RESUMO
Many breast cancer (BC) patients suffer from complications of metastatic disease. To form metastases, cancer cells must become migratory and coordinate both invasive and proliferative programs at distant organs. Here, we identify srGAP1 as a regulator of a proliferative-to-invasive switch in BC cells. High-resolution light-sheet microscopy demonstrates that BC cells can form actin-rich protrusions during extravasation. srGAP1low cells display a motile and invasive phenotype that facilitates their extravasation from blood vessels, as shown in zebrafish and mouse models, while attenuating tumor growth. Interestingly, a population of srGAP1low cells remain as solitary disseminated tumor cells in the lungs of mice bearing BC tumors. Overall, srGAP1low cells have increased Smad2 activation and TGF-ß2 secretion, resulting in increased invasion and p27 levels to sustain quiescence. These findings identify srGAP1 as a mediator of a proliferative to invasive phenotypic switch in BC cells in vivo through a TGF-ß2-mediated signaling axis.
Assuntos
Actinas , Fator de Crescimento Transformador beta2 , Animais , Linhagem Celular Tumoral , Regulação para Baixo , Camundongos , Peixe-ZebraRESUMO
Cancer cells disseminate and seed in distant organs, where they can remain dormant for many years before forming clinically detectable metastases. Here we studied how disseminated tumor cells sense and remodel the extracellular matrix (ECM) to sustain dormancy. ECM proteomics revealed that dormant cancer cells assemble a type III collagen-enriched ECM niche. Tumor-derived type III collagen is required to sustain tumor dormancy, as its disruption restores tumor cell proliferation through DDR1-mediated STAT1 signaling. Second-harmonic generation two-photon microscopy further revealed that the dormancy-to-reactivation transition is accompanied by changes in type III collagen architecture and abundance. Analysis of clinical samples revealed that type III collagen levels were increased in tumors from patients with lymph node-negative head and neck squamous cell carcinoma compared to patients who were positive for lymph node colonization. Our data support the idea that the manipulation of these mechanisms could serve as a barrier to metastasis through disseminated tumor cell dormancy induction.
Assuntos
Colágeno Tipo III , Neoplasias de Cabeça e Pescoço , Proliferação de Células , Matriz Extracelular , Humanos , Carcinoma de Células Escamosas de Cabeça e PescoçoRESUMO
While most primary tumors can be effectively treated, therapeutics fail to efficiently eliminate metastases. Metastases arise from cancer cells that leave the primary tumor and seed distant sites. Recent studies have shown that cancer cells disseminate early during tumor progression and can remain dormant for years before they resume growth. In these metastatic organs, cancer cells reside in microenvironments where they interact with other cells, but also with the extracellular matrix (ECM). The ECM was long considered to be an inert, non-cellular component of tissues, providing their architecture. However, in recent years, a growing body of evidence has shown that the ECM is a key driver of cancer progression, and it can exert effects on tumor cells, regulating their metastatic fate. ECM remodeling and degradation is required for the early steps of the metastatic cascade: invasion, tumor intravasation, and extravasation. Similarly, ECM molecules have been shown to be important for metastatic outgrowth. However, the role of ECM molecules on tumor dormancy and their contribution to the dormancy-supportive niches is not well understood. In this perspective article, we will summarize the current knowledge of ECM and its role in tumor metastasis and dormancy. We will discuss how a better understanding of the individual components of the ECM niche and their roles mediating the dormant state of disseminated tumor cells (DTCs) will advance the development of new therapies to target dormant cells and prevent metastasis outgrowth.
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Melanoma is the deadliest skin cancer. Despite improvements in the understanding of the molecular mechanisms underlying melanoma biology and in defining new curative strategies, the therapeutic needs for this disease have not yet been fulfilled. Herein, we provide evidence that the Activating Molecule in Beclin-1-Regulated Autophagy (Ambra1) contributes to melanoma development. Indeed, we show that Ambra1 deficiency confers accelerated tumor growth and decreased overall survival in Braf/Pten-mutated mouse models of melanoma. Also, we demonstrate that Ambra1 deletion promotes melanoma aggressiveness and metastasis by increasing cell motility/invasion and activating an EMT-like process. Moreover, we show that Ambra1 deficiency in melanoma impacts extracellular matrix remodeling and induces hyperactivation of the focal adhesion kinase 1 (FAK1) signaling, whose inhibition is able to reduce cell invasion and melanoma growth. Overall, our findings identify a function for AMBRA1 as tumor suppressor in melanoma, proposing FAK1 inhibition as a therapeutic strategy for AMBRA1 low-expressing melanoma.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Melanoma/genética , Melanoma/metabolismo , Animais , Autofagia/fisiologia , Proteína Beclina-1/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Modelos Animais de Doenças , Feminino , Quinase 1 de Adesão Focal/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Melanoma/patologia , Camundongos , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Fenótipo , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Transdução de Sinais , TranscriptomaRESUMO
Erythropoiesis involves complex interrelated molecular signals influencing cell survival, differentiation, and enucleation. Diseases associated with ineffective erythropoiesis, such as ß-thalassemias, exhibit erythroid expansion and defective enucleation. Clear mechanistic determinants of what make erythropoiesis effective are lacking. We previously demonstrated that exogenous transferrin ameliorates ineffective erythropoiesis in ß-thalassemic mice. In the current work, we utilize transferrin treatment to elucidate a molecular signature of ineffective erythropoiesis in ß-thalassemia. We hypothesize that compensatory mechanisms are required in ß-thalassemic erythropoiesis to prevent apoptosis and enhance enucleation. We identify pleckstrin-2-a STAT5-dependent lipid binding protein downstream of erythropoietin-as an important regulatory node. We demonstrate that partial loss of pleckstrin-2 leads to worsening ineffective erythropoiesis and pleckstrin-2 knockout leads to embryonic lethality in ß-thalassemic mice. In addition, the membrane-associated active form of pleckstrin-2 occurs at an earlier stage during ß-thalassemic erythropoiesis. Furthermore, membrane-associated activated pleckstrin-2 decreases cofilin mitochondrial localization in ß-thalassemic erythroblasts and pleckstrin-2 knockdown in vitro induces cofilin-mediated apoptosis in ß-thalassemic erythroblasts. Lastly, pleckstrin-2 enhances enucleation by interacting with and activating RacGTPases in ß-thalassemic erythroblasts. This data elucidates the important compensatory role of pleckstrin-2 in ß-thalassemia and provides support for the development of targeted therapeutics in diseases of ineffective erythropoiesis.
Assuntos
Apoptose , Núcleo Celular/patologia , Eritroblastos/patologia , Eritropoese , Proteínas de Membrana/fisiologia , Talassemia beta/patologia , Animais , Núcleo Celular/metabolismo , Eritroblastos/metabolismo , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Talassemia beta/etiologia , Talassemia beta/metabolismoRESUMO
The actin cytoskeleton is a dynamic network that regulates cellular behavior from development to disease. By rearranging the actin cytoskeleton, cells are capable of migrating and invading during developmental processes; however, many of these cellular properties are hijacked by cancer cells to escape primary tumors and disseminate to distant organs in the body. In this review article, we highlight recent work describing how cancer cells regulate the actin cytoskeleton to achieve efficient invasion and metastatic colonization. We also review new imaging technologies that are capable of revealing the complex architecture and regulation of the actin cytoskeleton during motility and invasion of tumor cells.
Assuntos
Actinas/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Animais , Ciclo Celular , Movimento Celular , Transição Epitelial-Mesenquimal , Humanos , Metástase NeoplásicaRESUMO
In the bone marrow (BM) microenvironment, where breast cancer (BC) disseminated tumour cells (DTCs) can remain dormant for decades, NG2+/Nestin+ mesenchymal stem cells (MSCs) promote hematopoietic stem cell quiescence. Here, we reveal that periarteriolar BM-resident NG2+/Nestin+ MSCs can also instruct BC DTCs to enter dormancy. NG2+/Nestin+ MSCs produce TGFß2 and BMP7 and activate a quiescence pathway dependent on TGFBRIII and BMPRII, which via p38-kinase result in p27 induction. Genetic depletion of MSCs or conditional knock-out of TGFß2 in MSCs using an NG2-CreER driver led to bone metastatic outgrowth of otherwise dormant p27+/Ki67- DTCs. Also ER+ BC patients without systemic recurrence displayed higher frequency of TGFß2 and BMP7 detection in the BM. Our results provide a direct proof that HSC dormancy niches control BC DTC dormancy and suggest that aging or extrinsic factors that affect the NG2+/Nestin+ MSC niche homeostasis may result in a break from dormancy and BC bone relapse.
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Neoplasias da Mama , Células-Tronco Mesenquimais , Medula Óssea/metabolismo , Neoplasias da Mama/genética , Feminino , Humanos , Células-Tronco Mesenquimais/metabolismo , Recidiva Local de Neoplasia/metabolismo , Nestina/metabolismo , Microambiente TumoralRESUMO
Trained immunity, a functional state of myeloid cells, has been proposed as a compelling immune-oncological target. Its efficient induction requires direct engagement of myeloid progenitors in the bone marrow. For this purpose, we developed a bone marrow-avid nanobiologic platform designed specifically to induce trained immunity. We established the potent anti-tumor capabilities of our lead candidate MTP10-HDL in a B16F10 mouse melanoma model. These anti-tumor effects result from trained immunity-induced myelopoiesis caused by epigenetic rewiring of multipotent progenitors in the bone marrow, which overcomes the immunosuppressive tumor microenvironment. Furthermore, MTP10-HDL nanotherapy potentiates checkpoint inhibition in this melanoma model refractory to anti-PD-1 and anti-CTLA-4 therapy. Finally, we determined MTP10-HDL's favorable biodistribution and safety profile in non-human primates. In conclusion, we show that rationally designed nanobiologics can promote trained immunity and elicit a durable anti-tumor response either as a monotherapy or in combination with checkpoint inhibitor drugs.
Assuntos
Inibidores de Checkpoint Imunológico/uso terapêutico , Imunidade , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/patologia , Nanotecnologia , Acetilmuramil-Alanil-Isoglutamina/metabolismo , Animais , Comportamento Animal , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Proliferação de Células/efeitos dos fármacos , Colesterol/metabolismo , Feminino , Células-Tronco Hematopoéticas/efeitos dos fármacos , Células-Tronco Hematopoéticas/metabolismo , Inibidores de Checkpoint Imunológico/farmacologia , Imunidade/efeitos dos fármacos , Imunoterapia , Lipoproteínas HDL/metabolismo , Camundongos Endogâmicos C57BL , Primatas , Distribuição Tecidual/efeitos dos fármacos , Microambiente Tumoral/efeitos dos fármacosRESUMO
In this review, we present recent findings on the dynamic nature of the tumour microenvironment (TME) and how intravital microscopy studies have defined TME components in a spatiotemporal manner. Intravital microscopy has shed light into the nature of the TME, revealing structural details of both tumour cells and other TME co-habitants in vivo, how these cells communicate with each other, and how they are organized in three-dimensional space to orchestrate tumour growth, invasion, dissemination and metastasis. We will review different imaging tools, imaging reporters and fate-mapping strategies that have begun to uncover the complexity of the TME in vivo.
Assuntos
Neoplasias/patologia , Microambiente Tumoral , Animais , Matriz Extracelular/metabolismo , Humanos , Microscopia Intravital , Metástase Neoplásica/fisiopatologia , Transdução de Sinais/fisiologia , Microambiente Tumoral/fisiologiaRESUMO
Liver sinusoidal endothelial cells (LSECs) possess fenestrae, which are key for the exchange between blood and hepatocytes. Alterations in their number or diameter have important implications for hepatic function in liver diseases. They are lost early in the development of hepatic fibrosis through a process called capillarization. In this study, we aimed to demonstrate whether in vitro dedifferentiated LSECs that have lost fenestrae are able to re-form these structures. Using stimulated emission depletion super-resolution microscopy in combination with transmission electron microscopy, we analyzed fenestrae formation in a model mimicking the capillarization process in vitro. Actin is known to be involved in fenestrae regulation in differentiated LSECs. Using cytochalasin D, an actin-depolymerizing agent, we demonstrated that dedifferentiated LSECs remain capable of forming fenestrae. Conclusion: We provide a new insight into the complex role of actin in fenestrae formation and in the control of their size and show that LSEC fenestrae re-formation is possible, suggesting that this process could be used during fibrosis regression to try to restore exchanges and hepatocyte functions.
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Advanced, metastatic melanomas frequently grow in subcutaneous tissues and portend a poor prognosis. Though subcutaneous tissues are largely composed of adipocytes, the mechanisms by which adipocytes influence melanoma are poorly understood. Using in vitro and in vivo models, we find that adipocytes increase proliferation and invasion of adjacent melanoma cells. Additionally, adipocytes directly transfer lipids to melanoma cells, which alters tumor cell metabolism. Adipocyte-derived lipids are transferred to melanoma cells through the FATP/SLC27A family of lipid transporters expressed on the tumor cell surface. Among the six FATP/SLC27A family members, melanomas significantly overexpress FATP1/SLC27A1. Melanocyte-specific FATP1 expression cooperates with BRAFV600E in transgenic zebrafish to accelerate melanoma development, an effect that is similarly seen in mouse xenograft studies. Pharmacologic blockade of FATPs with the small-molecule inhibitor Lipofermata abrogates lipid transport into melanoma cells and reduces melanoma growth and invasion. These data demonstrate that stromal adipocytes can drive melanoma progression through FATP lipid transporters and represent a new target aimed at interrupting adipocyte-melanoma cross-talk.Significance: We demonstrate that stromal adipocytes are donors of lipids that mediate melanoma progression. Adipocyte-derived lipids are taken up by FATP proteins that are aberrantly expressed in melanoma. Inhibition of FATPs decreases melanoma lipid uptake, invasion, and growth. We provide a mechanism for how stromal adipocytes drive tumor progression and demonstrate a novel microenvironmental therapeutic target. Cancer Discov; 8(8); 1006-25. ©2018 AACR.This article is highlighted in the In This Issue feature, p. 899.
Assuntos
Adipócitos/citologia , Proteínas de Transporte de Ácido Graxo/metabolismo , Ácidos Graxos/metabolismo , Melanoma/patologia , Proteínas Proto-Oncogênicas B-raf/genética , Células 3T3-L1 , Adipócitos/metabolismo , Animais , Animais Geneticamente Modificados , Transporte Biológico/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Técnicas de Cocultura , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Melanoma/tratamento farmacológico , Melanoma/metabolismo , Camundongos , Mutação , Invasividade Neoplásica , Transplante de Neoplasias , Compostos de Espiro/administração & dosagem , Compostos de Espiro/farmacologia , Tiadiazóis/administração & dosagem , Tiadiazóis/farmacologia , Microambiente Tumoral , Regulação para Cima , Peixe-ZebraRESUMO
Patterning of vertebrate melanophores is essential for mate selection and protection from UV-induced damage. Patterning can be influenced by circulating long-range factors, such as hormones, but it is unclear how their activity is controlled in recipient cells to prevent excesses in cell number and migration. The zebrafish wanderlust mutant harbors a mutation in the sheddase bace2 and exhibits hyperdendritic and hyperproliferative melanophores that localize to aberrant sites. We performed a chemical screen to identify suppressors of the wanderlust phenotype and found that inhibition of insulin/PI3Kγ/mTOR signaling rescues the defect. In normal physiology, Bace2 cleaves the insulin receptor, whereas its loss results in hyperactive insulin/PI3K/mTOR signaling. Insulin B, an isoform enriched in the head, drives the melanophore defect. These results suggest that insulin signaling is negatively regulated by melanophore-specific expression of a sheddase, highlighting how long-distance factors can be regulated in a cell-type-specific manner.
Assuntos
Secretases da Proteína Precursora do Amiloide/metabolismo , Padronização Corporal , Insulina/metabolismo , Melanóforos/fisiologia , Pigmentação , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/fisiologia , Secretases da Proteína Precursora do Amiloide/genética , Animais , Movimento Celular/fisiologia , Embrião não Mamífero/citologia , Embrião não Mamífero/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Insulina/genética , Melanóforos/citologia , Mutação , Fenótipo , Fosfatidilinositol 3-Quinases , Transdução de Sinais , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Peixe-Zebra/embriologia , Proteínas de Peixe-Zebra/genéticaRESUMO
BACKGROUND INFORMATION: Liver sinusoidal endothelial cells (LSECs) possess fenestrae, open transcellular pores with an average diameter of 100 nm. These fenestrae allow for the exchange between blood and hepatocytes. Alterations in their number or diameter in liver diseases have important implications for hepatic microcirculation and function. Although decades of studies, fenestrae are still observed into fixed cells and we have poor knowledge of their dynamics. RESULTS: Using stimulated emission depletion (STED) super-resolution microscopy, we have established a faster and simplest method to observe and quantify fenestrae. Indeed, using cytochalasin D, an actin depolymerising agent known to promote fenestrae formation, we measure the increase of fenestrae number. We adapted this methodology to develop an automated method to study fenestrae dynamics. Moreover, with two-colour STED analysis, we have shown that this approach could be useful to study LSECs fenestrae molecular composition. CONCLUSIONS: Our approach demonstrates that STED microscopy is suitable for LSEC fenestrae study. SIGNIFICANCE: This new way of analysing LSEC fenestrae will allow for expedited investigation of their dynamics, molecular composition and functions to better understand their function in liver pathophysiology.
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Células Endoteliais/fisiologia , Células Endoteliais/ultraestrutura , Processamento de Imagem Assistida por Computador/métodos , Fígado/fisiologia , Fígado/ultraestrutura , Microscopia Eletrônica de Varredura/métodos , Actinas/metabolismo , Animais , Células Cultivadas , Células Endoteliais/citologia , Fígado/citologia , Masculino , CamundongosRESUMO
Discoidin domain receptors, DDR1 and DDR2, are two members of collagen receptor family that belong to tyrosine kinase receptor subgroup. Unlike other matrix receptor-like integrins, these collagen receptors have not been extensively studied. However, more and more studies are focusing on their involvement in cancer. These two receptors are present in several subcellular localizations such as intercellular junction or along type I collagen fibers. Consequently, they are involved in multiple cellular functions, for instance, cell cohesion, proliferation, adhesion, migration and invasion. Furthermore, various signaling pathways are associated with these multiple functions. In this review, we highlight and characterize hallmarks of cancer in which DDRs play crucial roles. We discuss recent data from studies that demonstrate the involvement of DDRs in tumor proliferation, cancer mutations, drug resistance, inflammation, neo-angiogenesis and metastasis. DDRs could be potential targets in cancer and we conclude this review by discussing the different ways to inhibits them.
Assuntos
Receptores com Domínio Discoidina/metabolismo , Neoplasias/metabolismo , Animais , Resistencia a Medicamentos Antineoplásicos , Humanos , Modelos Biológicos , Neoplasias/irrigação sanguínea , Transdução de SinaisRESUMO
Interactions between tumor cells and tumor-associated macrophages play critical roles in the initiation of tumor cell motility. To capture the cellular interactions of the tumor microenvironment with high-resolution imaging, we directly visualized tumor cells and their interactions with macrophages in zebrafish. Live imaging in zebrafish revealed that macrophages are dynamic, yet maintain sustained contact with tumor cells. In addition, the recruitment of macrophages to tumor cells promotes tumor cell dissemination. Using a Cre/LoxP strategy, we found that macrophages transfer cytoplasm to tumor cells in zebrafish and mouse models. Remarkably, macrophage cytoplasmic transfer correlated with melanoma cell dissemination. We further found that macrophages transfer cytoplasm to tumor cells upon cell contact in vitro. Thus, we present a model in which macrophage/tumor cell contact allows for the transfer of cytoplasmic molecules from macrophages to tumor cells corresponding to increased tumor cell motility and dissemination.
Assuntos
Comunicação Celular/fisiologia , Macrófagos/patologia , Melanoma/patologia , Microambiente Tumoral/fisiologia , Animais , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Citoplasma/metabolismo , Camundongos , Invasividade Neoplásica , Peixe-ZebraRESUMO
Cell adhesion, migration, and invasion are involved in many physiological and pathological processes. For example, during metastasis formation, tumor cells have to cross anatomical barriers to invade and migrate through the surrounding tissue in order to reach blood or lymphatic vessels. This requires the interaction between cells and the extracellular matrix (ECM). At the cellular level, many cells, including the majority of cancer cells, are able to form invadosomes, which are F-actin-based structures capable of degrading ECM. Invadosomes are protrusive actin structures that recruit and activate matrix metalloproteinases (MMPs). The molecular composition, density, organization, and stiffness of the ECM are crucial in regulating invadosome formation and activation. In vitro, a gelatin assay is the standard assay used to observe and quantify invadosome degradation activity. However, gelatin, which is denatured collagen I, is not a physiological matrix element. A novel assay using type I collagen fibrils was developed and used to demonstrate that this physiological matrix is a potent inducer of invadosomes. Invadosomes that form along the collagen fibrils are known as linear invadosomes due to their linear organization on the fibers. Moreover, molecular analysis of linear invadosomes showed that the discoidin domain receptor 1 (DDR1) is the receptor involved in their formation. These data clearly demonstrate the importance of using a physiologically relevant matrix in order to understand the complex interactions between cells and the ECM.
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Podossomos/fisiologia , Actinas/metabolismo , Animais , Adesão Celular , Células Cultivadas , Colágeno Tipo I/metabolismo , Matriz Extracelular/metabolismo , Gelatina/metabolismo , HumanosRESUMO
Cutaneous melanoma is a type of cancer with an inherent potential for lymph node colonization, which is generally preceded by neolymphangiogenesis. However, sentinel lymph node removal does not necessarily extend the overall survival of patients with melanoma. Moreover, lymphatic vessels collapse and become dysfunctional as melanomas progress. Therefore, it is unclear whether (and how) lymphangiogenesis contributes to visceral metastasis. Soluble and vesicle-associated proteins secreted by tumours and/or their stroma have been proposed to condition pre-metastatic sites in patients with melanoma. Still, the identities and prognostic value of lymphangiogenic mediators remain unclear. Moreover, our understanding of lymphangiogenesis (in melanomas and other tumour types) is limited by the paucity of mouse models for live imaging of distal pre-metastatic niches. Injectable lymphatic tracers have been developed, but their limited diffusion precludes whole-body imaging at visceral sites. Vascular endothelial growth factor receptor 3 (VEGFR3) is an attractive 'lymphoreporter' because its expression is strongly downregulated in normal adult lymphatic endothelial cells, but is activated in pathological situations such as inflammation and cancer. Here, we exploit this inducibility of VEGFR3 to engineer mouse melanoma models for whole-body imaging of metastasis generated by human cells, clinical biopsies or endogenously deregulated oncogenic pathways. This strategy revealed early induction of distal pre-metastatic niches uncoupled from lymphangiogenesis at primary lesions. Analyses of the melanoma secretome and validation in clinical specimens showed that the heparin-binding factor midkine is a systemic inducer of neo-lymphangiogenesis that defines patient prognosis. This role of midkine was linked to a paracrine activation of the mTOR pathway in lymphatic endothelial cells. These data support the use of VEGFR3 reporter mice as a 'MetAlert' discovery platform for drivers and inhibitors of metastasis.
Assuntos
Citocinas/metabolismo , Vasos Linfáticos/metabolismo , Metástase Neoplásica/diagnóstico por imagem , Metástase Neoplásica/patologia , Imagem Corporal Total/métodos , Animais , Modelos Animais de Doenças , Progressão da Doença , Células Endoteliais/metabolismo , Feminino , Genes Reporter , Humanos , Linfangiogênese , Vasos Linfáticos/patologia , Masculino , Melanoma/diagnóstico por imagem , Melanoma/metabolismo , Melanoma/patologia , Camundongos , Midkina , Comunicação Parácrina , Prognóstico , Recidiva , Reprodutibilidade dos Testes , Serina-Treonina Quinases TOR/metabolismo , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/análise , Receptor 3 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: The most common and severe disease causing allele of Alpha 1-Antitrypsin Deficiency (1ATD) is Z-1AT. This protein aggregates in the endoplasmic reticulum, which is the main cause of liver disease in childhood. Based on recent evidences and on the frequency of liver disease occurrence in Z-1AT patients, it seems that liver disease progression is linked to still unknown genetic factors. METHODS: We used an innovative approach combining yeast genetic screens with next generation exome sequencing to identify and functionally characterize the genes involved in 1ATD associated liver disease. RESULTS: Using yeast genetic screens, we identified HRD1, an Endoplasmic Reticulum Associated Degradation (ERAD) associated protein, as an inducer of Z-mediated toxicity. Whole exome sequencing of 1ATD patients resulted in the identification of two variants associated with liver damages in Z-1AT homozygous cases: HFE H63D and HERPUD1 R50H. Functional characterization in Z-1AT model cell lines demonstrated that impairment of the ERAD machinery combined with the HFE H63D variant expression decreased both cell proliferation and cell viability, while Unfolded Protein Response (UPR)-mediated cell death was hyperstimulated. CONCLUSION: This powerful experimental pipeline allowed us to identify and functionally validate two genes involved in Z-1AT-mediated severe liver toxicity. This pilot study moves forward our understanding on genetic modifiers involved in 1ATD and highlights the UPR pathway as a target for the treatment of liver diseases associated with 1ATD. Finally, these findings support a larger scale screening for HERPUD1 R50H and HFE H63D variants in the sub-group of 1ATD patients developing significant chronic hepatic injuries (hepatomegaly, chronic cholestasis, elevated liver enzymes) and at risk developing liver cirrhosis.
Assuntos
Degradação Associada com o Retículo Endoplasmático/genética , Proteína da Hemocromatose , Cirrose Hepática , Fígado/metabolismo , Mutação de Sentido Incorreto , Deficiência de alfa 1-Antitripsina , Substituição de Aminoácidos , Linhagem Celular , Feminino , Proteína da Hemocromatose/genética , Proteína da Hemocromatose/metabolismo , Humanos , Fígado/patologia , Cirrose Hepática/genética , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Agregação Patológica de Proteínas/genética , Agregação Patológica de Proteínas/metabolismo , Agregação Patológica de Proteínas/patologia , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Deficiência de alfa 1-Antitripsina/genética , Deficiência de alfa 1-Antitripsina/metabolismoRESUMO
Invadosomes are actin-based structures involved in extracellular matrix degradation. Invadosomes is a term that includes podosomes and invadopodia, which decorate normal and tumour cells, respectively. They are mainly organised into dots or rosettes, and podosomes and invadopodia are often compared and contrasted. Various internal or external stimuli have been shown to induce their formation and/or activity. In this Commentary, we address the impact of the microenvironment and the role of matrix receptors on the formation, and dynamic and degradative activities of invadosomes. In particular, we highlight recent findings regarding the role of type I collagen fibrils in inducing the formation of a new linear organisation of invadosomes. We will also discuss invadosome plasticity more generally and emphasise its physio-pathological relevance.