Cellular and molecular mechanisms of skin wound healing.

Wound healing is a complex process that involves the coordinated actions of many different tissues and cell lineages. It requires tight orchestration of cell migration, proliferation, matrix deposition and remodelling, alongside inflammation and angiogenesis. Whereas small skin wounds heal in days, larger injuries resulting from trauma, acute illness or major surgery can take several weeks to heal, generally leaving behind a fibrotic scar that can impact tissue function. Development of therapeutics to prevent scarring and successfully repair chronic wounds requires a fuller knowledge of the cellular and molecular mechanisms driving wound healing. In this Review, we discuss the current understanding of the different phases of wound healing, from clot formation through re-epithelialization, angiogenesis and subsequent scar deposition. We highlight the contribution of different cell types to skin repair, with emphasis on how both innate and adaptive immune cells in the wound inflammatory response influence classically studied wound cell lineages, including keratinocytes, fibroblasts and endothelial cells, but also some of the less-studied cell lineages such as adipocytes, melanocytes and cutaneous nerves. Finally, we discuss newer approaches and research directions that have the potential to further our understanding of the mechanisms underpinning tissue repair.

Type V Collagen in Scar Tissue Regulates the Size of Scar after Heart Injury.

Scar tissue size following myocardial infarction is an independent predictor of cardiovascular outcomes, yet little is known about factors regulating scar size. We demonstrate that collagen V, a minor constituent of heart scars, regulates the size of heart scars after ischemic injury. Depletion of collagen V led to a paradoxical increase in post-infarction scar size with worsening of heart function. A systems genetics approach across 100 in-bred strains of mice demonstrated that collagen V is a critical driver of postinjury heart function. We show that collagen V deficiency alters the mechanical properties of scar tissue, and altered reciprocal feedback between matrix and cells induces expression of mechanosensitive integrins that drive fibroblast activation and increase scar size. Cilengitide, an inhibitor of specific integrins, rescues the phenotype of increased post-injury scarring in collagen-V-deficient mice. These observations demonstrate that collagen V regulates scar size in an integrin-dependent manner.

Exogenous Transforming Growth Factor-ß1 and Its Helminth-Derived Mimic Attenuate the Heart's Inflammatory Response to Ischemic Injury and Reduce Mature Scar Size.

Coronary reperfusion after acute ST-elevation myocardial infarction (STEMI) is standard therapy to salvage ischemic heart muscle. However, subsequent inflammatory responses within the infarct lead to further loss of viable myocardium. Transforming growth factor (TGF)-ß1 is a potent anti-inflammatory cytokine released in response to tissue injury. The aim of this study was to investigate the protective effects of TGF-ß1 after MI. In patients with STEMI, there was a significant correlation (P = 0.003) between higher circulating TGF-ß1 levels at 24 hours after MI and a reduction in infarct size after 3 months, suggesting a protective role of early increase in circulating TGF-ß1. A mouse model of cardiac ischemia reperfusion was used to demonstrate multiple benefits of exogenous TGF-ß1 delivered in the acute phase. It led to a significantly smaller infarct size (30% reduction, P = 0.025), reduced inflammatory infiltrate (28% reduction, P = 0.015), lower intracardiac expression of inflammatory cytokines IL-1ß and chemokine (C-C motif) ligand 2 (>50% reduction, P = 0.038 and 0.0004, respectively) at 24 hours, and reduced scar size at 4 weeks (21% reduction, P = 0.015) after reperfusion. Furthermore, a low-fibrogenic mimic of TGF-ß1, secreted by the helminth parasite Heligmosomoides polygyrus, had an almost identical protective effect on injured mouse hearts. Finally, genetic studies indicated that this benefit was mediated by TGF-ß signaling in the vascular endothelium.

Anti-Angiogenic and Anti-Scarring Dual Effect of Galectin-3 Inhibition in Mouse Models of Corneal Wound Healing.

Corneal scarring is the third leading cause of global blindness. Neovascularization of ocular tissues is a major predisposing factor in scar development. Although corneal transplantation is effective in restoring vision, some patients are at high risk for graft rejection due to the presence of blood vessels in the injured cornea. Current treatment options for controlling corneal scarring are limited, and outcomes are typically poor. In this study, topical application of a small-molecule inhibitor of galectin-3, GB1265, in mouse models of corneal wound healing, led to the reduction of the following in injured corneas: i) corneal angiogenesis; ii) corneal fibrosis; iii) infiltration of immune cells; and iv) expression of the proinflammatory cytokine IL-1ß. Four independent techniques (RNA sequencing, NanoString, real-time quantitative RT-PCR, and Western blot analysis) determined that decreased corneal opacity in the galectin-3 inhibitor-treated corneas was associated with decreases in the numbers of genes and signaling pathways known to promote fibrosis. These findings allowed for a high level of confidence in the conclusion that galectin-3 inhibition by the small-molecule inhibitor GB1265 has dual anti-angiogenic and anti-scarring effects. Targeting galectin-3 by GB1265 is, thus, attractive for the development of innovative therapies for a myriad of ocular and nonocular diseases characterized by pathologic angiogenesis and fibrosis.

p53 Regulates the Extent of Fibroblast Proliferation and Fibrosis in Left Ventricle Pressure Overload.

BACKGROUND: Cardiomyopathy is characterized by the pathological accumulation of resident cardiac fibroblasts that deposit ECM (extracellular matrix) and generate a fibrotic scar. However, the mechanisms that control the timing and extent of cardiac fibroblast proliferation and ECM production are not known, hampering the development of antifibrotic strategies to prevent heart failure. METHODS: We used the Tcf21 (transcription factor 21)MerCreMer mouse line for fibroblast-specific lineage tracing and p53 (tumor protein p53) gene deletion. We characterized cardiac physiology and used single-cell RNA-sequencing and in vitro studies to investigate the p53-dependent mechanisms regulating cardiac fibroblast cell cycle and fibrosis in left ventricular pressure overload induced by transaortic constriction. RESULTS: Cardiac fibroblast proliferation occurs primarily between days 7 and 14 following transaortic constriction in mice, correlating with alterations in p53-dependent gene expression. p53 deletion in fibroblasts led to a striking accumulation of Tcf21-lineage cardiac fibroblasts within the normal proliferative window and precipitated a robust fibrotic response to left ventricular pressure overload. However, excessive interstitial and perivascular fibrosis does not develop until after cardiac fibroblasts exit the cell cycle. Single-cell RNA sequencing revealed p53 null fibroblasts unexpectedly express lower levels of genes encoding important ECM proteins while they exhibit an inappropriately proliferative phenotype. in vitro studies establish a role for p53 in suppressing the proliferative fibroblast phenotype, which facilitates the expression and secretion of ECM proteins. Importantly, Cdkn2a (cyclin-dependent kinase inhibitor 2a) expression and the p16Ink4a-retinoblastoma cell cycle control pathway is induced in p53 null cardiac fibroblasts, which may eventually contribute to cell cycle exit and fulminant scar formation. CONCLUSIONS: This study reveals a mechanism regulating cardiac fibroblast accumulation and ECM secretion, orchestrated in part by p53-dependent cell cycle control that governs the timing and extent of fibrosis in left ventricular pressure overload.

Network modeling predicts personalized gene expression and drug responses in valve myofibroblasts cultured with patient sera.

Aortic valve stenosis (AVS) patients experience pathogenic valve leaflet stiffening due to excessive extracellular matrix (ECM) remodeling. Numerous microenvironmental cues influence pathogenic expression of ECM remodeling genes in tissue-resident valvular myofibroblasts, and the regulation of complex myofibroblast signaling networks depends on patient-specific extracellular factors. Here, we combined a manually curated myofibroblast signaling network with a data-driven transcription factor network to predict patient-specific myofibroblast gene expression signatures and drug responses. Using transcriptomic data from myofibroblasts cultured with AVS patient sera, we produced a large-scale, logic-gated differential equation model in which 11 biochemical and biomechanical signals were transduced via a network of 334 signaling and transcription reactions to accurately predict the expression of 27 fibrosis-related genes. Correlations were found between personalized model-predicted gene expression and AVS patient echocardiography data, suggesting links between fibrosis-related signaling and patient-specific AVS severity. Further, global network perturbation analyses revealed signaling molecules with the most influence over network-wide activity, including endothelin 1 (ET1), interleukin 6 (IL6), and transforming growth factor ß (TGFß), along with downstream mediators c-Jun N-terminal kinase (JNK), signal transducer and activator of transcription (STAT), and reactive oxygen species (ROS). Lastly, we performed virtual drug screening to identify patient-specific drug responses, which were experimentally validated via fibrotic gene expression measurements in valvular interstitial cells cultured with AVS patient sera and treated with or without bosentan-a clinically approved ET1 receptor inhibitor. In sum, our work advances the ability of computational approaches to provide a mechanistic basis for clinical decisions including patient stratification and personalized drug screening.

Automated spatially targeted optical micro proteomics identifies fibroblast- and macrophage-specific regulation of myocardial infarction scar maturation in rats.

Myocardial infarction (MI) results from occlusion of blood supply to the heart muscle causing death of cardiac muscle cells. Following myocardial infarction (MI), extracellular matrix deposition and scar formation mechanically stabilize the injured heart as damaged myocytes undergo necrosis and removal. Fibroblasts and macrophages are key drivers of post-MI scar formation, maturation, and ongoing long-term remodelling; however, their individual contributions are difficult to assess from bulk analyses of infarct scar. Here, we employ state-of-the-art automated spatially targeted optical micro proteomics (autoSTOMP) to photochemically tag and isolate proteomes associated with subpopulations of fibroblasts (SMA+) and macrophages (CD68+) in the context of the native, MI tissue environment. Over a time course of 6-weeks post-MI, we captured dynamic changes in the whole-infarct proteome and determined that some of these protein composition signatures were differentially localized near SMA+ fibroblasts or CD68+ macrophages within the scar region. These results link specific cell populations to within-infarct protein remodelling and illustrate the distinct metabolic and structural processes underlying the observed physiology of each cell type.

Metabolic disorders exacerbate the formation of glial scar after stroke.

Metabolic disorders are risk factors for stroke exacerbating subsequent complications. Rapidly after brain injury, a glial scar forms, preventing excessive inflammation and limiting axonal regeneration. Despite the growing interest in wound healing following brain injury, the formation of a glial scar in the context of metabolic disorders is poorly documented. In this study, we used db/db mice to investigate the impact of metabolic perturbations on brain repair mechanisms, with a focus on glial scarring. First, we confirmed the development of obesity, poor glucose regulation, hyperglycaemia and liver steatosis in these mice. Then, we observed that 3 days after a 30-min middle cerebral artery occlusion (MCAO), db/db mice had larger infarct area compared with their control counterparts. We next investigated reactive gliosis and glial scar formation in db/+ and db/db mice. We demonstrated that astrogliosis and microgliosis were exacerbated 3 days after stroke in db/db mice. Furthermore, we also showed that the synthesis of extracellular matrix (ECM) proteins (i.e., chondroitin sulphate proteoglycan, collagen IV and tenascin C) was increased in db/db mice. Consequently, we demonstrated for the first time that metabolic disorders impair reactive gliosis post-stroke and increase ECM deposition. Given that the damage size is known to influence glial scar, this study now raises the question of the direct impact of hyperglycaemia/obesity on reactive gliosis and glia scar. It paves the way to promote the development of new therapies targeting glial scar formation to improve functional recovery after stroke in the context of metabolic disorders.

Single-cell, single-nucleus, and spatial transcriptomics characterization of the immunological landscape in the healthy and PSC human liver.

BACKGROUND & AIMS: Primary sclerosing cholangitis (PSC) is an immune-mediated cholestatic liver disease for which there is an unmet need to understand the cellular composition of the affected liver and how it underlies disease pathogenesis. We aimed to generate a comprehensive atlas of the PSC liver using multi-omic modalities and protein-based functional validation. METHODS: We employed single-cell and single-nucleus RNA sequencing (47,156 cells and 23,000 nuclei) and spatial transcriptomics (one sample by 10x Visium and five samples with Nanostring GeoMx DSP) to profile the cellular ecosystem in 10 PSC livers. Transcriptomic profiles were compared to 24 neurologically deceased donor livers (107,542 cells) and spatial transcriptomics controls, as well as 18,240 cells and 20,202 nuclei from three PBC livers. Flow cytometry was performed to validate PSC-specific differences in immune cell phenotype and function. RESULTS: PSC explants with parenchymal cirrhosis and prominent periductal fibrosis contained a population of cholangiocyte-like hepatocytes that were surrounded by diverse immune cell populations. PSC-associated biliary, mesenchymal, and endothelial populations expressed chemokine and cytokine transcripts involved in immune cell recruitment. Additionally, expanded CD4+ T cells and recruited myeloid populations in the PSC liver expressed the corresponding receptors to these chemokines and cytokines, suggesting potential recruitment. Tissue-resident macrophages, by contrast, were reduced in number and exhibited a dysfunctional and downregulated inflammatory response to lipopolysaccharide and interferon-γ stimulation. CONCLUSIONS: We present a comprehensive atlas of the PSC liver and demonstrate an exhaustion-like phenotype of myeloid cells and markers of chronic cytokine expression in late-stage PSC lesions. This atlas expands our understanding of the cellular complexity of PSC and has potential to guide the development of novel treatments. IMPACT AND IMPLICATIONS: Primary sclerosing cholangitis (PSC) is a rare liver disease characterized by chronic inflammation and irreparable damage to the bile ducts, which eventually results in liver failure. Due to a limited understanding of the underlying pathogenesis of disease, treatment options are limited. To address this, we sequenced healthy and diseased livers to compare the activity, interactions, and localization of immune and non-immune cells. This revealed that hepatocytes lining PSC scar regions co-express cholangiocyte markers, whereas immune cells infiltrate the scar lesions. Of these cells, macrophages, which typically contribute to tissue repair, were enriched in immunoregulatory genes and demonstrated a lack of responsiveness to stimulation. These cells may be involved in maintaining hepatic inflammation and could be a target for novel therapies.

Integrin αV mediated activation of myofibroblast via mechanoparacrine of transforming growth factor ß1 in promoting fibrous scar formation after myocardial infarction.

BACKGROUND: Myocardial infarction (MI) dramatically changes the mechanical stress, which is intensified by the fibrotic remodeling. Integrins, especially the αV subunit, mediate mechanical signal and mechanoparacrine of transforming growth factor ß1 (TGF-ß1) in various organ fibrosis by activating CFs into myofibroblasts (MFBs). We investigated a possible role of integrin αV mediated mechanoparacrine of TGF-ß1 in MFBs activation for fibrous reparation in mice with MI. METHODS: Heart samples from MI, sham, or MI plus cilengitide (14 mg/kg, specific integrin αV inhibitor) treated mice, underwent functional and morphological assessments by echocardiography, and histochemistry on 7, 14 and 28 days post-surgery. The mechanical and ultrastructural changes of the fibrous scar were further evaluated by atomic mechanics microscope (AFM), immunofluorescence, second harmonic generation (SHG) imaging, polarized light and scanning electron microscope, respectively. Hydroxyproline assay was used for total collagen content, and western blot for protein expression profile examination. Fibroblast bioactivities, including cell shape, number, Smad2/3 signal and expression of extracellular matrix (ECM) related proteins, were further evaluated by microscopic observation and immunofluorescence in polyacrylamide (PA) hydrogel with adjustable stiffness, which was re-explored in fibroblast cultured on stiff matrix after silencing of integrin αV. The content of total and free TGF-ß1 was tested by enzyme-linked immunosorbent assay (ELISA) in both infarcted tissue and cell samples. RESULT: Increased stiffness with heterogeneity synchronized with integrin αV and alpha smooth muscle actin (α-SMA) positive MFBs accumulation in those less mature fibrous areas. Cilengitide abruptly reduced collagen content and disrupted collagen alignment, which also decreased TGF-ß1 bioavailability, Smad2/3 phosphorylation, and α-SMA expression in the fibrous area. Accordingly, fibroblast on stiff but not soft matrix exhibited obvious MFB phenotype, as evidenced by enlarged cell, hyperproliferation, well-developed α-SMA fibers, and elevated ECM related proteins, while silencing of integrin αV almost abolished this switch via attenuating paracrine of TGF-ß1 and nuclear translocation of Smad2/3. CONCLUSION: This study illustrated that increased tissue stiffness activates CFs into MFBs by integrin αV mediated mechanoparacrine of TGF-ß1, especially in immature scar area, which ultimately promotes fibrous scar maturation.

Lactate promotes microglial scar formation and facilitates locomotor function recovery by enhancing histone H4 lysine 12 lactylation after spinal cord injury.

Lactate-derived histone lactylation is involved in multiple pathological processes through transcriptional regulation. The role of lactate-derived histone lactylation in the repair of spinal cord injury (SCI) remains unclear. Here we report that overall lactate levels and lactylation are upregulated in the spinal cord after SCI. Notably, H4K12la was significantly elevated in the microglia of the injured spinal cord, whereas exogenous lactate treatment further elevated H4K12la in microglia after SCI. Functionally, lactate treatment promoted microglial proliferation, scar formation, axon regeneration, and locomotor function recovery after SCI. Mechanically, lactate-mediated H4K12la elevation promoted PD-1 transcription in microglia, thereby facilitating SCI repair. Furthermore, a series of rescue experiments confirmed that a PD-1 inhibitor or microglia-specific AAV-sh-PD-1 significantly reversed the therapeutic effects of lactate following SCI. This study illustrates the function and mechanism of lactate/H4K12la/PD-1 signaling in microglia-mediated tissue repair and provides a novel target for SCI therapy.

Elastin and collagen fibres in cutaneous wound healing.

Skin forms the outer barrier of the body. Upon injury, successful wound healing in normal skin restores tissue damage and counteracts the loss of extracellular matrix (ECM) proteins and cells. Collagens and elastin are the most abundant structural proteins of the ECM. In homeostasis, collagen type I is the prevalent form, but it is replaced by type III collagen upon wounding, and only later remodelled. In turn, unsuccessful healing results in scars, which tend to be inflexible and inelastic as compared to normal elastic dermis. Scar inelasticity may be due to the absence of mature elastin fibre formation and cross-linking. In this review, the available information on the process of formation of new collagen and elastic fibres during wound healing is analysed. The distinct roles of elastin and collagen proteins during healing are revisited and future research directions proposed which may help improve clinical management of open wounds and scars.

SHH induces macrophage oxidative phosphorylation and efferocytosis to promote scar formation.

Excessive scar formation such as hypertrophic scars and keloids, resulting from trauma or surgical procedures, present a widespread concern for causing disfigurement, discomfort, and functional limitations. Macrophages play pivotal roles in maintaining tissue homeostasis, orchestrating tissue development, repair, and immune responses, and its transition of function and phenotype plays a critical role in regulating the balance between inflammation and tissue regeneration, which is central to cutaneous scar formation. Recent evidence suggests the involvement of Sonic Hedgehog (SHH) in the induction of anti-inflammatory M2-like macrophage phenotypes within tumor microenvironments. In our study, we observed increased SHH expression in human hypertrophic scars, prompting an investigation into its influence on macrophage polarization, efferocytosis, and cutaneous scar formation. Our findings reveal that SHH can enhance oxidative phosphorylation (OXPHOS) in macrophages, augment macrophage efferocytosis, and promote M2 polarization, finally contributing to the progression of cutaneous scar formation. Notably, targeting SHH signaling with vismodegib exhibited promising potential in mitigating scar formation by reversing the effects of enhanced OXPHOS and M2 polarization in macrophages. In conclusion, this study underscores the critical roles of macrophage metabolism, particularly OXPHOS, efferocytosis and SHH signaling in cutaneous scar formation. Understanding these mechanisms provides new avenues for potential interventions and scar prevention strategies.

Interleukin-4 Receptor α Signaling in Myeloid Cells Controls Collagen Fibril Assembly in Skin Repair.

Activation of the immune response during injury is a critical early event that determines whether the outcome of tissue restoration is regeneration or replacement of the damaged tissue with a scar. The mechanisms by which immune signals control these fundamentally different regenerative pathways are largely unknown. We have demonstrated that, during skin repair in mice, interleukin-4 receptor α (IL-4Rα)-dependent macrophage activation controlled collagen fibril assembly and that this process was important for effective repair while having adverse pro-fibrotic effects. We identified Relm-α as one important player in the pathway from IL-4Rα signaling in macrophages to the induction of lysyl hydroxylase 2 (LH2), an enzyme that directs persistent pro-fibrotic collagen cross-links, in fibroblasts. Notably, Relm-ß induced LH2 in human fibroblasts, and expression of both factors was increased in lipodermatosclerosis, a condition of excessive human skin fibrosis. Collectively, our findings provide mechanistic insights into the link between type 2 immunity and initiation of pro-fibrotic pathways.

Inhibition of CK2 Diminishes Fibrotic Scar Formation and Improves Outcomes After Ischemic Stroke via Reducing BRD4 Phosphorylation.

Fibrotic scars play important roles in tissue reconstruction and functional recovery in the late stage of nervous system injury. However, the mechanisms underlying fibrotic scar formation and regulation remain unclear. Casein kinase II (CK2) is a protein kinase that regulates a variety of cellular functions through the phosphorylation of proteins, including bromodomain-containing protein 4 (BRD4). CK2 and BRD4 participate in fibrosis formation in a variety of tissues. However, whether CK2 affects fibrotic scar formation remains unclear, as do the mechanisms of signal regulation after cerebral ischemic injury. In this study, we assessed whether CK2 could modulate fibrotic scar formation after cerebral ischemic injury through BRD4. Primary meningeal fibroblasts were isolated from neonatal rats and treated with transforming growth factor-ß1 (TGF-ß1), SB431542 (a TGF-ß1 receptor kinase inhibitor) or TBB (a highly potent CK2 inhibitor). Adult SD rats were intraperitoneally injected with TBB to inhibit CK2 after MCAO/R. We found that CK2 expression was increased in vitro in the TGF-ß1-induced fibrosis model and in vivo in the MCAO/R injury model. The TGF-ß1 receptor kinase inhibitor SB431542 decreased CK2 expression in fibroblasts. The CK2 inhibitor TBB reduced the increases in proliferation, migration and activation of fibroblasts caused by TGF-ß1 in vitro, and it inhibited fibrotic scar formation, ameliorated histopathological damage, protected Nissl bodies, decreased infarct volume and alleviated neurological deficits after MCAO/R injury in vivo. Furthermore, CK2 inhibition decreased BRD4 phosphorylation both in vitro and in vivo. The findings of the present study suggested that CK2 may control BRD4 phosphorylation to regulate fibrotic scar formation, to affecting outcomes after ischemic stroke.

In vitro and in vivo Evaluation of Antifibrotic Properties of Verteporfin in a Composition of a Collagen Scaffold.

Extensive skin damage requires specialized therapy that stimulates regeneration processes without scarring. The possibility of using combination of a collagen gel application as a wound dressing and fibroblast attractant with verteporfin as an antifibrotic agent was examined in vivo and in vitro. In vitro effects of verteporfin on viability and myofibroblast markers expression were evaluated using fibroblasts isolated from human scar tissue. In vivo the collagen gel and verteporfin (individually and in combination) were applied into the wound to investigate scarring during skin regeneration: deviations in skin layer thickness, collagen synthesis, and extracellular matrix fibers were characterized. The results indicate that verteporfin reduces fibrotic phenotype by suppressing expression of the contractile protein Sm22α without inducing cell death. However, administration of verteporfin in combination with the collagen gel disrupts its ability to direct wound healing in a scarless manner, which may be related to incompatibility of the mechanisms by which collagen and verteporfin control regeneration.

Biological effect of laser-assisted scar healing (LASH) on standardized human three-dimensional wound healing skin models using fractional non-ablative 1540 nm Er:Glass or 1550 nm diode lasers.

PURPOSE: In postoperative wound healing after surgical operations or ablative laser treatments, recent studies suggest the timely use of non-ablative fractional laser treatments with the aim to improve wound healing and prevent pathological scar formation. However, the underlying molecular mechanisms are poorly understood. The aim of this study was to investigate the effects of laser-assisted scar healing (LASH) at the molecular level and to combine it with already established wound healing-promoting local treatments. METHODS: We irradiated full-thickness 3D skin models with a fractional ablative Er:YAG laser to set standardized lesions to the epidermal and upper dermal layer. Subsequently, LASH was induced by irradiating the models with either a fractional non-ablative 1540 nm Er:Glass or 1550 nm diode laser. In addition, we tested the combination of non-ablative fractional laser treatment and topical aftercare with a dexpanthenol-containing ointment (DCO). RESULTS: Histological analysis revealed that models irradiated with the 1540 nm Er:Glass or 1550 nm diode laser exhibited accelerated but not complete wound closure after 16 h. In contrast, additional topical posttreatment with DCO resulted in complete wound closure. At gene expression level, both non-ablative laser systems showed similar effects on epidermal differentiation and mild anti-inflammatory properties. The additional posttreatment with DCO enhanced the wound-healing effects of LASH, especially the upregulation of epidermal differentiation markers and anti-inflammatory cytokines at the gene expression level. CONCLUSION: This in vitro study deciphers the biological effects of LASH with a fractional non-ablative 1540 nm Er:Glass or a 1550 nm diode laser in 3D skin models. These data help to better understand the biological properties of the LASH technique and is important to optimize its application.

Human umbilical cord mesenchymal stem cells improve uterine incision healing after cesarean delivery in rats by modulating the TGF-ß/Smad signaling pathway.

OBJECTIVE: Although human umbilical cord-derived mesenchymal stem cells (HU-MSCs) have attracted increasing attention because of their pivotal functions in the process of wound healing, the underlying molecular mechanisms have been poorly understood. It has been shown that the TGF-ß/Smad signaling pathway plays an important role in the process of scar formation. The present study focused on exploring whether HU-MSCs improve uterine incision healing after cesarean delivery in rats via the TGF-ß/Smad signaling pathway. STUDY DESIGN: Pregnant rats were randomly assigned to three groups, including the NP group, incision-injected group (HU-MSCs1 group), and tail vein-injected group (HU-MSCs2 group), and 30 days after cesarean section, sampling was carried out to further explore the specific mechanisms from tissue and protein levels. RESULTS: HU-MSCs secretion could inhibit the fibrosis of scar tissue. We observed that the TGF-ß induced expression of TGF-ß1, Smad2, and Smad3 was attenuated upon HU-MSCs treatment in scar tissue, while the decrease in TGF-ß3 expression was enhanced by HU-MSCs. Furthermore, HU-MSCs treatment accelerated wound healing and attenuated collagen deposition in a damaged uterine rat model, leading to the promoting of uterine incision scarring. In addition, the expression of alpha-smooth muscle actin (a-SMA) was enhanced by HU-MSCs treatment. CONCLUSION: HU-MSCs transplantation promotes rat cesarean section uterine incision scar healing by modulating the TGF-ß/Smad signaling pathway.

Novel Fibrillar and Non-Fibrillar Collagens Involved in Fibrotic Scar Formation after Myocardial Infarction.

Following myocardial infarction (MI), adverse remodeling depends on the proper formation of fibrotic scars, composed of type I and III collagen. Our objective was to pinpoint the participation of previously unreported collagens in post-infarction cardiac fibrosis. Gene (qRT-PCR) and protein (immunohistochemistry followed by morphometric analysis) expression of fibrillar (types II and XI) and non-fibrillar (types VIII and XII) collagens were determined in RNA-sequencing data from 92 mice undergoing myocardial ischemia; mice submitted to permanent (non-reperfused MI, n = 8) or transient (reperfused MI, n = 8) coronary occlusion; and eight autopsies from chronic MI patients. In the RNA-sequencing analysis of mice undergoing myocardial ischemia, increased transcriptomic expression of collagen types II, VIII, XI, and XII was reported within the first week, a tendency that persisted 21 days afterwards. In reperfused and non-reperfused experimental MI models, their gene expression was heightened 21 days post-MI induction and positively correlated with infarct size. In chronic MI patients, immunohistochemistry analysis demonstrated their presence in fibrotic scars. Functional analysis indicated that these subunits probably confer tensile strength and ensure the cohesion of interstitial components. Our data reveal that novel collagens are present in the infarcted myocardium. These data could lay the groundwork for unraveling post-MI fibrotic scar composition, which could ultimately influence patient survivorship.

Nestin identifies a subpopulation of rat ventricular fibroblasts and participates in cell migration.

Fibroblast progenitor cells migrate to the endocardial region during cardiogenesis, and the migration of ventricular fibroblasts to the ischemically damaged region of the infarcted adult heart is a seminal event of reparative fibrosis. The intermediate filament protein nestin is implicated in cell migration and expression identified in a subpopulation of scar-derived myofibroblasts. The present study tested the hypothesis that fibroblast progenitor cells express nestin, and the intermediate filament protein drives the migratory phenotype of ventricular fibroblasts. Transcription factor 21 (Tcf21)- and Wilms tumor 1 (WT1)-fibroblast progenitor cells identified in the epicardial/endocardial regions of the E12.5- to E13.5-day embryonic mouse heart predominantly expressed nestin. Nuclear Tcf21/WT1 staining was identified in neonatal rat ventricular fibroblasts (NNVFbs), and a subpopulation coexpressed nestin. Nuclear Tcf21/WT1 expression persisted in adult rat ventricular fibroblasts, whereas nestin protein levels were downregulated. Nestin-expressing NNVFbs exhibited a unique phenotype as the subpopulation was refractory to cell cycle reentry in response to selective stimuli. Nestin(-)- and nestin(+)-scar-derived rat myofibroblasts plated in Matrigel unmasked a migratory phenotype characterized by the de novo formation of lumen-like structures. The elongated membrane projections emanating from scar myofibroblasts delineating the boundary of lumen-like structures expressed nestin. Lentiviral short-hairpin RNA (shRNA)-mediated nestin depletion inhibited the in vitro migratory response of NNVFbs as the wound radius was significantly larger compared with NNVFbs infected with the empty lentivirus. Thus, nestin represents a marker of embryonic Tcf21/WT1(+)-fibroblast progenitor cells. The neonatal rat heart contains a distinct subpopulation of nestin-immunoreactive Tcf21/WT1(+) fibroblasts refractory to cell cycle reentry, and the intermediate filament protein may preferentially facilitate ventricular fibroblast migration during physiological/pathological remodeling.NEW & NOTEWORTHY Tcf21/WT1(+)-fibroblast progenitor cells of the embryonic mouse heart predominantly express the intermediate filament protein nestin. A subpopulation of Tcf21/WT1(+)-neonatal rat ventricular fibroblasts express nestin and are refractory to selective stimuli influencing cell cycle reentry. Scar-derived myofibroblasts plated in Matrigel elicit the formation of lumen-like structures characterized by the appearance of nestin(+)-membrane projections. Lentiviral shRNA-mediated nestin depletion in a subpopulation of neonatal rat ventricular fibroblasts suppressed the migratory response following the in vitro scratch assay.