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RATIONALE: Asthma is a chronic inflammatory disease of the airways that involves crosstalk between myeloid-derived regulatory cells (MDRCs) and CD4+ T cells. Although small extracellular vesicles (sEVs) are known to mediate cell-cell communication, the role of sEV signaling via mitochondria in perpetuating asthmatic airway inflammation is unknown. OBJECTIVES: We investigated the effects of MDRC-derived exosomes on dysregulated T cell responses in asthmatics. METHODS: Small extracellular vesicles isolated from bronchoalveolar lavage fluid or airway MDRCs of mild to moderate asthmatics or healthy controls were co-cultured with autologous peripheral and airway CD4+ T lymphocytes. sEV internalization, sEV-mediated transfer of mitochondria targeted GFP to T cells, sEV mitochondrial signaling, and subsequent activation, proliferation and polarization of CD4+ T lymphocytes to Th1, Th2 and Th17 subsets were assessed. MEASUREMENTS AND MAIN RESULTS: Airway MDRC-derived sEVs from asthmatics mediated T cell receptor engagement and transfer of mitochondria that induced antigen-specific activation and polarization into Th17 and Th2 cells, drivers of chronic airway inflammation in asthma. CD4+ T cells internalized sEVs containing mitochondria predominantly by membrane fusion, and blocking mitochondrial oxidant signaling in MDRC-derived exosomes mitigated T cell activation. Reactive oxygen species-mediated signaling that elicited T cell activation in asthmatics was sEV-dependent. A Drp1-dependent mitochondrial fission in pro-inflammatory MDRCs promoted mitochondrial packaging within sEVs, which then co-localized with the polarized actin cytoskeleton and mitochondrial networks in the organized immune synapse of recipient T cells. CONCLUSIONS: Our studies indicate a previously unrecognized role for mitochondrial fission and exosomal mitochondrial transfer in dysregulated T cell activation and Th cell differentiation in asthma which could constitute a novel therapeutic target.
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Mitochondrial dysfunction has been associated with age-related diseases, including idiopathic pulmonary fibrosis (IPF). We provide evidence that implicates chronic elevation of the mitochondrial anion carrier protein, uncoupling protein-2 (UCP2), in increased generation of reactive oxygen species, altered redox state and cellular bioenergetics, impaired fatty acid oxidation, and induction of myofibroblast senescence. This pro-oxidant senescence reprogramming occurs in concert with conventional actions of UCP2 as an uncoupler of oxidative phosphorylation with dissipation of the mitochondrial membrane potential. UCP2 is highly expressed in human IPF lung myofibroblasts and in aged fibroblasts. In an aging murine model of lung fibrosis, the in vivo silencing of UCP2 induces fibrosis regression. These studies indicate a pro-fibrotic function of UCP2 in chronic lung disease and support its therapeutic targeting in age-related diseases associated with impaired tissue regeneration and organ fibrosis.
Assuntos
Fibrose Pulmonar Idiopática , Miofibroblastos , Proteína Desacopladora 2 , Idoso , Animais , Fibroblastos/metabolismo , Fibrose , Humanos , Fibrose Pulmonar Idiopática/metabolismo , Pulmão/metabolismo , Camundongos , Miofibroblastos/metabolismo , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Proteína Desacopladora 2/genética , Proteína Desacopladora 2/metabolismoRESUMO
Fibrosis is a leading cause of morbidity and mortality worldwide. Although fibrosis may involve different organ systems, transforming growth factor-ß (TGFß) has been established as a master regulator of fibrosis across organs. Pirfenidone and Nintedanib are the only currently-approved drugs to treat fibrosis, specifically idiopathic pulmonary fibrosis, but their mechanisms of action remain poorly understood. To identify novel drug targets and uncover potential mechanisms by which these drugs attenuate fibrosis, we performed an integrative 'omics analysis of transcriptomic and proteomic responses to TGFß1-stimulated lung fibroblasts. Significant findings were annotated as associated with pirfenidone and nintedanib treatment in silico via Coremine. Integrative 'omics identified a co-expressed transcriptomic and proteomic module significantly correlated with TGFß1 treatment that was enriched (FDR-p = 0.04) with genes associated with pirfenidone and nintedanib treatment. While a subset of genes in this module have been implicated in fibrogenesis, several novel TGFß1 signaling targets were identified. Specifically, four genes (BASP1, HSD17B6, CDH11, and TNS1) have been associated with pirfenidone, while five genes (CLINT1, CADM1, MTDH, SYDE1, and MCTS1) have been associated with nintedanib, and MYDGF has been implicated with treatment using both drugs. Using the Clue Drug Repurposing Hub, succinic acid was highlighted as a metabolite regulated by the protein encoded by HSD17B6. This study provides new insights into the anti-fibrotic actions of pirfenidone and nintedanib and identifies novel targets for future mechanistic studies.
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Antifibróticos/farmacologia , Biologia Computacional/métodos , Proteínas da Matriz Extracelular/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Fibrose Pulmonar Idiopática/genética , Fibrose Pulmonar Idiopática/patologia , Indóis/farmacologia , Piridonas/farmacologia , Fator de Crescimento Transformador beta/fisiologia , Proteínas Adaptadoras de Transporte Vesicular/genética , Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Antifibróticos/uso terapêutico , Caderinas/genética , Caderinas/metabolismo , Molécula 1 de Adesão Celular/genética , Molécula 1 de Adesão Celular/metabolismo , Feminino , Humanos , Fibrose Pulmonar Idiopática/tratamento farmacológico , Indóis/uso terapêutico , Interleucinas/genética , Interleucinas/metabolismo , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Piridonas/uso terapêutico , Racemases e Epimerases/genética , Racemases e Epimerases/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Tensinas/genética , Tensinas/metabolismoRESUMO
Multicellular organisms maintain structure and function of tissues/organs through emergent, self-organizing behavior. In this report, we demonstrate a critical role for lung mesenchymal stromal cell (L-MSC) aging in determining the capacity to form three-dimensional organoids or 'alveolospheres' with type 2 alveolar epithelial cells (AEC2s). In contrast to L-MSCs from aged mice, young L-MSCs support the efficient formation of alveolospheres when co-cultured with young or aged AEC2s. Aged L-MSCs demonstrated features of cellular senescence, altered bioenergetics, and a senescence-associated secretory profile (SASP). The reactive oxygen species generating enzyme, NADPH oxidase 4 (Nox4), was highly activated in aged L-MSCs and Nox4 downregulation was sufficient to, at least partially, reverse this age-related energy deficit, while restoring the self-organizing capacity of alveolospheres. Together, these data indicate a critical role for cellular bioenergetics and redox homeostasis in an organoid model of self-organization and support the concept of thermodynamic entropy in aging biology.
Many tissues in the body are capable of regenerating by replacing defective or worn-out cells with new ones. This process relies heavily on stem cells, which are precursor cells that lack a set role in the body and can develop into different types of cells under the right conditions. Tissues often have their own pool of stem cells that they use to replenish damaged cells. But as we age, this regeneration process becomes less effective. Many of our organs, such as the lungs, are lined with epithelial cells. These cells form a protective barrier, controlling what substances get in and out of the tissue. Alveoli are parts of the lungs that allow oxygen and carbon dioxide to move between the blood and the air in the lungs. And alveoli rely on an effective epithelial cell lining to work properly. To replenish these epithelial cells, alveoli have pockets, in which a type of epithelial cell, known as AEC2, lives. These cells can serve as stem cells, developing into a different type of cell under the right conditions. To work properly, AEC2 cells require close interactions with another type of cell called L-MSC, which supports the maintenance of other cells and also has the ability to differentiate into several other cell types. Both cell types can be found close together in these stem cell pockets. So far, it has been unclear how aging affects how these cells work together to replenish the epithelial lining of the alveoli. To investigate, Chanda et al. probed AEC2s and L-MSCs in the alveoli of young and old mice. The researchers collected both cell types from young (2-3 months) and aged (22-24 months) mice. Various combinations of these cells were grown to form 3D structures, mimicking how the cells grow in the lungs. Young L-MSCs formed normal 3D structures with both young and aged AEC2 cells. But aged L-MSCs developed abnormal, loose structures with AEC2 cells (both young and old cells). Aged L-MSCs were found to have higher levels of an enzyme (called Nox4) that produces oxidants and other 'pro-aging' factors, compared to young L-MSCs. However, reducing Nox4 levels in aged L-MSCs allowed these cells to form normal 3D structures with young AEC2 cells, but not aged AEC2 cells. These findings highlight the varying effects specific stem cells have, and how their behaviour is affected by pro-aging factors. Moreover, the pro-aging enzyme Nox4 shows potential as a therapeutic target downregulating its activity may reverse critical effects of aging in cells.
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Células Epiteliais Alveolares , Senescência Celular/fisiologia , Células-Tronco Mesenquimais , Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/fisiologia , Animais , Células Cultivadas , Masculino , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/fisiologia , Camundongos , NADPH Oxidase 4/genética , NADPH Oxidase 4/metabolismo , Organoides/citologia , Organoides/metabolismo , Estresse OxidativoRESUMO
Aging is a risk factor for progressive fibrotic disorders involving diverse organ systems, including the lung. Idiopathic pulmonary fibrosis, an age-associated degenerative lung disorder, is characterized by persistence of apoptosis-resistant myofibroblasts. In this report, we demonstrate that sirtuin-3 (SIRT3), a mitochondrial deacetylase, is downregulated in lungs of IPF human subjects and in mice subjected to lung injury. Over-expression of the SIRT3 cDNA via airway delivery restored capacity for fibrosis resolution in aged mice, in association with activation of the forkhead box transcription factor, FoxO3a, in fibroblasts, upregulation of pro-apoptotic members of the Bcl-2 family, and recovery of apoptosis susceptibility. While transforming growth factor-ß1 reduced levels of SIRT3 and FoxO3a in lung fibroblasts, cell non-autonomous effects involving macrophage secreted products were necessary for SIRT3-mediated activation of FoxO3a. Together, these findings reveal a novel role of SIRT3 in pro-resolution macrophage functions that restore susceptibility to apoptosis in fibroblasts via a FoxO3a-dependent mechanism.
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Fibrose Pulmonar Idiopática , Sirtuína 3 , Humanos , Animais , Camundongos , Sirtuína 3/genética , Pulmão/metabolismo , Fibrose , Fibrose Pulmonar Idiopática/metabolismo , Expressão GênicaRESUMO
Extracellular vesicles (EVs) are endosome and plasma membrane-derived nano-sized vesicles that participate in intercellular signaling. Although EV cargo may signal via multiple mechanisms, how signaling components on the surface of EVs mediate cellular signaling is less well understood. In this study, we show that fibroblast-derived EVs carry fibronectin on the vesicular surface, as evidenced by mass spectrometry-based proteomics (Sequential Window Acquisition of all Theoretical Mass Spectra) and flow-cytometric analyses. Fibroblasts undergoing replicative senescence or transforming growth factor ß1-induced senescence and fibroblasts isolated from human subjects with an age-related lung disorder, idiopathic pulmonary fibrosis, secreted higher numbers of EVs than their respective controls. Fibroblast-derived EVs induced an invasive phenotype in recipient fibroblasts. This invasive fibroblast phenotype was dependent on EV surface localization of fibronectin, interaction with the fibronectin receptor α5ß1 integrin, and activation of invasion-associated signaling pathways involving focal adhesion kinase and Src family kinases. EVs in the cellular supernatant, unbound to the extracellular matrix, were capable of mediating invasion signaling on recipient fibroblasts, supporting a direct interaction of EV surface fibronectin with the plasma membrane of recipient cells. Together, these studies uncover a novel mechanism of EV signaling of fibroblast invasion that may be relevant in the pathogenesis of fibrotic diseases and cancer.
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Vesículas Extracelulares/metabolismo , Fibroblastos/metabolismo , Fibronectinas/metabolismo , Movimento Celular/fisiologia , Células Cultivadas , Senescência Celular/fisiologia , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Humanos , Fibrose Pulmonar Idiopática/metabolismo , Integrina alfa5beta1/metabolismo , Transdução de Sinais/fisiologia , Fator de Crescimento Transformador beta1/metabolismo , Quinases da Família src/metabolismoRESUMO
Idiopathic pulmonary fibrosis (IPF) is a progressive and terminal lung disease with no known cure. IPF is a disease of aging, with median age of diagnosis over 65 years. Median survival is between 3 and 5 years after diagnosis. IPF is characterized primarily by excessive deposition of extracellular matrix (ECM) proteins by activated lung fibroblasts and myofibroblasts, resulting in reduced gas exchange and impaired pulmonary function. Growing evidence supports the concept of a pro-fibrotic environment orchestrated by underlying factors such as genetic predisposition, chronic injury and aging, oxidative stress, and impaired regenerative responses may account for disease development and persistence. Currently, two FDA approved drugs have limited efficacy in the treatment of IPF. Many of the genes and gene networks associated with lung development are induced or activated in IPF. In this review, we analyze current knowledge in the field, gained from both basic and clinical research, to provide new insights into the disease process, and potential approaches to treatment of pulmonary fibrosis.
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Fibrose Pulmonar/etiologia , Fibrose Pulmonar/metabolismo , Animais , Biomarcadores , Microambiente Celular , Suscetibilidade a Doenças , Proteínas de Homeodomínio/metabolismo , Humanos , Miofibroblastos/metabolismo , Fibrose Pulmonar/patologia , Transdução de Sinais , Células Estromais , Transativadores , Fator de Crescimento Transformador beta/metabolismoRESUMO
The lung is morphologically structured into a complex tree-like network with branched airways ending distally in a large number of alveoli for efficient oxygen exchange. At the cellular level, the adult lung consists of at least 40-60 different cell types which can be broadly classified into epithelial, endothelial, mesenchymal, and immune cells. Fibroblast growth factor 10 (Fgf10) located in the lung mesenchyme is essential to regulate epithelial proliferation and lineage commitment during embryonic development and post-natal life, and to drive epithelial regeneration after injury. The cells that express Fgf10 in the mesenchyme are progenitors for mesenchymal cell lineages during embryonic development. During adult lung homeostasis, Fgf10 is expressed in mesenchymal stromal niches, between cartilage rings in the upper conducting airways where basal cells normally reside, and in the lipofibroblasts adjacent to alveolar type 2 cells. Fgf10 protects and promotes lung epithelial regeneration after different types of lung injuries. An Fgf10-Hippo epithelial-mesenchymal crosstalk ensures maintenance of stemness and quiescence during homeostasis and basal stem cell (BSC) recruitment to further promote regeneration in response to injury. Fgf10 signaling is dysregulated in different human lung diseases including bronchopulmonary dysplasia (BPD), idiopathic pulmonary fibrosis (IPF), and chronic obstructive pulmonary disease (COPD), suggesting that dysregulation of the FGF10 pathway is critical to the pathogenesis of several human lung diseases.
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Asthma is a chronic inflammatory disease process involving the conductive airways of the human lung. The dysregulated inflammatory response in this disease process may involve multiple cell-cell interactions mediated by signaling molecules, including lipid mediators. Extracellular vesicles (EVs) are lipid membrane particles that are now recognized as critical mediators of cell-cell communication. Here, we compared the lipid composition and presence of specific lipid mediators in airway EVs purified from the bronchoalveolar lavage (BAL) fluid of healthy controls and asthmatic subjects with and without second-hand smoke (SHS) exposure. Airway exosome concentrations were increased in asthmatics, and correlated with blood eosinophilia and serum IgE levels. Frequencies of HLA-DR+ and CD54+ exosomes were also significantly higher in asthmatics. Lipidomics analysis revealed that phosphatidylglycerol, ceramide-phosphates, and ceramides were significantly reduced in exosomes from asthmatics compared to the non-exposed control groups. Sphingomyelin 34:1 was more abundant in exosomes of SHS-exposed asthmatics compared to healthy controls. Our results suggest that chronic airway inflammation may be driven by alterations in the composition of lipid mediators within airway EVs of human subjects with asthma.
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Asma/patologia , Vesículas Extracelulares/metabolismo , Adulto , Idoso , Líquido da Lavagem Broncoalveolar/química , Estudos de Casos e Controles , Ceramidas/metabolismo , Análise Discriminante , Regulação para Baixo , Exossomos/metabolismo , Feminino , Antígenos HLA-DR/metabolismo , Humanos , Imunoglobulina E/sangue , Molécula 1 de Adesão Intercelular/metabolismo , Masculino , Pessoa de Meia-Idade , Fosfatidilgliceróis/metabolismo , Esfingomielinas/metabolismo , Poluição por Fumaça de TabacoRESUMO
Chronic inflammation involving both innate and adaptive immune cells is implicated in the pathogenesis of asthma. Intercellular communication is essential for driving and resolving inflammatory responses in asthma. Emerging studies suggest that extracellular vesicles (EVs) including exosomes facilitate this process. In this report, we have used a range of approaches to show that EVs contain markers of mitochondria derived from donor cells which are capable of sustaining a membrane potential. Further, we propose that these participate in intercellular communication within the airways of human subjects with asthma. Bronchoalveolar lavage fluid of both healthy volunteers and asthmatics contain EVs with encapsulated mitochondria; however, the % HLA-DR+ EVs containing mitochondria and the levels of mitochondrial DNA within EVs were significantly higher in asthmatics. Furthermore, mitochondria are present in exosomes derived from the pro-inflammatory HLA-DR+ subsets of airway myeloid-derived regulatory cells (MDRCs), which are known regulators of T cell responses in asthma. Exosomes tagged with MitoTracker Green, or derived from MDRCs transduced with CellLight Mitochondrial GFP were found in recipient peripheral T cells using a co-culture system, supporting direct exosome-mediated cell-cell transfer. Importantly, exosomally transferred mitochondria co-localize with the mitochondrial network and generate reactive oxygen species within recipient T cells. These findings support a potential novel mechanism of cell-cell communication involving exosomal transfer of mitochondria and the bioenergetic and/or redox regulation of target cells.
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Asma/patologia , Exossomos/patologia , Mitocôndrias/patologia , Células Mieloides/patologia , Comunicação Celular , DNA Mitocondrial/análise , Antígenos HLA-DR/análise , Humanos , Oxirredução , Espécies Reativas de Oxigênio/análiseRESUMO
The lung harbors its basal stem/progenitor cells (BSCs) in the protected environment of the cartilaginous airways. After major lung injuries, BSCs are activated and recruited to sites of injury. Here, we show that during homeostasis, BSCs in cartilaginous airways maintain their stem cell state by downregulating the Hippo pathway (resulting in increased nuclear Yap), which generates a localized Fgf10-expressing stromal niche; in contrast, differentiated epithelial cells in non-cartilaginous airways maintain quiescence by activating the Hippo pathway and inhibiting Fgf10 expression in airway smooth muscle cells (ASMCs). However, upon injury, surviving differentiated epithelial cells spread to maintain barrier function and recruit integrin-linked kinase to adhesion sites, which leads to Merlin degradation, downregulation of the Hippo pathway, nuclear Yap translocation, and expression and secretion of Wnt7b. Epithelial-derived Wnt7b, then in turn, induces Fgf10 expression in ASMCs, which extends the BSC niche to promote regeneration.
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Diferenciação Celular/fisiologia , Fator 10 de Crescimento de Fibroblastos/metabolismo , Pulmão/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Regeneração/fisiologia , Células-Tronco/citologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Células Epiteliais/citologia , Via de Sinalização Hippo , Camundongos Transgênicos , Miócitos de Músculo Liso/citologia , Fosfoproteínas/metabolismoRESUMO
DNA methylation is a major epigenetic event that affects not only cellular gene expression but that also has the potential to influence bacterial and viral DNA in their host-dependent functions. Adeno-associated virus (AAV) genome contains a high degree of CpG sequences capable of methylation in its terminal repeat sequences, which are the sole elements retained in AAV-based vectors used in gene therapy. The present study determined the influence of methylation status of the host cell on wild type (wt) AAV integration and recombinant (r) AAV transgene expression in HeLa cells. Results of the study indicated that hypo-methylation significantly enhanced both wtAAV chromosomal integration and transgene expression of rAAV. A direct influence of methylation on AAV integration was further confirmed by methylating the AAVS1 integration sites prior to viral infection with DNA trans-complementation assay. These results signify the importance of epigenetic status of target cells as one of the key factors in long-term transgene expression in AAV gene therapy.
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The current treatment options for multiple myeloma (MM) osteolytic lesions are mainly combinations of chemotherapy and other small-molecule inhibitors, but toxic side effects still remain a major concern. Studies have shown that osteoclast activity is enhanced in MM patients through increased expression of receptor activator of nuclear factor κB ligand (RANKL), triggering RANK signaling on osteoclast precursors, which results in aggressive bone resorption. Furthermore, osteoprotegerin (OPG), a decoy receptor for RANKL, and the osteogenic potential of mesenchymal stem cells (MSCs) are significantly decreased in myeloma patients with multiple bone lesions. Thus, the use of OPG as a therapeutic molecule would greatly decrease osteolytic damage and reduce morbidity. However, in addition to inhibiting osteoclast activation, OPG binds to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), thereby rendering the tumor cells resistant to TRAIL-induced apoptosis and limiting the use of OPG for therapy. The present study developed a bone-disseminated myeloma disease model in mouse and successfully tested a cell therapy approach using MSCs, genetically engineered to express OPG variants that retain the capacity to bind RANKL, but do not bind TRAIL. Our results of skeletal remodeling following this regenerative stem cell therapy with OPG variants indicated a significant protection against myeloma-induced osteolytic bone damage in areas of major myeloma skeletal dissemination, suggesting the potential of this therapy for treating osteolytic damage in myeloma patients.
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Cellular plasticity and de-differentiation are hallmarks of tissue/organ regenerative capacity in diverse species. Despite a more restricted capacity for regeneration, humans with age-related chronic diseases, such as cancer and fibrosis, show evidence of a recapitulation of developmental gene programs. We have previously identified a resident population of mesenchymal stromal cells (MSCs) in the terminal airways-alveoli by bronchoalveolar lavage (BAL) of human adult lungs. In this study, we characterized MSCs from BAL of patients with stable and progressive idiopathic pulmonary fibrosis (IPF), defined as <5% and ≥10% decline, respectively, in forced vital capacity over the preceding 6-month period. Gene expression profiles of MSCs from IPF subjects with progressive disease were enriched for genes regulating lung development. Most notably, genes regulating early tissue patterning and branching morphogenesis were differentially regulated. Network interactive modeling of a set of these genes indicated central roles for TGF-ß and SHH signaling. Importantly, fibroblast growth factor-10 (FGF-10) was markedly suppressed in IPF subjects with progressive disease, and both TGF-ß1 and SHH signaling were identified as critical mediators of this effect in MSCs. These findings support the concept of developmental gene re-activation in IPF, and FGF-10 deficiency as a potentially critical factor in disease progression.
Assuntos
Reprogramação Celular , Fibrose Pulmonar Idiopática/patologia , Células-Tronco Mesenquimais/patologia , Líquido da Lavagem Broncoalveolar/citologia , Progressão da Doença , Regulação para Baixo/genética , Fator 10 de Crescimento de Fibroblastos/metabolismo , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Genes Controladores do Desenvolvimento , Proteínas Hedgehog/metabolismo , Humanos , Fibrose Pulmonar Idiopática/genética , Imuno-Histoquímica , Pulmão/patologia , Células-Tronco Mesenquimais/metabolismo , Reprodutibilidade dos Testes , Transdução de Sinais/genética , Fator de Crescimento Transformador beta/metabolismo , Regulação para Cima/genéticaRESUMO
OBJECTIVES: The distant organs to which breast cancer preferentially metastasizes are of significant clinical importance. METHODS: We explored the relationship between the clinicopathologic factors and the common sites of distant metastasis in 531 consecutive patients with advanced breast cancer. RESULTS: Breast cancer subtype as a variable was significantly associated with all five common sites of relapse by multivariate analysis. The luminal tumors were remarkable for their significant bone-seeking phenotype and were less frequently observed in lung, brain, and pleural metastases and less likely to be associated with multiorgan relapse. The HER2 subtype demonstrated a significant liver-homing characteristic. African Americans were significantly less likely to have brain-only metastasis in patients with brain relapse. CONCLUSIONS: These findings further articulate that breast cancer subtypes differ not only in tumor characteristics but also in their metastatic behavior, thus raising the possibility that this knowledge could potentially be used in determining the appropriate strategy for follow-up of patients with newly diagnosed breast cancer.
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Neoplasias Ósseas/secundário , Neoplasias Encefálicas/secundário , Neoplasias da Mama/patologia , Neoplasias Hepáticas/secundário , Neoplasias Pulmonares/secundário , Neoplasias Pleurais/secundário , Neoplasias da Mama/classificação , Causalidade , Estudos de Coortes , Feminino , Humanos , Metástase Neoplásica , Recidiva Local de Neoplasia/patologia , Recidiva Local de Neoplasia/secundárioRESUMO
UNLABELLED: Osteolytic bone damage is a major cause of morbidity in several metastatic pathologies. Current therapies using bisphosphonates provide modest improvement, but cytotoxic side effects still occur prompting the need to develop more effective therapies to target aggressive osteoclastogenesis. Increased levels of receptor activator of NF-κB ligand (TNFSF11/RANKL), leading to RANKL-RANK signaling, remain the key axis for osteoclast activation and bone resorption. Osteoprotegerin (TNFRSF11B/OPG), a decoy receptor for RANKL, is significantly decreased in patients who present with bone lesions. Despite its potential in inhibiting osteoclast activation, OPG also binds to TNF-related apoptosis-inducing ligand (TNFSF10/TRAIL), making tumor cells resistant to apoptosis. Toward uncoupling the events of TRAIL binding of OPG and to improve its utility for bone remodeling without inducing tumor resistance to apoptosis, OPG mutants were developed by structural homology modeling based on interactive domain identification and by superimposing models of OPG, TRAIL, and its receptor DR5 (TNFRSF10B) to identify regions of OPG for rational design. The OPG mutants were purified and extensively characterized for their ability to decrease osteoclast damage without affecting tumor apoptosis pathway both in vitro and in vivo, confirming their potential in bone remodeling following cancer-induced osteolytic damage. IMPLICATIONS: OPG variants were developed that lack TRAIL binding, yet retain RANKL binding and suggest new possibilities for therapeutic targeting in osteolytic malignancies.
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Neoplasias Ósseas/metabolismo , Remodelação Óssea , Osteoprotegerina/metabolismo , Neoplasias da Próstata/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Animais , Apoptose/fisiologia , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Humanos , Masculino , Camundongos , Camundongos Nus , Osteoclastos/metabolismo , Osteoclastos/patologia , Osteólise , Neoplasias da Próstata/patologia , Transdução de SinaisRESUMO
Acquired resistance to androgen receptor (AR)-targeted therapies compels the development of novel treatment strategies for castration-resistant prostate cancer (CRPC). Here, we report a profound effect of endostatin on prostate cancer cells by efficient intracellular trafficking, direct interaction with AR, reduction of nuclear AR level, and down-regulation of AR-target gene transcription. Structural modeling followed by functional analyses further revealed that phenylalanine-rich α1-helix in endostatin-which shares structural similarity with noncanonical nuclear receptor box in AR-antagonizes AR transcriptional activity by occupying the activation function (AF)-2 binding interface for coactivators and N-terminal AR AF-1. Together, our data suggest that endostatin can be recognized as an endogenous AR inhibitor that impairs receptor function through protein-protein interaction. These findings provide new insights into endostatin whose antitumor effect is not limited to inhibiting angiogenesis, but can be translated to suppressing AR-mediated disease progression in CRPC.
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Antagonistas de Androgênios/metabolismo , Endostatinas/farmacologia , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Sítios de Ligação , Núcleo Celular/metabolismo , Humanos , MasculinoRESUMO
Anterior Gradient Protein (AGR-2) is reported to be over-expressed in many epithelial cancers and promotes metastasis. A clear-cut mechanism for its observed function(s) has not been previously identified. We found significant upregulation of AGR-2 expression in a bone metastatic prostate cancer cell line, PC3, following culturing in bone marrow-conditioned medium. Substantial AGR-2 expression was also confirmed in prostate cancer tissue specimens in patients with bone lesions. By developing stable clones of PC3 cells with varying levels of AGR-2 expression, we identified that abrogation of AGR-2 significantly reduced cellular attachment to fibronectin, collagen I, collagen IV, laminin I and fibrinogen. Loss of cellular adhesion was associated with sharp decrease in the expression of α4, α5, αV, ß3 and ß4 integrins. Failure to undergo apoptosis following detachment is a hallmark of epithelial cancer metastasis. The AGR-2-silenced PC3 cells showed higher resistance to Tumor necrosis factor-related apoptosis- inducing ligand (TRAIL) induced apoptosis in vitro. This observation was also supported by significantly reduced Caspase-3 expression in AGR-2-silenced PC3 cells, which is a key effector of both extrinsic and intrinsic death signaling pathways. These data suggest that AGR-2 influence prostate cancer metastasis by regulation of cellular adhesion and apoptosis.
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Neoplasias da Próstata/metabolismo , Proteínas/metabolismo , Animais , Medula Óssea/metabolismo , Medula Óssea/patologia , Adesão Celular/efeitos dos fármacos , Adesão Celular/genética , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células , Análise por Conglomerados , Expressão Gênica , Perfilação da Expressão Gênica , Inativação Gênica , Humanos , Integrinas/genética , Integrinas/metabolismo , Masculino , Camundongos , Mucoproteínas , Metástase Neoplásica , Proteínas Oncogênicas , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Proteínas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/farmacologiaRESUMO
Elevated levels of plasmacytoid dendritic cells (pDC) have been reported in breast cancer patients, but the significance remains undefined. Using three immunocompetent mouse models of breast cancer bone metastasis, we identified a key role for pDC in facilitating tumor growth through immunosuppression and aggressive osteolysis. Following infiltration of macrophages upon breast cancer dissemination, there was a steady increase in pDC within the bone, which resulted in a sustained Th2 response along with elevated levels of regulatory T cells and myeloid-derived suppressor cells. Subsequently, pDC and CD4(+) T cells, producing osteolytic cytokines, increased with tumor burden, causing severe bone damage. Microcomputed tomography and histology analyses of bone showed destruction of femur and tibia. The therapeutic significance of this finding was confirmed by depletion of pDC, which resulted in decreased tumor burden and bone loss by activating tumor-specific cytolytic CD8(+) T cells and decreasing suppressor cell populations. Thus, pDC depletion may offer a novel adjuvant strategy to therapeutically influence breast cancer bone metastasis.