The diagnostic value and associated molecular mechanism study for fibroblast-related mitochondrial genes on keloid.

PURPOSE: This study aims to reveal the mechanism of fibroblast-related mitochondrial genes on keloid formation and explore promising signature genes for keloid diagnosis. METHOD: The distribution of fibroblasts between the keloid sample and control sample based on three keloid datasets, followed by the differentially expressed genes (DEGs) investigation and associated enrichment analysis. Then, hub genes were explored based on DEGs, mitochondrial genes from an online database, as well as fibroblast-related genes that were revealed by WCGNA. Subsequently, signature genes were screened through machine learning, and their diagnostic value was validated by nomogram. Moreover, the targeted drugs and related transcriptional regulation of these genes were analyzed. Finally, the verification analysis was performed on signature genes using qPCR analysis. RESULT: A total of totally 329 DEGs were revealed based on three datasets, followed by enrichment analysis. WGCNA revealed a total of 258 fibroblast-related genes, which were primarily assembled in functions like muscle tissue development. By using machine learning, we screened four signature genes (ACSF2, ALDH1B1, OCIAD2, and SIRT4) based on eight hub genes (fibroblast-related mitochondrial genes). Nomogram and validation analyses confirmed the well-diagnostic performance of these four genes in keloid. Immune infiltration and drug correlation analyses showed that SIRT4 was significantly associated with immune cell type 2 T helper cells and molecular drug cyclosporin. All these findings provided new perspectives for the clinical diagnosis and therapy of keloid. CONCLUSION: The fibroblast-related mitochondrial genes including SIRT4, OCIAD2, ALDH1B1, and ACSF2 were novel signature genes for keloid diagnosis, offering novel targets and strategies for diagnosis and therapy of keloid.

Phosphodiesterase 4 is overexpressed in keloid epidermal scars and its inhibition reduces keratinocyte fibrotic alterations.

BACKGROUND: Epidermal remodeling and hypertrophy are hallmarks of skin fibrotic disorders, and keratinocyte to mesenchymal (EMT)-like transformations drive epidermis alteration in skin fibrosis such as keloids and hypertrophic scars (HTS). While phosphodiesterase 4 (PDE4) inhibitors have shown effectiveness in various fibrotic disorders, their role in skin fibrosis is not fully understood. This study aimed to explore the specific role of PDE4B in epidermal remodeling and hypertrophy seen in skin fibrosis. METHODS: In vitro experiments examined the effects of inhibiting PDE4A-D (with Roflumilast) or PDE4B (with siRNA) on TGFß1-induced EMT differentiation and dedifferentiation in human 3D epidermis. In vivo studies investigated the impact of PDE4 inhibition on HOCl-induced skin fibrosis and epidermal hypertrophy in mice, employing both preventive and therapeutic approaches. RESULTS: The study found increased levels of PDE4B (mRNA, protein) in keloids > HTS compared to healthy epidermis, as well as in TGFß-stimulated 3D epidermis. Keloids and HTS epidermis exhibited elevated levels of collagen Iα1, fibronectin, αSMA, N-cadherin, and NOX4 mRNA, along with decreased levels of E-cadherin and ZO-1, confirming an EMT process. Inhibition of both PDE4A-D and PDE4B prevented TGFß1-induced Smad3 and ERK1/2 phosphorylation and mesenchymal differentiation in vitro. PDE4A-D inhibition also promoted mesenchymal dedifferentiation and reduced TGFß1-induced ROS and keratinocyte senescence by rescuing PPM1A, a Smad3 phosphatase. In vivo, PDE4 inhibition mitigated HOCl-induced epidermal hypertrophy in mice in both preventive and therapeutic settings. CONCLUSIONS: Overall, the study supports the potential of PDE4 inhibitors, particularly PDE4B, in treating skin fibrosis, including keloids and HTS, shedding light on their functional role in this condition.

The fibroblast heterogeneity across keloid, normal and tumor samples from single-cell resolution.

Keloids are defined as a benign dermal fibroproliferative disorder, with excessive fibroblast proliferation, and excessive overproduction of collagen. Although the heterogeneity during keloid development has been extensively studied, the heterogeneity across different skin states is still unclear. So, a global comparison across skin states is needed. In this study, we collected samples from 5 states of skin, including melanoma, cutaneous squamous cell carcinoma, keloid skin, scar skin, and healthy control samples. The heterogeneity of cell types and subtypes was analyzed and compared across 5 states, and we observed significant differences among them. Our results showed a cancer-like fibroblast, which is not in normal samples, may play an important role in antigen processing and presentation. We also noticed that the mesenchymal fibroblast increased in keloid samples, which highly expressed POSTN. And POSTN may participate in epithelial-mesenchymal transition and collagen overexpression to promote keloid growth. These findings help to understand the alteration among different skin states and provide potential genetic basis for keloid therapies.

Comprehensive Insights into Keloid Pathogenesis and Advanced Therapeutic Strategies.

Keloid scars, characterized by abnormal fibroproliferation and excessive extracellular matrix (ECM) production that extends beyond the original wound, often cause pruritus, pain, and hyperpigmentation, significantly impacting the quality of life. Keloid pathogenesis is multifactorial, involving genetic predisposition, immune response dysregulation, and aberrant wound-healing processes. Central molecular pathways such as TGF-ß/Smad and JAK/STAT are important in keloid formation by sustaining fibroblast activation and ECM deposition. Conventional treatments, including surgical excision, radiation, laser therapies, and intralesional injections, yield variable success but are limited by high recurrence rates and potential adverse effects. Emerging therapies targeting specific immune pathways, small molecule inhibitors, RNA interference, and mesenchymal stem cells show promise in disrupting the underlying mechanisms of keloid pathogenesis, potentially offering more effective and lasting treatment outcomes. Despite advancements, further research is essential to fully elucidate the precise mechanisms of keloid formation and to develop targeted therapies. Ongoing clinical trials and research efforts are vital for translating these scientific insights into practical treatments that can markedly enhance the quality of life for individuals affected by keloid scars.

Identification of novel immune-related signatures for keloid diagnosis and treatment: insights from integrated bulk RNA-seq and scRNA-seq analysis.

BACKGROUND: Keloid is a disease characterized by proliferation of fibrous tissue after the healing of skin tissue, which seriously affects the daily life of patients. However, the clinical treatment of keloids still has limitations, that is, it is not effective in controlling keloids, resulting in a high recurrence rate. Thus, it is urgent to identify new signatures to improve the diagnosis and treatment of keloids. METHOD: Bulk RNA seq and scRNA seq data were downloaded from the GEO database. First, we used WGCNA and MEGENA to co-identify keloid/immune-related DEGs. Subsequently, we used three machine learning algorithms (Randomforest, SVM-RFE, and LASSO) to identify hub immune-related genes of keloid (KHIGs) and investigated the heterogeneous expression of KHIGs during fibroblast subpopulation differentiation using scRNA-seq. Finally, we used HE and Masson staining, quantitative reverse transcription-PCR, western blotting, immunohistochemical, and Immunofluorescent assay to investigate the dysregulated expression and the mechanism of retinoic acid in keloids. RESULTS: In the present study, we identified PTGFR, RBP5, and LIF as KHIGs and validated their diagnostic performance. Subsequently, we constructed a novel artificial neural network molecular diagnostic model based on the transcriptome pattern of KHIGs, which is expected to break through the current dilemma faced by molecular diagnosis of keloids in the clinic. Meanwhile, the constructed IG score can also effectively predict keloid risk, which provides a new strategy for keloid prevention. Additionally, we observed that KHIGs were also heterogeneously expressed in the constructed differentiation trajectories of fibroblast subtypes, which may affect the differentiation of fibroblast subtypes and thus lead to dysregulation of the immune microenvironment in keloids. Finally, we found that retinoic acid may treat or alleviate keloids by inhibiting RBP5 to differentiate pro-inflammatory fibroblasts (PIF) to mesenchymal fibroblasts (MF), which further reduces collagen secretion. CONCLUSION: In summary, the present study provides novel immune signatures (PTGFR, RBP5, and LIF) for keloid diagnosis and treatment, and identifies retinoic acid as potential anti-keloid drugs. More importantly, we provide a new perspective for understanding the interactions between different fibroblast subtypes in keloids and the remodeling of their immune microenvironment.

Liposomes Loaded with 5-Fluorouracil Can Improve the Efficacy in Pathological Scars.

Introduction: Pathological scars, such as hypertrophic scars and keloids, are characterized by the proliferation of fibroblasts and the deposition of collagen that often cause pruritus, pain, and disfigurement. Due to their high incidence and deformity, pathological scars have resulted in severe physical and psychological trauma for patients. Intralesional injection of 5-fluorouracil (5-Fu) is a recommended option for treating pathological scars. However, the efficacy of 5-Fu injection was limited and unstable due to limited drug penetration and short retention time. Methods: Liposomes are promising carriers that have advantages, such as high biocompatibility, controlled release property, and enhanced clinical efficacy. Here, we constructed a transdermal 5-Fu-loaded liposome (5-Fu-Lip) to provide a more effective and safer modality to scar treatment. Results: Compared to 5-Fu, 5-Fu-Lip showed superior ability in inhibiting primary keloid fibroblasts proliferation, migration, and collagen deposition, and also significantly inhibited human umbilical vein endothelial cells (HUVECs) proliferation and microvessel construction. In vivo experiments demonstrated that 5-Fu-Lip can significantly reduce the severity of hypertrophic scars in a rabbit ear wounding model. Discussion: 5-Fu-Lip provides a promising strategy to improve drug efficacy, which has great potential in the treatment of pathological scars.

SIRT6 Inhibits the Proliferation and Collagen Synthesis of Keloid Fibroblasts through MAPK/ERK Pathway.

BACKGROUND: Keloid, a fibroproliferative disorder, significantly impacts patients' quality of life, yet effective therapies remain elusive. This study explored the role of silent information regulator 6 (SIRT6) in modulating the proliferation, invasion, and collagen synthesis of keloid fibroblasts. METHODS: Keloid and normal skin specimens were collected, and fibroblasts were isolated from the keloid tissue. SIRT6 recombinant adenovirus (Ad) was constructed to infect keloid fibroblasts to overexpress SIRT6. This study entails three groups: Control group, adenovirus-Negative Control (Ad-NC) group, and Ad-SIRT6 group. SIRT6 protein and mRNA levels were measured via Western blotting and Quantitative reverse transcription polymerase chain reaction (qRT-PCR), respectively. Cell viability was determined using 5-ethynyl-2'-deoxyuridine (EdU) assay. Flow cytometry was exploited to measure cell apoptosis. To investigate cell migration, wound healing assay and Transwell assay were employed. Western blotting was also utilized to study the expression levels of apoptotic proteins, collagen deposition-related proteins, and Mitogen-Activated Protein Kinases (MAPK)/extracellular regulated protein kinases (ERK) pathway-related proteins. RESULTS: Compared to the control and Ad-NC groups, the Ad-SIRT6 group exhibited significantly elevated SIRT6 level; diminished cell proliferation, migration and invasion; reduced protein levels of α-smooth muscle actin (α-SMA), collagen I, collagen III, phospho SMAD Family Member 3 (p-Smad3), transforming growth factor-ß 1 (TGF-ß1), and MAPK/ERK pathway proteins (phospho extracellular signal-regulated protein kinase 1/2 (p-ERK1/2), phospho MAP kinase-ERK kinase (p-MEK) and phospho-c-Raf (p-c-Raf)). Treatment with epidermal growth factor (EGF), an MAPK/ERK pathway agonists, reversed the inhibitory effect of SIRT6 on cell activity and inhibited apoptosis in keloid fibroblasts. CONCLUSION: SIRT6 overexpression in keloid fibroblasts attenuates proliferation, invasion, and collagen synthesis, while fostering apoptosis, likely through the suppression of MAPK/ERK pathway activity. This suggests a potential therapeutic target for keloid treatment.

Remdesivir alleviates skin fibrosis by suppressing TGF-ß1 signaling pathway.

Fibrotic skin diseases, such as keloids, are pathological results of aberrant tissue healing and are characterized by overgrowth of dermal fibroblasts. Remdesivir (RD), an antiviral drug, has been reported to have pharmacological activities in a wide range of fibrotic diseases. However, whether RD function on skin fibrosis remains unclear. Therefore, in our study, we explored the potential effect and mechanisms of RD on skin fibrosis both in vivo and in vitro. As expected, the results demonstrated that RD alleviated BLM-induced skin fibrosis and attenuates the gross weight of keloid tissues in vivo. Further studies suggested that RD suppressed fibroblast activation and autophagy both in vivo and in vitro. In addition, mechanistic research showed that RD attenuated fibroblasts activation by the TGF-ß1/Smad signaling pathway and inhibited fibroblasts autophagy by the PI3K/Akt/mTOR signaling pathway. In summary, our results demonstrate therapeutic potential of RD for skin fibrosis in the future.

Unlocking the potential of tranilast: Targeting fibrotic signaling pathways for therapeutic benefit.

Fibrosis is the excessive deposition of extracellular matrix in an organ or tissue that results from an impaired tissue repair in response to tissue injury or chronic inflammation. The progressive nature of fibrotic diseases and limited treatment options represent significant healthcare challenges. Despite the substantial progress in understanding the mechanisms of fibrosis, a gap persists translating this knowledge into effective therapeutics. Here, we discuss the critical mediators involved in fibrosis and the role of tranilast as a potential antifibrotic drug to treat fibrotic conditions. Tranilast, an antiallergy drug, is a derivative of tryptophan and has been studied for its role in various fibrotic diseases. These include scleroderma, keloid and hypertrophic scars, liver fibrosis, renal fibrosis, cardiac fibrosis, pulmonary fibrosis, and uterine fibroids. Tranilast exerts antifibrotic effects by suppressing fibrotic pathways, including TGF-ß, and MPAK. Because it disrupts fibrotic pathways and has demonstrated beneficial effects against keloid and hypertrophic scars, tranilast could be used to treat other conditions characterized by fibrosis.

The biodistribution of triamcinolone acetonide injections in severe keloids: an exploratory three-dimensional fluorescent cryomicrotome study.

Intralesional corticosteroid injections are a first-line treatment for keloids; yet clinical treatment results are highly variable and often suboptimal. Variation in triamcinolone acetonide (TAC) biodistribution may be an important reason for the variable effects of TAC treatment in keloids. In this exploratory study we investigated the biodistribution of TAC in keloids and normal skin using different drug delivery techniques. Fluorescent-labeled TAC suspension was administered into keloids and normal skin with a hypodermic needle and an electronic pneumatic jet injector. TAC biodistribution was represented by the fluorescent TAC volume and 3D biodistribution shape of TAC, using a 3D-Fluorescence-Imaging Cryomicrotome System. Twenty-one keloid and nine normal skin samples were analyzed. With needle injections, the mean fluorescent TAC volumes were 990 µl ± 479 in keloids and 872 µl ± 227 in normal skin. With the jet injector, the mean fluorescent TAC volumes were 401 µl ± 252 in keloids and 249 µl ± 67 in normal skin. 3D biodistribution shapes of TAC were highly variable in keloids and normal skin. In conclusion, TAC biodistribution in keloids is highly variable for both needle and jet injection. This may partly explain the variable treatment effects of intralesional TAC in keloids. Future research is needed to confirm this preliminary finding and to optimize drug delivery in keloids.

Diagnostic and prognostic significance of keloid-like collagen remodeling patterns in the extracellular matrix of colorectal cancer.

Background: The desmoplastic reaction is considered a promising prognostic parameter for colorectal cancer. However, intermediate desmoplastic reaction is characterized by sizeable stromal heterogeneity, including both small amounts of keloid-like collagen (KC) in the fibrotic stroma and thick tufts of KC circumferentially surrounding cancer nests and occupying most of the fields of view. The present study aimed to evaluate the diagnostic and prognostic significance of KC histophenotyping with a quantitative visual assessment of its presence in the stroma of the invasive margin of TNM (The "tumor-node-metastasis" classification) stage II/III colorectal cancer (CRC). Methods and results: 175 resected tumors from patients with TNM stage II/III CRC were examined. Keloid-like collagen was assessed according to Ueno H. criteria. KC was assessed at the primary tumor invasive margin using Hematoxylin & Eosin and Masson's trichrome staining. The cut-off point for KC was examined using "the best cutoff approach by log-rank test." Using a cutoff point of 30%, we histologically divided fibrous stroma in the invasive area into two groups: "type A"-KC ≤ 0.3 and "type B"-KC>0.3. Type A stroma was observed in 48% of patients, type B-in 52%. The association between collagen amount and 5-year recurrence-free survival (5-RFS) was assessed using Cox regression analysis. Kaplan-Meier analysis and log-rank tests were used to assess the significance of survival analysis. Analysis of categorical variables showed that increased KC in CRC stroma predicted adverse outcomes for 5-RFS (hazard ratio [HR] = 3.143, 95%, confidence interval [CI] = 1.643-6.012, p = 0.001). Moreover, in Kaplan-Meier analysis, the log-rank test showed that type B exhibited worse 5-RFS than type A (p = 0.000). Conclusion: KC is an independent predictor of 5-year overall and RFS in patients with TNM stage II/III CRC treated with surgery, with worse survival rates when the amount of KC increases by >30%.

Ethyl Pyruvate Decreases Collagen Synthesis and Upregulates MMP Activity in Keloid Fibroblasts and Keloid Spheroids.

Keloids, marked by abnormal cellular proliferation and excessive extracellular matrix (ECM) accumulation, pose significant therapeutic challenges. Ethyl pyruvate (EP), an inhibitor of the high-mobility group box 1 (HMGB1) and TGF-ß1 pathways, has emerged as a potential anti-fibrotic agent. Our research evaluated EP's effects on keloid fibroblast (KF) proliferation and ECM production, employing both in vitro cell cultures and ex vivo patient-derived keloid spheroids. We also analyzed the expression levels of ECM components in keloid tissue spheroids treated with EP through immunohistochemistry. Findings revealed that EP treatment impedes the nuclear translocation of HMGB1 and diminishes KF proliferation. Additionally, EP significantly lowered mRNA and protein levels of collagen I and III by attenuating TGF-ß1 and pSmad2/3 complex expression in both human dermal fibroblasts and KFs. Moreover, metalloproteinase I (MMP-1) and MMP-3 mRNA levels saw a notable increase following EP administration. In keloid spheroids, EP induced a dose-dependent reduction in ECM component expression. Immunohistochemical and western blot analyses confirmed significant declines in collagen I, collagen III, fibronectin, elastin, TGF-ß, AKT, and ERK 1/2 expression levels. These outcomes underscore EP's antifibrotic potential, suggesting its viability as a therapeutic approach for keloids.

Development and validation of novel keloid-derived immortalized fibroblast cell lines.

Keloids are a common connective tissue disorder with an ill-understood etiopathogenesis and no effective treatment. This is exacerbated because of the absence of an animal model. Patient-derived primary keloid cells are insufficient as they age through passaging and have a limited supply. Therefore, there is an unmet need for development of a cellular model that can consistently and faithfully represent keloid's pathognomic features. In view of this, we developed keloid-derived immortalized fibroblast (KDIF) cell lines from primary keloid fibroblasts (PKF) by transfecting the human telomerase reverse transcriptase (hTERT) gene. The TERT gene encodes the catalytic subunit of the telomerase enzyme, which is responsible for maintaining the cellular replicative potential (cellular immortalization). Primary fibroblasts from keloid-specific lesional (peripheral, middle, and top) as well as extralesional sites were isolated and evaluated for cell line development and comparative cellular characteristics by employing qRT-PCR and immunofluorescence staining. Moreover, the immortalized behavior of KDIF cell lines was evaluated by comparing with cutaneous fibrosarcoma and dermatofibrosarcoma protuberans cell lines. Stable KDIF cell lines with elevated expression of hTERT exhibited the cellular characteristics of site-specific keloid fibroblasts. Histochemical staining for ß-galactosidase revealed a significantly lower number of ß-gal-positive cells in all three KDIF cell lines compared with that in PKFs. The cell growth curve pattern was studied over 10 passages for all three KDIF cell lines and was compared with the control groups. The results showed that all three KDIF cell lines grew significantly faster and obtained a fast growing characteristic as compared to primary keloid and normal fibroblasts. Phenotypic behavior in growth potential is an indication of hTERT-mediated immortalized transformation. Cell migration analysis revealed that the top and middle KDIF cell lines exhibited similar migration trend as site-specific PKFs. Notably, peripheral KDIF cell line showed significantly enhanced cell migration in comparison to the primary peripheral fibroblasts. All KDIF cell lines expressed Collagen I protein as a keloid-associated fibrotic marker. Functional testing with triamcinolone inhibited cell migration in KDIF. ATCC short tandem repeat profiling validated the KDIF as keloid representative cell line. In summary, we provide the first novel KDIF cell lines. These cell lines overcome the limitations related to primary cell passaging and tissue supply due to immortalized features and present an accessible and consistent experimental model for keloid research.

BMP1 Promotes Keloid by Inducing Fibroblast Inflammation and Fibrogenesis.

Keloid is a typical fibrotic and inflammatory skin disease with unclear mechanisms and few therapeutic targets. In this study, we found that BMP1 was significantly increased in a collagen high-expressing subtype of fibroblast by reanalyzing a public single-cell RNA-sequence data set of keloid. The number of BMP1-positive fibroblast cells was increased in keloid fibrotic loci. Increased levels of BMP1 were further validated in the skin tissues and fibroblasts from keloid patients. Additionally, a positive correlation between BMP1 and the Keloid Area and Severity Index was found in keloid patients. In vitro analysis revealed collagen production, the phosphorylation levels of p65, and the IL-1ß secretion decreased in BMP1 interfered keloid fibroblasts. Besides, the knockdown of BMP1 inhibited the growth and migration of keloid fibroblast cells. Mechanistically, BMP1 inhibition downregulated the noncanonical TGF-ß pathways, including p-p38 and p-ERK1/2 signaling. Furthermore, we found the delivery of BMP1 siRNAs could significantly alleviate keloid in human keloid-bearing nude mice. Collectively, our results indicated that BMP1 exhibited various pathogenic effects on keloids as promoting cell proliferation, migration, inflammation, and ECM deposition of fibroblast cells by regulating the noncanonical TGF-ß/p38 MAPK, and TGF-ß/ERK pathways. BMP1-lowing strategies may appear as a potential new therapeutic target for keloid.

The role of α7nAchR and PD-L1 in neuroimmune regulation of keloid treatment.

Keloid formation, characterized by aberrant fibroproliferation and immune dysregulation, remains a challenging clinical concern. This study aims to elucidate the neuroimmune mechanisms underlying keloid pathogenesis and explores the efficacy of a combined treatment approach involving modulation of the α7 nicotinic acetylcholine receptor (α7nAchR), a key player in neural transmission, and programmed death ligand 1 (PD-L1), an immune checkpoint molecule, for keloid intervention. A key innovation lies in the identification of signal peptide-CUB-EGF-like domain-containing protein 3 (SCUBE3) as a potential target gene influenced by this dual treatment. We elucidate the underlying mechanism, wherein the hypoxic keloid microenvironment fosters an upsurge in SCUBE3 secretion. Subsequently, SCUBE3 forms complexes with TGF-ß, initiating the activation of the PI3K/AKT/NF-κB signaling pathway. Notably, SCUBE3 is secreted in the form of exosomes, thereby exerting a profound influence on the differentiation of T cells and macrophages within the keloid milieu. This research not only provides a comprehensive understanding of the molecular mechanisms involved but also offers a promising avenue for the development of targeted therapies to address keloid-associated fibrosis and immune dysregulation. In conclusion, the combined inhibition of α7nAchR and PD-L1 represents a promising therapeutic strategy with SCUBE3 as a pivotal molecular target in the complex landscape of keloid pathophysiology.

Serum deprivation protein response intervenes in the proliferation, motility, and extracellular matrix production in keloid fibroblasts by blocking the amplification of TGF-ß1/SMAD signal cascade via ERK1/2.

Keloid formation has been linked to abnormal fibroblast function, such as excessive proliferation and extracellular matrix (ECM) production. Serum deprivation protein response (SDPR) is a crucial regulator of cellular function under diverse pathological conditions, yet its role in keloid formation remains unknown. The current work investigated the function of SDPR in regulating the proliferation, motility, and ECM production of keloid fibroblasts (KFs), as well as to decipher the mechanisms involved. Analysis of RNA sequencing data from the GEO database demonstrated significant down-regulation of SDPR in KF compared to normal fibroblasts (NFs). This down-regulation was also observed in clinical keloid specimens and isolated KFs. Overexpression of SDPR suppressed the proliferation, motility, and ECM production of KFs, while depletion of SDPR exacerbated the enhancing impact of TGF-ß1 on the proliferation, motility, and ECM production of NFs. Mechanistic studies revealed that SDPR overexpression repressed TGF-ß/Smad signal cascade activation in KFs along with decreased levels of phosphorylated Samd2/3, while SDPR depletion exacerbated TGF-ß/Smad activation in TGF-ß1-stimulated NFs. SDPR overexpression also repressed ERK1/2 activation in KFs, while SDPR depletion exacerbated ERK1/2 activation in TGF-ß1-stimulated NFs. Inhibition of ERK1/2 abolished SDPR-depletion-induced TGF-ß1/Smad activation, cell proliferation, motility, and ECM production in NFs. In conclusion, SDPR represses the proliferation, motility, and ECM production in KFs by blocking the TGF-ß1/Smad pathway in an ERK1/2-dependent manner. The findings highlight the role of SDPR in regulating abnormal behaviors of fibroblasts associated with keloid formation and suggest it as a potential target for anti-keloid therapy development.

STAT3-induced lncRNA GNAS-AS1 accelerates keloid formation by mediating the miR-196a-5p/CXCL12/STAT3 axis in a feedback loop.

Keloids are pathological scar tissue resulting from skin trauma or spontaneous formation, often accompanied by itching and pain. Although GNAS antisense RNA 1 (GNAS-AS1) shows abnormal upregulation in keloids, the underlying molecular mechanism is unclear. The levels of genes and proteins in clinical tissues from patients with keloids and human keloid fibroblasts (HKFs) were measured using quantitative reverse transcription PCR, western blot and enzyme-linked immunosorbent assay. The features of HKFs, including proliferation and migration, were evaluated using cell counting kit 8 and a wound healing assay. The colocalization of GNAS-AS1 and miR-196a-5p in HKFs was measured using fluorescence in situ hybridization. The relationships among GNAS-AS1, miR-196a-5p and C-X-C motif chemokine ligand 12 (CXCL12) in samples from patients with keloids were analysed by Pearson correlation analysis. Gene interactions were validated by chromatin immunoprecipitation and luciferase reporter assays. GNAS-AS1 and CXCL12 expression were upregulated and miR-196a-5p expression was downregulated in clinical tissues from patients with keloids. GNAS-AS1 knockdown inhibited proliferation, migration, and extracellular matrix (ECM) accumulation of HKFs, all of which were reversed by miR-196a-5p downregulation. Signal transducer and activator of transcription 3 (STAT3) induced GNAS-AS1 transcription through GNAS-AS1 promoter interaction, and niclosamide, a STAT3 inhibitor, decreased GNAS-AS1 expression. GNAS-AS1 positively regulated CXCL12 by sponging miR-196-5p. Furthermore, CXCL12 knockdown restrained STAT3 phosphorylation in HKFs. Our findings revealed a feedback loop of STAT3/GNAS-AS1/miR-196a-5p/CXCL12/STAT3 that promoted HKF proliferation, migration and ECM accumulation and affected keloid progression.

Ferrostatin-1 inhibits fibroblast fibrosis in keloid by inhibiting ferroptosis.

Background: Keloid is a chronic proliferative fibrotic disease caused by abnormal fibroblasts proliferation and excessive extracellular matrix (ECM) production. Numerous fibrotic disorders are significantly influenced by ferroptosis, and targeting ferroptosis can effectively mitigate fibrosis development. This study aimed to investigate the role and mechanism of ferroptosis in keloid development. Methods: Keloid tissues from keloid patients and normal skin tissues from healthy controls were collected. Iron content, lipid peroxidation (LPO) level, and the mRNA and protein expression of ferroptosis-related genes including solute carrier family 7 member 11 (SLC7A11), glutathione peroxidase 4 (GPX4), transferrin receptor (TFRC), and nuclear factor erythroid 2-related factor 2 (Nrf2) were determined. Mitochondrial morphology was observed using transmission electron microscopy (TEM). Keloid fibroblasts (KFs) were isolated from keloid tissues, and treated with ferroptosis inhibitor ferrostatin-1 (fer-1) or ferroptosis activator erastin. Iron content, ferroptosis-related marker levels, LPO level, mitochondrial membrane potential, ATP content, and mitochondrial morphology in KFs were detected. Furthermore, the protein levels of α-smooth muscle actin (α-SMA), collagen I, and collagen III were measured to investigate whether ferroptosis affect fibrosis in KFs. Results: We found that iron content and LPO level were substantially elevated in keloid tissues and KFs. SLC7A11, GPX4, and Nrf2 were downregulated and TFRC was upregulated in keloid tissues and KFs. Mitochondria in keloid tissues and KFs exhibited ferroptosis-related pathology. Fer-1 treatment reduced iron content, restrained ferroptosis and mitochondrial dysfunction in KFs, Moreover, ferrostatin-1 restrained the protein expression of α-SMA, collagen I, and collagen III in KFs. Whereas erastin treatment showed the opposite results. Conclusion: Ferroptosis exists in keloid. Ferrostatin-1 restrained ECM deposition and fibrosis in keloid through inhibiting ferroptosis, and erastin induced ECM deposition and fibrosis through intensifying ferroptosis.

Enhanced bioenergetic cellular activity with metabolic switch to aerobic glycolysis in Keloid and Folliculitis Keloidalis Nuchae.

Keloid scars and folliculitis keloidalis nuchae (FKN) are benign fibroproliferative dermal lesions of unknown aetiology and ill-defined treatment, which typically present in genetically susceptible individuals. Their pathognomonic hallmarks include local aggressive invasive behaviour plus high recurrence post-therapy. In view of this, we investigated proliferative and key parameters of bioenergetic cellular characteristics of site-specific keloid-derived fibroblasts (intra(centre)- and peri(margin)-lesional) and FKN compared to normal skin and normal flat non-hypertrophic scar fibroblasts as negative controls.The results showed statistically significant (P < 0.01) and variable growth dynamics with increased proliferation and migration in keloid fibroblasts, while FKN fibroblasts showed a significant (P < 0.001) increase in proliferation but similar migration profile to controls. A statistically significant metabolic switch towards aerobic glycolysis in the fibroblasts from the disease conditions was noted. Furthermore, an increase in basal glycolysis with a concomitant increase in the cellular maximum glycolytic capacity was also demonstrated in perilesional keloid and FKN fibroblasts (P < 0.05). Mitochondrial function parameters showed increased oxidative phosphorylation in the disease conditions (P < 0.05) indicating functional mitochondria. These findings further suggest that Keloids and FKN demonstrate a switch to a metabolic phenotype of aerobic glycolysis. Increased glycolytic flux inhibition is a potential mechanistic basis for future therapy.

No Association of Polymorphisms in the Genes Encoding Interleukin-6 and Interleukin-6 Receptor Subunit Alpha with the Risk of Keloids in Polish Patients.

A keloid is a benign fibroproliferative hypertrophy of scar tissue that extends outside the original wound and invades adjacent healthy skin. Keloid formation is thought to be a complex process including overactivity of the interleukin-6 signaling pathway and genetic susceptibility. The aim of the study was to investigate possible associations between rs1800797, rs1800796, and rs1800795 polymorphisms in the promoter of the IL6 gene encoding interleukin-6 and the rs2228145 polymorphism in the IL6R gene encoding the interleukin-6 receptor subunit alpha with the predisposition to keloids in Polish patients. The genetic polymorphisms were identified either using Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) or sequencing of samples of genomic DNA extracted from blood leukocytes of 86 adult patients with keloids and 100 newborns comprising a control group. No significant differences in the distributions of IL6 or IL6R alleles or genotypes were found between keloid patients and newborn controls. There were also no significant differences between both groups in the distribution of IL6 haplotypes. The IL6 rs1800797, rs1800796 and rs1800795 and IL6R rs2228145 polymorphisms were not found to predispose individuals in the study group to keloids. IL6 promoter haplotypes were not found to be associated with a higher risk of keloids in the studied group.