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1.
Plast Reconstr Surg ; 144(2): 372-384, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31348346

RESUMO

BACKGROUND: We investigated expression of prorenin receptor, angiotensin-converting enzyme, angiotensin II receptor 1, and angiotensin II receptor 2 by the embryonic stem cell-like population on the endothelium of the microvessels and perivascular cells within keloid-associated lymphoid tissues. METHODS: Immunohistochemical staining for prorenin receptor, angiotensin-converting enzyme, angiotensin II receptor 1, and angiotensin II receptor 2 was performed on 11 formalin-fixed, paraffin-embedded sections of keloid tissue samples. Immunofluorescence staining was performed on three keloid tissue samples by co-staining with OCT4, CD34, ERG, and tryptase. Real-time quantitative polymerase chain reaction was performed on five keloid tissue samples and four keloid-derived primary cell lines. Western blotting was performed on the four keloid-derived primary cell lines for mRNA and protein expression of these proteins, respectively. RESULTS: Immunohistochemical and immunofluorescence staining showed expression of prorenin receptor, angiotensin-converting enzyme, angiotensin II receptor 1, and angiotensin II receptor 2 in all 11 keloid tissue samples. Prorenin receptor and angiotensin II receptor 1 were expressed on the endothelium and the pericyte layer of the microvessels and perivascular cells, angiotensin II receptor 2 was localized to the endothelium of the microvessels and the tryptase-positive perivascular cells, and angiotensin-converting enzyme was localized to the endothelium of the microvessel, within the keloid-associated lymphoid tissues. Real-time quantitative polymerase chain reaction showed transcripts of prorenin receptor, angiotensin-converting enzyme, and angiotensin II receptor 1 in the keloid tissue samples and keloid-derived primary cell lines, whereas angiotensin II receptor 2 was detected in keloid tissue samples only. Western blotting confirmed the presence of prorenin receptor, angiotensin-converting enzyme, and angiotensin II receptor 1 in the keloid-derived primary cell lines. CONCLUSION: Prorenin receptor, angiotensin-converting enzyme, angiotensin II receptor 1, and angiotensin II receptor 2 were expressed by the embryonic stem cell-like population within the keloid-associated lymphoid tissues, suggesting that this primitive population may be a potential therapeutic target by modulation of the renin-angiotensin system.


Assuntos
Células-Tronco Embrionárias/metabolismo , Queloide/metabolismo , Sistema Renina-Angiotensina/fisiologia , Adolescente , Adulto , Antígenos CD34/metabolismo , Linhagem Celular , Criança , Pré-Escolar , Feminino , Imunofluorescência , Humanos , Queloide/patologia , Masculino , Pessoa de Meia-Idade , Fator 3 de Transcrição de Octâmero/metabolismo , Peptidil Dipeptidase A/metabolismo , RNA Mensageiro/metabolismo , Receptor Tipo 1 de Angiotensina/metabolismo , Receptor Tipo 2 de Angiotensina/metabolismo , Receptores de Superfície Celular/metabolismo , Regulador Transcricional ERG/metabolismo , Adulto Jovem
2.
Life Sci ; 232: 116637, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31288014

RESUMO

Keloid is characterized by overactive fibroblasts. Forkhead box M1 (FOXM1) is transcription factor that plays important roles in the progression of fibrosis. However, the role of FOXM1 in keloid has not been elucidated. In the present study, we examined the expression levels of FOXM1 in clinical keloid tissue specimens and primary keloid fibroblasts (KFs). The results showed that FOXM1 levels were significantly increased in both keloid tissues and KFs. To further investigate the biological functions of FOXM1, FOXM1 was knocked down in KFs by transfection with small interfering RNA targeting FOXM1 (si-FOXM1). Knockdown of FOXM1 inhibited transforming growth factor-ß1 (TGF-ß1)-induced cell proliferation and migration of KFs. Besides, the increased expressions of collagen (coll I), connective tissue growth factor (CTGF), and α-smooth muscle actin (α-SMA) in TGF-ß1-induced KFs were suppressed by si-FOXM1 transfection. Furthermore, TGF-ß1-induced increase in p-Smad2 and p-Smad3 expressions was attenuated by FOXM1 knockdown. These data indicated that knockdown of FOXM1 inhibited TGF-ß1-induced KFs activation and extracellular matrix (ECM) accumulation, which was attributed to the inhibition of TGF-ß1/Smad pathway.


Assuntos
Proteína Forkhead Box M1/deficiência , Queloide/metabolismo , Actinas/metabolismo , Adolescente , Adulto , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Células Cultivadas , Colágeno/metabolismo , Colágeno Tipo I/metabolismo , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Feminino , Fibroblastos/metabolismo , Proteína Forkhead Box M1/genética , Proteína Forkhead Box M1/metabolismo , Técnicas de Silenciamento de Genes/métodos , Humanos , Queloide/genética , Masculino , Fosforilação , RNA Interferente Pequeno/genética , Transdução de Sinais , Proteína Smad2/antagonistas & inibidores , Proteína Smad2/metabolismo , Proteína Smad3/antagonistas & inibidores , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Fator de Crescimento Transformador beta1/metabolismo
3.
Plast Reconstr Surg ; 144(1): 58e-67e, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31246819

RESUMO

BACKGROUND: Fibroproliferative disorders result in excessive scar formation, are associated with high morbidity, and cost billions of dollars every year. Of these, keloid disease presents a particularly challenging clinical problem because the cutaneous scars progress beyond the original site of injury. Altered mechanotransduction has been implicated in keloid development, but the mechanisms governing scar progression into the surrounding tissue remain unknown. The role of mechanotransduction in keloids is further complicated by the differential mechanical properties of keloids and the surrounding skin. METHODS: The authors used human mechanical testing, finite element modeling, and immunohistologic analyses of human specimens to clarify the complex interplay of mechanical stress, strain, and stiffness in keloid scar progression. RESULTS: Changes in human position (i.e., standing, sitting, and supine) are correlated to dynamic changes in local stress/strain distribution, particularly in regions with a predilection for keloids. Keloids are composed of stiff tissue, which displays a fibrotic phenotype with relatively low proliferation. In contrast, the soft skin surrounding keloids is exposed to high mechanical strain that correlates with increased expression of the caveolin-1/rho signaling via rho kinase mechanotransduction pathway and elevated inflammation and proliferation, which may lead to keloid progression. CONCLUSIONS: The authors conclude that changes in human position are strongly correlated with mechanical loading of the predilection sites, which leads to increased mechanical strain in the peripheral tissue surrounding keloids. Furthermore, increased mechanical strain in the peripheral tissue, which is the site of keloid progression, was correlated with aberrant expression of caveolin-1/ROCK signaling pathway. These findings suggest a novel mechanism for keloid progression.


Assuntos
Caveolina 1/fisiologia , Queloide , Mecanotransdução Celular/fisiologia , Transdução de Sinais/fisiologia , Estresse Mecânico , Quinases Associadas a rho/fisiologia , Adulto , Idoso , Estudos de Casos e Controles , Feminino , Análise de Elementos Finitos , Humanos , Queloide/metabolismo , Queloide/fisiopatologia , Masculino , Pessoa de Meia-Idade , Postura/fisiologia , Adulto Jovem
4.
Int J Mol Sci ; 20(10)2019 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-31137604

RESUMO

Previous studies described the involvement of extracellular signal-related kinase (ERK) in systemic fibrotic diseases, but the role of ERK in cutaneous scarring is unknown. Although hypoxia drives tissue fibrosis by activating hypoxia-inducible factor-1α (HIF-1α), the specific roles of hypoxia and associated ERK phosphorylation in abnormal fibroblast activity during cutaneous scarring are unclear. Here, we investigated whether pathologic myofibroblast-like keloid fibroblast activity is promoted by hypoxia-induced epithelial-mesenchymal transition mediated by ERK activation. ERK phosphorylation was significantly increased in keloid tissue and fibroblasts. Human dermal fibroblasts cultured under hypoxia (1% O2) expressed phosphorylated ERK and exhibited activation of p38 mitogen-activated protein kinase signaling. Hypoxic human dermal fibroblasts showed increased protein and mRNA levels of epithelial-mesenchymal transition markers. Furthermore, administration of an ERK inhibitor (SCH772984) reduced the hypoxia-induced elevation of collagen type I levels in human dermal fibroblasts. Therefore, ERK may be a promising therapeutic target in profibrogenic diseases.


Assuntos
Transição Epitelial-Mesenquimal , Fibroblastos/metabolismo , Queloide/metabolismo , Sistema de Sinalização das MAP Quinases , Oxigênio/metabolismo , Cicatrização , Hipóxia Celular , Células Cultivadas , Fibroblastos/patologia , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Queloide/patologia , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
5.
Mol Med Rep ; 19(6): 5251-5262, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31059100

RESUMO

Keloids are benign fibrous overgrowths that occur as a result of abnormal wound healing following cutaneous injury. MicroRNAs (miRNAs/miRs) are short non­coding RNAs that serve critical roles in numerous important biological processes, such as cell proliferation, differentiation and apoptosis. However, their role in keloid development remains largely unknown. In the present study, the role of miR­30a­5p, a miRNA regulated by Trichostatin A (TSA), in apoptosis within cultured keloid fibroblasts was investigated. An MTT assay was used to detect the proliferation of cultured keloid fibroblasts treated with TSA. Cell apoptosis and cell cycle phases were analyzed using flow cytometry. In addition, an miRNA microarray was performed to compare expression profiles between cultured keloid fibroblasts treated with or without 1,000 nM TSA. Reverse transcription­quantitative polymerase chain reaction analysis was conducted to estimate miRNA expression levels. The direct target of miR­30a­5p was identified using a dual­luciferase reporter assay. Western blotting was employed to assess protein expression levels in keloid fibroblasts. The results demonstrated that TSA inhibited the proliferation of keloid fibroblasts in a time­ and dose­dependent manner. The miRNA microarray revealed alterations in the expression of numerous miRNA sequences in response to TSA when compared with controls. Notably, the expression of miR­30a­5p was downregulated in keloid tissues. In addition, overexpression of miR­30a­5p induced apoptosis by targeting B­cell lymphoma 2, which was similar to that observed in response to TSA. These results provide important information regarding a novel miR­30a­5p­mediated signaling pathway induced by TSA treatment, and suggest a potential use for TSA and miR­30a­5p as effective therapeutic strategies for keloids.


Assuntos
Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/farmacologia , MicroRNAs/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Regiões 3' não Traduzidas , Antagomirs/metabolismo , Análise por Conglomerados , Colágeno/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Queloide/metabolismo , Queloide/patologia , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/genética , Pele/metabolismo , Pele/patologia
6.
Eur J Pharmacol ; 854: 282-288, 2019 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-31034822

RESUMO

Keloids are characterized by excessive proliferation of fibroblasts and invasion of surrounding healthy skin. High levels of Nitric Oxide (NO) are thought to be the crucial factor within the micro-environment in promoting keloid formation. However, the effects and mechanisms of NO on the proliferation of Keloid Fibroblasts (KDFs) remain unclear. In this study, we investigated the effect of NO on KDFs proliferation by Sodium Nitroprusside (SNP), an NO donor. Our results show that SNP significantly enhanced KDFs proliferation. Moreover, with prolonged treatment with SNP after cell confluence, the growth of KDFs escape contact inhibition and experience significant pile up growth. Furthermore, PTIO, an NO scavenger, attenuated SNP-enhanced cell proliferation effectively. The mechanism involved in SNP-induced KDFs proliferation was soluble Guanylyl Cyclase (sGC) and cGMP independent. ODQ, a specific sGC inhibitor, failed to suppress SNP-enhanced KDFs proliferation. 8-Bromo-c GMP, a cell-permeable cGMP analogue, could not stimulate KDFs proliferation. Erk and Akt provide important signaling for cell growth. U0126 and LY294002, inhibitors of Erk and Akt respectively, block SNP-enhanced KDFs proliferation effectively. As expected, a Western blot showed that SNP up-regulated the phosphorylation levels of Erk and Akt. Moreover, it decreased the expression of p27, a cell cycle inhibitor. Our results reveal that SNP induced KDFs proliferation and loss contact inhibition led to pile up growth via activation of the Erk and Akt pathways, as well as a decreased expression of p27. Thus, we speculate that the pathological feature of continuous expansion in keloids is caused by NO-induced KDFs sustained growth.


Assuntos
Queloide/metabolismo , Queloide/patologia , Óxido Nítrico/metabolismo , Proliferação de Células/efeitos dos fármacos , GMP Cíclico/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Humanos , Doadores de Óxido Nítrico/farmacologia , Nitroprussiato/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos
7.
Biomed Res Int ; 2019: 8214923, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30956986

RESUMO

The aim of this study was to detect abnormally expressed microRNA (miRNA) in keloids and to study their functions. The differential expression of miRNAs in keloids and normal tissue was detected by gene microarray. MiRNA expression was verified by real-time PCR. A luciferase reporter gene assay, western blot, and real-time PCR were used to detect the effect of miR-194-3p on RUNX2. An MTT assay and a transwell assay were used to detect the effect of miR-194-3p in both primary cultured fibroblasts and HKF cells. Related proteins were analysed by western blot and real-time PCR. The expression of miR-194-3p was lower in keloids, and MiR-194-3p was shown to target RUNX2 directly. MiR-194-3p inhibited the proliferation and migration of fibroblasts through the inhibition of CDK4 and MMP2. MiR-194-3p and RUNX2 may become new targets for the prevention and treatment of keloids.


Assuntos
Movimento Celular , Proliferação de Células , Subunidade alfa 1 de Fator de Ligação ao Core/biossíntese , Fibroblastos/metabolismo , Regulação da Expressão Gênica , Queloide/metabolismo , MicroRNAs/biossíntese , Linhagem Celular , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Feminino , Fibroblastos/patologia , Humanos , Queloide/genética , Queloide/patologia , Masculino , MicroRNAs/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa
8.
Dermatol Surg ; 45(4): 536-546, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30829753

RESUMO

BACKGROUND: Ablative fractional laser-assisted therapy is increasingly used to facilitate drug delivery and intensify clinical efficacy of topically applied drugs. OBJECTIVE: To evaluate the effectiveness of combined ablative fractional CO2 laser and topically applied 5-fluorouracil (5-FU) or verapamil hydrochloride in the treatment of hypertrophic scars (HTSs) and keloids and to examine their possible effects on TGF-ß1 expression. PATIENTS AND METHODS: Thirty patients with HTSs and keloids were randomly treated with combined CO2 laser followed by topical verapamil or 5-FU application or CO2 laser monotherapy. All patients received 4 treatments at 1-month intervals. Subjective and objective assessment was obtained using the Vancouver Scar Scale (VSS). Histological changes and immunohistochemical staining for TGF-ß1 were performed. RESULTS: Compared with baseline, there was a significant reduction in the VSS 1 month after the last treatment session in all groups (p < .05). Laser-assisted 5-FU delivery tended to show a higher extent of improvement in scar characteristics than laser-assisted verapamil hydrochloride delivery, without significance. No significant side effects were reported in all patient groups. TGF-ß1 expression was significantly decreased after laser sessions. CONCLUSION: Combined fractional CO2 laser and topical 5-FU or verapamil hydrochloride offer a safe therapy for HTSs and keloids.


Assuntos
Cicatriz Hipertrófica/terapia , Fluoruracila/administração & dosagem , Queloide/terapia , Terapia a Laser , Lasers de Gás/uso terapêutico , Verapamil/administração & dosagem , Administração Tópica , Cicatriz Hipertrófica/metabolismo , Terapia Combinada , Fracionamento da Dose de Radiação , Humanos , Imuno-Histoquímica , Queloide/metabolismo , Fator de Crescimento Transformador beta1/biossíntese , Resultado do Tratamento
9.
Am J Dermatopathol ; 41(3): 193-204, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30801341

RESUMO

Keloids are defined histopathologically as an inflammatory disorder characterized by exhibiting numerous fibroblasts, abnormal vascularization, increased number of proinflammatory immune cells as well as uncontrolled cell proliferation, and exacerbated and disorganized deposition of extracellular matrix (ECM) molecules. Importantly, many of these ECM molecules display N- and O-linked glycan residues and are considered as potential targets for galectin-1 (Gal-1) and galectin-3 (Gal-3). Nevertheless, the presence and localization of Gal-1 and Gal-3 as well as the interactions with some of their binding partners in keloid tissues have not been considered. Here, we show that in the dermal thickening of keloids, versican, syndecan-1, fibronectin, thrombospondin-1, tenascin C, CD44, integrin ß1, and N-cadherin were immunolocalized in the elongated fibroblasts that were close to the immune cell infiltrate, attached to collagen bundles, and around the microvasculature and in some immune cells. We also show that Gal-1 and Gal-3 were present in the cytoplasm and along the cell membrane of some fibroblasts and immune and endothelial cells of the dermal thickening. We suggest that Gal-1 and Gal-3, in concert with some of the ECM molecules produced by fibroblasts and by immune cells, counteract the inflammatory response in keloids. We also proposed that Gal-1 and Gal-3 through their binding partners may form a supramolecular structure at the cell surface of fibroblasts, immune cells, endothelial cells, and in the extracellular space that might influence the fibroblast morphology, adhesion, proliferation, migration, and survival as well as the inflammatory responses.


Assuntos
Derme/química , Fibroblastos/química , Galectina 1/análise , Galectina 3/análise , Queloide/metabolismo , Adolescente , Adulto , Biomarcadores/análise , Derme/patologia , Feminino , Fibroblastos/patologia , Humanos , Imuno-Histoquímica , Queloide/patologia , Masculino , Ligação Proteica , Adulto Jovem
10.
Analyst ; 144(6): 1866-1875, 2019 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-30734778

RESUMO

A keloid is a type of unusually raised scar. Unlike other raised scars, keloids form larger sizes than the wound site due to overgrowth, generally related to various biological factors. To date, only a few diagnostic and therapeutic methods for keloids have been reported. The high recurrence rates and undesirable side effects of keloids, at the end stage, encourage the invention of novel diagnostic tools, in order to cure keloids at an earlier stage. In this review, we summarize the general information about keloid diagnosis, keloid biomarkers, and recently reported fluorescent probes that can sense the key biomarkers of keloids. The focused description of fluorescent probes for keloid biomarkers and the author's perspective give useful insights in order to design the next-generation diagnostic sensing system for keloids.


Assuntos
Biomarcadores/metabolismo , Corantes Fluorescentes/metabolismo , Queloide/diagnóstico , Queloide/metabolismo , Humanos
11.
BMB Rep ; 52(3): 202-207, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30638178

RESUMO

Keloids are the most common pathological form of trauma healing, with features that seriously affect appearance and body function, are difficult to treat and have a high recurrence rate. Emerging evidence suggests that miRNAs are involved in a variety of pathological processes and play an important role in the process of fibrosis. In this study, we investigated the function and regulatory network of miR-152-5p in keloids. The miRNA miR-152-5p is frequently downregulated in keloid tissue and primary cells compared to normal skin tissue and fibroblasts. In addition, the downregulation of miR-152-5p is significantly associated with the proliferation, migration and apoptosis of keloid cells. Overexpression of miR-152-5p significantly inhibits the progression of fibrosis in keloids. Smad3 is a direct target of miR-152-5p, and knockdown of Smad3 also inhibits fibrosis progression, consistent with the overexpression of miR-152-5p. The interaction between miR-152-5p and Smad3 occurs through the Erk1/2 and Akt pathways and regulates collagen3 production. In summary, our study demonstrates that miR-152-5p/Smad3 regulatory pathways involved in fibrotic progression may be a potential therapeutic target of keloids. [BMB Reports 2019; 52(3): 202-207].


Assuntos
Queloide/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Proteína Smad3/biossíntese , Adolescente , Adulto , Apoptose/genética , Proliferação de Células/genética , Regulação para Baixo , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibroblastos/fisiologia , Humanos , Queloide/metabolismo , Queloide/patologia , Masculino , Transdução de Sinais/genética , Pele/metabolismo , Pele/patologia , Proteína Smad3/genética , Proteína Smad3/metabolismo
12.
Life Sci ; 219: 272-282, 2019 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-30597173

RESUMO

AIMS: Keloids are a dermal fibrotic disease whose etiology remains totally unknown and for which there is no successful treatment. Mechanical tension, in addition, is closely associated with the germination and development of keloids. In this study, we investigated the influence of human keloid-derived mesenchymal stem cells (KD-MSCs) on cell proliferation, collagen synthesis, and expressions of integrin αvß3 under tension. MAIN METHODS: KD-MSCs and human normal skin-derived mesenchymal stem cells (NS-MSCs) were isolated and cultured in stem cell medium with a gradual increase in the serum concentration. Cell proliferation and collagen synthesis were detected by Cell Counting Kit-8 (CCK-8) assay and hydroxyproline content analysis under tension respectively. We investigated the messenger RNA expressions of nine integrin subunits, including integrin units α2, α3, α5, αv, α8, α10, α11, ß1, and ß3, in KD-MSCs stimulated with tension. Identification of differentially expressed genes was performed by Western blot analysis and immunocytochemistry staining. KEY FINDINGS: We obtained high-purity KD-MSCs and NS-MSCs using the culture method of decreasing serum concentration gradient gradually. Furthermore, we found that tension enhances cell proliferation and collagen synthesis and promotes expressions of integrin αvß3 in KD-MSCs. In addition, blocking experiments showed that increased integrin αvß3 expression affects cell proliferation and collagen synthesis of KD-MSCs under tension. SIGNIFICANCE: Our results suggest that integrin αvß3 receptor may be sensitive molecules of mechanical tension and could contribute to the occurrence and development of keloids. It could lead to novel targets for therapeutic intervention, treatment, and prevention of recurrence for keloid disorders.


Assuntos
Colágeno/metabolismo , Integrina alfaVbeta3/metabolismo , Queloide/metabolismo , Células-Tronco Mesenquimais/metabolismo , Adolescente , Adulto , Western Blotting , Proliferação de Células , Feminino , Citometria de Fluxo , Regulação da Expressão Gênica , Humanos , Hidroxiprolina/metabolismo , Masculino , Reação em Cadeia da Polimerase em Tempo Real , Pele/citologia , Pele/metabolismo , Estresse Mecânico , Regulação para Cima
13.
Acta Biochim Biophys Sin (Shanghai) ; 51(2): 185-196, 2019 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-30668826

RESUMO

Keloids (KDs) and hypertrophic scars (HSs), two forms of pathological scars, seriously affect the physical and psychological health of patients. Despite many similarities with HSs, KDs are characterized by invasion and a high rate of recurrence after surgery, features they share in common with tumors. The underlying molecular mechanisms of this phenomenon have not been fully elucidated. In this study, we used microRNA (miRNA) array analysis to search for invasion-associated miRNAs in KDs. The expression of miR-188-5p in KDs, HSs, normal skin (NS) tissues, and cell lines was measured by quantitative real-time polymerase chain reaction. Furthermore, cell proliferation, migration, and invasion were detected in KD fibroblasts (KFs) and HS fibroblasts (HSFs), and interrelated proteins were ascertained by western blot analysis. It was found that miR-188-5p was significantly decreased in KD tissue compared with HS and NS tissues. Upregulated expression of miR-188-5p suppressed KF proliferation, migration, and invasion; and decreased expression of miR-188-5p also promoted HSF proliferation, migration, and invasion. The protein levels of MMP-2, MMP-9, PI3K, and p-Akt in miR-188-5p mimic-transfected KFs were repressed. In contrast, after transfection with miR-188-5p inhibitor, the protein levels of MMP-2, MMP-9, PI3K, and p-Akt were higher than the control in HSFs. Treatment with PI3K/Akt inhibitor LY294002 in KFs with miR-188-5p inhibitor did not further reduce their proliferation, migration, and invasion. The upregulation of MMP-2 and MMP-9 by miR-188-5p inhibitor could be abolished by LY294002. These findings together demonstrate a tumor-suppressive role of miR-188-5p in KD proliferation and invasion via PI3K/Akt/MMP-2/9 signaling, indicating that miR-188-5p may be a potential prognostic marker and therapeutic target for KDs.


Assuntos
Movimento Celular/genética , Proliferação de Células/genética , Perfilação da Expressão Gênica , Queloide/genética , MicroRNAs/genética , Transdução de Sinais/genética , Adolescente , Adulto , Células Cultivadas , Feminino , Humanos , Queloide/metabolismo , Queloide/patologia , Masculino , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Adulto Jovem
14.
Mol Med Rep ; 19(2): 919-926, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30569148

RESUMO

An increasing number of studies have demonstrated that tumor necrosis factor­stimulated gene­6 (TSG­6) has a key role in the progression of fibrosis; however, the exact effects of TSG­6 in keloid fibroblasts (KFs) remain unknown. The aim of the current study was to investigate the role of TSG­6 in the pathogenesis of keloids. Primary fibroblasts from 10 patients with keloid were cultured and transfected with pLVX­Puro or pLVX­Puro­TSG­6. Alterations of TSG­6 expression were then determined by reverse transcription­polymerase chain reaction (RT­PCR) and regulation was observed in KFs transfected with pLVX­Puro­TSG­6. Compared with the control group, transfection with pLVX­Puro­TSG­6 induced growth suppression, cell apoptosis and G2/M arrest in KFs. In addition, the mitochondrial apoptosis pathway was activated in KFs transfected with pLVX­Puro­TSG­6. These findings indicate that TSG­6 is a novel regulator of keloid fibrogenesis, and thus could be used/targeted TSG­6 as a promising treatment for keloid.


Assuntos
Moléculas de Adesão Celular/metabolismo , Queloide/metabolismo , Queloide/patologia , Adolescente , Adulto , Apoptose/fisiologia , Moléculas de Adesão Celular/genética , Pontos de Checagem do Ciclo Celular/fisiologia , Proliferação de Células/fisiologia , Criança , Pré-Escolar , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibrose/metabolismo , Fibrose/patologia , Pontos de Checagem da Fase G2 do Ciclo Celular/fisiologia , Humanos , Queloide/genética , Masculino , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Transdução de Sinais/fisiologia , Transfecção/métodos , Adulto Jovem
15.
Medicine (Baltimore) ; 97(37): e12167, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30212944

RESUMO

To study the effect of knocking down wingless-related MMTV integration site 2 (Wnt2) expression by RNAi on the growth and signaling pathways of ex vitro-cultured keloid fibroblasts (KFB).Human KFB were isolated from 10 keloid patient specimens. The KFB cells were then transfected with 4 pairs of small interfering RNA (siRNA) targeting human Wnt2, respectively. Reverse transcriptase-polymerase chain reaction and Western blot analysis were conducted to verify the knock down of Wnt2, and the expression of ß-catenin glycogen synthase kinase-3ß (GSK-3ß) and cyclin D1 were examined.siRNA Wnt2 transfection (siWnt2) resulted in the significant inhibition of Wnt2 expression at both the mRNA and protein levels. The expression of ß-catenin, GSK-3ß, p-GSK-3ß, and cyclin D1 at the protein level also decreased in siWnt2 cells. siWnt2 resulted in a substantially slower growth and significant delay in cell doubling time of the KFB cells compared with control groups. Further, the siRNA knock down of GSK-3ß and ß-catenin resulted in slower proliferation rates, respectively.Wnt2 siRNA has an inhibitive effect on keloid fibroblast proliferation, which may be a potential therapeutic approach for keloid and other human fibrotic diseases.


Assuntos
Fibroblastos/metabolismo , Queloide/metabolismo , Interferência de RNA/fisiologia , Transdução de Sinais/fisiologia , Proteína Wnt2/biossíntese , Adolescente , Adulto , Western Blotting , Criança , Pré-Escolar , Ciclina D1/biossíntese , Feminino , Quinases da Glicogênio Sintase/biossíntese , Humanos , Masculino , Pessoa de Meia-Idade , RNA Interferente Pequeno , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Adulto Jovem
16.
Medicine (Baltimore) ; 97(29): e11529, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30024539

RESUMO

BACKGROUND: Hyperbaric oxygen therapy (HBOT) has been widely used in the clinical setting. In this study, HBOT therapy was evaluated for its ability to ameliorate the epithelial-to-mesenchymal transition (EMT) phenomenon in keloid tissue. METHODS: Keloid patients were randomly divided into two groups: keloid patients (K group, 9 patients) and keloid patients receiving HBOT (O group, 9 patients). A third group with normal skin (S group, 9 patients) was established for control. Before HBOT and surgery, a laser Doppler flowmeter was used to measure the keloid blood supply of patients in the O group. Hematoxylin and eosin (H&E) staining was used to observe morphology. E-cadherin, ZO-1, vimentin, fibronectin, vascular endothelial growth factor (VEGF), and hypoxia inducible factor (HIF)-1α were measured by immunofluorescence staining and Western blot analysis. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to evaluate the mRNA expression level of these factors as well. RESULTS: In the O group, keloid blood perfusion was significantly reduced after patients received HBOT. Compared with the K group, lower expression levels of vimentin, vibronectin, VEGF, and HIF-1α were observed in the O group, whereas the expression of E-cadherin and ZO-1 was significantly higher. The mRNA expression of E-cadherin and ZO-1 was also increased after HBOT. CONCLUSIONS: The expression levels of factors related to the EMT phenomenon were significantly reversed in keloid patients after they received HBOT, indicating that HBOT may be an effective therapy against the EMT phenomenon in keloid patients.


Assuntos
Transição Epitelial-Mesenquimal/fisiologia , Oxigenação Hiperbárica/métodos , Queloide/terapia , Adolescente , Adulto , Western Blotting , Caderinas/metabolismo , Feminino , Fibronectinas/metabolismo , Imunofluorescência , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Queloide/metabolismo , Queloide/fisiopatologia , Fluxometria por Laser-Doppler , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Resultado do Tratamento , Fator A de Crescimento do Endotélio Vascular/metabolismo , Vimentina/metabolismo , Adulto Jovem , Proteína da Zônula de Oclusão-1/metabolismo
18.
Mol Med Rep ; 18(2): 1660-1665, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29901093

RESUMO

Keloid scarring is a type of fibroproliferative disease with a high recurrence rate. However, no effective treatment is currently available. Combined therapy with recombinant lentivirus­mediated Drosophila melanogaster deoxyribonucleoside kinase (Dm­dNK) and prodrug has been widely studied and used for cancer treatment. Due to the similarities between keloid scars and tumors, the aim of the present study was to investigate the efficacy of a Dm­dNK/nucleoside analog system for the treatment of keloid scars. Recombinant lentivirus expression of the Dm­dNK suicide gene was assessed. Western blotting was used to examine the protein expression of lentivirus mediated Dm­dNK in keloid fibroblasts. Enzyme activity assays were conducted using [3H]­labeled substrates. Furthermore, cytotoxicity and bystander effects were evaluated using MTT assays. The expression of green fluorescent protein was observed using fluorescence microscope and results indicated that there was no notable difference in lentivirus infectivity between the multiplicity of infection (MOI) of 1 and 10 in cells. Notably, western blotting revealed that Dm­dNK was stably expressed in keloid fibroblasts and the enzymatic activity assays revealed that the enzyme was activated following introduction into the keloid fibroblasts via the lentivirus. The cytotoxicity and bystander effects of Dm­dNK combined with cytotoxic nucleoside analogs were both observed in Dm­dNK+ keloid fibroblasts. These results demonstrated that the lentivirus­mediated Dm­dNK therapy may be effective in treating keloid fibroblasts, which provides some evidence for the use of Dm­dNK/prodrug therapy for keloid treatment in vivo in the future.


Assuntos
Arabinonucleosídeos/farmacologia , Bromodesoxiuridina/análogos & derivados , Fibroblastos/efeitos dos fármacos , Vetores Genéticos/química , Proteínas de Insetos/genética , Queloide/terapia , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Timidina/análogos & derivados , Adulto , Animais , Apoptose/efeitos dos fármacos , Bromodesoxiuridina/farmacologia , Efeito Espectador , Proliferação de Células/efeitos dos fármacos , Terapia Combinada/métodos , Drosophila melanogaster/química , Drosophila melanogaster/enzimologia , Ensaios Enzimáticos , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Genes Reporter , Vetores Genéticos/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Proteínas de Insetos/metabolismo , Queloide/genética , Queloide/metabolismo , Queloide/patologia , Lentivirus/genética , Lentivirus/metabolismo , Masculino , Pessoa de Meia-Idade , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Cultura Primária de Células , Timidina/farmacologia , Transgenes
19.
Mol Med Rep ; 18(2): 1628-1636, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29845237

RESUMO

Keloids are a type of abnormal scar tissue. MicroRNAs (miRNAs) exhibit a pivotal role in the regulation of cell proliferation and metastasis of keloids. miRNA microarray revealed that miR­637 was one of the most frequently altered miRNAs in keloids. Furthermore, up-regulation of miR­637 inhibited cell proliferation and metastasis by targeting mothers against decapentaplegic homolog (Smad)3, one of the important proteins that affects the formation of keloids. Further studies demonstrated that miR­637 regulated the proliferation and metastasis of human keloid fibroblast (HKF) cells by mediating the Smad3 signaling pathway. Overall, the present findings suggest that miR­637 may be a promising therapeutic target in keloids.


Assuntos
Fibroblastos/metabolismo , Queloide/genética , MicroRNAs/genética , Proteína Smad3/genética , Adulto , Sequência de Bases , Estudos de Casos e Controles , Movimento Celular , Proliferação de Células , Feminino , Fibroblastos/patologia , Regulação da Expressão Gênica , Humanos , Queloide/metabolismo , Queloide/patologia , Masculino , MicroRNAs/classificação , MicroRNAs/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Proteína Smad3/antagonistas & inibidores , Proteína Smad3/metabolismo
20.
Int J Mol Sci ; 19(5)2018 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-29695130

RESUMO

Keloids occur after failure of the wound healing process; inflammation persists, and various treatments are ineffective. Keloid pathogenesis is still unclear. We have previously analysed the gene expression profiles in keloid tissue and found that HtrA1 was markedly up-regulated in the keloid lesions. HtrA1 is a serine protease suggested to play a role in the pathogenesis of various diseases, including age-related macular degeneration and osteoarthritis, by modulating extracellular matrix or cell surface proteins. We analysed HtrA1 localization and its role in keloid pathogenesis. Thirty keloid patients and twelve unrelated patients were enrolled for in situ hybridization, immunohistochemical, western blot, and cell proliferation analyses. Fibroblast-like cells expressed more HtrA1 in active keloid lesions than in surrounding lesions. The proportion of HtrA1-positive cells in keloids was significantly higher than that in normal skin, and HtrA1 protein was up-regulated relative to normal skin. Silencing HtrA1 gene expression significantly suppressed cell proliferation. HtrA1 was highly expressed in keloid tissues, and the suppression of the HtrA1 gene inhibited the proliferation of keloid-derived fibroblasts. HtrA1 may promote keloid development by accelerating cell proliferation and remodelling keloid-specific extracellular matrix or cell surface molecules. HtrA1 is suggested to have an important role in keloid pathogenesis.


Assuntos
Regulação da Expressão Gênica , Serina Peptidase 1 de Requerimento de Alta Temperatura A/genética , Queloide/genética , Queloide/patologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores , Biópsia , Proliferação de Células , Células Cultivadas , Feminino , Fibroblastos/metabolismo , Técnicas de Silenciamento de Genes , Humanos , Imuno-Histoquímica , Queloide/metabolismo , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/genética , Pele/metabolismo , Pele/patologia , Regulação para Cima , Adulto Jovem
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