in that diseased lung primarily by the production of several soluble factors, such as TGF-β, PDGF, VEGF, and thrombospondin 1, which can differentiate resident fibroblast into myofibroblasts (170, 181, 195). Regardless of the source of lung fibroblasts, myofibroblasts, which resemble smooth muscle cells in terms of their contractile ability and expression of α-SMA, are considered the key cells in PF development.

Myofibroblasts are the primary effectors responsible for the excessive production of collagen and other extracellular matrix proteins in fibrotic lungs (104, 198). These contractile fibroblasts express α-SMA and abnormally proliferate in PF. They play a significant role in the occurrence and progression of PF by synthesizing and secreting large amounts of ECM components, such as collagen (I, III, IV, V, and VI), fibronectin, and laminin (199–201), making them critical in regulating the progression of PF. Myofibroblasts have also been found to secrete or release various proteins, lipids, and nucleic acid molecules that contribute to the pathological characteristics of other cell types in fibrotic lungs (129).

The accumulation of myofibroblasts is considered a marker of PF (202). Current research indicates that myofibroblasts involved in PF originate from several sources, including the proliferation and differentiation of resident fibroblasts, the recruitment of circulating fibroblasts to injury sites in organs, endothelial-mesenchymal transformation, and epithelial-mesenchymal transformation (203–205). The synthesis of pathogenic collagen by myofibroblasts, as the main effector of tissue fibrosis, and the process of MMT are essential regardless of the etiology of fibrosis (3, 206–208). Myofibroblast transdifferentiation is a marker of the fibrotic response. Evidence suggests that macrophages are involved in regulating fibrotic responses, with pulmonary myofibroblasts being the primary target for the development of new therapies for IPF (104, 198).

As described earlier, fibrosis is defined by the excessive accumulation of fibrous connective tissue in and around inflamed or damaged tissue, which can lead to permanent scarring, organ malfunction, and, ultimately, death, as seen in end-stage liver disease, kidney disease, IPF, and heart failure (91, 209). The development of PF involves genes and molecular pathways that primarily participate in pre- and postnatal lung development (210, 211). The key pathophysiological events of IPF include repetitive alveolar epithelial cell injury, the presence or absence of local inflammation, impaired epithelial-mesenchymal crosstalk, and subsequent fibroblast-to-myofibroblast activation (212–214). These mechanisms are mediated by abnormally activated signaling molecules that drive the process of fibrosis, such as TGF-β, Wnt/β-catenin, hedgehog, Notch, and fibroblast growth factor signaling pathways, with the TGF-β signaling pathway being the most critical (215, 216). While most of these pathways are inactive in the adult organism, they become active during tissue regeneration, and the chronic pathological activation of these signaling pathways is associated with injury restoration processes in all organs, including the lungs (210, 217, 218). Furthermore, a recent study demonstrated that nintedanib, one of the FDA-approved anti-fibrotic drugs, modulates TGF-β, VEGF, and Wnt/β-catenin signaling pathways, further supporting the central role of these pathways in PF development (219) ( Figure 3 ).

MMT has been shown to contribute to interstitial fibrosis in patients with chronic renal allograft injury, a mouse model of unilateral ureteral obstruction (UUO), and progressive chronic kidney disease (1). Macrophages expressing CD68+ and α-SMA+ markers play a significant role in collagen production, particularly collagen I, and are associated with lung injury and interstitial fibrosis (12, 196, 289). MMT cells with M2 phenotype have been found to contribute to PF in animal models, including the lungs of rats with unilateral ureteral obstruction (UUO) (1, 196, 289). Eplerenone reduced the accumulation of MMT cells in the lung. In UUO rat lung fibrosis, UUO-induced lung injury, and fibrosis, MMT cells were found to account for the myofibroblast group, confirming that MMT plays a role in PF. These MMT cells in the lung exhibited an apparent M2 phenotype, indicating that the MMT process may be an important pathway leading to PF (12).

MMT plays a crucial role in the progression of chronic inflammation to pathological fibrosis, and the severity of interstitial fibrosis is closely related to the number of MMT cells (1, 51, 196, 289). MMT contributes to an increase in the population of myofibroblasts in the lungs, which have a strong proliferative capacity and further promote the proliferation of fibroblasts. Myofibroblasts, a subset of activated fibroblasts, are primarily responsible for organ deformation by inducing the deposition of fibrous collagen during tissue fibrosis (290). Upon transdifferentiation, myofibroblasts secrete various components of the extracellular matrix, including collagen, leading to excessive deposition of extracellular matrix in the lungs, a key pathological characteristic of PF. This excessive deposition disrupts the normal alveolar structure, resulting in alveolar collapse and reduced lung function.

The pro-fibrotic cytokine TGF-β1 is an essential initiator of organ inflammation and fibrosis by activating the downstream Smad signaling cascade, especially the Smad3 signaling cascade (6). Smad3 is a crucial transcription factor for classical TGF-β1 signal transduction (234, 291). The inhibition of MMT by targeting cytokines such as TGF-β1 or blocking the Smad3 signal pathway can slow down the process of PF. Moreover, the non-receptor tyrosine kinase Src, which can be activated by TGF-β1, has been closely associated with tissue fibrosis. Inhibition of Src has been shown to block MMT in animal models and reduce the severity of PF induced by bleomycin (292–294). However, further research is needed to fully understand the role of MMT in Src-mediated PF and explore the potential of Src-targeted therapy for blocking MMT and treating PF.

In summary, MMT plays an essential role in the process of PF, which accelerates the process of PF by promoting the transdifferentiation of macrophages into myofibroblasts. Inhibiting the MMT process represents a potential therapeutic target for anti-fibrotic treatment. Future studies should focus on elucidating the regulatory mechanisms of MMT and its specific role in PF to provide novel insights and treatment strategies for PF. A comprehensive treatment approach considering various factors, including inflammation control, inhibition of the fibrotic process, and improvement of lung function, is essential for effectively managing PF.

Lung cancer is the leading cause of death worldwide. For decades, it has remained the second most common cancer and the leading cause of cancer deaths, accounting for about 11.4% of new cancer cases and 18% of cancer deaths globally in 2020. Cancer‐associated fibroblasts (CAFs) are essential in tumor microenvironment (TME) driven cancer progression. CAFs are the most prominent stromal components (295). CAFs, a subtype of myofibroblasts, contribute to the malignancy and advancement of cancer (296). Cancer cells possess heterogeneity, versatility, and adaptability, resulting in primary and secondary drug resistance (297). The degree of macrophage-myofibroblast transition (MMT) has been found to be closely associated with the prognosis of certain cancers (297). MMT is an essential source of CAFs in non-small cell lung cancer (NSCLC). The hematopoietic transcription factor Runx1 has been identified as a critical regulator of MMT in cancer patients. Inhibition of Runx1, macrophage-specific and systemic, effectively blocks MMT-driven tumor formation in vivo, making it a potential therapeutic target for eliminating pro-tumor CAFs in patients with NSCLC (298).

Myofibroblasts can secrete various growth factors and cytokines, such as TGF-β and PDGF, which can stimulate the proliferation and migration of tumor cells and promote the progress of cancer. The TGF-β/Smad3 signal pathway is a critical regulatory factor promoting tumor microenvironment (299–301). It is essential to initiate MMT in chronic inflammatory diseases, including cancer. The MMT process and tumor growth in lung cancer are tightly regulated by Smad3 (302). TGF-β/Smad3 signal transduction is a key regulatory factor in the tumorigenic microenvironment. Recent evidence indicates that TGF-β can trigger the M1/M2 polarization of TAMs by activating Smad2/3 and PI3K/AKT pathways, thus enhancing the transcription of tumorigenic effectors such as IL-10, VEGFA, and CXCR4 (303). However, targeting Smad3 also inhibits T cell anti-cancer immunity, highlighting the complexity of potential therapeutic strategies (5, 207, 293, 304).

MMT is a critical pathophysiological process within the tumor microenvironment, leading to the generation of myofibroblasts that secrete inflammatory factors and fibrosis-related proteins in tumor tissues, promoting inflammation and fibrosis changes in the tumor microenvironment (305). Co-expression of TAM markers (CD68) and CAF markers (α-SMA) has been observed in lung, renal, and prostate cancers, indicating the presence of MMT in these types of cancer (1, 2, 196, 301). An interesting phenomenon in MMT is the further differentiation of TAMs into CAFs. Silencing Smad3 specifically in macrophages effectively inhibits MMT and consequently impedes CAF-mediated cancer progression. These findings highlight the significance of macrophage Smad3 in regulating CAFs through MMT, providing a specific therapeutic target for cancer immunotherapy (5). Given the critical role of MMT in cancer progression, inhibiting MMT may become a new target for cancer treatment. By blocking the process of MMT, the support of the tumor microenvironment can be weakened, the proliferation and migration of cancer cells can be inhibited, and the prognosis of cancer can be improved. Therefore, it is significant to study the mechanism and intervention strategy of MMT for developing new cancer treatment methods and improving cancer prognosis.