RESUMO
BACKGROUND: Radiation-induced fibrosis (RIF) is an important late complication of radiation therapy, and the resulting damaging effects of RIF can significantly impact reconstructive outcomes. There is currently a paucity of effective treatment options available, likely due to the continuing knowledge gap surrounding the cellular mechanisms involved. In this study, detailed analyses of irradiated and non-irradiated human skin samples were performed incorporating histological and single-cell transcriptional analysis to identify novel features guiding development of skin fibrosis following radiation injury. METHODS: Paired irradiated and contralateral non-irradiated skin samples were obtained from six female patients undergoing post-oncologic breast reconstruction. Skin samples underwent histological evaluation, immunohistochemistry, and biomechanical testing. Single-cell RNA sequencing was performed using the 10X single cell platform. Cells were separated into clusters using Seurat in R. The SingleR classifier was applied to ascribe cell type identities to each cluster. Differentially expressed genes characteristic to each cluster were then determined using non-parametric testing. RESULTS: Comparing irradiated and non-irradiated skin, epidermal atrophy, dermal thickening, and evidence of thick, disorganized collagen deposition within the extracellular matrix of irradiated skin were readily appreciated on histology. These histologic features were associated with stiffness that was higher in irradiated skin. Single-cell RNA sequencing revealed six predominant cell types. Focusing on fibroblasts/stromal lineage cells, five distinct transcriptional clusters (Clusters 0-4) were identified. Interestingly, while all clusters were noted to express Cav1, Cluster 2 was the only one to also express Cav2. Immunohistochemistry demonstrated increased expression of Cav2 in irradiated skin, whereas Cav1 was more readily identified in non-irradiated skin, suggesting Cav1 and Cav2 may act antagonistically to modulate fibrotic cellular responses. CONCLUSION: In response to radiation therapy, specific changes to fibroblast subpopulations and enhanced Cav2 expression may contribute to fibrosis. Altogether, this study introduces a novel pathway of caveolin involvement which may contribute to fibrotic development following radiation injury.
Assuntos
Caveolina 1 , Fibroblastos , Análise de Célula Única , Pele , Humanos , Feminino , Fibroblastos/efeitos da radiação , Fibroblastos/metabolismo , Caveolina 1/metabolismo , Caveolina 1/genética , Caveolina 1/biossíntese , Pele/efeitos da radiação , Pele/patologia , Pele/metabolismo , Neoplasias da Mama/radioterapia , Neoplasias da Mama/patologia , Caveolina 2/metabolismo , Caveolina 2/genética , Lesões por Radiação/patologia , Lesões por Radiação/metabolismo , Fibrose , Pessoa de Meia-IdadeRESUMO
OBJECTIVES: Caveolin family proteins, including caveolin-1 (Cav-1), caveolin-2 (Cav-2), and caveolin-3 (Cav-3), are identified as the principal protein components of caveolae in mammalian cells. Circulating form of caveolin family proteins can be used as a good potential biomarker for predicting disease. METHODS: To investigate the clinical significance of the serological levels of caveolin family proteins in patients with systemic lupus erythematosus (SLE), we evaluated the soluble serum levels of caveolin family proteins in patients with SLE by enzyme-linked immunosorbent assay (ELISA) and assessed their associations with various known clinical variables. RESULTS: The major findings of our study are as follows: Cav-2 was not detected in serum of SLE patients and normal controls (NCs). Serum Cav-1 and Cav-3 levels were higher in SLE patients compared with NCs. There were no significant correlations between serum Cav-1 and Cav-3 levels and SLE disease activity. Further analysis showed that serum Cav-3 may be more valuable as a marker than serum Cav-1 in SLE patients. CONCLUSION: Serum levels of Cav-1 and Cav-3 might have a diagnostic role in patients with SLE. However, their predictive and prognostic value was not determined. Further studies are necessary to determine the potential clinical significance of these assays in SLE.
Assuntos
Biomarcadores , Caveolinas , Lúpus Eritematoso Sistêmico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Biomarcadores/sangue , Biomarcadores/metabolismo , Caveolina 1/biossíntese , Caveolina 1/sangue , Caveolina 3/biossíntese , Caveolina 3/sangue , Caveolinas/biossíntese , Caveolinas/sangue , Criança , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/diagnóstico , Lúpus Eritematoso Sistêmico/metabolismo , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Adulto JovemRESUMO
Macrophages are immune cells that play crucial roles in host defense against pathogens by triggering their exceptional phagocytic and inflammatory functions. Macrophages that reside in healthy tissues also accomplish important tasks to preserve organ homeostasis, including lipid uptake/efflux or apoptotic-cell clearance. Both homeostatic and inflammatory functions of macrophages require the precise stability of lipid-rich microdomains located at the cell membrane for the initiation of downstream signaling cascades. Caveolin-1 (Cav-1) is the main protein responsible for the biogenesis of caveolae and plays an important role in vascular inflammation and atherosclerosis. The Liver X receptors (LXRs) are key transcription factors for cholesterol efflux and inflammatory gene responses in macrophages. Although the role of Cav-1 in cellular cholesterol homeostasis and vascular inflammation has been reported, the connection between LXR transcriptional activity and Cav-1 expression and function in macrophages has not been investigated. Here, using gain and loss of function approaches, we demonstrate that LXR-dependent transcriptional pathways modulate Cav-1 expression and compartmentation within the membrane during macrophage activation. As a result, Cav-1 participates in LXR-dependent cholesterol efflux and the control of inflammatory responses. Together, our data show modulation of the LXR-Cav-1 axis could be exploited to control exacerbated inflammation and cholesterol overload in the macrophage during the pathogenesis of lipid and immune disorders, such as atherosclerosis.
Assuntos
Caveolina 1/biossíntese , Colesterol/metabolismo , Receptores X do Fígado/biossíntese , Macrófagos/metabolismo , Transportador 1 de Cassete de Ligação de ATP/metabolismo , Animais , Anti-Inflamatórios , Apolipoproteína A-I/metabolismo , Aterosclerose/metabolismo , Caveolina 1/genética , Membrana Celular/metabolismo , Detergentes , Expressão Gênica , Proteínas de Fluorescência Verde/metabolismo , Humanos , Inflamação , Camundongos , Camundongos Endogâmicos C57BL , Células RAW 264.7 , Transdução de Sinais , Transcrição GênicaRESUMO
The extracellular matrix (ECM) is important for normal development and disease states, including inflammation and fibrosis. To understand the complex regulation of ECM, we performed a suppressor screening using Caenorhabditis elegans expressing the mutant ROL-6 collagen protein. One cuticle mutant has a mutation in dpy-23 that encodes the µ2 adaptin (AP2M1) of clathrin-associated protein complex II (AP-2). The subsequent suppressor screening for dpy-23 revealed the lon-2 mutation. LON-2 functions to regulate body size through negative regulation of the tumor growth factor-beta (TGF-ß) signaling pathway responsible for ECM production. RNA-seq analysis showed a dominant change in the expression of collagen genes and cuticle components. We noted an increase in the cav-1 gene encoding caveolin-1, which functions in clathrin-independent endocytosis. By knockdown of cav-1, the reduced TGF-ß signal was significantly restored in the dpy-23 mutant. In conclusion, the dpy-23 mutation upregulated cav-1 expression in the hypodermis, and increased CAV-1 resulted in a decrease of TßRI. Finally, the reduction of collagen expression including rol-6 by the reduced TGF-ß signal influenced the cuticle formation of the dpy-23 mutant. These findings could help us to understand the complex process of ECM regulation in organism development and disease conditions.
Assuntos
Complexo 2 de Proteínas Adaptadoras/genética , Proteínas de Caenorhabditis elegans/biossíntese , Proteínas de Caenorhabditis elegans/genética , Caveolina 1/biossíntese , Colágeno/biossíntese , Matriz Extracelular/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Complexo 2 de Proteínas Adaptadoras/metabolismo , Animais , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/metabolismo , Caveolina 1/genética , Colágeno/genética , Endocitose/genética , Glipicanas/genética , Interferência de RNA , Receptor do Fator de Crescimento Transformador beta Tipo I/metabolismo , Transdução de Sinais/fisiologiaRESUMO
The integral membrane protein caveolin-1 (CAV1) plays a central role in radioresistance-mediating tumor-stroma interactions of advanced prostate cancer (PCa). Among the tumor-stroma, endothelial cells (EC) evolved as critical determinants of the radiation response. CAV1 deficiency in angiogenic EC was already shown to account for increased apoptosis rates of irradiated EC. This study explores the potential impact of differential CAV1 levels in EC on the acid sphingomyelinase (ASMase)/ceramide pathway as a key player in the regulation of EC apoptosis upon irradiation and cancer cell radioresistance. Enhanced apoptosis sensitivity of CAV1-deficient EC was associated with increased ASMase activity, ceramide generation, formation of large lipid platforms, and finally an altered p38 mitogen-activated protein kinase (MAPK)/heat-shock protein 27 (HSP27)/AKT (protein kinase B, PKB) signaling. CAV1-deficient EC increased the growth delay of LNCaP and PC3 PCa cells upon radiation treatment in direct 3D spheroid co-cultures. Exogenous C6 and C16 ceramide treatment in parallel increased the growth delay of PCa spheroids and induced PCa cell apoptosis. Analysis of the respective ceramide species in PCa cells with increased CAV1 levels like those typically found in radio-resistant advanced prostate tumors further revealed an upregulation of unsaturated C24:1 ceramide that might scavenge the effects of EC-derived apoptosis-inducing C16 ceramide. Higher ASMase as well as ceramide levels could be confirmed by immunohistochemistry in human advanced prostate cancer specimen bearing characteristic CAV1 tumor-stroma alterations. Conclusively, CAV1 critically regulates the generation of ceramide-dependent (re-)organization of the plasma membrane that in turn affects the radiation response of EC and adjacent PCa cells. Understanding the CAV1-dependent crosstalk between tumor cells and the host-derived tumor microvasculature and its impact on radiosensitivity may allow to define a rational strategy for overcoming tumor radiation resistance improving clinical outcomes by targeting CAV1.
Assuntos
Caveolina 1/metabolismo , Ceramidas/metabolismo , Células Endoteliais/efeitos da radiação , Neoplasias da Próstata/patologia , Neoplasias da Próstata/radioterapia , Esfingomielina Fosfodiesterase/metabolismo , Células Estromais/patologia , Caveolina 1/biossíntese , Caveolina 1/deficiência , Comunicação Celular/fisiologia , Comunicação Celular/efeitos da radiação , Linhagem Celular Tumoral , Ceramidas/biossíntese , Ceramidas/farmacologia , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Humanos , Sistema de Sinalização das MAP Quinases/efeitos da radiação , Masculino , Células PC-3 , Neoplasias da Próstata/metabolismo , Tolerância a Radiação , Transdução de Sinais , Células Estromais/metabolismo , Microambiente TumoralRESUMO
The mTOR pathway is one of the key signal cascades in the pathogenesis of idiopathic pulmonary fibrosis. Previous studies have mainly focused on this pathway in the fibroblasts and/or myofibroblasts, but not in the epithelial cells. In this study, we sought to investigate the role of the mTOR pathway in lung epithelial cells in lung fibrosis. Using Sftpc-mTORSL1+IT transgenic mice, in which active mTOR is conditionally expressed in lung epithelial cells, we assessed the effects of chronically activated mTOR in lung epithelial cells on lung phenotypes as well as bleomycin-induced lung fibrosis. Furthermore, we isolated alveolar epithelial cell type 2 from mice and performed RNA sequencing. Sftpc-mTORSL1+IT transgenic mice had no obvious abnormal findings, but, after bleomycin administration, showed more severe fibrotic changes and lower lung compliance than control mice. RNA sequencing revealed Angptl4 (angiopoietin-like protein 4) as a candidate downstream gene of the mTOR pathway. In vitro studies revealed that ANGPTL4, as well as mTOR, promoted tight junction vulnerability and epithelial-mesenchymal transition. mTOR activation in lung epithelial cells promoted lung fibrosis and the expression of ANGPTL4, a novel downstream target of the mTOR pathway, which could be related to the etiology of fibrosis.
Assuntos
Células Epiteliais Alveolares/enzimologia , Transição Epitelial-Mesenquimal/fisiologia , Fibrose Pulmonar Idiopática/enzimologia , Pulmão/enzimologia , Serina-Treonina Quinases TOR/fisiologia , Células A549 , Células Epiteliais Alveolares/patologia , Proteína 4 Semelhante a Angiopoietina/biossíntese , Proteína 4 Semelhante a Angiopoietina/genética , Animais , Bleomicina/toxicidade , Caveolina 1/biossíntese , Caveolina 1/genética , Ativação Enzimática , Humanos , Fibrose Pulmonar Idiopática/induzido quimicamente , Fibrose Pulmonar Idiopática/patologia , Pulmão/patologia , Masculino , Camundongos , Camundongos Transgênicos , Fenótipo , Interferência de RNA , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Proteínas Recombinantes/metabolismo , Transdução de Sinais/fisiologia , Serina-Treonina Quinases TOR/genética , Proteína da Zônula de Oclusão-1/biossíntese , Proteína da Zônula de Oclusão-1/genéticaRESUMO
Chronic hypoxia (CH) augments depolarization-induced pulmonary vasoconstriction through superoxide-dependent, Rho kinase-mediated Ca2+ sensitization. Nicotinamide adenine dinucleotide phosphate oxidase and EGFR (epidermal growth factor receptor) signaling contributes to this response. Caveolin-1 regulates the activity of a variety of proteins, including EGFR and nicotinamide adenine dinucleotide phosphate oxidase, and membrane cholesterol is an important regulator of caveolin-1 protein interactions. We hypothesized that derangement of these membrane lipid domain components augments depolarization-induced Ca2+ sensitization and resultant vasoconstriction after CH. Although exposure of rats to CH (4 wk, â¼380 mm Hg) did not alter caveolin-1 expression in intrapulmonary arteries or the incidence of caveolae in arterial smooth muscle, CH markedly reduced smooth muscle membrane cholesterol content as assessed by filipin fluorescence. Effects of CH on vasoreactivity and superoxide generation were examined using pressurized, Ca2+-permeabilized, endothelium-disrupted pulmonary arteries (â¼150 µm inner diameter) from CH and control rats. Depolarizing concentrations of KCl evoked greater constriction in arteries from CH rats than in those obtained from control rats, and increased superoxide production as assessed by dihydroethidium fluorescence only in arteries from CH rats. Both cholesterol supplementation and the caveolin-1 scaffolding domain peptide antennapedia-Cav prevented these effects of CH, with each treatment restoring membrane cholesterol in CH arteries to control levels. Enhanced EGF-dependent vasoconstriction after CH similarly required reduced membrane cholesterol. However, these responses to CH were not associated with changes in EGFR expression or activity, suggesting that cholesterol regulates this signaling pathway downstream of EGFR. We conclude that alterations in membrane lipid domain signaling resulting from reduced cholesterol content facilitate enhanced depolarization- and EGF-induced pulmonary vasoconstriction after CH.
Assuntos
Cálcio/fisiologia , Caveolina 1/biossíntese , Colesterol/fisiologia , Hipóxia/fisiopatologia , Lipídeos de Membrana/fisiologia , Músculo Liso Vascular/metabolismo , Artéria Pulmonar/fisiopatologia , Vasoconstrição/fisiologia , Animais , Caveolina 1/genética , Doença Crônica , Receptores ErbB/fisiologia , Hipóxia/metabolismo , Masculino , Potenciais da Membrana , Artéria Pulmonar/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/fisiologia , Superóxidos/metabolismoRESUMO
BACKGROUND: Hashimoto's thyroiditis (HT) is characterized by lymphocytic infiltration of the thyroid parenchyma, which ultimately leads to tissue destruction and loss of function. Caveolin-1 (Cav-1) is an essential structural constituent of lipid rafts in the plasma membrane of cells and is reported to be significantly reduced in thyrocytes from HT patients. However, the mechanism of Cav-1 involvement in HT pathogenesis is still largely unclear. METHODS: Cav-1 expression in thyroid tissues from HT patients and euthyroid nodular goiter tissues was detected by immunohistochemistry staining. Cav-1 knockdown and overexpression were constructed by lentiviral transfection in the human thyroid follicular epithelial cell (TFC) line of Nthy-ori 3-1. The mRNA expression levels of chemokines in TFCs were determined by quantitative real-time PCR (qPCR). Cav-1 and peroxisome proliferator-activated receptor gamma (PPARγ) levels were analysed by qPCR and Western blot analysis. The migration ability of peripheral blood mononuclear cells (PBMCs) was detected by the Transwell assay. RESULTS: In this study, Cav-1 and PPARγ expression was reduced in the thyroid tissues from HT patients. In vitro experiments showed that the expressions of chemokine (C-C motif) ligand 5 (CCL5) and migration of PBMCs were markedly increased, while the level of PPARγ was significantly decreased after the lentivirus-mediated knockdown of Cav-1 in Nthy-ori 3-1 cells. Interestingly, pioglitazone, a PPARγ agonist, not only upregulated PPARγ and Cav-1 proteins significantly, but also effectively reversed the Cav-1-knockdown-induced upregulation of CCL5 in Nthy-ori 3-1 cells and reduced the infiltration of lymphocytes. CONCLUSION: The inhibition of Cav-1 upregulated the CCL5 expression and downregulated the PPARγ expression in TFC while pioglitazone, a PPARγ agonist, reversed the detrimental consequence. This outcome might be a potential target for the treatment of lymphocyte infiltration into the thyroid gland and HT development.
Assuntos
Caveolina 1/biossíntese , Quimiocina CCL5/biossíntese , Doença de Hashimoto/metabolismo , PPAR gama/biossíntese , Caveolina 1/genética , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Movimento Celular/fisiologia , Quimiocina CCL5/genética , Expressão Gênica , Doença de Hashimoto/genética , Doença de Hashimoto/patologia , Humanos , PPAR gama/agonistas , PPAR gama/genética , Pioglitazona/farmacologia , Glândula Tireoide/efeitos dos fármacos , Glândula Tireoide/metabolismoRESUMO
Ischemic stroke is a major cause of death and motor disabilities all over the world. It is a muti-factorial disorder associated with inflammatory, apoptotic, and oxidative responses. Nateglinide (NAT), an insulinotropic agent used for the treatment of type 2 diabetes mellitus, recently showed potential anti-inflammatory and anti-apoptotic effects. The aim of our study was to elucidate the unique neuroprotective role of NAT in the middle cerebral artery occlusion (MCAO)-induced stroke in rats. Fifty-six male rats were divided to 4 groups (n = 14 in each group): the sham-operated group, sham receiving NAT (50 mg/kg/day, p.o) group, ischemia/reperfusion (IR) group, and IR receiving NAT group (50 mg/kg/day, p.o). MCAO caused potent deficits in motor and behavioral functions of the rats. Significant increase in inflammatory and apoptotic biomarkers has been observed in rats' hippocampi. Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) pathway was significantly stimulated causing activation of series inflammatory biomarkers ending up neuro-inflammatory milieu. Pretreatment with NAT preserved rats' normal behavioral and motor functions. Moreover, NAT opposed the expression of hypoxia-inducible factor-1α (HIF-1α) resulting in downregulation of more inflammatory mediators namely, NF-κB, tumor necrosis factor-ß (TNF-ß), and the anti-survival gene PMAIP-1. NAT stimulated caveolin-1 (Cav-1) which prevented expression of oxidative biomarkers, nitric oxide (NO), and myeloperoxidase (MPO) and hamper the activation of apoptotic biomarker caspase-3. In conclusion, our work postulated that NAT exhibited its neuroprotective effects in rats with ischemic stroke via attenuation of different unique oxidative, apoptotic, and inflammatory pathways.
Assuntos
Isquemia Encefálica/metabolismo , Caveolina 1/biossíntese , Hipocampo/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Nateglinida/uso terapêutico , Receptores de Superfície Celular/metabolismo , Animais , Isquemia Encefálica/patologia , Isquemia Encefálica/prevenção & controle , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/fisiologia , Hipocampo/efeitos dos fármacos , Hipocampo/patologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Mediadores da Inflamação/antagonistas & inibidores , Mediadores da Inflamação/metabolismo , Masculino , Nateglinida/farmacologia , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/uso terapêutico , Desempenho Psicomotor/efeitos dos fármacos , Desempenho Psicomotor/fisiologia , Ratos , Ratos Wistar , Receptores de Superfície Celular/antagonistas & inibidores , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologia , Traumatismo por Reperfusão/prevenção & controle , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologiaRESUMO
In this study we aim to demonstrate the value of monoclonal Caveolin 1 expression in distinguishing between malignant pleural mesothelioma and pulmonary adenocarcinoma. Total of 129 cases, consisting of 68 cases of malignant pleural mesothelioma (51 epitheloid, 12 biphasic, and 5 sarcomatoid type) and 61 cases of pulmonary adenocarcinoma were examined and stained with monoclonal Caveolin-1. Caveolin 1 expression with a membranous and /or cytoplasmic pattern was detected only in 32.35% (n:22/68) of malignant pleural mesothelioma and 6.5% (n:4/61) of pulmonary adenocarcinoma cases. This finding suggests that the choice of poly/monoclonal antibody for Caveolin 1 in the differential diagnosis of malignant pleural mesothelioma and pulmonary adenocarcinoma is important.
Assuntos
Adenocarcinoma de Pulmão/diagnóstico , Biomarcadores Tumorais/análise , Caveolina 1/biossíntese , Neoplasias Pulmonares/diagnóstico , Mesotelioma Maligno/diagnóstico , Neoplasias Pleurais/diagnóstico , Idoso , Anticorpos Monoclonais , Caveolina 1/análise , Diagnóstico Diferencial , Feminino , Humanos , Imuno-Histoquímica/métodos , Masculino , Pessoa de Meia-IdadeRESUMO
Caveolin-1, the major protein component of caveolae, plays vital functions in tumorigenesis and metastasis. Previous evidence demonstrated the positive role of Caveolin-1 in the regulation of endothelial cell differentiation and the involvement of Caveolin-1 in vascular endothelial growth factor (VEGF) mediated angiogenesis. The correlation of Caveolin-1 expression and angiogenesis is not yet elucidated in osteosarcoma. This study aimed to investigate the expression levels of Caveolin-1 and VEGF in osteosarcoma and their associations with clinicopathological data. This study included 66 formalin-fixed and paraffin embedded osteosarcoma tissue samples. The expression levels of Caveolin-1 and VEGF were assessed by immunohistochemistry. Then associations with clinicopathological variables and the correlation between both markers were evaluated statistically. We also investigated the expression of Caveolin-1 and VEGF values in gene microarrays of osteosarcoma patients and cell lines by using GEO data sets on https://www.ncbi.nlm.nih.gov. Caveolin-1 and VEGF were expressed in 19.6% and 77.3%, respectively. Caveolin-1 expression was associated positively with osteoblastic histological subtype (P < 0.0001). VEGF expression showed positive association with patient age, histological grade and clinical stage (P = 0.031, P = 0.024 and P < 0.001; respectively). An inverse correlation between Caveolin-1 and VEGF expressions in osteosarcoma was found (r = 0.2 P = 0.04). In silico analysis of Caveolin-1 and VEGF expression supported our results. Our results suggest that Caveolin-1 may act as a tumor suppressor in osteosarcoma. Down-regulation of Caveolin-1 can be used as an indicator for poor prognosis in osteosarcoma patients. Meanwhile, overexpression of VEGF is a predictor of pulmonary metastasis and poor prognosis.
Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias Ósseas/patologia , Caveolina 1/biossíntese , Neoplasias Pulmonares/secundário , Osteossarcoma/secundário , Fator A de Crescimento do Endotélio Vascular/biossíntese , Adolescente , Biomarcadores Tumorais/análise , Neoplasias Ósseas/metabolismo , Criança , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Masculino , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Osteossarcoma/metabolismo , PrognósticoRESUMO
Idiopathic pulmonary fibrosis (IPF) is a devastating and fatal disease and characterized by increased deposition of extracellular matrix proteins and scar formation in the lung, resulting from alveolar epithelial damage and accumulation of inflammatory cells. Evidence suggests that Caveolin-1 (Cav-1), a major component of caveolae which regulates cell signaling and endocytosis, is a potential target to treat fibrotic diseases, although the mechanisms and responsible cell types are unclear. We show that Cav-1 expression was downregulated both in alveolar epithelial type I cells in bleomycin-injured mouse lungs and in lung sections from IPF patients. Increased expression of IL-1ß and caspase-1 has been observed in IPF patients, indicating inflammasome activation associated with IPF. Gene transfer of a plasmid expressing Cav-1 using transthoracic electroporation reduced infiltration of neutrophils and monocytes/macrophages and protected from subsequent bleomycin-induced pulmonary fibrosis. Overexpression of Cav-1 suppressed bleomycin- or silica-induced activation of caspase-1 and maturation of pro-IL-1ß to secrete cleaved IL-1ß both in mouse lungs and in primary type I cells. These results demonstrate that gene transfer of Cav-1 downregulates inflammasome activity and protects from subsequent bleomycin-mediated pulmonary fibrosis. This indicates a pivotal regulation of Cav-1 in inflammasome activity and suggests a novel therapeutic strategy for patients with IPF.
Assuntos
Células Epiteliais Alveolares/metabolismo , Bleomicina/efeitos adversos , Caveolina 1 , Terapia Genética , Fibrose Pulmonar Idiopática , Inflamassomos , Animais , Bleomicina/farmacologia , Caveolina 1/biossíntese , Caveolina 1/genética , Eletroporação , Técnicas de Transferência de Genes , Humanos , Fibrose Pulmonar Idiopática/induzido quimicamente , Fibrose Pulmonar Idiopática/genética , Fibrose Pulmonar Idiopática/metabolismo , Fibrose Pulmonar Idiopática/terapia , Inflamassomos/genética , Inflamassomos/metabolismo , CamundongosRESUMO
Caveolin-1 (Cav-1) expression is increased in hepatic stellate cells (HSC) upon liver cirrhosis and it functions as an integral membrane protein of lipid rafts and caveolae that regulates and integrates multiple signals as a platform. This study aimed to evaluate the role of Cav-1 in HSC. Thus, the effects of exogenous expression of Cav-1 in GRX cells, a model of activated HSC, were determined. Here, we demonstrated through evaluating well-known HSC activation markers - such as α-smooth muscle actin, collagen I, and glial fibrillary acidic protein - that up regulation of Cav-1 induced GRX to a more activated phenotype. GRXEGFP-Cav1 presented an increased migration, an altered adhesion pattern, a reorganization f-actin cytoskeleton, an arrested cell cycle, a modified cellular ultrastructure, and a raised endocytic flux. Based on this, GRX EGFP-Cav1 represents a new cellular model that can be an important tool for understanding of events related to HSC activation. Furthermore, our results reinforce the role of Cav-1 as a molecular marker of HSC activation.
Assuntos
Caveolina 1/biossíntese , Pontos de Checagem do Ciclo Celular , Expressão Gênica , Células Estreladas do Fígado/metabolismo , Caveolina 1/genética , Linhagem Celular , Células Estreladas do Fígado/citologia , HumanosRESUMO
The sinusoidal endothelial cells present in the liver (LSECs) are tipically characterized by the presence of pores (fenestrae). During some pathological conditions LSECs undergo "capillarization", a process characterized by loss of fenestrations and acquisition of a vascular phenotype. In chronic liver disease capillarization has been reported to precede the development of fibrosis. LSECs modification in the setting of HCV infection is currently poorly investigated. Considering that HCV accounts for important changes in hepatocytes and in view of the intimate connection between hepatocytes and LSECs, here we set out to study in great detail the LSECs modifications in individuals with HCV-dependent chronic hepatitis. Electron microscopy analysis, and evaluation of CD32, CD31 and caveolin-1 expression showed that in HCV infection LSECs display major morphological changes but maintain their phenotypical identity. Capillarization was observed only in cases at initial stages of fibrosis. Our findings showed that the severity of LSECs modifications appears to be correlated with hepatocytes damage and fibrosis stage providing novel insight in the pathogenesis of HCV-chronic hepatitis.
Assuntos
Células Endoteliais , Hepacivirus/metabolismo , Hepatite C Crônica , Fígado , Adulto , Idoso , Caveolina 1/biossíntese , Células Endoteliais/metabolismo , Células Endoteliais/ultraestrutura , Feminino , Regulação da Expressão Gênica , Hepatite C Crônica/metabolismo , Hepatite C Crônica/patologia , Humanos , Fígado/metabolismo , Fígado/ultraestrutura , Masculino , Pessoa de Meia-Idade , Molécula-1 de Adesão Celular Endotelial a Plaquetas/biossíntese , Receptores de IgG/biossínteseRESUMO
Objective- Arteriovenous fistulae (AVF) are the most common access created for hemodialysis; however, many AVF fail to mature and require repeated intervention, suggesting a need to improve AVF maturation. Eph-B4 (ephrin type-B receptor 4) is the embryonic venous determinant that is functional in adult veins and can regulate AVF maturation. Cav-1 (caveolin-1) is the major scaffolding protein of caveolae-a distinct microdomain that serves as a mechanosensor at the endothelial cell membrane. We hypothesized that Cav-1 function is critical for Eph-B4-mediated AVF maturation. Approach and Results- In a mouse aortocaval fistula model, both Cav-1 mRNA and protein were increased in the AVF compared with control veins. Cav-1 KO (knockout) mice showed increased fistula wall thickening ( P=0.0005) and outward remodeling ( P<0.0001), with increased eNOS (endothelial NO synthase) activity compared with WT (wild type) mice. Ephrin-B2/Fc inhibited AVF outward remodeling in WT mice but not in Cav-1 KO mice and was maintained in Cav-1 RC (Cav-1 endothelial reconstituted) mice (WT, P=0.0001; Cav-1 KO, P=0.7552; Cav-1 RC, P=0.0002). Cavtratin-a Cav-1 scaffolding domain peptide-decreased AVF wall thickness in WT mice and in Eph-B4 het mice compared with vehicle alone (WT, P=0.0235; Eph-B4 het, P=0.0431); cavtratin also increased AVF patency (day 42) in WT mice ( P=0.0275). Conclusions- Endothelial Cav-1 mediates Eph-B4-mediated AVF maturation. The Eph-B4-Cav-1 axis regulates adaptive remodeling during venous adaptation to the fistula environment. Manipulation of Cav-1 function may be a translational strategy to enhance AVF patency.
Assuntos
Derivação Arteriovenosa Cirúrgica , Caveolina 1/fisiologia , Receptor EphB4/fisiologia , Transdução de Sinais/fisiologia , Veia Cava Inferior/fisiologia , Animais , Aorta Abdominal/cirurgia , Cavéolas/metabolismo , Caveolina 1/biossíntese , Caveolina 1/deficiência , Caveolina 1/genética , Caveolina 1/farmacologia , Células Cultivadas , Avaliação Pré-Clínica de Medicamentos , Hemorreologia , Humanos , Pulmão/citologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/antagonistas & inibidores , Óxido Nítrico Sintase Tipo III/fisiologia , Fragmentos de Peptídeos/farmacologia , Remodelação Vascular/fisiologia , Veia Cava Inferior/cirurgiaRESUMO
BACKGROUND: Breast cancer remains the most common female malignancy and metastasis is the leading cause of death in breast cancer patients. Oldenlandia diffusa has been empirically and extensively used as an adjuvant therapy for metastatic breast cancer patients in Traditional Chinese Medicine (TCM) with proven efficacy. However, its anti-metastasis mechanism has been poorly revealed. METHODS: Multiple molecular biology experiments as well as network pharmacology, bioinformatics analysis were conducted to investigate the anti-metastasis mechanism of Oldenlandia diffusa in breast cancer. RESULTS: We demonstrated that ethanol extract of Oldenlandia diffusa (EEOD) significantly inhibited proliferation and induced apoptosis of high-metastatic breast cancer cell lines MDA-MB-231 and MDA-MB-453, while having no obvious cytotoxic effect on multiple nonmalignant cells. Furthermore, EEOD remarkably suppressed the migration and invasion capacities of the above breast cancer cells by modulating the matrix metalloproteinases (MMPs) and the epithelial-mesenchymal transition (EMT) pathway. More importantly, EEOD also significantly inhibited breast cancer metastasis in zebrafish xenotransplantation model in vivo. Network pharmacology and bioinformatics analysis further demonstrated that EEOD yielded 12 candidate compounds and 225 potential targets, and shared 85 putative targets associated with breast cancer metastasis. Mechanistically, RNA sequencing and experimental validation results suggested that EEOD might inhibit breast cancer metastasis by attenuating the expression of caveolin-1 (Cav-1) as overexpression of Cav-1 could weaken the anti-metastasis efficacy of EEOD. CONCLUSIONS: Overall, our findings proved that EEOD could inhibit breast cancer metastasis by attenuating the expression of Cav-1, highlighting the use of EEOD as an adjunctive therapy for metastatic breast cancer patients. This study also provides novel insights into network pharmacology and bioinformatics analysis as effective tools to illuminate the scientific basis of TCM.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Caveolina 1/biossíntese , Sistemas de Liberação de Medicamentos/métodos , Medicamentos de Ervas Chinesas/farmacologia , Oldenlandia , Animais , Animais Geneticamente Modificados , Antineoplásicos Fitogênicos/isolamento & purificação , Antineoplásicos Fitogênicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Caveolina 1/antagonistas & inibidores , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Medicamentos de Ervas Chinesas/isolamento & purificação , Medicamentos de Ervas Chinesas/uso terapêutico , Feminino , Humanos , Ensaios Antitumorais Modelo de Xenoenxerto/métodos , Peixe-ZebraRESUMO
Exercise has been regarded as an effective rehabilitation strategy to facilitate motor and cognitive functional recovery after stroke, even though the complex effects associated with exercise-induced repair of cerebral ischemic injury are not fully elucidated. The enhancement of angiogenesis and neurogenesis, and the improvement of synaptic plasticity following moderate exercise are conducive to functional recovery after ischemic damage. Our previous studies have confirmed the angiogenesis and neurogenesis through the caveolin-1/VEGF pathway in MCAO rats. As an essential neurotrophic factor, BDNF has multiple effects on ischemic injury. In this study, we attempted to determine an additional mechanism of treadmill exercise-mediated motor and cognitive functional recovery through the caveolin-1/VEGF pathway associated with BDNF in the ischemic penumbra of MCAO mice. We found that mice exposed to treadmill exercise after the MCAO operation showed a significant up-regulation in expression of caveolin-1, VEGF, BDNF, synapsin I and CYFIP1 proteins, numbers of cells positive for BrdU/CD34, BDNF, BrdU/NeuN, BrdU/Synapsin I and CYFIP1 expression were increased, which support the reduction in neurological deficit and infarction volume, as well as improved synaptic morphology and spatial learning abilities, compared with the non-exercise mice. However, the caveolin-1 inhibitor, daidzein, resulted in increase in neurological deficit and infarction volume. The selective VEGFR2 inhibitor, PD173074, significantly induced larger infarction volume and neurological injury, and decreased the expression of BDNF in the ischemic penumbra. These findings indicate that exercise improves angiogenesis, neurogenesis and synaptic plasticity to ameliorate motor and cognitive impairment after stroke partially through the caveolin-1/VEGF pathway, which is associated with the coregulator factor, BDNF.
Assuntos
Isquemia Encefálica/metabolismo , Caveolina 1/biossíntese , Cognição/fisiologia , Condicionamento Físico Animal/métodos , Recuperação de Função Fisiológica/fisiologia , Fator A de Crescimento do Endotélio Vascular/biossíntese , Animais , Isquemia Encefálica/terapia , Caveolina 1/antagonistas & inibidores , Cognição/efeitos dos fármacos , Teste de Esforço/métodos , Infarto da Artéria Cerebral Média/metabolismo , Infarto da Artéria Cerebral Média/terapia , Injeções Subcutâneas , Isoflavonas/administração & dosagem , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Destreza Motora/efeitos dos fármacos , Destreza Motora/fisiologia , Pirimidinas/administração & dosagem , Recuperação de Função Fisiológica/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidoresRESUMO
Emerging evidence has suggested that targeting glycolysis may be a promising strategy for cancer treatment. Betulinic acid (BA) is a natural pentacyclic terpene that has been reported to be active in inhibiting various malignancies. Here, we showed that BA could inhibit aerobic glycolysis activity in breast cancer cell lines MCF-7 and MDA-MB-231 by hampering lactate production, glucose uptake and extracellular acidification rate (ECAR), as well as suppressing aerobic glycolysis-related proteins including c-Myc, lactate dehydrogenase A (LDH-A) and p-PDK1/PDK1 (pyruvate dehydrogenase kinase 1). Mechanistic studies validated Caveolin-1 (Cav-1) as one of key targets of BA in suppressing aerobic glycolysis, as BA administration resulted in Cav-1 upregulation, whereas silencing Cav-1 abrogated the inhibitory effect of BA on aerobic glycolysis. Further investigations demonstrated that BA suppressed aerobic glycolysis in breast cancer cells by regulating the Cav-1/NF-κB/c-Myc pathway. More meaningfully, BA significantly inhibited breast cancer growth and glycolytic activity in both the transgenic MMTV-PyVT+/- breast cancer spontaneous model and the zebrafish breast cancer xenotransplantation model without any detectable side effects in vivo. Taken together, our study sheds novel insights into BA as a promising candidate drug for suppressing aerobic glycolysis, highlighting Cav-1 as a potential molecular target of BA and aerobic glycolysis regulation.
Assuntos
Neoplasias da Mama/metabolismo , Caveolina 1/biossíntese , Glicólise/efeitos dos fármacos , NF-kappa B/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Triterpenos/farmacologia , Animais , Antineoplásicos Fitogênicos/farmacologia , Antineoplásicos Fitogênicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Feminino , Glicólise/fisiologia , Humanos , Células MCF-7 , Camundongos , Camundongos Knockout , NF-kappa B/antagonistas & inibidores , Triterpenos Pentacíclicos , Proteínas Proto-Oncogênicas c-myc/antagonistas & inibidores , Distribuição Aleatória , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Triterpenos/uso terapêutico , Peixe-Zebra , Ácido BetulínicoRESUMO
Dendritic and synaptic plasticity in the penumbra are important processes and are considered to be therapeutic targets of ischemic stroke. Treadmill exercise is known to be a beneficial treatment following stroke. However, its effects and potential mechanism in promoting dendritic and synaptic plasticity remain unknown. We have previously demonstrated that the caveolin-1/VEGF signaling pathway plays a positive role in angiogenesis and neurogenesis. Here, we further investigated the effects of treadmill exercise on promoting dendritic and synaptic plasticity in the penumbra and whether they involve the caveolin-1/VEGF signaling pathway. A middle cerebral artery occlusion (MCAO) animal model was established, and rats were randomly divided into eleven groups. At 2â¯days after MCAO, rats were subjected to treadmill exercise for 7 or 28â¯days. Daidzein (a specific inhibitor of caveolin-1, 0.4â¯mg/kg) was used to confirm the effect of caveolin-1/VEGF signaling on exercise-mediated dendritic and synaptic plasticity. Neurobehavioral performance, tissue morphology and infarct volumes were detected by Modified Neurology Severity Score (mNSS), Hematoxylin-eosin (HE), and Nissl staining, while neural plasticity and its molecular mechanism were examined by Golgi-Cox staining, transmission electron microscopy, western blot analysis and immunofluorescence. We found that treadmill exercise promoted dendritic plasticity in the penumbra, consistent with the significant increase in caveolin-1 and VEGF expression; improved neurological recovery; and reduced infarct volume. In contrast to the positive effects of the treadmill, a caveolin-1 inhibitor abrogated the dendritic and synaptic plasticity. Furthermore, we observed that treadmill exercise-induced improved dendritic and synaptic plasticity were significantly inhibited by the caveolin-1 inhibitor, consistent with the lower expression of caveolin-1 and VEGF, as well as the worse neurobehavioral state. The findings indicate that treadmill exercise ameliorates focal cerebral ischemia/reperfusion-induced neurological deficit by promoting dendritic and synaptic plasticity via upregulating caveolin-1/VEGF signaling pathways.
Assuntos
Caveolina 1/biossíntese , Dendritos/patologia , Terapia por Exercício/métodos , Doenças do Sistema Nervoso/terapia , Plasticidade Neuronal , Traumatismo por Reperfusão/terapia , Sinapses/patologia , Fator A de Crescimento do Endotélio Vascular/biossíntese , Animais , Caveolina 1/antagonistas & inibidores , Caveolina 1/genética , Infarto da Artéria Cerebral Média/complicações , Infarto da Artéria Cerebral Média/patologia , Infarto da Artéria Cerebral Média/terapia , Masculino , Doenças do Sistema Nervoso/etiologia , Doenças do Sistema Nervoso/psicologia , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica , Traumatismo por Reperfusão/complicações , Traumatismo por Reperfusão/patologia , Transdução de Sinais , Fator A de Crescimento do Endotélio Vascular/genéticaRESUMO
Spinal cord injury (SCI) causes long-term disability and has no effective clinical treatment. After SCI, extracellular adenosine triphosphate (ATP) leads to an influx of extracellular Ca2+, and this Ca2+ overload causes neuronal toxicosis and apoptosis. The biological functions of leptin have been widely investigated in the central nervous system. In this study, we discovered that the administration of leptin could improve locomotor recovery following SCI. The aim of this study was to determine the neuroprotective mechanism of leptin in vivo and in vitro. The neuronal apoptosis and Ca2+ imaging signal induced by ATP were suppressed by leptin, due to elevated caveolin-1 expression. In vivo two-photon observations revealed that leptin reduced the neuronal Ca2+ imaging signal in the exposed spinal cords of live Thy1-YFP mice. In conclusion, leptin promotes locomotor functional recovery and suppresses neuronal impairment after SCI, suggesting that leptin has a promising clinical therapeutic value for treatment of SCI.