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Cutaneous squamous cell carcinoma (cSCC) ranks as the second most prevalent skin tumour (excluding melanoma). However, the molecular mechanisms driving cSCC progression remain elusive. This study aimed to investigate GBP1 expression in cSCC and elucidate its potential molecular mechanisms underlying cSCC development. GBP1 expression was assessed across public databases, cell lines and tissue samples. Various assays, including clone formation, CCK8 and EdU were employed to evaluate cell proliferation, while wound healing and transwell assays determined cell migration and invasion. Subcutaneous tumour assays were conducted to assess in vivo tumour proliferation, and molecular mechanisms were explored through western blotting, immunofluorescence and immunoprecipitation. Results identified GBP1 as an oncogene in cSCC, with elevated expression in both tumour tissues and cells, strongly correlating with tumour stage and grade. In vitro and in vivo investigations revealed that increased GBP1 expression significantly enhanced cSCC cell proliferation, migration and invasion. Mechanistically, GBP1 interaction with SP1 promoted STAT3 activation, contributing to malignant behaviours. In conclusion, the study highlights the crucial role of the GBP1/SP1/STAT3 signalling axis in regulating tumour progression in cSCC. These findings provide valuable insights into the molecular mechanisms of cSCC development and offer potential therapeutic targets for interventions against cSCC.
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Carcinoma de Células Escamosas , Movimento Celular , Proliferação de Células , Proteínas de Ligação ao GTP , Invasividade Neoplásica , Fator de Transcrição STAT3 , Neoplasias Cutâneas , Fator de Transcrição Sp1 , Fator de Transcrição STAT3/metabolismo , Humanos , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/genética , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/genética , Fator de Transcrição Sp1/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Proteínas de Ligação ao GTP/genética , Linhagem Celular Tumoral , Animais , Camundongos , Transdução de Sinais , Feminino , Camundongos NusRESUMO
Fibrosarcoma is a highly malignant type of soft tissue sarcoma that currently lacks effective treatment options. Polypeptide N-acetylgalactosaminyltransferase 12 (GALNT12) belongs to the uridine diphosphate N-acetylgalactosamine gene family, which is involved in numerous biological processes of diseases, such as tumor progression. Its upregulated expression is closely associated with the development of colorectal cancer. However, research on the role of GALNT12 in fibrosarcoma is currently limited. The present study aimed to assess the expression and biological function of GALNT12 in fibrosarcoma. Patient data and tissue samples were collected and public datasets were obtained from the Gene Expression Omnibus (GSE24369 and GSE21124). Immunofluorescence assays were performed to observe the cellular localization of GALNT12. GALNT12 expression was measured using reverse transcription-quantitative PCR, western blotting and immunohistochemistry. Small interfering RNAs were constructed to knock down GALNT12 expression in HT-1080 cells. Cell Counting Kit-8 and EdU assays were used to assess fibrosarcoma cell proliferation. Wound healing and Transwell assays were used to detect migration. Gene set enrichment analysis was performed to identify key pathways. Paired and unpaired Student's t-test, Fisher's exact test and one-way ANOVA (followed by Tukey's Honest Significant Difference test) were used to analyze the data. It was demonstrated that GALNT12 expression was upregulated in both fibrosarcoma cell lines and tissue samples and predicted poor patient prognosis. In vitro experiments demonstrated that high GALNT12 expression levels significantly increased HT-1080 cell proliferation and migration. Furthermore, it was demonstrated that high GALNT12 expression levels were closely associated with the yes1 associated transcriptional regulator (YAP1) signaling pathway. Knockdown of GALNT12 inhibited YAP1 nuclear translocation, which affected activation of key downstream genes including AMOTL2, BIRC5 and CYR61. Therefore, the present study demonstrated that GALNT12 promoted fibrosarcoma progression. GALNT12 could be a potential biomarker for this disease and may potentially provide new ideas for targeted therapy of fibrosarcoma in the future.
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Thermal injury repair is a complex process during which the maintenance of the proliferation and migration of human skin fibroblasts (HSFs) exert a crucial role. MicroRNAs have been proven to exert an essential function in repairing skin burns. This study delves into the regulatory effects of miR-24-3p on the migration and proliferation of HSFs that have sustained a thermal injury, thereby, providing deeper insight into thermal injury repair pathogenesis. The PPAR-ß protein expression level progressively increased in a time-dependent manner on the 12th, 24th and 48th hour following the thermal injury of the HSFs. The knockdown of PPAR-ß markedly suppressed the proliferation of and migration of HSF. Following thermal injury, the knockdown also promoted the inflammatory cytokine IL-6, TNF-α, PTGS-2 and P65 expression. PPAR-ß contrastingly exhibited an opposite trend. A targeted relationship between PPAR-ß and miR-24-3p was predicted and verified. miR-24-3p inhibited thermal injured HSF proliferation and migration and facilitated inflammatory cytokine expression through the regulation of PPAR-ß. p65 directly targeted the transcriptional precursor of miR-24 and promoted miR-24 expression. A negative correlation between miR-24-3p expression level and PPAR-ß expression level in rats' burnt dermal tissues was observed. Our findings reveal that miR-24-3p is conducive to rehabilitating the denatured dermis, which may be beneficial in providing effective therapy of skin burns.
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Queimaduras , MicroRNAs , PPAR beta , Animais , Queimaduras/genética , Proliferação de Células , Citocinas/metabolismo , Fibroblastos/metabolismo , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , NF-kappa B/metabolismo , PPAR beta/genética , PPAR beta/metabolismo , RatosRESUMO
Preservation of denatured dermis exerts promotive functions in wound healing and improves the appearance and function of skin. Angiogenesis is crucial for wound healing during burn injury. However, the potential molecular mechanism of angiogenesis in the recovery after burn injury remains to be elucidated. Herein, RNA chromatin immunoprecipitation (ChIP) sequencing analysis revealed upregulation of long intergenic non-coding RNA 00174 (linc00174) in the post-burn tissues. linc00174 overexpression promoted angiogenic activities of human umbilical vein endothelial cells (HUVECs) in the heat-denatured cell model, characterized by the promotion of cell proliferation, migration, and tube formation. Mechanistically, linc00174 directly bound to enhancer of zeste homolog 2 (EZH2), thus stimulating the protein level of trimethylation at lysine 27 of histone H3 (H3K27me3). Moreover, inhibition of EZH2 resulted in downregulation of ZNF24 and Runx1, as well as a decline of vascular endothelial growth factor A (VEGFA). Furthermore, EZH2 modulated epigenetic repression of ZNF24 and Runx1 through the promoter of H3K27me3. Additionally, ZNF24 and Runx1 both functioned as transcriptional inhibitors of VEGFA. Taken together, these findings uncover that linc00174 epigenetically inhibits ZNF24 and Runx1 expression through binding to EZH2, thus attenuating the suppression of VEGFA, contributing to the facilitation of angiogenesis during the recovery of heat-denatured endothelial cells.
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OBJECTIVE: Thermal injury repair is a complex process during which maintaining the proliferation of human dermis fibroblasts (HDFs) and synthesis of extracellular matrix (ECM) plays a critical role. In the present study, we analyzed potential molecular markers and the probable association between differentially-expressed lncRNAs and protein-coding genes within denatured dermis following thermal injury, attempting to provide further insights to thermal injury repair pathogenesis. METHODS AND MAIN RESULTS: We found that the expression of 3940 lncRNAs was increased, while that of 1438 lncRNAs was reduced in the denatured dermis following thermal injury when compared to normal tissue. Of them, 338 were upregulated and 154 were downregulated by more than 128 times. Via cross-check with another microarray profile analysis on differentially-expressed lncRNAs after thermal injury, LINC00302 was found to be downregulated after thermal injury; more importantly, this skin-specially expressed lncRNA is located near a series of genes related to multiple skin inflammation and skin barrier-associated genomes. LINC00302 overexpression promoted the cell viability and the protein levels of α-SMA and Collagen I in HDFs. CONCLUSIONS: In conclusion, mRNAs and lncRNAs could be differentially expressed in the denatured dermis following thermal injury. mRNA and lncRNA regulatory signaling pathways could participate in thermal injury repair pathogenesis. More importantly, LINC00302 may play a critical role in thermal injury repair.
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Actinas/genética , Queimaduras/genética , Colágeno Tipo I/genética , Derme/metabolismo , Fibroblastos/metabolismo , Regulação da Expressão Gênica , RNA Longo não Codificante/genética , RNA Mensageiro/genética , Actinas/metabolismo , Queimaduras/metabolismo , Linhagem Celular , Sobrevivência Celular/genética , Colágeno Tipo I/metabolismo , Regulação para Baixo , Expressão Gênica , Perfilação da Expressão Gênica , Técnicas de Introdução de Genes , Humanos , RNA Longo não Codificante/metabolismo , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Regulação para CimaRESUMO
Lymphangiogenesis has been regarded as a physiological response to pathologic stimuli. The abnormal proliferation of lymphatic endothelial cell (LECs) and lymphangiogenesis is involved in the development of lymphatic disorders. Reportedly, VEGFC/VEGFR3 plays a key role in lymphangiogenesis; moreover, VEGFC/VEGFR3 exerts their cellular effects through activation of Ca2+ signaling in several cell types. Herein, we demonstrated that VEGFC significantly up-regulated LEC proliferation through VEGFR3; moreover, VEGFC/VEGFR3 induced Ca2+ signaling activation. By using online tools, miR-128 and miR-3916 were predicted as candidate upstream miRNAs which might target VEGFC/VEGFR3. As verified using Immunoblotting assays, miR-128 significantly regulated the protein levels of VEGFC/VEGFR3, whereas miR-3916 only slightly modulated VEGFC and VEGFR3 proteins. Contrary to VEGFC, miR-128 overexpression remarkably suppressed LEC proliferation, Ca2+ release and ERK1/2-Akt signaling; moreover, the effect of VEGFC could be partially attenuated by miR-128. In summary, miR-128 interacts with the 3'-UTR of VEGFC and VEGFR3 to inhibit their expression, thus suppressing LEC proliferation through Ca2+ and ERK1/2-Akt signaling. Taken together, we provided novel experimental basis for miRNA-regulated LEC proliferation through Ca2+ signaling.
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Sinalização do Cálcio/genética , Células Endoteliais/citologia , MicroRNAs/genética , Fator C de Crescimento do Endotélio Vascular/genética , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/genética , Sequência de Bases , Cálcio/metabolismo , Linhagem Celular , Proliferação de Células/genética , Humanos , Fator C de Crescimento do Endotélio Vascular/metabolismo , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
Pressure ulcers (PUs) are a complex and serious clinical problem. Deep tissue injury (DTI) is either the outcome or the trigger of deep PUs. However, the cellular and molecular mechanisms that contribute to the pathogenesis of deep PUs remain unclear. In this study, the degeneration characteristics and increased autophagy and apoptosis were observed in deep PU muscle tissues. Muscular proteome of deep PU revealed that a total of 520 proteins were differentially expressed, particularly, JAK2 was down-regulated. Intriguingly, expression of JAK2 in C2C12 myoblasts exposed to oxygen-glucose deprivation and reoxygenation (OGD/R) insult was also distinctly reduced. Ex vivo, we transfected C2C12 myoblasts with lentivirus carrying the JAK2 plasmid and found that JAK2-overexpressed myoblasts exhibited a decrease in autophagy and apoptosis after OGD/R treatment, as well as less cell death. Finally, Western blot analysis determined that p-JAK2, p-AKT, p-mTOR and p-ERK1/2 levels were significantly elevated, accompanied by JAK2 overexpression but without p-STAT3, and inhibition of the AKT and ERK1/2 pathway resulted in elevated apoptosis and/or autophagy. These results demonstrated that JAK2 may play an important protective role in muscular ischemia and reperfusion injury during DTI development by inhibition of autophagy and apoptosis through the AKT and ERK1/2 pathways.
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Denatured dermis is a part of the dermis in deep burn wound and has the ability to restore normal morphology and function. In our previous study, we revealed that miR-29a downregulation in denatured dermis may help burn wound healing in the later phase, and further enhance type I collagen synthesis. LIN28A, a highly-conserved RNA binding protein expressed during embryogenesis, plays roles in development, pluripotency, metabolism, as well as tissue repair in adults. In the present study, we investigated the functional roles of LIN28A in human skin fibroblasts (HSFs) and extracellular matrix (ECM), and the interaction between miR-29a and LIN28A. In recent years, long non-coding RNAs have been reported to play a key role in normal development and physiology, as well as in disease development. By using online tools, we screened out several candidate lncRNAs of miR-29a, among which XIST was inversely regulated by miR-29a. XIST, one of the first found cancer-associated lncRNAs, has been frequently reported to play major role in several biological processes. Further, we evaluated the roles and mechanism of XIST in HSF proliferation, migration, and ECM synthesis. Through regulation of miR-29a/LIN28A, XIST knockdown suppressed HSF proliferation, migration, and ECM synthesis. In denatured dermis tissues, XIST, and LIN28A expression was upregulated, miR-29a expression was downregulated. Taken together, promoting XIST expression in denatured dermis, thus to inhibit miR-29a and promote LIN28A expression, further promote HSF proliferation, migration, and ECM synthesis presents a promising strategy for denatured dermis repair.
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Queimaduras/genética , MicroRNAs/genética , RNA Longo não Codificante/genética , Proteínas de Ligação a RNA/genética , Movimento Celular , Proliferação de Células , Células Cultivadas , Derme/citologia , Fibroblastos/citologia , Células HEK293 , Temperatura Alta , HumanosRESUMO
OBJECTIVE: Electrical injuries to the fingers account for the majority of total severe burns that occur each year. While several types of flaps have been used in covering finger defects, all have limitations or disadvantages. The purpose of this study was to introduce our clinical experiences of using the lateral tarsal artery (LTA) flap to successfully restore fingers after electrical injury. PATIENTS AND METHODS: From 2005 to 2012, 10 patients with 14 severe electrical burns to their fingers, including six thumbs and four index and four middle fingers, were treated with LTA flap. The wound size ranged from 2.0×3.0 cm to 3.5×5.0 cm. The flap with free tendon graft was used to repair the tendon defect in four cases, free nerve graft was used to repair the feeling defect in two cases, and the flap with nerve was used to repair the feeling defect in two cases. All the patients were followed up for 3 months to 2 years. RESULTS: All skin flaps adhered successfully and there were no complications. All patients were satisfied with the esthetic appearance and functional outcome of the finger reconstruction. CONCLUSION: LTA flap is a reliable method to restore fingers after severe electrical injuries.
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INTRODUCTION: High-tension electricity can cause devastating injuries that may result in abdominal wall loss, visceral damage, and sometimes major threat to life. The visceral organ may be exposed after debridement and require flap cover, but the tensile strength of abdominal wall may be lack even if flap transplanted. METHODS: From April 2007 through May 2015, 5 patients with severe abdominal electrical injury were treated at our hospital. Exploratory laparotomy was performed based on their clinical manifestations and debridement findings of abdominal wall at early stage, and decision regarding technique for reconstruction of abdominal wall was based on an assessment of the location and extent of the defect. Medical records were reviewed for these data. RESULTS: Clinical evaluation and debridement findings of the abdomen revealed 4 patients with suspicious visceral damage. Laparotomy was performed in 4 cases, and revealed obvious lesion in 3 cases, including segmental necrosis of small intestine, partial necrosis of diaphragm, left liver and gastric wall, and greater omentum. Five patients underwent abdominal wall reconstruction using island retrograde latissimus dorsi myocutaneous flap or free/island composite anterolateral thigh myocutaneous flap. All flaps survived, abdominal bulging occurred in 3 cases after follow-up of 12 to 36 months. CONCLUSIONS: The clinical manifestations and wound features of abdomen collectively suggest a possible requirement of laparotomy for severe abdominal electrical burns. Retrograde latissimus dorsi myocutaneous flap or composite anterolateral thigh myocutaneous flap is an effective option for reconstruction of abdominal wall loss, the long-term complication of abdominal bulging, however, remains a significant clinical challenge.
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Traumatismos Abdominais/cirurgia , Traumatismos por Eletricidade/cirurgia , Laparotomia , Procedimentos de Cirurgia Plástica , Traumatismos Abdominais/etiologia , Traumatismos Abdominais/patologia , Adulto , Desbridamento , Traumatismos por Eletricidade/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Retalhos Cirúrgicos/patologia , Fatores de TempoRESUMO
OBJECTIVE: In the present study, we aimed to present our experience of an effective two-stage surgical approach using scrotal skin flap for patients with penile skin defects following severe burn injury. MATERIALS AND METHODS: A total of 17 patients with penile skin defects underwent scheduled two-stage reconstruction using scrotal skin flap from January 2004 to October 2016. Patients, who were selected as eligible candidates for scrotal flap, exhibited a wide range of indications, including iatrogenic injuries (e.g., diathermy treatment after circumcision), scalding, flame injuries and electrical burns. During the first stage, the denuded penis secondary to debridement was temporally embedded in scrotal skin flap through an intrascrotal tunnel created between the skin and darto's fascia. During the second stage, the skin around the penis was separated and divided from the scrotum after incising the scrotal skin at the ventral penile base. After a mean follow-up of 30 months, the reconstructed penises yielded satisfactory aesthetic outcomes and maintained erectile function. RESULTS: The mean age of patients was 35 years (19-55 years), and the median follow-up was 30 months (12-60 months). No major perioperative complication occurred except for dehiscence of scrotal skin after 2nd stage in three patients, and split-thickness skin grafts were applied for recovery of scrotum. Of 17 cases, 15 patients (88.2%) were in satisfactory cosmetic appearance. All patients regained penile sensation and normal voiding function in standing position. The International Index of Erectile Function (IIEF) was used to assess sexual function, with 13 patients (76.5%) reported normal erectile function, while the rest 4 reported mild erectile dysfunction. With regards to intercourse satisfaction domain, encouraging results indicated a total of 17 patients resumed sexual intercourse after surgery. In overall satisfaction domain, 11/17 (64.7%) reported a satisfaction from "very" to "moderately" with their overall sex life and sexual relationship with their partners, while only one patient reported "moderately dissatisfied" with his overall sex life. CONCLUSIONS: We report a series of particular cases, including rare iatrogenic injury (diathermy treatment after circumcision) and severe compound electrical injury. Meanwhile, we show our successful experience that scrotal skin flap could be used as an effective surgical alternative to cover severe burn injury wound in male genitalia. The scrotal flap, with higher anti-infection ability and flexible contexture, is available for recovering penile skin defects following severe burn injury resulting in good aesthetic and sexual outcomes.
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Queimaduras/cirurgia , Pênis/lesões , Pênis/cirurgia , Procedimentos de Cirurgia Plástica/métodos , Escroto/cirurgia , Retalhos Cirúrgicos , Adulto , Queimaduras/etiologia , Queimaduras por Corrente Elétrica/cirurgia , Circuncisão Masculina/efeitos adversos , Diatermia/efeitos adversos , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Orgasmo , Satisfação do Paciente , Ereção Peniana , Recuperação de Função Fisiológica , Comportamento Sexual , Micção , Adulto JovemRESUMO
OBJECTIVE: To analyze the etiology and clinical characteristics of Marjolin's ulcer, and to explore its prevention and treatment. METHODS: Medical records of 187 patients with Marjolin's ulcers admitted to the Department of Burns and Reconstructive Surgery of Xiangya Hospital of Central South University from January 1998 to May 2015 were retrospectively analyzed, including gender, age of onset of initial injury or primary disease, age of onset of Marjolin's ulcer, initial injury or primary disease, length of latency, lengths of pre- and post-ulceration periods, lesion site, lesion type, lesion area, local scar tension, histopathological type, degree of carcinoma cell differentiation, bone invasion and lymphadenopathy, treatment, and prognosis. The relationships between the age of onset of initial injury or primary disease and the length of latency, and the length of pre-ulceration period and the length of post-ulceration period were assessed by Spearman correlation analysis. The recurrence rates were processed with Fisher's exact test. RESULTS: (1) Among the patients, the ratio of male to female was nearly 1.6:1.0. The age of onset of initial injury or primary disease was 0.17-78.00 (17±18) years, and the age of onset of Marjolin's ulcers was 18-84 (49±14) years. (2) The most common initial injury among the patients was flame burn. The length of latency was 0.25-74.00 (32±16) years, and the lengths of pre- and post-ulceration periods were 0-73.00 (26±19) years and 0.08-59.00 (6±11) years respectively. The common lesion sites were the lower limbs and head and face. The rodent ulcer was the most common lesion type, and the lesion area was 1-625 (69±110) cm(2). There were obviously negative correlations between the age of onset of initial injury or primary disease and the length of latency, as well as the length of pre-ulceration period and the length of post-ulceration period (with r values respectively -0.71 and -0.50, P values below 0.01). The pathological scars of strong tension around lesions were seen in 176 cases. (3) The major histopathological type was squamous cell carcinoma, with high cell differentiation in most cases. (4) Bone invasion of carcinoma cells was observed in 59 cases. Lymph node enlargement was observed in 100 cases, and lymph node metastasis was observed in 18 cases. (5) Twenty patients did not receive any surgery, while 167 patients were treated by surgery with lesion extended resection as the main method. According to the condition of wound after the lesion extended resection, the wounds were mainly repaired by skin grafting and transplantation of local skin flap. The majority of wounds in 139 patients who underwent lesion extended resection were repaired in one surgery. Twenty-eight patients out of 104 followed-up cases had recurrence after surgery, mainly seen on head and face, upper limb, lower limb, and buttock, and there was no significant difference among them (P>0.05). The recurrence time of most patients was longer than 6 months after cure. CONCLUSIONS: Patients with Marjolin's ulcers in younger age of onset of initial injury or primary disease tend to have longer latency, during which the shorter the pre-ulceration period is, the longer the post-ulceration period will be. Marjolin's ulcers are prone to occur in scar sites with large tension. Early treatment of high tension scar and scar ulcer is important in prevention, and surgery is the optimal treatment for Marjolin's ulcers. Regular follow-up should be carried out owning to recurrence rate in certain degree after surgery.
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Carcinoma de Células Escamosas/patologia , Cicatriz/patologia , Neoplasias Cutâneas/patologia , Úlcera Cutânea/patologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Queimaduras/patologia , Carcinoma de Células Escamosas/cirurgia , Criança , Pré-Escolar , Face/patologia , Feminino , Humanos , Lactente , Extremidade Inferior/patologia , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Retrospectivos , Neoplasias Cutâneas/cirurgia , Transplante de Pele , Úlcera Cutânea/cirurgia , Retalhos Cirúrgicos , Adulto JovemRESUMO
OBJECTIVE: To study the expression levels of annexin A1 (ANXA1), GATA-3, and T-bet in T lymphocytes of peripheral blood in burned mice with sepsis at early stage, and to analyze their immune regulatory mechanisms. METHODS: Seven-hundred and eighty male mice of clean grade were divided into sham injury group (n=60, sham injured on the back by immersing in 37 â warm water for 10 s), burn group (n=240, inflicted with 20% TBSA deep partial- thickness burn on the back by immersing in 100 â hot water for 10 s), sepsis group (n=240, intraperitoneally injected with 6 mg/kg lipopolysaccharide), and burn+ sepsis group (n=240) according to the random number table. Mice of burn+ sepsis group were treated as that in burn group at first, and then they were treated as that in sepsis group. (1) Immediately after injury, six mice in sham injury group were selected to collect lymphocyte suspension of peripheral blood (1 tube each mouse) according to the random number table. According to the random number table, 6 mice of each of the other three groups were respectively selected at post injury hour (PIH) 12, 24, 48, and 72 for the collection of lymphocyte suspension from peripheral blood (1 tube each mouse). Each tube of cell suspension was equally divided into two parts. Fluorescein isothiocyanate (FITC)-labeled human anti-mouse CD4 monoclonal antibody and phycoerythrin (PE)-labeled human anti-mouse interferon-γ monoclonal antibody were added to one part of cell suspension to mark helper T lymphocyte 1 (Th1). FITC-labeled human anti-mouse CD4 monoclonal antibody and PE-labeled human anti-mouse interleukin-4 (IL-4) monoclonal antibody were added to the other part of cell suspension to mark Th2. The percentages of Th1 and Th2 were determined with flow cytometer, and the ratio of Th1 to Th2 was calculated. (2) According to the random number table, 18 mice in sham injury group were selected immediately after injury for the collection of lymphocyte suspension of peripheral blood (1 tube each mouse), and 18 mice of each of the other 3 groups were respectively selected at PIH 12, 24, 48, and 72 to collect the lymphocyte suspension of peripheral blood (1 tube each mouse). The mRNA expression levels of ANXA1, GATA-3, and T-bet were determined by real-time fluorescent quantitative reverse transcription-PCR. (3) Immediately after injury, 36 mice in sham injury group were selected to collect lymphocyte suspension of peripheral blood (1 tube each mouse) according to the random number table, and then 36 tubes of cell suspension were divided into 6 batches (6 tubes each batch). Each one of 6 kinds of antibody combinations: antibodies for labeling Th1 and Th2 in combination with PE-anthocyanin 7 labeled human anti-mouse ANXA1 monoclonal antibody, PE-anthocyanin 7 labeled human anti-mouse GATA-3 monoclonal antibody, and PE-anthocyanin 7 labeled human anti-mouse T-bet monoclonal antibody was added to 1 tube of cell suspension at each batch. According to the random number table, 36 mice of each of the other 3 groups were respectively selected at PIH 12, 24, 48, and 72 for the collection of lymphocyte suspension of peripheral blood (1 tube each mouse), and then 36 tubes of cell suspension at each time point were divided into 6 batches for marking with 3 kinds of surface markers of Th1 and Th2 (6 tubes each batch). Each one of above-mentioned 6 kinds of antibodies was added to 1 tube of cell suspension at each time point for each batch. The percentages of ANXA1, GATA-3, and T-bet positive cells in Th1 and Th2 were determined with flow cytometer. Data were processed with one-way analysis of variance, analysis of variance of factorial design, and SNK test. The relationship between the percentages of ANXA1 positive cell and the percentages of GATA-3 positive cell in Th1 and Th2, and mRNA expression level of ANXA1 and mRNA expression level of GATA-3 in lymphocytes were assessed by linear correlation analysis. RESULTS: (1) Compared with those in sham injury group immediately after injury, the percentages of Th1 and Th2 and the ratio of Th1 to Th2 of mice in burn group were significantly decreased from PIH 24 on, with P values below 0.05; the percentages of Th1 and Th2 and the ratios of Th1 to Th2 of mice in sepsis group and burn+ sepsis group were significantly decreased from PIH 12 on, with P values below 0.05. (2) Compared with those in sham injury group immediately after injury, the mRNA expression levels of ANXA1 and GATA-3 in lymphocyte of mice in burn group were significantly decreased from PIH 24 on, with P values below 0.05; the mRNA expression level of T-bet was significantly decreased at PIH 24 but significantly increased at PIH 48 and 72, with P values below 0.05. Compared with those in sham injury group immediately after injury, the mRNA expression levels of ANXA1 and GATA-3 in lymphocytes of mice in sepsis group were significantly decreased from PIH 12 on, and the mRNA expression level of T-bet was increased significantly from PIH 12 on, with P values below 0.05; the mRNA expression levels of ANXA1, GATA-3, and T-bet in lymphocytes of mice in burn+ sepsis group were significantly decreased from PIH 12 on, with P values below 0.05, reaching the nadir at PIH 72 (0.50±0.04, 0.45±0.03, 0.21±0.05, respectively). (3) A significant positive correlation was observed between ANXA1 mRNA expression level and GATA-3 mRNA expression level in lymphocytes of peripheral blood (r=0.862, P<0.05). (4) Compared with those in sham injury group immediately after injury, the percentages of ANXA1 and GATA-3 positive cellsin Th1 and Th2 of mice in burn group were significantly lowered from PIH 24 on, and the percentage of T-bet positive cells was significantly decreased at PIH 24, but it was increased from PIH 48 on, with P values below 0.05. The percentages of ANXA1 and GATA-3 positive cells in Th1 and Th2 of mice in sepsis group were continuously decreased from PIH 12 on, which were lower at most time points than those in sham injury group immediately after injury, with P values below 0.05. The percentages of T-bet positive cells in Th1 and Th2 of mice in sepsis group were significantly increased since PIH 12 as compared with those in sham injury group immediately after injury, with P values below 0.05. The percentages of ANXA1, GATA-3, and T-bet positive cells in Th1 and Th2 of mice in burn+ sepsis group were continuously lowered from PIH 12, with significantly statistical differences at most time points as compared with those in sham injury group immediately after injury, with P values below 0.05. (5) The percentages of GATA-3 positive cells in Th1 and Th2 were significantly positively correlated with those of ANXA1 (with r values respectively 0.747 and 0.787, P values below 0.05). CONCLUSIONS: The expression levels of ANXA1, GATA-3, and T-bet were continuously lowered in burned mice with sepsis, and it may play an important role in Th1/Th2 balance switching to Th2 bias and immunosuppressive process.
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Queimaduras/imunologia , Queimaduras/metabolismo , Fator de Transcrição GATA3/biossíntese , Sepse/sangue , Linfócitos T/metabolismo , Fatores de Transcrição/biossíntese , Animais , Biomarcadores , Fator de Transcrição GATA3/genética , Humanos , Interferon gama/biossíntese , Interferon gama/genética , Interleucina-4/metabolismo , Masculino , Camundongos , RNA Mensageiro , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição/genéticaRESUMO
Our previous study has suggested that downregulated microRNA (miR)-29a in denatured dermis might be involved in burn wound healing. However, the exact role of miR-29a in healing of burn injury still remains unclear. Here, we found that expression of miR-29a was notably upregulated in denatured dermis tissues and skin fibroblast cells after thermal injury, and thereafter gradually downregulated compared with control group. By contrast, the expression of collagen, type I, alpha 2 (COL1A2) and vascular endothelial growth factor (VEGF-A) were first reduced and subsequently upregulated in denatured dermis tissues and skin fibroblast cells after thermal injury. We further identified COL1A2 as a novel target of miR-29a, which is involved in type I collagen synthesis, and showed that miR-29a negatively regulated the expression level of COL1A2 in skin fibroblast cells. In addition, VEGF-A, another target gene of miR-29a, was also negatively mediated by miR-29a in skin fibroblast cells. Inhibition of miR-29a expression significantly promoted the proliferation and migration of skin fibroblast cells after thermal injury, and knockdown of COL1A2 and VEGF-A reversed the effects of miR-29a on the proliferation and migration of skin fibroblast cells. Furthermore, we found that Notch2/Jagged2 signaling was involved in miR-29a response to burn wound healing. Our findings suggest that downregulated miR-29a in denatured dermis may help burn wound healing in the later phase, probably via upregulation of COL1A2 and VEGF-A expression, which can further enhance type I collagen synthesis and angiogenesis.
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OBJECTIVE: To investigate the clinical efficacy of free latissimus dorsi musculocutaneous flaps in repairing large and deep skin and soft tissue defects around the knee joints. METHODS: Twenty-five patients with large and deep skin and soft tissue defects around the knee joints were hospitalized from March 2005 to March 2014. The area of defects around the knee joints ranged from 10 cm × 8 cm to 43 cm × 23 cm. The free latissimus dorsi musculocutaneous flaps were used to repair the defects, with the area ranging from 12 cm × 10 cm to 45 cm × 25 cm. The thoracodorsal artery and its concomitant vein of the musculocutaneous flap were anastomosed to the descending branch of the lateral circumflex femoral artery and its concomitant vein respectively to reconstruct blood supply. Split-thickness skin grafts around the flap donor sites were harvested to cover the muscle surface of the musculocutaneous flaps. The flap donor sites were closed directly with suture, and the skin donor sites were healed by dressing change. RESULTS: All the 25 flaps survived without vascular crisis. The flaps were in satisfactory appearance. The flap donor sites were healed with linear scar. All the patients were followed up for 3 to 6 months. At last, they were able to stand up and walk. CONCLUSIONS: The free latissimus dorsi musculocutaneous flap transplantation is an effective treatment for the repair of large and deep soft tissue defects around the knee joints, and the descending branch of lateral circumflex femoral artery and its concomitant vein are the appropriate recipient vessels.
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Retalho Miocutâneo , Transplante de Pele , Lesões dos Tecidos Moles/cirurgia , Músculos Superficiais do Dorso , Cicatriz , Humanos , Articulação do Joelho , Resultado do Tratamento , Veias , CicatrizaçãoRESUMO
OBJECTIVE: To observe the effect of free lateral upper arm perforator flap in repairing wound on hand or foot due to electrical burn. METHODS: Six patients with full-thickness wounds on hand or foot resulting from electrical burn were hospitalized from June 2010 to June 2013. The wounds ranged from 6.0 cm ×4.0 cm to 8.5 cm×7.5 cm in area. Free lateral upper arm perforator flaps were used to repair these defects, with flap area ranging from 9 cm ×4 cm to 12 cm × 9 cm. The donor sites in five cases were closed by suturing; the other one donor site was closed by transplantation of full-thickness skin from abdomen. RESULTS: One flap used to repair the wound in middle finger failed due to failure of venous return, and it was repaired with full-thickness skin harvested from abdomen after dressing change. The other five flaps survived resulting in good elasticity and matched appearance of the recipient area without obvious bulkiness. Patients were followed up for 6 to 24 months. The function of the injured hands or feet recovered well, and the results of function evaluation of five hands were excellent in 2 cases, good in 2 cases, and poor in 1 case. Little scar formation with no contraction or function impairment was observed on donor site, and the result was satisfactory. CONCLUSIONS: Free lateral upper arm perforator flap, with long vessel and less adipose tissue, is suitable for repairing small but deep wound on hand or foot due to electrical burn.
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Queimaduras por Corrente Elétrica/cirurgia , Retalho Perfurante , Transplante de Pele/métodos , Adulto , Idoso , Braço/cirurgia , Traumatismos do Pé/cirurgia , Traumatismos da Mão/cirurgia , Humanos , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
BACKGROUND: The aim of this study is to identify the differentially expressed proteins in circulating polymorphonuclear neutrophils (PMN) from scalded bacteremia rabbits infected with Staphylococcus aureus to provide a basis to reveal the pathogenesis of burns and sepsis. METHODS: Rabbits were subjected to sham burn (A), A + bacterial challenge (B), 30% scald injury (C), or C + bacterial challenge (D). Bacterial challenge was inflicted as an injection of 2.0 x 10(8) cfu S. aureus 18 h after burn procedure. Animals were sacrificed 24 h after burn. PMN were isolated, and the differential proteins in the PMN from these animals were identified by two-dimensional electrophoresis coupled with MALDI-TOF-MS; two proteins were confirmed by Western blotting. RESULTS: Twenty-one differential protein spots were found, and seven differential proteins were identified. Among the identified proteins, the expression levels of protein disulfide-isomerase and thiol-specific antioxidant protein were down-regulated in groups C and D, and two protein spots of annexin I were identified, one of which was down-regulated and another up-regulated in groups C and D. CONCLUSIONS: Preliminary proteome changes in PMN from rabbits experiencing scald injury and S. aureus sepsis were revealed, which possibly play an important role in the inflammation and pathogenesis of sepsis after scald injury.
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Queimaduras/metabolismo , Neutrófilos/metabolismo , Proteômica , Infecções Estafilocócicas/metabolismo , Animais , Anexina A1/biossíntese , Anexina A1/genética , Western Blotting , Queimaduras/complicações , Queimaduras/patologia , Bases de Dados de Proteínas , Ecocardiografia , Processamento de Imagem Assistida por Computador , Masculino , Mapeamento de Peptídeos , Peroxirredoxinas/biossíntese , Peroxirredoxinas/genética , Isomerases de Dissulfetos de Proteínas/biossíntese , Isomerases de Dissulfetos de Proteínas/genética , Proteínas/química , Coelhos , Sepse/complicações , Sepse/metabolismo , Sepse/patologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Infecções Estafilocócicas/complicações , Infecções Estafilocócicas/patologia , Tripsina/químicaRESUMO
BACKGROUND: Increased susceptibility to infection has been related to impairment of lymphocyte-regulated immune responses after severe burn. The aim of this study is to identify the differential expression of proteins in circulating lymphocytes from scald injury and Pseudomonas aeruginosa sepsis in rabbits to provide a basis for pathogenesis of burns and sepsis. METHODS: Rabbits were subjected to sham burn (A), 30% scald (B), A+bacterial challenge (C) or B+bacterial challenge (D). Bacterial challenge was inflicted by an injection of 2.0x10(8) CFU P. aeruginosa (ATCC27853) in the auricular vein 22 h after the burn procedure. The animals were sacrificed 24 h later. Lymphocytes were isolated, and the differential proteins in the lymphocytes from the experimental and control animals were identified by two-dimensional electrophoresis (2-DE) coupled with matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF-MS), two of which were confirmed by Western blotting. RESULTS: Nineteen differential protein spots were found by 2-DE and 12 spots (11 proteins) were identified. Differential expression of peroxiredoxin and annexin I was validated by Western blotting. Among the identified proteins, the expression levels of cofilin, cyclophilin A, ubiquitin, nucleoside diphosphate kinase, glutamate dehydrogenase and annexin I were down-regulated in group B, excessively down-regulated in group D, but mildly in group C, and peroxiredoxin was up-regulated in groups B and D. CONCLUSIONS: Proteome changes in lymphocytes from P. aeruginosa sepsis in the scalded rabbits were revealed, which are related to immune suppression and the pathogenesis of sepsis after scald injury.
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Proteínas Sanguíneas/metabolismo , Queimaduras/imunologia , Linfócitos/metabolismo , Infecções por Pseudomonas/imunologia , Pseudomonas aeruginosa , Animais , Western Blotting/métodos , Queimaduras/microbiologia , Tolerância Imunológica , Masculino , Proteômica/métodos , Coelhos , Sepse/imunologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodosRESUMO
OBJECTIVE: To study the proteomic change in lymphocytes of rabbits with scald injury and Staphylococcus aureus (SA) invasion. METHODS: Twenty-four rabbits were divided into four groups as follows: control group, scald group, scald with SA invasion 2 hs group, and scald with SA invasion 6 hs group, according to random number table, with 6 rabbits in each group. Except for rabbits in control group (sham scald at 37 degrees C), rabbits in the other 3 groups were subjected to 30% TBSA full-thickness scald. Rabbits in SA invasion 2 and 6 hs groups were injected with 2 mL (1.0 x 10(8) CFU/mL) SA suspension, which was in the log growth phase, via auricle vein 18 hs and 22 hs after injury. Whole blood samples were collected from carotid artery of rabbits in 4 groups 24 hs after scald. Lymphocytes were isolated and its extracted proteins were analyzed by two-dimensional gel electrophoresis coupled with mass spectroscopy. RESULTS: About 1030 protein spots of lymphocytes were detected in each group. Compared with that of control group, 19 protein spots were found to be differentially expressed in the other 3 groups, and 11 spots (10 proteins) were identified. Expression levels of cofilin, cyclophilin A, ubiquitin, nucleoside diphosphate kinase, glutamate dehydrogenase and annexin 1 were down-regulated, but expression level of peroxiredoxin was up-regulated obviously. CONCLUSIONS: There is obvious proteomic change in lymphocytes of scalded rabbits or of scalded rabbits invaded by SA, and it may relate to immune suppression and sepsis after injury.
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Queimaduras/metabolismo , Linfócitos/metabolismo , Proteoma , Infecções Estafilocócicas/metabolismo , Animais , Queimaduras/microbiologia , Masculino , Coelhos , Infecções Estafilocócicas/etiologiaRESUMO
OBJECTIVE: To study the effect of severe burn and Pseudomonas aeruginosa sepsis on the proteomics of lymphocytes (LCs) of rabbits. METHODS: Twenty-four rabbits were divided into four groups, i.e. control, severe scald, severe scald and 2-hour sepsis, severe scald and 6-hour sepsis (6 rabbits in each group). The scald in rabbits was third degree in depth involving 30% of total body surface area (TBSA). The sepsis model was reproduced by intravenous injection of a suspension of Pseudomonas aeruginosa (ATCC 27853, 6 x 10(12)cfu/L) 1 ml/kg 24 hours after scald. The rabbits in control group were treated with warm water of 37 centigrade. Peripheral blood was obtained from the carotid artery 24 hours after scald, or 2 hours after sepsis, or 6 hours after sepsis. The LCs in each blood sample were separated, disrupted and the total proteins of LCs were extracted. The proteins were separated by two dimensional gel electrophoresis. The gels were stained with Coomassie brilliant blue and then were scanned. The images were analyzed by PD quest software. The protein spots of discrepant expression were sieved and then analyzed by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS). The peptide mass finger printing (PMFs) were obtained and were input into the data bank of proteins for identification of the proteins. RESULTS: The average spots of 6 gels were 1 051+/-21 (control), 1 026+/-30 (severe scald), 1 078+/- 36 (2-hour sepsis) and 1 065+/-31 (6-hour sepsis), and the average matching rate were 91% (control), 89% (severe scald), 92% (2-hour sepsis) and 94% (6-hour sepsis), respectively. No difference was found in the protein expression of LCs between 2-hour sepsis group and 6-hour sepsis group, but the protein expression of LCs in severe scald group, 2-hour sepsis group and 6-hour sepsis group differed when compared with control group. Nineteen protein spots expressed discrepancy were sieved and their PMFs were obtained. Twelve protein spots (including 11 proteins) were identified, including Cofilin, peptidyl- prolyl cis-trans isomerase cyclophilin A, ubiquitin, nucleoside diphosphate kinase, glutamate dehydrogenase, selenium binding protein I, beta-actin, peroxiredoxin-6, annexin I, actin-3, cellular retinoic-acid binding protein. CONCLUSION: The proteomics of peripheral blood LCs alters in rabbits with severe burn and Pseudomonas aeruginosa sepsis. The proteins with discrepant expression included 11 proteins, which are related with the folding, assembling, transportation and degradation of proteins, signal transmission, inflammation, immunization, energy metabolism, the proliferation, differentiation and apoptosis of cells. These proteins might be associated with the pathogenesis of severe burn and Pseudomonas aeruginosa sepsis.