RESUMEN
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.
Asunto(s)
Fibroblastos , Genes Mitocondriales , Queloide , Queloide/genética , Queloide/patología , Queloide/diagnóstico , Humanos , Fibroblastos/metabolismo , Genes Mitocondriales/genética , Aprendizaje Automático , Perfilación de la Expresión Génica , Masculino , FemeninoRESUMEN
BACKGROUND: Fibrosis with unrelieved chronic inflammation is an important pathological change in keloids. Mitochondrial autophagy plays a crucial role in reducing inflammation and inhibiting fibrosis. Adipose stem cell-derived exosomes, a product of adipose stem cell paracrine secretion, have pharmacological effects, such as anti-inflammatory and antiapoptotic effects, and mediate autophagy. Therefore, this study aims to investigate the function and mechanism of adipose stem cell exosomes in the treatment of keloids. METHOD: We isolated adipose stem cell exosomes under normoxic and hypoxic condition to detect their effects on keloid fibroblast proliferation, migration, and collagen synthesis. Meanwhile, 740YPDGFR (PI3K/AKT activator) was applied to detect the changes in autophagic flow levels and mitochondrial morphology and function in keloid fibroblasts. We constructed a human keloid mouse model by transplanting human keloid tissues into six-week-old (20-22 g; female) BALB/c nude mice, meanwhile, we applied adipose stem cell exosomes to treat the mouse model and observed the retention and effect of ADSC exosomes in vivo. RESULTS: ADSC exosomes can inhibit the PI3K/AKT/mTOR signaling pathway. The exosomes of ADSCs decreased the inflammatory level of KFs, enhanced the interaction between P62 and LC3, and restored the mitochondrial membrane potential. In the human keloid mouse model, ADSC exosomes can exist stably, promote mitochondrial autophagy in keloid tissue, improve mitochondrial morphology, reduce inflammatory reaction and fibrosis. Meanwhile, At the same time, the exosomes derived from hypoxic adipose stem cells have played a more effective role in both in vitro and in vivo experiments. CONCLUSIONS: Adipose stem cell exosomes inhibited the PI3K/AKT/mTOR pathway, activated mitochondrial autophagy, and alleviated keloid scars.
Asunto(s)
Autofagia , Exosomas , Queloide , Mitocondrias , Fosfatidilinositol 3-Quinasas , Proteínas Proto-Oncogénicas c-akt , Transducción de Señal , Serina-Treonina Quinasas TOR , Queloide/metabolismo , Queloide/terapia , Queloide/patología , Exosomas/metabolismo , Exosomas/trasplante , Animales , Serina-Treonina Quinasas TOR/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Humanos , Fosfatidilinositol 3-Quinasas/metabolismo , Ratones , Mitocondrias/metabolismo , Femenino , Ratones Endogámicos BALB C , Ratones Desnudos , Tejido Adiposo/metabolismo , Tejido Adiposo/citología , Células Madre/metabolismo , Células Madre/citología , Proliferación Celular , Fibroblastos/metabolismoRESUMEN
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.
Asunto(s)
Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Fibrosis , Queloide , Queratinocitos , Inhibidores de Fosfodiesterasa 4 , Humanos , Queloide/metabolismo , Queloide/patología , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/genética , Queratinocitos/metabolismo , Queratinocitos/efectos de los fármacos , Inhibidores de Fosfodiesterasa 4/farmacología , Animales , Ratones , Epidermis/metabolismo , Epidermis/patología , Factor de Crecimiento Transformador beta1/metabolismo , Transición Epitelial-Mesenquimal/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , MasculinoRESUMEN
Keloids are characterized histologically by excessive fibroblast proliferation and connective tissue deposition, and clinically by scar tissue extending beyond the original site of skin injury. These scars can cause pruritus, pain, physical disfigurement, anxiety, and depression. As a result, keloid patients often have a diminished quality of life with a disproportionate burden on ethnic minorities. Despite advances in understanding keloid pathology, there is no effective Food and Drug Administration (FDA)-approved pharmacotherapy. Recent studies have highlighted the possible pathologic role of T helper (Th)17 cells and interleukin (IL)-17 in keloid formation, as well as their implication in other inflammatory disorders. This systematic review characterizes the role of Th17 cells and IL-17 in keloid pathogenesis, highlighting this pathway as a potential therapeutic target. Adhering to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, we conducted a comprehensive search on PubMed, Embase, MEDLINE, and Web of Science databases on June 5, 2024. The search included terms related to Th17 cells, IL-17, and keloids. Thirteen studies met the inclusion criteria, comprising basic science and bioinformatic studies focusing on Th17 cells and IL-17. Key findings include increased Th17 cell infiltration and IL-17 expression in keloids, IL-17's role in amplifying the inflammatory and fibrotic response via the promotion of IL-6 expression, and IL-17's involvement in upregulating fibrotic markers via SDF-1 and HIF-1α pathways. IL-17 also activates the transforming growth factor beta (TGF-ß)/Smad pathway in keloid fibroblasts. Th17 cells and IL-17 significantly contribute to the inflammatory and fibrotic processes in keloid pathogenesis. Therefore, targeting the IL-17 pathway offers a potential new therapeutic target to improve keloid patients' outcomes. Future research could further elucidate the role of Th17 cells and IL-17 in keloid pathogenesis and assess the safety and efficacy of targeting this pathway in human studies.
Asunto(s)
Interleucina-17 , Queloide , Células Th17 , Humanos , Queloide/inmunología , Queloide/patología , Células Th17/inmunología , Interleucina-17/metabolismo , Interleucina-17/inmunología , Transducción de Señal/inmunología , Piel/patología , Piel/inmunologíaRESUMEN
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.
Asunto(s)
Queloide , Queloide/terapia , Queloide/patología , Queloide/etiología , Humanos , Animales , Transducción de Señal , Matriz Extracelular/metabolismo , Factor de Crecimiento Transformador beta/metabolismo , Cicatrización de HeridasRESUMEN
There is a pressing medical need for improved treatments in skin fibrosis including keloids and hypertrophic scars (HTS). This study aimed to characterize the role of phosphodiesterase 4 (PDE4), specifically PDE4B in fibrotic skin remodeling in vitro and in vivo. In vitro, effects of PDE4A-D (Roflumilast) or PDE4B (siRNA) inhibition on TGFß1-induced myofibroblast differentiation and dedifferentiation were studied in normal (NHDF) and keloid (KF) human dermal fibroblasts. In vivo, the role of PDE4 on HOCl-induced skin fibrosis in mice was addressed in preventive and therapeutic protocols. PDE4B (mRNA, protein) was increased in Keloid > HTS compared to healthy skin and in TGFß-stimulated NHDF and KF. In Keloid > HTS, collagen Iα1, αSMA, TGFß1 and NOX4 mRNA were all elevated compared to healthy skin confirming skin fibrosis. In vitro, inhibition of PDE4A-D and PDE4B similarly prevented TGFß1-induced Smad3 and ERK1/2 phosphorylation and myofibroblast differentiation, elevated NOX4 protein and proliferation in NHDF. PDE4A-D inhibition enabled myofibroblast dedifferentiation and curbed TGFß1-induced reactive oxygen species and fibroblast senescence. In KF PDE4A-D inhibition restrained TGFß1-induced Smad3 and ERK1/2 phosphorylation, myofibroblast differentiation and senescence. Mechanistically, PDE4A-D inhibition rescued from TGFß1-induced loss in PPM1A, a Smad3 phosphatase. In vivo, PDE4 inhibition mitigated HOCl-induced skin fibrosis in mice in preventive and therapeutic protocols. The current study provides novel evidence evolving rationale for PDE4 inhibitors in skin fibrosis (including keloids and HTS) and delivered evidence for a functional role of PDE4B in this fibrotic condition.
Asunto(s)
Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Fibroblastos , Fibrosis , Queloide , Inhibidores de Fosfodiesterasa 4 , Piel , Factor de Crecimiento Transformador beta1 , Queloide/patología , Queloide/metabolismo , Humanos , Animales , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Fibroblastos/metabolismo , Fibroblastos/efectos de los fármacos , Fibroblastos/patología , Ratones , Inhibidores de Fosfodiesterasa 4/farmacología , Factor de Crecimiento Transformador beta1/metabolismo , Piel/patología , Piel/metabolismo , Piel/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Masculino , Células Cultivadas , NADPH Oxidasa 4/metabolismo , NADPH Oxidasa 4/antagonistas & inhibidores , NADPH Oxidasa 4/genética , Ácido Hipocloroso/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Proteína smad3/metabolismo , Proliferación Celular/efectos de los fármacos , FemeninoRESUMEN
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.
Asunto(s)
Fibroblastos , Queloide , Neoplasias Cutáneas , Humanos , Queloide/patología , Queloide/metabolismo , Queloide/genética , Fibroblastos/metabolismo , Fibroblastos/patología , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/metabolismo , Neoplasias Cutáneas/genética , Análisis de la Célula Individual/métodos , Piel/patología , Piel/metabolismo , Moléculas de Adhesión Celular/metabolismo , Moléculas de Adhesión Celular/genética , Transición Epitelial-Mesenquimal/genética , Colágeno/metabolismo , MasculinoRESUMEN
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.
Asunto(s)
Queloide , RNA-Seq , Queloide/genética , Queloide/diagnóstico , Queloide/patología , Queloide/inmunología , Queloide/tratamiento farmacológico , Humanos , Transcriptoma/genética , Perfilación de la Expresión Génica , Fibroblastos/metabolismo , Fibroblastos/patología , Fibroblastos/inmunología , Redes Reguladoras de Genes , Tretinoina/farmacología , Tretinoina/uso terapéutico , Análisis de la Célula Individual/métodos , Diferenciación Celular/genética , Análisis de Secuencia de ARN/métodos , Aprendizaje Automático , Análisis de Expresión Génica de una Sola CélulaRESUMEN
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.
Asunto(s)
Proliferación Celular , Cicatriz Hipertrófica , Fibroblastos , Fluorouracilo , Células Endoteliales de la Vena Umbilical Humana , Queloide , Liposomas , Fluorouracilo/administración & dosificación , Fluorouracilo/farmacología , Fluorouracilo/química , Conejos , Animales , Liposomas/química , Humanos , Cicatriz Hipertrófica/tratamiento farmacológico , Fibroblastos/efectos de los fármacos , Queloide/tratamiento farmacológico , Queloide/patología , Proliferación Celular/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Colágeno/química , Movimiento Celular/efectos de los fármacos , Administración CutáneaRESUMEN
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.
Asunto(s)
Proliferación Celular , Colágeno , Fibroblastos , Queloide , Sistema de Señalización de MAP Quinasas , Sirtuinas , Humanos , Sirtuinas/metabolismo , Sirtuinas/genética , Queloide/patología , Queloide/metabolismo , Fibroblastos/metabolismo , Fibroblastos/patología , Colágeno/biosíntesis , Colágeno/metabolismo , Apoptosis/genética , Movimiento Celular , Masculino , Femenino , Células Cultivadas , AdultoRESUMEN
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.
Asunto(s)
Adenosina Monofosfato , Alanina , Fibroblastos , Fibrosis , Transducción de Señal , Piel , Factor de Crecimiento Transformador beta1 , Animales , Transducción de Señal/efectos de los fármacos , Factor de Crecimiento Transformador beta1/metabolismo , Fibrosis/tratamiento farmacológico , Alanina/análogos & derivados , Alanina/farmacología , Alanina/uso terapéutico , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Adenosina Monofosfato/análogos & derivados , Adenosina Monofosfato/farmacología , Adenosina Monofosfato/metabolismo , Ratones , Piel/efectos de los fármacos , Piel/patología , Piel/metabolismo , Humanos , Autofagia/efectos de los fármacos , Queloide/tratamiento farmacológico , Queloide/metabolismo , Queloide/patología , Antivirales/farmacología , Serina-Treonina Quinasas TOR/metabolismo , Bleomicina , Fosfatidilinositol 3-Quinasas/metabolismo , Masculino , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Smad/metabolismoRESUMEN
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.
Asunto(s)
Proliferación Celular , Fibroblastos , Foliculitis , Glucólisis , Queloide , Humanos , Queloide/metabolismo , Queloide/patología , Fibroblastos/metabolismo , Fibroblastos/patología , Foliculitis/metabolismo , Foliculitis/patología , Mitocondrias/metabolismo , Mitocondrias/patología , Células Cultivadas , Fosforilación Oxidativa , Movimiento Celular , Adulto , Piel/patología , Piel/metabolismo , Metabolismo Energético , Femenino , MasculinoRESUMEN
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.
Asunto(s)
Queloide , Triamcinolona Acetonida , Queloide/tratamiento farmacológico , Queloide/patología , Humanos , Triamcinolona Acetonida/farmacocinética , Triamcinolona Acetonida/administración & dosificación , Adulto , Femenino , Distribución Tisular , Masculino , Persona de Mediana Edad , Inyecciones Intralesiones , Piel/metabolismo , Piel/patología , Piel/diagnóstico por imagen , Crioultramicrotomía/métodos , Adulto Joven , Imagenología Tridimensional , Sistemas de Liberación de Medicamentos/métodosRESUMEN
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%.
Asunto(s)
Colágeno , Neoplasias Colorrectales , Matriz Extracelular , Queloide , Humanos , Neoplasias Colorrectales/patología , Neoplasias Colorrectales/metabolismo , Masculino , Femenino , Pronóstico , Persona de Mediana Edad , Colágeno/metabolismo , Anciano , Matriz Extracelular/patología , Matriz Extracelular/metabolismo , Queloide/patología , Queloide/metabolismo , Adulto , Anciano de 80 o más Años , Tasa de Supervivencia , Estudios de SeguimientoRESUMEN
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.
Asunto(s)
Proliferación Celular , Quimiocina CXCL12 , Fibroblastos , Queloide , MicroARNs , ARN Largo no Codificante , Factor de Transcripción STAT3 , Queloide/metabolismo , Queloide/genética , Queloide/patología , Humanos , MicroARNs/metabolismo , MicroARNs/genética , Factor de Transcripción STAT3/metabolismo , Factor de Transcripción STAT3/genética , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , Quimiocina CXCL12/metabolismo , Quimiocina CXCL12/genética , Fibroblastos/metabolismo , Movimiento Celular , Retroalimentación Fisiológica , Cromograninas/genética , Cromograninas/metabolismo , Masculino , Femenino , Subunidades alfa de la Proteína de Unión al GTP Gs/genética , Subunidades alfa de la Proteína de Unión al GTP Gs/metabolismo , Transducción de Señal , Adulto , Células Cultivadas , Regulación hacia ArribaRESUMEN
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.
Asunto(s)
Ciclohexilaminas , Ferroptosis , Fibroblastos , Fibrosis , Queloide , Factor 2 Relacionado con NF-E2 , Fenilendiaminas , Fosfolípido Hidroperóxido Glutatión Peroxidasa , Humanos , Ferroptosis/efectos de los fármacos , Queloide/patología , Queloide/metabolismo , Queloide/tratamiento farmacológico , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Fibroblastos/patología , Ciclohexilaminas/farmacología , Fibrosis/metabolismo , Fibrosis/patología , Fenilendiaminas/farmacología , Factor 2 Relacionado con NF-E2/metabolismo , Factor 2 Relacionado con NF-E2/genética , Fosfolípido Hidroperóxido Glutatión Peroxidasa/metabolismo , Fosfolípido Hidroperóxido Glutatión Peroxidasa/genética , Masculino , Peroxidación de Lípido/efectos de los fármacos , Femenino , Adulto , Hierro/metabolismo , Sistema de Transporte de Aminoácidos y+/metabolismo , Sistema de Transporte de Aminoácidos y+/genética , Receptores de Transferrina/metabolismo , Receptores de Transferrina/genética , Piperazinas/farmacología , Actinas/metabolismo , Actinas/genética , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Mitocondrias/patología , Potencial de la Membrana Mitocondrial/efectos de los fármacosRESUMEN
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.
Asunto(s)
Fibroblastos , Queloide , Piruvatos , Esferoides Celulares , Humanos , Queloide/metabolismo , Queloide/patología , Fibroblastos/metabolismo , Fibroblastos/efectos de los fármacos , Piruvatos/farmacología , Esferoides Celulares/efectos de los fármacos , Esferoides Celulares/metabolismo , Metaloproteinasa 1 de la Matriz/metabolismo , Metaloproteinasa 1 de la Matriz/genética , Factor de Crecimiento Transformador beta1/metabolismo , Proteína HMGB1/metabolismo , Proteína HMGB1/genética , Colágeno/metabolismo , Colágeno/biosíntesis , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Metaloproteinasa 3 de la Matriz/metabolismo , Metaloproteinasa 3 de la Matriz/genética , Matriz Extracelular/metabolismo , Matriz Extracelular/efectos de los fármacos , Colágeno Tipo I/metabolismo , Colágeno Tipo I/genética , Proteína Smad2/metabolismo , Proteína Smad2/genética , Proteína smad3/metabolismo , Regulación hacia Arriba/efectos de los fármacos , MasculinoRESUMEN
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.
Asunto(s)
Movimiento Celular , Proliferación Celular , Matriz Extracelular , Fibroblastos , Queloide , Sistema de Señalización de MAP Quinasas , Proteínas Smad , Factor de Crecimiento Transformador beta1 , Humanos , Queloide/patología , Queloide/metabolismo , Queloide/genética , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Proliferación Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Factor de Crecimiento Transformador beta1/metabolismo , Matriz Extracelular/metabolismo , Matriz Extracelular/efectos de los fármacos , Proteínas Smad/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/fisiología , Transducción de Señal , Células Cultivadas , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Masculino , Femenino , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , AdultoRESUMEN
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.
Asunto(s)
Proteína Morfogenética Ósea 1 , Fibroblastos , Inflamación , Queloide , Queloide/metabolismo , Queloide/patología , Queloide/genética , Humanos , Fibroblastos/metabolismo , Fibroblastos/patología , Proteína Morfogenética Ósea 1/metabolismo , Proteína Morfogenética Ósea 1/genética , Animales , Ratones , Inflamación/metabolismo , Inflamación/patología , Inflamación/genética , Masculino , Ratones Desnudos , Proliferación Celular , Femenino , Movimiento Celular , Fibrosis , Adulto , Factor de Crecimiento Transformador beta/metabolismo , Sistema de Señalización de MAP QuinasasRESUMEN
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.