Fibroblasts, once perceived as a uniform cell type, are now recognized as a mosaic of distinct populations with specialized roles in tissue homeostasis and pathology. Here we provide a global overview of the expanding compendium of fibroblast cell types and states, their diverse lineage origins and multifaceted functions across various human organs. By integrating insights from developmental biology, lineage tracing and single-cell technologies, we highlight the complex nature of fibroblasts. We delve into their origination from embryonic mesenchyme and tissue-resident populations, elucidating lineage-specific behaviours in response to physiological cues. Furthermore, we highlight the pivotal role of fibroblasts in orchestrating tissue repair, connective tissue remodelling and immune modulation across diverse pathologies. This knowledge is essential to develop novel fibroblast-targeted therapies to restore steady-state fibroblast function and advance regenerative medicine strategies across multiple diseases.

Reactive oxygen species (ROS) play a dual role in cancer progression, acting as both signaling molecules and drivers of oxidative damage. Emerging evidence highlights the intricate interplay between ROS, microRNAs (miRNAs), and exosomes within the tumor microenvironment (TME), forming a regulatory axis that modulates immune responses, angiogenesis, and therapeutic resistance. In particular, oxidative stress not only stimulates exosome biogenesis but also influences the selective packaging of redox-sensitive miRNAs (miR-21, miR-155, and miR-210) via RNA-binding proteins such as hnRNPA2B1 and SYNCRIP. These miRNAs, delivered through exosomes, alter gene expression in recipient cells and promote tumor-supportive phenotypes such as M2 macrophage polarization, CD8+ T-cell suppression, and endothelial remodeling. This review systematically explores how this ROS-miRNA-exosome axis orchestrates communication across immune and stromal cell populations under hypoxic and inflammatory conditions. Particular emphasis is placed on the role of NADPH oxidases, hypoxia-inducible factors, and autophagy-related mechanisms in regulating exosomal output. In addition, we analyze the therapeutic relevance of natural products and herbal compounds-such as curcumin, resveratrol, and ginsenosides-which have demonstrated promising capabilities to modulate ROS levels, miRNA expression, and exosome dynamics. We further discuss the clinical potential of leveraging this axis for cancer therapy, including strategies involving mesenchymal stem cell-derived exosomes, ferroptosis regulation, and miRNA-based immune modulation. Incorporating insights from spatial transcriptomics and single-cell analysis, this review provides a mechanistic foundation for the development of exosome-centered, redox-modulating therapeutics. Ultimately, this work aims to guide future research and drug discovery efforts toward integrating herbal medicine and redox biology in the fight against cancer.

Cancer-associated fibroblasts (CAFs) are integral components of the tumor microenvironment playing key roles in tumor progression, metastasis, and therapeutic resistance. However, challenges persist in understanding their heterogeneity, origin, and functional diversity. One major obstacle is the lack of standardized naming conventions for CAF subpopulations, with current systems failing to capture their full complexity. Additionally, the identification of CAFs is hindered by the absence of specific biomarkers, limiting the precision of diagnostic and therapeutic strategies. In vitro culture conditions often fail to maintain the in vivo characteristics of CAFs, which complicates their study and the translation of findings to clinical practice. Although current detection methods, such as antibodies, mRNA probes, and single-cell transcriptomics, offer insights into CAF biology, they lack standardization and fail to provide reliable quantitative measures. Furthermore, the dynamic interactions between CAFs, tumor cells, and immune cells within the TME remain insufficiently understood, and the role of CAFs in immune evasion and therapy resistance is an area of ongoing research. Understanding how CAFs influence drug resistance and the immune response is essential for developing more effective cancer therapies. This review aims to provide an in-depth analysis of the challenges in CAF research, propose future research directions, and emphasize the need for improved CAF-targeted therapeutic strategies. By addressing these gaps, it seeks to highlight the potential of CAFs as targets for overcoming therapeutic resistance and enhancing the efficacy of cancer treatments.

Systemic Sclerosis (SSc) is a systemic autoimmune disease of unknown etiology characterized by the development of frequently progressive cutaneous and internal organ fibrosis accompanied by severe vascular alterations. The pathogenesis of SSc is highly complex and, despite extensive investigation, has not been fully elucidated. Numerous studies have suggested that unknown etiologic factors cause multiple alterations in genetically receptive hosts, leading to SSc development and progression. These events may be functionally and pathologically interconnected and include: 1) Structural and functional microvascular and endothelial cell abnormalities; 2) Severe oxidative stress and high reactive oxygen species (3); Frequently progressive cutaneous and visceral fibrosis; 4) Transdifferentiation of various cell types into activated myofibroblasts, the cells ultimately responsible for the fibrotic process; 5) Establishment of a chronic inflammatory process in various affected tissues; 6) Release of cytokines, chemokines, and growth factors from the inflammatory cells; 7) Abnormalities in humoral and cellular immunity with the production of specific autoantibodies; and 8) Epigenetic alterations including changes in multiple non-coding RNAs. These events manifest with different levels of intensity in the affected organs and display remarkable individual variability, resulting in a wide heterogeneity in the extent and severity of clinical manifestations. Here, we will review some of the recent studies related to SSc pathogenesis.

Tendon ruptures in humans have often been studied during the chronic phase of injury, particularly in the context of rotator cuff disease. However, the early response to acute tendon ruptures remains less investigated. Quadriceps tendons, which require prompt surgical treatment, offer a model to investigate this early response. Therefore, this study aimed to explore the early cellular changes in ruptured compared to healthy human quadriceps tendons. Quadriceps tendon samples were collected from patients undergoing fracture repair (healthy) or tendon repair surgery (collected 7-8 days post-injury). Nuclei were isolated for single-nucleus RNA sequencing, and comprehensive transcriptomic analysis was conducted. The transcriptomes of 12,808 nuclei (7268 from healthy and 5540 from ruptured quadriceps tendons) were profiled, revealing 12 major cell types and several cell subtypes and states. Rupture samples showed increased expression of genes related to extracellular matrix organisation and cell cycle signalling, and a decrease in expression of genes in lipid metabolism pathways. These changes were predominantly driven by gene expression changes in the fibroblast, vascular endothelial cell (VEC), mural cell, and macrophage populations: fibroblasts shift to an activated phenotype upon rupture and there is an increase in the proportion of capillary and dividing VECs. A diverse immune environment was observed, with a shift from homeostatic to activated macrophages following rupture. Cell-cell interactions increased in number and diversity in rupture, and primarily involved fibroblast and VEC populations. Collectively, this transcriptomic analysis suggests that fibroblasts and endothelial cells are key orchestrators of the early injury response within ruptured quadriceps tendon. KEY POINTS: Tendon ruptures in humans have regularly been studied during the chronic phase of injury, but less is known about the early injury response after acute tendon ruptures. This study explored the early cellular changes in ruptured compared to healthy human quadriceps tendons at single-cell resolution. Fibroblasts and endothelial cells seem to be the key orchestrators of the early injury response within ruptured quadriceps tendon. Therefore, these cell types are obvious targets for interventions to enhance tendon healing. Overall, this study highlights that the development of more effective therapeutic options for tendon injury requires better understanding of the cellular, extracellular, and mechanical landscape of tendon tissue.

Skeletal muscle fibrosis is strongly associated with the differentiation of its resident multipotent fibro/adipogenic progenitors (FAPs) toward the myofibroblast phenotype. Although transforming growth factor type β (TGF-β) signaling is well-known for driving FAPs differentiation and fibrosis, due to its pleiotropic functions its complete inhibition is not suitable for treating fibrotic disorders such as muscular dystrophies. Here, we describe that TGF-β operates through the mechanosensitive transcriptional regulators Yes-associated protein (YAP)/ transcriptional coactivator with PDZ-binding motif (TAZ) to determine the myofibroblast fate of FAPs and skeletal muscle fibrosis. Spatial transcriptomics analyses of dystrophic and acute injured muscles showed that areas with active fibrosis and TGF-β signaling displayed high YAP/TAZ activity. Using a TGF-β-driven fibrotic mouse model, we found that activation of YAP/TAZ in activated FAPs is associated with the fibrotic process. Mechanistically, primary culture of FAPs reveals the remarkable ability of TGF-β1 to activate YAP/TAZ through its canonical SMAD3 pathway. Moreover, inhibition of YAP/TAZ, either by disrupting its activity (with Verteporfin) or cellular mechanotransduction (with the Rho inhibitor C3 or soft matrices), decreased TGF-β1-dependent FAPs differentiation into myofibroblasts. In vivo, administration of Verteporfin in mice limits the deposition of collagen and fibronectin, and the activation of FAPs during the development of fibrosis. Overall, our work provides robust evidence for considering YAP/TAZ as a potential target in muscular fibroproliferative disorders.NEW & NOTEWORTHY The understanding of the nuclear factors governing the differentiation of muscular fibro/adipogenic progenitors (FAPs) into myofibroblasts is in its infancy. Here, we comprehensively elucidate the status, regulation, and role of the mechanotransducers Yes-associated protein (YAP)/transcriptional coactivator with PDZ-binding motif (TAZ) in the muscular fibrotic process. Our findings reveal that inhibiting cellular mechanotransduction limits FAP differentiation and the extent of muscular fibrosis exerted by transforming growth factor type β (TGF-β). This research shed new lights on the molecular mechanisms dictating the cell fate of FAPs and the muscular fibrosis.

Functional revascularization is key to stroke recovery and requires remodeling and regeneration of blood vessels around which is located the brain's only stromal compartment. Stromal progenitor cells (SPCs) are critical for tissue regeneration following injury in many organs, yet their identity in the brain remains elusive. Here we show that the perivascular niche of brain SPCs includes pericytes, venular smooth muscle cells and perivascular fibroblasts that together help cerebral microvasculature regenerate following experimental stroke. Ischemic injury triggers amplification of pericytes and perivascular fibroblasts in the infarct region where they associate with endothelial cells inside a reactive astrocyte border. Fate-tracking of Hic1+ SPCs uncovered a transient functional and transcriptional phenotype of stroke-activated pericytes and perivascular fibroblasts. Both populations of these cells remained segregated, displaying distinct angiogenic and fibrogenic profiles. Therefore, pericytes and perivascular fibroblasts are distinct subpopulations of SPCs in the adult brain that coordinate revascularization and scar formation after injury.

Tendinopathy is a condition characterized by persistent tendon pain, structural damage, and compromised functionality. Presently, the treatment for tendinopathy remains a formidable challenge, partly because of its unclear pathogenesis. Tendon stem/progenitor cells (TSPCs) are essential for tendon homeostasis, regeneration, remodeling, and repair. An innovative theory has been previously proposed, with insufficient evidence, that the erroneous differentiation of TSPCs may constitute one of the fundamental mechanisms underpinning tendinopathy. Over the past few years, there has been accumulating evidence for plausibility of this theory. In this review, we delve into alterations in the differentiation potential of TSPCs and the underlying mechanisms in the context of injury-induced tendinopathy, diabetic tendinopathy, and age-related tendinopathy to provide updated evidence on the erroneous differentiation theory. Despite certain limitations inherent in the existing body of evidence, the erroneous differentiation theory emerges as a promising and highly pertinent avenue for understanding tendinopathy. In the future, advanced methodologies will be harnessed to further deepen comprehension of this theory, paving the way for prospective developments in clinical therapies targeting TSPCs for the management of tendinopathy.

Gynecologic cancer, a prevalent and debilitating disease affecting women worldwide, is characterized by the uncontrolled proliferation of cells in the reproductive organs. The complex etiology of gynecologic cancer encompasses multiple subtypes, including cervical, ovarian, uterine, vaginal, and vulvar cancers. Despite optimal treatment strategies, which typically involve cytoreductive surgery and platinum-based chemotherapy, gynecologic cancer frequently exhibits recalcitrant relapse and poor prognosis. Recent studies have underscored the significance of the tumor microenvironment in ovarian carcinogenesis, particularly with regards to the discovery of aberrant genomic, transcriptomic, and proteomic profiles. Within this context, cancer-associated fibroblasts (CAFs) emerge as a crucial component of the stromal cell population, playing a pivotal role in oncogenesis and cancer progression. CAF-derived exosomes, small extracellular vesicles capable of conveying biological information between cells, have been implicated in a range of tumor-related processes, including tumorigenesis, cell proliferation, metastasis, drug resistance, and immune responses. Furthermore, aberrant expression of CAF-derived exosomal noncoding RNAs and proteins has been found to strongly correlate with clinical and pathological characteristics of gynecologic cancer patients. Our review provides a novel perspective on the role of CAF-derived exosomes in gynecologic cancer, highlighting their potential as diagnostic biomarkers and therapeutic targets.

Background/Objectives: Desmoid-type fibromatosis (DTF) is a locally invasive tumor composed of myofibroblast-like cells and collagen; it does not metastasize but can cause significant local morbidity. Most sporadic cases are associated with mutations in the CTNNB1 gene, which encodes beta-catenin. Various treatments have been used with differing efficacy and toxicity profiles. At our institution, pegylated liposomal doxorubicin (PLD) has become the preferred treatment for patients with DTF. We aim to describe our experience using PLD in patients with DTF who require treatment. Methods: A retrospective review of 61 DTF patients (41 females, 20 males) treated between 2000 and 2023 was conducted to assess the efficacy and toxicity of PLD. Results: Of the 26 patients treated with PLD, 23 had follow-up clinical data to assess benefit. Twenty-one showed clinical benefit, and only one progressed. Two patients did not benefit from PLD due to infusion reactions and chose alternative therapies. The primary side effect of PLD was hand-foot syndrome (HFS), but dose reduction and extended intervals allowed most patients to tolerate treatment. Other treatments, such as methotrexate, vinblastine/vinorelbine, and sorafenib, also showed activity but had significant toxicities, including severe HFS, malaise, and hypertension. Interestingly, 31 out of 61 patients had a pre-existing history of psychiatric conditions (primarily depression and anxiety), and 6 of 41 women had personal or family history of polycystic ovary syndrome (PCOS). Additionally, 15 patients had obesity, and 4 had hypothyroidism. Conclusions: PLD is an effective and well-tolerated treatment for DTF, with good clinical responses at lower, tolerable doses. The association of pre-existing psychiatric diagnoses, PCOS, and obesity warrants further investigation.

A hallmark of chronic and inflammatory diseases is the formation of a fibrotic and stiff extracellular matrix (ECM), typically associated with abnormal, leaky microvascular capillaries. Mechanisms explaining how the microvasculature responds to ECM alterations remain unknown. Here, we used a microphysiological model of capillaries on a chip mimicking the characteristics of healthy or fibrotic collagen to test the hypothesis that perivascular cells mediate the response of vascular capillaries to mechanical and structural changes in the human ECM. Capillaries engineered in altered fibrotic collagen had abnormal migration of perivascular cells, reduced pericyte differentiation, increased leakage, and higher regulation of inflammatory/remodeling genes, all regulated via NOTCH3, a known mediator of endothelial-perivascular cell communication. Capillaries engineered either with endothelial cells alone or with perivascular cells silenced for NOTCH3 expression showed a minimal response to ECM alterations. These findings reveal a previously unknown mechanism of vascular response to changes in the ECM in health and disease.

Myocardial fibrosis, a condition linked to several cardiovascular diseases, is associated with a poor prognosis. Stem cell therapy has emerged as a potential treatment option and the application of stem cell therapy has been studied extensively. However, a comprehensive bibliometric analysis of these studies has yet to be conducted.

To map thematic trends, analyze research hotspots, and project future directions of stem cell-based myocardial fibrosis therapy.

We conducted a bibliometric and visual analysis of studies in the Web of Science Core Collection using VOSviewer and Microsoft Excel. The dataset included 1510 articles published between 2001 and 2024. Countries, organizations, authors, references, keywords, and co-citation networks were examined to identify evolving research trends.

Our findings revealed a steady increase in the number of publications, with a projected increase to over 200 publications annually by 2030. Initial research focused on stem cell-based therapy, particularly for myocardial infarction and heart failure. More recently, there has been a shift toward cell-free therapy, involving extracellular vesicles, exosomes, and microRNAs. Key research topics include angiogenesis, inflammation, apoptosis, autophagy, and oxidative stress.

This analysis highlights the evolution of stem cell therapies for myocardial fibrosis, with emerging interest in cell-free approaches. These results are expected to guide future scientific exploration and decision-making.

Within the intricate milieu of colorectal cancer (CRC) tissues, cancer-associated fibroblasts (CAFs) act as pivotal orchestrators, wielding considerable influence over tumor progression. This review endeavors to dissect the multifaceted functions of CAFs within the realm of CRC, thereby highlighting their indispensability in fostering CRC malignant microenvironment and indicating the development of CAFs-targeted therapeutic interventions. Through a comprehensive synthesis of current knowledge, this review delineates insights into CAFs-mediated modulation of cancer cell proliferation, invasiveness, immune evasion, and neovascularization, elucidating the intricate web of interactions that sustain the pro-tumor metabolism and secretion of multiple factors. Additionally, recognizing the high level of heterogeneity within CAFs is crucial, as they encompass a range of subtypes, including myofibroblastic CAFs, inflammatory CAFs, antigen-presenting CAFs, and vessel-associated CAFs. Innovatively, the symbiotic relationship between CAFs and the intestinal microbiota is explored, shedding light on a novel dimension of CRC pathogenesis. Despite remarkable progress, the orchestrated dynamic functions of CAFs remain incompletely deciphered, underscoring the need for continued research endeavors for therapeutic advancements in CRC management.

Pericytes function to maintain tissue homeostasis by regulating capillary blood flow and maintaining endothelial barrier function. Pericyte dysfunction is associated with various pathologies and has recently been found to aid cancer progression. Despite having critical functions in health and disease, pericytes remain an understudied population due to a lack of model systems which accurately reflect in vivo biology. In this study we developed a protocol to isolate and culture murine lung, brain, bone, and liver pericytes, that maintains their known phenotypes and functions. We demonstrate that pericytes, being inherently plastic, benefit from controlled oxygen tension culture conditions, aiding their expansion ex vivo. Primary pericytes grown in physiologically relevant oxygen tensions (10% O2 for lung; 5% O2 for brain, bone, and liver) also better retain pericyte phenotypes indicated by stable expression of characteristic transcriptional and protein markers. In functional tube formation assays, pericytes were observed to significantly associate with endothelial junctions. Importantly, we identified growth conditions that limit expression of the plasticity factor Klf4 to prevent spontaneous phenotypic switching in vitro. Additionally, we were able to induce pathological pericyte phenotypic switching in response to metastatic stimuli to accurately recapitulate in vivo biology. Here, we present a robust method for studying pericyte biology in both physiology and disease.

Pulmonary fibrosis, including systemic sclerosis-associated interstitial lung disease (SSc-ILD), involves myofibroblasts and SPP1hi macrophages as drivers of fibrosis. Single-cell RNA sequencing has delineated fibroblast and macrophages transcriptomes, but limited insight into transcriptional control of profibrotic gene programs. To address this challenge, we analyzed multiomic snATAC/snRNA-seq on explanted SSc-ILD and donor control lungs. The neural network tool ChromBPNet inferred increased TF binding at single base pair resolution to profibrotic genes, including CTHRC1 and ADAM12, in fibroblasts and SPP1 and CCL18 in macrophages. The novel algorithm HALO confirmed AP-1, RUNX, and EGR TF activity controlling profibrotic gene programs and established TF-regulatory element-gene networks. This TF action atlas provides comprehensive insights into the transcriptional regulation of fibroblasts and macrophages in healthy and fibrotic human lungs.

Transcriptomic data demonstrated that fibroblasts are heterogeneous with functionally diverse subpopulations. Although fibroblasts are key effector cells of fibrotic diseases such as systemic sclerosis (SSc), they have not yet been characterised spatially at the cellular level. Here, we aimed to investigate fibroblast subpopulations using imaging mass cytometry (IMC) as a proteomic-based, spatially resolved omics approach.

We applied IMC to deconvolute the heterogeneity of 49 969 cells including 6501 fibroblasts at the single-cell level, to analyse their spatial distribution and to characterise their cellular niches in skin sections of patients with SSc and controls in situ.

We identified 13 different subpopulations of fibroblasts in SSc and control skin, the proportion increases in five fibroblast subpopulations (myofibroblasts, FAPhigh, S1PR+, Thy1+;ADAM12high;PU.1high and ADAM12+;GLI1+ fibroblasts) and decreases in three subpopulations (TFAMhigh, PI16+;FAP+ and Thy1+;ADAM12low fibroblasts). Several fibroblast subpopulations demonstrated spatial enrichment and altered cellular interactions in SSc. The proportion of S1PR+-fibroblast positively correlated with more extensive skin fibrosis, whereas high numbers of PI16+;FAP--fibroblasts were associated with milder skin fibrosis. The frequency of aberrant cellular interaction between S1PR+ and ADAM12+;GLI1+-fibroblasts also positively associated with the extent of skin fibrosis in SSc.

Using IMC, we demonstrated profound changes in composition and localisation of the majority of fibroblast subpopulations in SSc skin. These findings may provide a rationale for specific targeting of deregulated fibroblast subpopulations in SSc. Quantification of S1PR+-fibroblast and PI16+;FAP--fibroblasts may offer potential for patient stratification according to severity of skin fibrosis.

Fibrosis is a pathological condition that in the muscular context is linked to primary diseases such as dystrophies, laminopathies, neuromuscular disorders, and volumetric muscle loss following traumas, accidents, and surgeries. Although some basic mechanisms regarding the role of myofibroblasts in the progression of muscle fibrosis have been discovered, our knowledge of the complex cell-cell, and cell-matrix interactions occurring in the fibrotic microenvironment is still rudimentary. Recently, vascular dysfunction has been emerging as a key hallmark of fibrosis through a process called endothelial-mesenchymal transition (EndoMT). Nevertheless, no effective therapeutic options are currently available for the treatment of muscle fibrosis. This lack is partially due to the absence of advanced in vitro models that can recapitulate the 3D architecture and functionality of a vascularized muscle microenvironment in a human context. These models could be employed for the identification of novel targets and for the screening of potential drugs blocking the progression of the disease. In this review, we explore the potential of 3D human muscle models in studying the role of endothelial cells and EndoMT in muscle fibrotic tissues and identify limitations and opportunities for optimizing the next generation of these microphysiological systems. Starting from the biology of muscle fibrosis and EndoMT, we highlight the synergistic links between different cell populations of the fibrotic microenvironment and how to recapitulate them through microphysiological systems.

Systemic sclerosis (SSc) is a chronic disease characterized by fibrosis and vascular abnormalities in the skin and internal organs, including the lung. SSc-associated pulmonary fibrosis (SSc-PF) is the leading cause of death in SSc patients. Pericytes are key regulators of vascular integrity and endothelial function. The role that pericytes play in SSc-PF remains unclear. We compared the transcriptome of pericytes from SSc-PF lungs (SScL) to pericytes from normal lungs (NORML). We identified 1,179 differentially expressed genes in SScL pericytes. Pathways enriched in SScL pericytes included prostaglandin, PI3K-AKT, calcium, and vascular remodeling signaling. Decreased cyclic AMP production and altered phosphorylation of AKT in response to prostaglandin E2 in SScL pericytes demonstrate the functional consequence of changes in the prostaglandin pathway that may contribute to fibrosis. The transcriptomic signature of SSc lung pericytes suggests that they promote vascular dysfunction and contribute to the loss of protection against lung inflammation and fibrosis.

This lecture delves into the pivotal role of adipose tissue in obesity and its response to weight loss, particularly via bariatric surgery. Adipose tissue, responsible for storing excess energy, undergoes significant changes during obesity, marked by inflammation and fibrosis. Bariatric surgery, serving as a model, allow the exploration of adipose tissue remodeling post-weight loss, inducing metabolic and fibro-inflammatory shifts. Despite successful weight loss, inflammation and fibrosis persist, as evidenced by changes in immune cells, altered cytokine profiles and the accumulation of extracellular matrix (ECM). Unfortunately, these lingering effects impair the normal adipose tissue function. In this context, adipose progenitors, an heterogenous resident population of mesenchymal stromal cells, display functions important to fibrosis development, capable of differentiating into myofibroblasts and contributing to ECM deposition. Particularly, a distinct subpopulation of adipose progenitors with high CD9 expression (CD9high) is associated with fibrosis and insulin resistance in human obesity. The persistence of fibrosis post-weight loss poses challenges, correlating with metabolic dysfunction despite improved glucose tolerance. A comprehensive understanding of the mechanisms driving adipose tissue remodeling and fibrosis post-weight loss is imperative for the development of effective treatments for obesity. The intricate interplay between adipose tissue, inflammation, and fibrosis underscores the necessity for further in-depth research to elucidate these mechanisms and formulate targeted therapies for obesity-related complications.

Tendon adhesion is a common complication after tendon injury with the development of accumulated fibrotic tissues without effective anti-fibrotic therapies, resulting in severe disability. Macrophages are widely recognized as a fibrotic trigger during peritendinous adhesion formation. However, different clusters of macrophages have various functions and receive multiple regulation, which are both still unknown. In our current study, multi-omics analysis including single-cell RNA sequencing and proteomics was performed on both human and mouse tendon adhesion tissue at different stages after tendon injury. The transcriptomes of over 74 000 human single cells were profiled. As results, we found that SPP1+ macrophages, RGCC+ endothelial cells, ACKR1+ endothelial cells and ADAM12+ fibroblasts participated in tendon adhesion formation. Interestingly, despite specific fibrotic clusters in tendon adhesion, FOLR2+ macrophages were identified as an antifibrotic cluster by in vitro experiments using human cells. Furthermore, ACKR1 was verified to regulate FOLR2+ macrophages migration at the injured peritendinous site by transplantation of bone marrow from Lysm-Cre;R26RtdTomato mice to lethally irradiated Ackr1-/- mice (Ackr1-/- chimeras; deficient in ACKR1) and control mice (WT chimeras). Compared with WT chimeras, the decline of FOLR2+ macrophages was also observed, indicating that ACKR1 was specifically involved in FOLR2+ macrophages migration. Taken together, our study not only characterized the fibrosis microenvironment landscape of tendon adhesion by multi-omics analysis, but also uncovered a novel antifibrotic cluster of macrophages and their origin. These results provide potential therapeutic targets against human tendon adhesion.

Cancer-associated fibroblasts (CAFs), a phenotypically and functionally heterogeneous stromal cell, are one of the most important components of the tumour microenvironment. Previous studies have consolidated it as a promising target against cancer. However, variable therapeutic efficacy-both protumor and antitumor effects have been observed not least owing to the strong heterogeneity of CAFs. Over the past 10 years, advances in single-cell RNA sequencing (scRNA-seq) technologies had a dramatic effect on biomedical research, enabling the analysis of single cell transcriptomes with unprecedented resolution and throughput. Specifically, scRNA-seq facilitates our understanding of the complexity and heterogeneity of diverse CAF subtypes. In this review, we discuss the up-to-date knowledge about CAF heterogeneity with a focus on scRNA-seq perspective to investigate the emerging strategies for integrating multimodal single-cell platforms. Furthermore, we summarized the clinical application of scRNA-seq on CAF research. We believe that the comprehensive understanding of the heterogeneity of CAFs form different visions will generate innovative solutions to cancer therapy and achieve clinical applications.

The largest mammalian organ, skin, consisting of a dermal connective tissue layer that underlies and supports the epidermis, acts as a protective barrier that excludes external pathogens and disseminates sensory signals emanating from the local microenvironment. Dermal connective tissue is comprised of a collagen-rich extracellular matrix (ECM) that is produced by connective tissue fibroblasts resident within the dermis. When wounded, a tissue repair program is induced whereby fibroblasts, in response to alterations in the microenvironment, produce new ECM components, resulting in the formation of a scar. Failure to terminate the normal tissue repair program causes fibrotic conditions including: hypertrophic scars, keloids, and the systemic autoimmune connective tissue disease scleroderma (systemic sclerosis, SSc). Histological and single-cell RNA sequencing (scRNAseq) studies have revealed that fibroblasts are heterogeneous and highly plastic. Understanding how this diversity contributes to dermal homeostasis, wounding, fibrosis, and cancer may ultimately result in novel anti-fibrotic therapies and personalized medicine. This review summarizes studies supporting this concept.

Systemic sclerosis (SSc) is a connective tissue disease that can serve as a model to study vascular changes in response to inflammation, autoimmunity, and fibrotic remodeling. Although microvascular changes are the earliest histopathologic manifestation of SSc, the vascular pathophysiology remains poorly understood.

We applied spatial proteomic approaches to deconvolute the heterogeneity of vascular cells at the single-cell level in situ and characterize cellular alterations of the vascular niches of patients with SSc. Skin biopsies of patients with SSc and control individuals were analyzed by imaging mass cytometry, yielding a total of 90 755 cells including 2987 endothelial cells and 4096 immune cells.

We identified 7 different subpopulations of blood vascular endothelial cells (VECs), 2 subpopulations of lymphatic endothelial cells, and 3 subpopulations of pericytes. A novel population of CD34+;αSMA+ (α-smooth muscle actin);CD31+ VECs was more common in SSc, whereas endothelial precursor cells were decreased. Co-detection by indexing and tyramide signal amplification confirmed these findings. The microenvironment of CD34+;αSMA+;CD31+ VECs was enriched for immune cells and myofibroblasts, and CD34+;αSMA+;CD31+ VECs expressed markers of endothelial-to-mesenchymal transition. The density of CD34+;αSMA+;CD31+ VECs was associated with clinical progression of fibrosis in SSc.

Using spatial proteomics, we unraveled the heterogeneity of vascular cells in control individuals and patients with SSc. We identified CD34+;αSMA+;CD31+ VECs as a novel endothelial cell population that is increased in patients with SSc, expresses markers for endothelial-to-mesenchymal transition, and is located in close proximity to immune cells and myofibroblasts. CD34+;αSMA+;CD31+ VEC counts were associated with clinical outcomes of progressive fibrotic remodeling, thus providing a novel cellular correlate for the crosstalk of vasculopathy and fibrosis.

Currently, pancreatic cancer (PC) is one of the most lethal malignant tumors. PC is typically diagnosed at a late stage, exhibits a poor response to conventional treatment, and has a bleak prognosis. Unfortunately, PC's survival rate has not significantly improved since the 1960s. Cancer-associated fibroblasts (CAFs) are a key component of the pancreatic tumor microenvironment (TME). They play a vital role in maintaining the extracellular matrix and facilitating the intricate communication between cancer cells and infiltrated immune cells. Exploring therapeutic approaches targeting CAFs may reverse the current landscape of PC therapy. In recent years, nano-drug delivery systems have evolved rapidly and have been able to accurately target and precisely release drugs with little or no toxicity to the whole body. In this review, we will comprehensively discuss the origin, heterogeneity, potential targets, and recent advances in the nano-drug delivery system of CAFs in PC. We will also propose a novel integrated treatment regimen that utilizes a nano-drug delivery system to target CAFs in PC, combined with radiotherapy and immunotherapy. Additionally, we will address the challenges that this regimen currently faces.

Diabetes is not only an endocrine but also a vascular disease. Vascular defects are usually seen as consequence of diabetes. However, at the level of the pancreatic islet, vascular alterations have been described before symptom onset. Importantly, the cellular and molecular mechanisms underlying these early vascular defects have not been identified, neither how these could impact the function of islet endocrine cells. In this review, we will discuss the possibility that dysfunction of the mural cells of the microvasculature-known as pericytes-underlies vascular defects observed in islets in pre-symptomatic stages. Pericytes are crucial for vascular homeostasis throughout the body, but their physiological and pathophysiological functions in islets have only recently started to be explored. A previous study had already raised interest in the "microvascular" approach to this disease. With our increased understanding of the crucial role of the islet microvasculature for glucose homeostasis, here we will revisit the vascular aspects of islet function and how their deregulation could contribute to diabetes pathogenesis, focusing in particular on type 1 diabetes (T1D).

Acute skeletal muscle injury is common in sports. The injured muscle cannot fully recover due to fibrosis resulting from myofibroblasts. Understanding the origin of fibroblasts is, therefore, important for the development of anti-fibrotic therapies. Accumulating evidence shows that a mechanism called macrophage-myofibroblast transition (MMT) can lead to tissue or organ fibrosis, yet it is still unclear whether MMT exists in skeletal muscle and the exact mechanisms.

Single-cell transcriptome of mice skeletal muscle after acute injury was analyzed with a specific attention on the process of MMT. Cell-cell interaction network, pseudotime trajectory analysis, Gene Ontology (GO), and Kyoto Genome Encyclopedia (KEGG) were conducted. A series of experiments in vivo and in vitro were launched for verification.

Single cell transcriptomic analysis indicated that, following acute injury, there were much interactions between macrophages and myofibroblasts. A detailed analysis on macrophages indicated that, CD68+α-SMA+ cells, which represented the status of MMT, mainly appeared at five days post-injury. KEGG/GO analysis underlined the involvement of complement system, within which C3ar1, C1qa, C1qb, and C1qc were up-regulated. Trajectory analysis also confirmed a potential shift from macrophages to myofibroblasts. These findings were verified by histological study in mice skeletal muscle, that there were much MMT cells at five days, declined gradually, and vanished 14 days after trauma, when there was remarkable fibrosis formation within the injured muscle. Moreover, C3a stimulation could directly induce MMT in BMDMs.

Fibrosis following acute injury is disastrous to skeletal muscle, but the origin of myofibroblasts remains unclear. We proved that, following acute injury, macrophage-myofibroblast transition happened in skeletal muscle, which may contribute to fibrosis formation. This phenomenon mainly occurred at five days post-injury. The complement system can activate MMT. More evidence is needed to directly support the pro-fibrotic role of MMT in skeletal muscle fibrosis after acute injury.

Skeletal muscle stem cells (MuSCs) are recognised as functionally heterogeneous. Cranial MuSCs are reported to have greater proliferative and regenerative capacity when compared with those in the limb. A comprehensive understanding of the mechanisms underlying this functional heterogeneity is lacking. Here, we have used clonal analysis, live imaging and single cell transcriptomic analysis to identify crucial features that distinguish extraocular muscle (EOM) from limb muscle stem cell populations. A MyogeninntdTom reporter showed that the increased proliferation capacity of EOM MuSCs correlates with deferred differentiation and lower expression of the myogenic commitment gene Myod. Unexpectedly, EOM MuSCs activated in vitro expressed a large array of extracellular matrix components typical of mesenchymal non-muscle cells. Computational analysis underscored a distinct co-regulatory module, which is absent in limb MuSCs, as driver of these features. The EOM transcription factor network, with Foxc1 as key player, appears to be hardwired to EOM identity as it persists during growth, disease and in vitro after several passages. Our findings shed light on how high-performing MuSCs regulate myogenic commitment by remodelling their local environment and adopting properties not generally associated with myogenic cells.

Skin injury always results in fibrotic, non-functional scars in adults. Although multiple factors are well-known contributors to scar formation, the precise underlying mechanisms remain elusive. This review aims to elucidate the intricacies of the wound healing process, summarize the known factors driving skin cells in wounds toward a scarring fate, and particularly to discuss the impact of fibroblast heterogeneity on scar formation. To the end, we explore potential therapeutic interventions used in the treatment of scarring wounds.

We aimed to determine whether and how the combination of acetazolamide and remote ischemic preconditioning (RIPC) reduced the incidence and severity of acute mountain sickness (AMS).

This is a prospective, randomized, open-label, blinded endpoint (PROBE) study involving 250 healthy volunteers. Participants were randomized (1:1:1:1:1) to following five groups: Ripc (RIPC twice daily, 6 days), Rapid-Ripc (RIPC four times daily, 3 days), Acetazolamide (twice daily, 2 days), Combined (Acetazolamide plus Rapid-Ripc), and Control group. After interventions, participants entered a normobaric hypoxic chamber (equivalent to 4000 m) and stayed for 6 h. The primary outcomes included the incidence and severity of AMS, and SpO2 after hypoxic exposure. Secondary outcomes included systolic and diastolic blood pressure, and heart rate after hypoxic exposure. The mechanisms of the combined regime were investigated through exploratory outcomes, including analysis of venous blood gas, complete blood count, human cytokine antibody array, ELISA validation for PDGF-AB, and detection of PDGF gene polymorphisms.

The combination of acetazolamide and RIPC exhibited powerful efficacy in preventing AMS, reducing the incidence of AMS from 26.0 to 6.0% (Combined vs Control: RR 0.23, 95% CI 0.07-0.70, P = 0.006), without significantly increasing the incidence of adverse reactions. Combined group also showed the lowest AMS score (0.92 ± 1.10). Mechanistically, acetazolamide induced a mild metabolic acidosis (pH 7.30 ~ 7.31; HCO3- 18.1 ~ 20.8 mmol/L) and improved SpO2 (89 ~ 91%) following hypoxic exposure. Additionally, thirty differentially expressed proteins (DEPs) related to immune-inflammatory process were identified after hypoxia, among which PDGF-AB was involved. Further validation of PDGF-AB in all individuals showed that both acetazolamide and RIPC downregulated PDGF-AB before hypoxic exposure, suggesting a possible protective mechanism. Furthermore, genetic analyses demonstrated that individuals carrying the PDGFA rs2070958 C allele, rs9690350 G allele, or rs1800814 G allele did not display a decrease in PDGF-AB levels after interventions, and were associated with a higher risk of AMS.

The combination of acetazolamide and RIPC exerts a powerful anti-hypoxic effect and represents an innovative and promising strategy for rapid ascent to high altitudes. Acetazolamide improves oxygen saturation. RIPC further aids acetazolamide, which synergistically regulates PDGF-AB, potentially involved in the pathogenesis of AMS.

ClinicalTrials.gov NCT05023941.

Fibrosis is associated with compromised muscle functionality in Duchenne muscular dystrophy (DMD). We report observations with tissues from dystrophic patients and mice supporting a model to explain fibrosis in DMD, which relies on the crosstalk between the complement and the WNT signaling pathways and the functional interactions of two cellular types. Fibro-adipogenic progenitors and macrophages, which populate the inflamed dystrophic muscles, act as a combinatorial source of WNT activity by secreting distinct subunits of the C1 complement complex. The resulting aberrant activation of the WNT signaling in responsive cells, such as fibro-adipogenic progenitors, contributes to fibrosis. Indeed, pharmacological inhibition of the C1r/s subunits in a murine model of DMD mitigated the activation of the WNT signaling pathway, reduced the fibrogenic characteristics of the fibro-adipogenic progenitors, and ameliorated the dystrophic phenotype. These studies shed new light on the molecular and cellular mechanisms responsible for fibrosis in muscular dystrophy and open to new therapeutic strategies.

Despite their importance in tissue maintenance and repair, fibroblast diversity and plasticity remain poorly understood. Using single-cell RNA sequencing, we uncover distinct sclerotome-derived fibroblast populations in zebrafish, including progenitor-like perivascular/interstitial fibroblasts, and specialized fibroblasts such as tenocytes. To determine fibroblast plasticity in vivo, we develop a laser-induced tendon ablation and regeneration model. Lineage tracing reveals that laser-ablated tenocytes are quickly regenerated by preexisting fibroblasts. By combining single-cell clonal analysis and live imaging, we demonstrate that perivascular/interstitial fibroblasts actively migrate to the injury site, where they proliferate and give rise to new tenocytes. By contrast, perivascular fibroblast-derived pericytes or specialized fibroblasts, including tenocytes, exhibit no regenerative plasticity. Active Hedgehog (Hh) signaling is required for the proliferation of activated fibroblasts to ensure efficient tenocyte regeneration. Together, our work highlights the functional diversity of fibroblasts and establishes perivascular/interstitial fibroblasts as tenocyte progenitors that promote tendon regeneration in a Hh signaling-dependent manner.

Heterogeneous dermal fibroblasts are the main components that constitute the dermis. Distinct fibroblast subgroups show specific characteristics and functional plasticity that determine dermal structure during skin development and wound healing. Although researchers have described the roles of fibroblast subsets, this is not completely understood. We review recent evidence supporting understanding about the heterogeneity of fibroblasts. We summarize the origins and the identified profiles of fibroblast subpopulations. The characteristics of fibroblast subpopulations in both healthy and diseased states are highlighted, and the potential of subpopulations to be involved in wound healing in different ways was discussed. Additionally, we review the plasticity of subpopulations and the underlying signalling mechanisms. This review may provide greater insights into potential novel therapeutic targets and tissue regeneration strategies for the future.

Adipose tissue is the largest endocrine organ and an accepted contributor to overall energy homeostasis. There is strong evidence linking increased adiposity to the development of 13 types of cancer. With increased adiposity comes metabolic dysfunction and insulin resistance, and increased systemic insulin and glucose support the growth of many cancers, including those of the colon and endometrium. There is also an important direct crosstalk between adipose tissue and various organs. For instance, the healthy development and function of the mammary gland, as well as the development, growth, and progression of breast cancer, are heavily impacted by the breast adipose tissue in which breast epithelial cells are embedded. Cells of the adipose tissue are responsive to external stimuli, including overfeeding, leading to remodeling and important changes in the secretion of factors known to drive the development and growth of cancers. Loss of factors like adiponectin and increased production of leptin, endotrophin, steroid hormones, and inflammatory mediators have been determined to be important mediators of the obesity-cancer link. Obesity is also associated with a structural remodeling of the adipose tissue, including increased localized fibrosis and disrupted angiogenesis that contribute to the development and progression of cancers. Furthermore, tumor cells feed off the adipose tissue, where increased lipolysis within adipocytes leads to the release of fatty acids and stromal cell aerobic glycolysis leading to the increased production of lactate. Both have been hypothesized to support the higher energetic demands of cancer cells. Here, we aim to provide an update on the state of the literature revolving around the role of the adipose tissue in cancer initiation and progression.

The management of preretinal fibrovascular membranes, a devastating complication of advanced diabetic retinopathy (DR), remains challenging. We characterized the molecular profile of cell populations in these fibrovascular membranes to identify potentially new therapeutic targets. Preretinal fibrovascular membranes were surgically removed from patients and submitted for single-cell RNA-Seq (scRNA-Seq). Differential gene expression was implemented to define the transcriptomics profile of these cells and revealed the presence of endothelial, inflammatory, and stromal cells. Endothelial cell reclustering identified subclusters characterized by noncanonical transcriptomics profile and active angiogenesis. Deeper investigation of the inflammatory cells showed a subcluster of macrophages expressing proangiogenic cytokines, presumably contributing to angiogenesis. The stromal cell cluster included a pericyte-myofibroblast transdifferentiating subcluster, indicating the involvement of pericytes in fibrogenesis. Differentially expressed gene analysis showed that Adipocyte Enhancer-binding Protein 1, AEBP1, was significantly upregulated in myofibroblast clusters, suggesting that this molecule may have a role in transformation. Cell culture experiments with human retinal pericytes (HRP) in high-glucose condition confirmed the molecular transformation of pericytes toward myofibroblastic lineage. AEBP1 siRNA transfection in HRP reduced the expression of profibrotic markers in high glucose. In conclusion, AEBP1 signaling modulates pericyte-myofibroblast transformation, suggesting that targeting AEBP1 could prevent scar tissue formation in advanced DR.

Although two targeted therapies have received recent approval for systemic sclerosis (SSc)-associated interstitial lung disease, they do not show major disease-modifying activity, highlighting the need for novel therapies and innovative paradigms. To that end, cellular therapies may represent a new opportunity for the treatment of SSc. The purpose of this review is to provide an up-to-date overview of emerging cell-based disease-modifying therapies in SSc.

Initial small studies in patients with severe refractory systemic lupus erythematosus (SLE) using engineered regulatory cells show promising results. CD19-directed CAR-T have shown promising results in one case report of refractory diffuse cutaneous SSc patients. T cells engineered to express a chimeric autoantibody receptor (CAAR-T cells) may be even more relevant via the specific elimination of auto-reactive B cells. Targeting pro-fibrotic or senescence-related pathways may also constitute promising approaches in SSc.

Building on the classification of the clinical phenotype and prediction of clinical trajectory based on individual patients' autoantigen and/or autoantibody profile, cellular therapies targeting the same autoantigen or related autoreactive cells may represent an unprecedented opportunity to implement personalized medicine in SSc.

The capacity to survive and thrive in conditions of limited resources and high inflammation is a major driver of tumor malignancy. Here we identified slow-cycling ADAM12+PDGFRα+ mesenchymal stromal cells (MSCs) induced at the tumor margins in mouse models of melanoma, pancreatic cancer and prostate cancer. Using inducible lineage tracing and transcriptomics, we demonstrated that metabolically altered ADAM12+ MSCs induced pathological angiogenesis and immunosuppression by promoting macrophage efferocytosis and polarization through overexpression of genes such as Gas6, Lgals3 and Csf1. Genetic depletion of ADAM12+ cells restored a functional tumor vasculature, reduced hypoxia and acidosis and normalized CAFs, inducing infiltration of effector T cells and growth inhibition of melanomas and pancreatic neuroendocrine cancer, in a process dependent on TGF-β. In human cancer, ADAM12 stratifies patients with high levels of hypoxia and innate resistance mechanisms, as well as factors associated with a poor prognosis and drug resistance such as AXL. Altogether, our data show that depletion of tumor-induced slow-cycling PDGFRα+ MSCs through ADAM12 restores antitumor immunity.

Cancer-associated fibroblasts (CAFs), enriched in the tumor stroma, have received increasing attention because of their multifaceted effects on tumorigenesis, development, metastasis, and treatment resistance in malignancies. CAFs contributed to suppressive microenvironment via different mechanisms, while CAFs also exerted some antitumor effects. Therefore, CAFs have been considered promising therapeutic targets for their remarkable roles in malignant tumors. However, patients with malignancies failed to benefit from current CAFs-targeted drugs in many clinical trials, which suggests that further in-depth investigation into CAFs is necessary. Here, we summarize and outline the heterogeneity and plasticity of CAFs mainly by exploring their origin and activation, highlighting the regulation of CAFs in the tumor microenvironment during tumor evolution, as well as the critical roles performed by CAFs in tumor immunity. In addition, we summarize the current immunotherapies targeting CAFs, and conclude with a brief overview of some prospects for the future of CAFs research in the end. Video Abstract.