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1.
EMBO Rep ; 24(12): e57042, 2023 Dec 06.
Artigo em Inglês | MEDLINE | ID: mdl-37971863

RESUMO

Extracellular vesicles released by tumors (tEVs) disseminate via circulatory networks and promote microenvironmental changes in distant organs favoring metastatic seeding. Despite their abundance in the bloodstream, how hemodynamics affect the function of circulating tEVs remains unsolved. We demonstrated that efficient uptake of tEVs occurs in venous endothelial cells that are subjected to hemodynamics. Low flow regimes observed in veins partially reroute internalized tEVs toward non-acidic and non-degradative Rab14-positive endosomes, at the expense of lysosomes, suggesting that endothelial mechanosensing diverts tEVs from degradation. Subsequently, tEVs promote the expression of pro-angiogenic transcription factors in low flow-stimulated endothelial cells and favor vessel sprouting in zebrafish. Altogether, we demonstrate that low flow regimes potentiate the pro-tumoral function of circulating tEVs by promoting their uptake and rerouting their trafficking. We propose that tEVs contribute to pre-metastatic niche formation by exploiting endothelial mechanosensing in specific vascular regions with permissive hemodynamics.


Assuntos
Vesículas Extracelulares , Neoplasias , Animais , Células Endoteliais , Peixe-Zebra , Vesículas Extracelulares/metabolismo , Hemodinâmica , Neoplasias/patologia , Angiogênese
2.
Am J Hum Genet ; 104(2): 319-330, 2019 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-30639322

RESUMO

ZMIZ1 is a coactivator of several transcription factors, including p53, the androgen receptor, and NOTCH1. Here, we report 19 subjects with intellectual disability and developmental delay carrying variants in ZMIZ1. The associated features include growth failure, feeding difficulties, microcephaly, facial dysmorphism, and various other congenital malformations. Of these 19, 14 unrelated subjects carried de novo heterozygous single-nucleotide variants (SNVs) or single-base insertions/deletions, 3 siblings harbored a heterozygous single-base insertion, and 2 subjects had a balanced translocation disrupting ZMIZ1 or involving a regulatory region of ZMIZ1. In total, we identified 13 point mutations that affect key protein regions, including a SUMO acceptor site, a central disordered alanine-rich motif, a proline-rich domain, and a transactivation domain. All identified variants were absent from all available exome and genome databases. In vitro, ZMIZ1 showed impaired coactivation of the androgen receptor. In vivo, overexpression of ZMIZ1 mutant alleles in developing mouse brains using in utero electroporation resulted in abnormal pyramidal neuron morphology, polarization, and positioning, underscoring the importance of ZMIZ1 in neural development and supporting mutations in ZMIZ1 as the cause of a rare neurodevelopmental syndrome.


Assuntos
Deficiências do Desenvolvimento/genética , Deficiência Intelectual/genética , Mutação Puntual , Fatores de Transcrição/genética , Alelos , Animais , Criança , Pré-Escolar , Deficiências do Desenvolvimento/patologia , Feminino , Humanos , Lactente , Deficiência Intelectual/patologia , Masculino , Camundongos , Síndrome , Fatores de Transcrição/química , Fatores de Transcrição/metabolismo
3.
Hum Mol Genet ; 26(13): 2565-2576, 2017 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-28379387

RESUMO

The association of primary Sjögren's syndrome (pSS) with Major Histocompatibility Complex (MHC) alleles is quintessential of MHC-disease associations. Indeed, although disease associations with classical HLA class I and II alleles/haplotypes are amply documented, further dissection is often prevented by the strong linkage disequilibrium across the entire MHC complex. Here we study the association of pSS, not with HLA genes, but with the non-conventional MHC encoded class I gene, MICA (MHC class I chain-related gene A). MICA is selectively expressed within epithelia, and is the major ligand for the activatory receptor, NKG2D, both attributes relevant to pSS' etiology. MICA-pSS association was studied in two independent (French and UK) cohorts representing a total of 959 cases and 1,043 controls. MICA*008 allele was shown to be significantly associated with pSS (pcor=2.61 × 10-35). A multivariate logistic regression showed that this association was independent of all major known MHC-linked risk loci/alleles, as well as other relevant candidate loci that are in linkage disequilibrium with MICA*008 i.e. HLA-B*08:01, rs3131619 (T), MICB*008, TNF308A, HLA-DRB1*03:01 and HLA-DRB1*15:01 (P = 1.84 × 10-04). Furthermore, independently of the MICA*008 allele, higher levels of soluble MICA proteins were detected in sera of pSS patients compared to healthy controls. This study hence defines MICA as a new, MHC-linked, yet HLA-independent, pSS risk locus and opens a new front in our understanding of the still enigmatic pathophysiology of this disease. The fact that the soluble MICA protein is further amplified in MICA*008 carrying individuals, might also be relevant in other auto-immune diseases and cancer.


Assuntos
Antígenos de Histocompatibilidade Classe I/genética , Síndrome de Sjogren/genética , Adulto , Alelos , Feminino , Frequência do Gene/genética , Predisposição Genética para Doença , Antígenos HLA-B/genética , Cadeias HLA-DRB1/genética , Haplótipos , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Desequilíbrio de Ligação , Complexo Principal de Histocompatibilidade/genética , Masculino , Pessoa de Meia-Idade , Polimorfismo Genético , População Branca/genética
4.
J Hum Genet ; 64(7): 689-694, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31068678

RESUMO

Cathepsin C (CatC) is a cysteine protease involved in a variety of immune and inflammatory pathways such as activation of cytotoxicity of various immune cells. Homozygous or compound heterozygous variants in the CatC coding gene CTSC cause different conditions that have in common severe periodontitis. Periodontitis may occur as part of Papillon-Lefèvre syndrome (PLS; OMIM#245000) or Haim-Munk syndrome (HMS; OMIM#245010), or may present as an isolated finding named aggressive periodontitis (AP1; OMIM#170650). AP1 generally affects young children and results in destruction of the periodontal support of the primary dentition. In the present study we report exome sequencing of a three generation consanguineous Turkish family with a recessive form of early-onset AP1. We identified a novel homozygous missense variant in exon 2 of CTSC (NM_148170, c.G302C, p.Trp101Ser) predicted to disrupt protein structure and to be disease causing. This is the first described CTSC variant specific to the nonsyndromic AP1 form. Given the broad phenotypic spectrum associated with CTSC variants, reporting this novel variant gives new insights on genotype/phenotype correlations and might improve diagnosis of patients with early-onset AP1.


Assuntos
Periodontite Agressiva/genética , Catepsina C/genética , Mutação de Sentido Incorreto , Adolescente , Adulto , Periodontite Agressiva/diagnóstico por imagem , Catepsina C/química , Criança , Feminino , Estudos de Associação Genética , Homozigoto , Humanos , Masculino , Turquia , Sequenciamento do Exoma
6.
Ann Rheum Dis ; 77(11): 1675-1687, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30030262

RESUMO

OBJECTIVES: The objective of the present study was to explain why two siblings carrying both the same homozygous pathogenic mutation for the autoinflammatory disease hyper IgD syndrome, show opposite phenotypes, that is, the first being asymptomatic, the second presenting all classical characteristics of the disease. METHODS: Where single omics (mainly exome) analysis fails to identify culprit genes/mutations in human complex diseases, multiomics analyses may provide solutions, although this has been seldom used in a clinical setting. Here we combine exome, transcriptome and proteome analyses to decipher at a molecular level, the phenotypic differences between the two siblings. RESULTS: This multiomics approach led to the identification of a single gene-STAT1-which harboured a rare missense variant and showed a significant overexpression of both mRNA and protein in the symptomatic versus the asymptomatic sister. This variant was shown to be of gain of function nature, involved in an increased activation of the Janus kinase/signal transducer and activator of transcription signalling (JAK/STAT) pathway, known to play a critical role in inflammatory diseases and for which specific biotherapies presently exist. Pathway analyses based on information from differentially expressed transcripts and proteins confirmed the central role of STAT1 in the proposed regulatory network leading to an increased inflammatory phenotype in the symptomatic sibling. CONCLUSIONS: This study demonstrates the power of a multiomics approach to uncover potential clinically actionable targets for a personalised therapy. In more general terms, we provide a proteogenomics analysis pipeline that takes advantage of subject-specific genomic and transcriptomic information to improve protein identification and hence advance individualised medicine.


Assuntos
Genes Modificadores , Deficiência de Mevalonato Quinase/genética , Fator de Transcrição STAT1/genética , Adulto , Exoma , Feminino , Perfilação da Expressão Gênica/métodos , Humanos , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Fenótipo , Polimorfismo de Nucleotídeo Único , Proteômica/métodos
7.
Blood ; 128(15): 1979-1986, 2016 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-27549307

RESUMO

Graft-versus-host disease (GVHD) is among the most challenging complications in unrelated donor hematopoietic cell transplantation (HCT). The highly polymorphic MHC class I chain-related gene A, MICA, encodes a stress-induced glycoprotein expressed primarily on epithelia. MICA interacts with the invariant activating receptor NKG2D, expressed by cytotoxic lymphocytes, and is located in the MHC, next to HLA-B Hence, MICA has the requisite attributes of a bona fide transplantation antigen. Using high-resolution sequence-based genotyping of MICA, we retrospectively analyzed the clinical effect of MICA mismatches in a multicenter cohort of 922 unrelated donor HLA-A, HLA-B, HLA-C, HLA-DRB1, and HLA-DQB1 10/10 allele-matched HCT pairs. Among the 922 pairs, 113 (12.3%) were mismatched in MICA MICA mismatches were significantly associated with an increased incidence of grade III-IV acute GVHD (hazard ratio [HR], 1.83; 95% confidence interval [CI], 1.50-2.23; P < .001), chronic GVHD (HR, 1.50; 95% CI, 1.45-1.55; P < .001), and nonelapse mortality (HR, 1.35; 95% CI, 1.24-1.46; P < .001). The increased risk for GVHD was mirrored by a lower risk for relapse (HR, 0.50; 95% CI, 0.43-0.59; P < .001), indicating a possible graft-versus-leukemia effect. In conclusion, when possible, selecting a MICA-matched donor significantly influences key clinical outcomes of HCT in which a marked reduction of GVHD is paramount. The tight linkage disequilibrium between MICA and HLA-B renders identifying a MICA-matched donor readily feasible in clinical practice.


Assuntos
Doença Enxerto-Hospedeiro , Antígenos HLA/genética , Transplante de Células-Tronco Hematopoéticas , Antígenos de Histocompatibilidade Classe I/genética , Teste de Histocompatibilidade , Desequilíbrio de Ligação , Doença Aguda , Adolescente , Adulto , Idoso , Aloenxertos , Criança , Pré-Escolar , Doença Crônica , Feminino , Doença Enxerto-Hospedeiro/epidemiologia , Doença Enxerto-Hospedeiro/etiologia , Doença Enxerto-Hospedeiro/genética , Doença Enxerto-Hospedeiro/prevenção & controle , Humanos , Incidência , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Subfamília K de Receptores Semelhantes a Lectina de Células NK/genética , Estudos Retrospectivos
8.
Nat Commun ; 15(1): 3297, 2024 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-38740748

RESUMO

Despite abundant evidence demonstrating that platelets foster metastasis, anti-platelet agents have low therapeutic potential due to the risk of hemorrhages. In addition, whether platelets can regulate metastasis at the late stages of the disease remains unknown. In this study, we subject syngeneic models of metastasis to various thrombocytopenic regimes to show that platelets provide a biphasic contribution to metastasis. While potent intravascular binding of platelets to tumor cells efficiently promotes metastasis, platelets further support the outgrowth of established metastases via immune suppression. Genetic depletion and pharmacological targeting of the glycoprotein VI (GPVI) platelet-specific receptor in humanized mouse models efficiently reduce the growth of established metastases, independently of active platelet binding to tumor cells in the bloodstream. Our study demonstrates therapeutic efficacy when targeting animals bearing growing metastases. It further identifies GPVI as a molecular target whose inhibition can impair metastasis without inducing collateral hemostatic perturbations.


Assuntos
Plaquetas , Metástase Neoplásica , Glicoproteínas da Membrana de Plaquetas , Animais , Plaquetas/metabolismo , Plaquetas/efeitos dos fármacos , Humanos , Camundongos , Glicoproteínas da Membrana de Plaquetas/metabolismo , Glicoproteínas da Membrana de Plaquetas/genética , Linhagem Celular Tumoral , Feminino , Camundongos Endogâmicos C57BL
9.
Sci Adv ; 10(37): eado5545, 2024 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-39270020

RESUMO

Inositol 1,4,5-trisphosphate (IP3) receptor type 1 (ITPR1), 2 (ITPR2), and 3 (ITPR3) encode the IP3 receptor (IP3R), a key player in intracellular calcium release. In four unrelated patients, we report that an identical ITPR3 de novo variant-NM_002224.3:c.7570C>T, p.Arg2524Cys-causes, through a dominant-negative effect, a complex multisystemic disorder with immunodeficiency. This leads to defective calcium homeostasis, mitochondrial malfunction, CD4+ lymphopenia, a quasi-absence of naïve CD4+ and CD8+ cells, an increase in memory cells, and a distinct TCR repertoire. The calcium defect was recapitulated in Jurkat knock-in. Site-directed mutagenesis displayed the exquisite sensitivity of Arg2524 to any amino acid change. Despite the fact that all patients had severe immunodeficiency, they also displayed variable multisystemic involvements, including ectodermal dysplasia, Charcot-Marie-Tooth disease, short stature, and bone marrow failure. In conclusion, unlike previously reported ITPR1-3 deficiencies leading to narrow, mainly neurological phenotypes, a recurrent dominant ITPR3 variant leads to a multisystemic disease, defining a unique role for IP3R3 in the tetrameric IP3R complex.


Assuntos
Receptores de Inositol 1,4,5-Trifosfato , Humanos , Receptores de Inositol 1,4,5-Trifosfato/genética , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Masculino , Feminino , Cálcio/metabolismo , Criança , Mutação , Células Jurkat , Pré-Escolar , Genes Dominantes , Linhagem , Fenótipo
10.
Ann Rheum Dis ; 72(6): 1071-9, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23087182

RESUMO

OBJECTIVE: To evaluate whether miR-20a belonging to the cluster miR-17-92 is a negative regulator of inflammation in rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS) by modulating expression of apoptosis signal-regulating kinase (ASK) 1, a key component of the toll-like receptors 4 pathway, upstream of p38 mitogen-activated protein kinase. METHODS: Evaluation of miR-20a and ASK1 mRNA was performed by RT-qPCR. ASK1 protein expression was assessed by western blotting. Overexpression of miR-20a was performed by transfection of RA FLS and THP-1 cells with miR-20a mimics. Interleukin (IL)-6, CXCL-10, IL-1ß and TNF-α release were measured by ELISA. The role of miR-20a in vivo was assessed by IL-6 release from macrophages obtained from mice injected intraperitoneally with vectorised miR-20a mimics. RESULTS: We showed that stimulation of RA FLS with lipopolysacharide (LPS) and bacterial lipoproteins (BLP) induces a drop in expression of miR-20a and that this decrease is associated with an upregulation of ASK1 expression. Using transfection of Ask1 3'UTR reporters, we demonstrate that Ask1 is a direct target of miR-20a. Overexpression of miR-20a led to a global decrease in ASK1 protein in BLP- and LPS-activated cells indicating that miR-20a regulates the expression of ASK1 at the translational level. Transfection of miR-20a mimics decreases IL-6 and CXCL10 release by RA FLS and IL-1ß and TNF-α by activated THP-1 cells but only in response to LPS. Last, injection of vectorised miR-20a mimics to mice led to a global decrease in ASK1 protein expression and IL-6 secretion in LPS-activated macrophages. CONCLUSIONS: Our data point toward an important role for miR-20a in the regulation of pro-inflammatory cytokines release, by controlling ASK1 expression in RA FLS.


Assuntos
Artrite Reumatoide/fisiopatologia , Citocinas/metabolismo , MAP Quinase Quinase Quinase 5/metabolismo , Macrófagos/metabolismo , MicroRNAs/fisiologia , RNA Mensageiro/análise , Membrana Sinovial/citologia , Animais , Artrite Reumatoide/metabolismo , Células Cultivadas , Células HEK293 , Humanos , Camundongos , Membrana Sinovial/metabolismo , Membrana Sinovial/fisiopatologia
11.
STAR Protoc ; 4(3): 102397, 2023 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-37393615

RESUMO

Adipose tissue plays a central role in age-related diseases. While RNAseq protocols exist for many tissues, few data have been generated with this technology to explore gene expression in adipocytes, particularly during aging. Here, we present a protocol to analyze the transcriptional changes that occur in adipose tissue during normal and accelerated aging in mouse models. We describe steps for genotyping, diet control, euthanasia, and dissection. We then detail RNA purification and genome-wide data generation and analysis. For complete details on the use and execution of this protocol, please refer to De Cauwer et al. (2022) iScience. Sep 16;25(10):105149.


Assuntos
Adipócitos , Transcriptoma , Animais , Camundongos , Transcriptoma/genética , Tecido Adiposo , Envelhecimento/genética , Modelos Animais de Doenças
12.
iScience ; 25(10): 105149, 2022 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-36185376

RESUMO

Age-related diseases are major concern in developed countries. To avoid disabilities that accompany increased lifespan, pharmaceutical approaches are considered. Therefore, appropriate animal models are required for a better understanding of aging processes and potential in vivo assays to evaluate the impact of molecules that may delay the occurrence of age-related diseases. Few mouse models exhibiting pathological aging exist, but currently, none of them reproducibly mimics human diseases like osteoporosis, cognitive dysfunctions or sarcopenia that can be seen in some, but not all, elders. Here, we describe the premature aging phenotypes of Dicer-deficient mature animals, which exhibit an overall deterioration of many organs and tissues (skin, heart, and adipose tissue) ultimately leading to a significant reduction of their lifespan. Molecular characterization of transcriptional responses focused on the adipose tissue suggested that both canonical and non-canonical functions of DICER are involved in this process and highlight potential actionable pathways to revert it.

13.
Nat Med ; 28(5): 989-998, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35288692

RESUMO

The identity of histocompatibility loci, besides human leukocyte antigen (HLA), remains elusive. The major histocompatibility complex (MHC) class I MICA gene is a candidate histocompatibility locus. Here, we investigate its role in a French multicenter cohort of 1,356 kidney transplants. MICA mismatches were associated with decreased graft survival (hazard ratio (HR), 2.12; 95% confidence interval (CI): 1.45-3.11; P < 0.001). Both before and after transplantation anti-MICA donor-specific antibodies (DSA) were strongly associated with increased antibody-mediated rejection (ABMR) (HR, 3.79; 95% CI: 1.94-7.39; P < 0.001; HR, 9.92; 95% CI: 7.43-13.20; P < 0.001, respectively). This effect was synergetic with that of anti-HLA DSA before and after transplantation (HR, 25.68; 95% CI: 3.31-199.41; P = 0.002; HR, 82.67; 95% CI: 33.67-202.97; P < 0.001, respectively). De novo-developed anti-MICA DSA were the most harmful because they were also associated with reduced graft survival (HR, 1.29; 95% CI: 1.05-1.58; P = 0.014). Finally, the damaging effect of anti-MICA DSA on graft survival was confirmed in an independent cohort of 168 patients with ABMR (HR, 1.71; 95% CI: 1.02-2.86; P = 0.041). In conclusion, assessment of MICA matching and immunization for the identification of patients at high risk for transplant rejection and loss is warranted.


Assuntos
Transplante de Rim , Rejeição de Enxerto/genética , Sobrevivência de Enxerto/genética , Antígenos de Histocompatibilidade Classe I/genética , Humanos
14.
Sci Transl Med ; 14(628): eabj7521, 2022 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-34698500

RESUMO

The drivers of critical coronavirus disease 2019 (COVID-19) remain unknown. Given major confounding factors such as age and comorbidities, true mediators of this condition have remained elusive. We used a multi-omics analysis combined with artificial intelligence in a young patient cohort where major comorbidities were excluded at the onset. The cohort included 47 "critical" (in the intensive care unit under mechanical ventilation) and 25 "non-critical" (in a non-critical care ward) patients with COVID-19 and 22 healthy individuals. The analyses included whole-genome sequencing, whole-blood RNA sequencing, plasma and blood mononuclear cell proteomics, cytokine profiling, and high-throughput immunophenotyping. An ensemble of machine learning, deep learning, quantum annealing, and structural causal modeling were used. Patients with critical COVID-19 were characterized by exacerbated inflammation, perturbed lymphoid and myeloid compartments, increased coagulation, and viral cell biology. Among differentially expressed genes, we observed up-regulation of the metalloprotease ADAM9. This gene signature was validated in a second independent cohort of 81 critical and 73 recovered patients with COVID-19 and was further confirmed at the transcriptional and protein level and by proteolytic activity. Ex vivo ADAM9 inhibition decreased severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) uptake and replication in human lung epithelial cells. In conclusion, within a young, otherwise healthy, cohort of individuals with COVID-19, we provide the landscape of biological perturbations in vivo where a unique gene signature differentiated critical from non-critical patients. We further identified ADAM9 as a driver of disease severity and a candidate therapeutic target.


Assuntos
COVID-19 , Proteínas ADAM , Inteligência Artificial , Humanos , Unidades de Terapia Intensiva , Proteínas de Membrana , Respiração Artificial , SARS-CoV-2
15.
RMD Open ; 7(3)2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34725261

RESUMO

OBJECTIVE: To explore at the molecular level the phenotype of a patient suffering an autoinflammatory syndrome which was diagnosed as familial cold autoinflammatory syndrome type 2 (FCAS-2). To explore the functions of Nlrp12 in inflammation using mouse models. METHODS: Whole exome sequencing and Nlrp12 targeted resequencing were performed on DNA isolated from the patient and her family members. In vivo and ex vivo models of inflammation (urate crystals-dependent acute joint inflammation and urate crystals-induced peritonitis) were analysed in Nlrp12-deficient and Nlrp12-competent mice. RESULTS: A rare missense NLRP12 variant (c.857C>T, p.P286L) was identified in the patient and her healthy relatives. Nlrp12-deficient mice exhibit reduced systemic inflammation and neutrophilic infiltration. CONCLUSION: Nlrp12 mediates proinflammatory functions in mice. In humans, the identification of Nlrp12 variants must be cautiously interpreted depending on clinical and paraclinical data to diagnose FCAS-2.


Assuntos
Artrite , Síndromes Periódicas Associadas à Criopirina , Animais , Síndromes Periódicas Associadas à Criopirina/diagnóstico , Síndromes Periódicas Associadas à Criopirina/genética , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Camundongos , Mutação de Sentido Incorreto , Fenótipo
16.
Sci Rep ; 11(1): 13144, 2021 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-34162963

RESUMO

Tumor progression and metastatic dissemination are driven by cell-intrinsic and biomechanical cues that favor the growth of life-threatening secondary tumors. We recently identified pro-metastatic vascular regions with blood flow profiles that are permissive for the arrest of circulating tumor cells. We have further established that such flow profiles also control endothelial remodeling, which favors extravasation of arrested CTCs. Yet, how shear forces control endothelial remodeling is unknown. In the present work, we aimed at dissecting the cellular and molecular mechanisms driving blood flow-dependent endothelial remodeling. Transcriptomic analysis of endothelial cells revealed that blood flow enhanced VEGFR signaling, among others. Using a combination of in vitro microfluidics and intravital imaging in zebrafish embryos, we now demonstrate that the early flow-driven endothelial response can be prevented upon specific inhibition of VEGFR tyrosine kinase and subsequent signaling. Inhibitory targeting of VEGFRs reduced endothelial remodeling and subsequent metastatic extravasation. These results confirm the importance of VEGFR-dependent endothelial remodeling as a driving force of CTC extravasation and metastatic dissemination. Furthermore, the present work suggests that therapies targeting endothelial remodeling might be a relevant clinical strategy in order to impede metastatic progression.


Assuntos
Endotélio Vascular/fisiologia , Hemorreologia , Migração Transendotelial e Transepitelial , Animais , Animais Geneticamente Modificados , Velocidade do Fluxo Sanguíneo/efeitos dos fármacos , Embrião não Mamífero/irrigação sanguínea , Embrião não Mamífero/fisiologia , Regulação Neoplásica da Expressão Gênica , Ontologia Genética , Células Endoteliais da Veia Umbilical Humana , Humanos , Técnicas In Vitro , Microscopia Intravital , Microfluídica , Microscopia Confocal , Células Neoplásicas Circulantes , Quinazolinas/farmacologia , Quinazolinas/uso terapêutico , RNA Neoplásico/biossíntese , RNA Neoplásico/genética , Transdução de Sinais/fisiologia , Sunitinibe/farmacologia , Sunitinibe/uso terapêutico , Migração Transendotelial e Transepitelial/efeitos dos fármacos , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/fisiologia , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/fisiologia , Peixe-Zebra/embriologia
17.
Leukemia ; 35(5): 1463-1474, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33833385

RESUMO

B-cell receptor (BCR) signaling is crucial for the pathophysiology of most mature B-cell lymphomas/leukemias and has emerged as a therapeutic target whose effectiveness remains limited by the occurrence of mutations. Therefore, deciphering the cellular program activated downstream this pathway has become of paramount importance for the development of innovative therapies. Using an original ex vivo model of BCR-induced proliferation of chronic lymphocytic leukemia cells, we generated 108 temporal transcriptional and proteomic profiles from 1 h up to 4 days after BCR activation. This dataset revealed a structured temporal response composed of 13,065 transcripts and 4027 proteins, comprising a leukemic proliferative signature consisting of 430 genes and 374 proteins. Mathematical modeling of this complex cellular response further highlighted a transcriptional network driven by 14 early genes linked to proteins involved in cell proliferation. This group includes expected genes (EGR1/2, NF-kB) and genes involved in NF-kB signaling modulation (TANK, ROHF) and immune evasion (KMO, IL4I1) that have not yet been associated with leukemic cells proliferation. Our study unveils the BCR-activated proliferative genetic program in primary leukemic cells. This approach combining temporal measurements with modeling allows identifying new putative targets for innovative therapy of lymphoid malignancies and also cancers dependent on ligand-receptor interactions.


Assuntos
Linfócitos B/metabolismo , Proliferação de Células/genética , Leucemia Linfocítica Crônica de Células B/genética , Receptores de Antígenos de Linfócitos B/genética , Idoso , Feminino , Humanos , Leucemia Linfocítica Crônica de Células B/metabolismo , Ativação Linfocitária/genética , Masculino , Pessoa de Meia-Idade , Proteoma/genética , Proteômica/métodos , Transdução de Sinais/genética , Transcrição Gênica/genética
18.
Theranostics ; 10(5): 2158-2171, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32104502

RESUMO

Rationale: The role of Monosodium Urate (MSU) crystals in gout pathophysiology is well described, as is the major impact of IL-1ß in the inflammatory reaction that constitutes the hallmark of the disease. However, despite the discovery of the NLRP3 inflammasome and its role as a Pattern Recognition Receptor linking the detection of a danger signal (MSU) to IL-1ß secretion in vitro, the precise mechanisms leading to joint inflammation in gout patients are still poorly understood. Methods: Acute urate crystal inflammation was obtained by subcutaneous injections of MSU crystals in mice. Symptoms were followed by scoring, cytokine quantification by ELISA and western blot, gene expression by RT-qPCR and RNAseq; Magnetic Resonance Imaging was also used to assess inflammation. Results: We provide an extensive clinical, biological and molecular characterization of an acute uratic inflammation mouse model which accurately mimics human gout. We report the efficacy of topical imiquimod treatment and its impact on Interferon-dependent down modulation of Il-1ß gene expression in this experimental model. Conclusion: Our work reveals several key features of MSU-dependent inflammation and identifies novel therapeutic opportunities for gout patients.


Assuntos
Gota/tratamento farmacológico , Imiquimode/farmacologia , Inflamação/induzido quimicamente , Interleucina-1beta/efeitos dos fármacos , Ácido Úrico/efeitos adversos , Doença Aguda , Adjuvantes Imunológicos/administração & dosagem , Adjuvantes Imunológicos/farmacologia , Adjuvantes Imunológicos/uso terapêutico , Administração Tópica , Animais , Antioxidantes/administração & dosagem , Antioxidantes/efeitos adversos , Citocinas/metabolismo , Modelos Animais de Doenças , Gota/metabolismo , Gota/patologia , Imiquimode/administração & dosagem , Imiquimode/uso terapêutico , Inflamação/diagnóstico , Inflamação/imunologia , Injeções Subcutâneas , Imageamento por Ressonância Magnética/métodos , Camundongos , Camundongos Knockout , Ácido Úrico/administração & dosagem
19.
J Exp Med ; 217(12)2020 12 07.
Artigo em Inglês | MEDLINE | ID: mdl-32766723

RESUMO

The Nck-associated protein 1-like (NCKAP1L) gene, alternatively called hematopoietic protein 1 (HEM-1), encodes a hematopoietic lineage-specific regulator of the actin cytoskeleton. Nckap1l-deficient mice have anomalies in lymphocyte development, phagocytosis, and neutrophil migration. Here we report, for the first time, NCKAP1L deficiency cases in humans. In two unrelated patients of Middle Eastern origin, recessive mutations in NCKAP1L abolishing protein expression led to immunodeficiency, lymphoproliferation, and hyperinflammation with features of hemophagocytic lymphohistiocytosis. Immunophenotyping showed an inverted CD4/CD8 ratio with a major shift of both CD4+ and CD8+ cells toward memory compartments, in line with combined RNA-seq/proteomics analyses revealing a T cell exhaustion signature. Consistent with the core function of NCKAP1L in the reorganization of the actin cytoskeleton, patients' T cells displayed impaired early activation, immune synapse morphology, and leading edge formation. Moreover, knockdown of nckap1l in zebrafish led to defects in neutrophil migration. Hence, NCKAP1L mutations lead to broad immune dysregulation in humans, which could be classified within actinopathies.


Assuntos
Síndromes de Imunodeficiência/complicações , Inflamação/complicações , Transtornos Linfoproliferativos/complicações , Proteínas de Membrana/metabolismo , Actinas/metabolismo , Animais , Degranulação Celular , Proliferação de Células , Criança , Citotoxicidade Imunológica , Família , Feminino , Homozigoto , Humanos , Síndromes de Imunodeficiência/imunologia , Sinapses Imunológicas/metabolismo , Lactente , Inflamação/imunologia , Inflamação/patologia , Ativação Linfocitária/imunologia , Transtornos Linfoproliferativos/imunologia , Masculino , Proteínas de Membrana/química , Proteínas de Membrana/deficiência , Proteínas de Membrana/genética , Mutação/genética , Linhagem , Fenótipo , Síndrome , Peixe-Zebra
20.
Bone Marrow Transplant ; 55(7): 1367-1378, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32286503

RESUMO

Graft-versus-host disease (GVHD) and cytomegalovirus (CMV)-related complications are leading causes of mortality after unrelated-donor hematopoietic cell transplantation (UD-HCT). The non-conventional MHC class I gene MICB, alike MICA, encodes a stress-induced polymorphic NKG2D ligand. However, unlike MICA, MICB interacts with the CMV-encoded UL16, which sequestrates MICB intracellularly, leading to immune evasion. Here, we retrospectively analyzed the impact of mismatches in MICB amino acid position 98 (MICB98), a key polymorphic residue involved in UL16 binding, in 943 UD-HCT pairs who were allele-matched at HLA-A, -B, -C, -DRB1, -DQB1 and MICA loci. HLA-DP typing was further available. MICB98 mismatches were significantly associated with an increased incidence of acute (grade II-IV: HR, 1.20; 95% CI, 1.15 to 1.24; P < 0.001; grade III-IV: HR, 2.28; 95% CI, 1.56 to 3.34; P < 0.001) and chronic GVHD (HR, 1.21; 95% CI, 1.10 to 1.33; P < 0.001). MICB98 matching significantly reduced the effect of CMV status on overall mortality from a hazard ratio of 1.77 to 1.16. MICB98 mismatches showed a GVHD-independent association with a higher incidence of CMV infection/reactivation (HR, 1.84; 95% CI, 1.34 to 2.51; P < 0.001). Hence selecting a MICB98-matched donor significantly reduces the GVHD incidence and lowers the impact of CMV status on overall survival.


Assuntos
Infecções por Citomegalovirus , Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Aminoácidos , Infecções por Citomegalovirus/epidemiologia , Infecções por Citomegalovirus/prevenção & controle , Doença Enxerto-Hospedeiro/etiologia , Doença Enxerto-Hospedeiro/prevenção & controle , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Humanos , Incidência , Estudos Retrospectivos
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