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1.
Clin Immunol ; 255: 109732, 2023 10.
Artículo en Inglés | MEDLINE | ID: mdl-37562721

RESUMEN

Subcutaneous panniculitis-like T-cell lymphoma (SPTCL) is a rare primary cutaneous non-Hodgkin lymphoma involving CD8+ T cells, the genetic underpinnings of which remain incompletely understood. Here we report two unrelated patients with B cell Expansion with NF-κB and T cell Anergy (BENTA) disease and a novel presentation of SPTCL. Patient 1 presented early in life with recurrent infections and B cell lymphocytosis, linked to a novel gain-of-function (GOF) CARD11 mutation (p.Lys238del). He developed SPTCL-like lesions and membranoproliferative glomerulonephritis by age 2, treated successfully with cyclosporine. Patient 2 presented at 13 months with splenomegaly, lymphadenopathy, and SPTCL with evidence of hemophagocytic lymphohistiocytosis. Genetic analysis revealed two in cis germline GOF CARD11 variants (p.Glu121Asp/p.Gly126Ser). Autologous bone marrow transplant resulted in SPTCL remission despite persistent B cell lymphocytosis. These cases illuminate an unusual pathological manifestation for BENTA disease, suggesting that CARD11 GOF mutations can manifest in cutaneous CD4+and CD8+ T cell malignancies.


Asunto(s)
Síndromes de Inmunodeficiencia , Linfocitosis , Linfoma de Células T , Paniculitis , Masculino , Humanos , Preescolar , Linfocitos T CD8-positivos/patología , Paniculitis/genética , Paniculitis/patología , Paniculitis/terapia , Linfoma de Células T/genética , Linfoma de Células T/terapia
2.
J Infect Dis ; 224(12): 2010-2019, 2021 12 15.
Artículo en Inglés | MEDLINE | ID: mdl-34673956

RESUMEN

BACKGROUND: Characterizing the longevity and quality of cellular immune responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) enhances understanding of coronavirus disease 2019 (COVID-19) immunity that influences clinical outcomes. Prior studies suggest SARS-CoV-2-specific T cells are present in peripheral blood 10 months after infection. Analysis of the function, durability, and diversity of cellular response long after natural infection, over a range of ages and disease phenotypes, is needed to identify preventative and therapeutic interventions. METHODS: We identified participants in our multisite longitudinal, prospective cohort study 12 months after SARS-CoV-2 infection representing a range of disease severity. We investigated function, phenotypes, and frequency of T cells specific for SARS-CoV-2 using intracellular cytokine staining and spectral flow cytometry, and compared magnitude of SARS-CoV-2-specific antibodies. RESULTS: SARS-CoV-2-specific antibodies and T cells were detected 12 months postinfection. Severe acute illness was associated with higher frequencies of SARS-CoV-2-specific CD4 T cells and antibodies at 12 months. In contrast, polyfunctional and cytotoxic T cells responsive to SARS-CoV-2 were identified in participants over a wide spectrum of disease severity. CONCLUSIONS: SARS-CoV-2 infection induces polyfunctional memory T cells detectable at 12 months postinfection, with higher frequency noted in those who experienced severe disease.


Asunto(s)
COVID-19/inmunología , COVID-19/virología , Memoria Inmunológica , Células T de Memoria , SARS-CoV-2/inmunología , Subgrupos de Linfocitos T/inmunología , Adulto , Anticuerpos Antivirales , Antígenos Virales , Biomarcadores , COVID-19/diagnóstico , COVID-19/epidemiología , Femenino , Humanos , Inmunidad Celular , Estudios Longitudinales , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Índice de Severidad de la Enfermedad , Subgrupos de Linfocitos T/metabolismo , Factores de Tiempo
3.
Cell Immunol ; 353: 104129, 2020 07.
Artículo en Inglés | MEDLINE | ID: mdl-32473470

RESUMEN

BENTA (B cell Expansion with NF-κB and T cell Anergy) is a novel lymphoproliferative disorder caused by germline, gain-of-function (GOF) mutations in the lymphocyte-restricted scaffolding protein CARD11. Similar somatic CARD11 mutations are found in lymphoid malignancies such as diffuse large B cell lymphoma (DLBCL). Normally, antigen receptor (AgR) engagement converts CARD11 into an active conformation that nucleates a signalosome required for IκB kinase (IKK) activation and NF-κB nuclear translocation. However, GOF CARD11 mutants drive constitutive NF-κB activity without AgR stimulation. Here we show that unlike wild-type CARD11, GOF CARD11 mutants can form large, peculiar cytosolic protein aggregates we term mCADS (mutant CARD11 dependent shells). MALT1 and phospho-IKK are reliably colocalized with mCADS, indicative of active signaling. Moreover, endogenous mCADS are detectable in ABC-DLBCL lines harboring similar GOF CARD11 mutations. The unique aggregation potential of GOF CARD11 mutants may represent a novel therapeutic target for treating BENTA or DLBCL.


Asunto(s)
Proteínas Adaptadoras de Señalización CARD/genética , Agregación Celular/fisiología , Mutación con Ganancia de Función , Guanilato Ciclasa/genética , Proteínas Reguladoras de la Apoptosis/metabolismo , Proteínas Adaptadoras de Señalización CARD/metabolismo , Guanilato Ciclasa/metabolismo , Humanos , Quinasa I-kappa B/metabolismo , Células Jurkat , Activación de Linfocitos , Linfocitos/metabolismo , Linfocitos/fisiología , Linfoma de Células B Grandes Difuso/genética , Linfoma de Células B Grandes Difuso/metabolismo , Linfoma de Células B Grandes Difuso/patología , Proteína 1 de la Translocación del Linfoma del Tejido Linfático Asociado a Mucosas/metabolismo , FN-kappa B/metabolismo , Transducción de Señal
4.
J Allergy Clin Immunol ; 143(4): 1482-1495, 2019 04.
Artículo en Inglés | MEDLINE | ID: mdl-30170123

RESUMEN

BACKGROUND: Caspase activation and recruitment domain 11 (CARD11) encodes a scaffold protein in lymphocytes that links antigen receptor engagement with downstream signaling to nuclear factor κB, c-Jun N-terminal kinase, and mechanistic target of rapamycin complex 1. Germline CARD11 mutations cause several distinct primary immune disorders in human subjects, including severe combined immune deficiency (biallelic null mutations), B-cell expansion with nuclear factor κB and T-cell anergy (heterozygous, gain-of-function mutations), and severe atopic disease (loss-of-function, heterozygous, dominant interfering mutations), which has focused attention on CARD11 mutations discovered by using whole-exome sequencing. OBJECTIVES: We sought to determine the molecular actions of an extended allelic series of CARD11 and to characterize the expanding range of clinical phenotypes associated with heterozygous CARD11 loss-of-function alleles. METHODS: Cell transfections and primary T-cell assays were used to evaluate signaling and function of CARD11 variants. RESULTS: Here we report on an expanded cohort of patients harboring novel heterozygous CARD11 mutations that extend beyond atopy to include other immunologic phenotypes not previously associated with CARD11 mutations. In addition to (and sometimes excluding) severe atopy, heterozygous missense and indel mutations in CARD11 presented with immunologic phenotypes similar to those observed in signal transducer and activator of transcription 3 loss of function, dedicator of cytokinesis 8 deficiency, common variable immunodeficiency, neutropenia, and immune dysregulation, polyendocrinopathy, enteropathy, X-linked-like syndrome. Pathogenic variants exhibited dominant negative activity and were largely confined to the CARD or coiled-coil domains of the CARD11 protein. CONCLUSION: These results illuminate a broader phenotypic spectrum associated with CARD11 mutations in human subjects and underscore the need for functional studies to demonstrate that rare gene variants encountered in expected and unexpected phenotypes must nonetheless be validated for pathogenic activity.


Asunto(s)
Proteínas Adaptadoras de Señalización CARD/genética , Proteínas Adaptadoras de Señalización CARD/inmunología , Guanilato Ciclasa/genética , Guanilato Ciclasa/inmunología , Enfermedades del Sistema Inmune/genética , Enfermedades del Sistema Inmune/inmunología , Adulto , Femenino , Humanos , Masculino , Mutación , Fenotipo
5.
PLoS Pathog ; 11(8): e1005102, 2015 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-26285145

RESUMEN

Human T lymphotropic virus type 1 (HTLV-1) trans-activator/oncoprotein, Tax, impacts a multitude of cellular processes, including I-κB kinase (IKK)/NF-κB signaling, DNA damage repair, and mitosis. These activities of Tax have been implicated in the development of adult T-cell leukemia (ATL) in HTLV-1-infected individuals, but the underlying mechanisms remain obscure. IKK and its upstream kinase, TGFß-activated kinase 1 (TAK1), contain ubiquitin-binding subunits, NEMO and TAB2/3 respectively, which interact with K63-linked polyubiquitin (K63-pUb) chains. Recruitment to K63-pUb allows cross auto-phosphorylation and activation of TAK1 to occur, followed by TAK1-catalyzed IKK phosphorylation and activation. Using cytosolic extracts of HeLa and Jurkat T cells supplemented with purified proteins we have identified ubiquitin E3 ligase, ring finger protein 8 (RNF8), and E2 conjugating enzymes, Ubc13:Uev1A and Ubc13:Uev2, to be the cellular factors utilized by Tax for TAK1 and IKK activation. In vitro, the combination of Tax and RNF8 greatly stimulated TAK1, IKK, IκBα and JNK phosphorylation. In vivo, RNF8 over-expression augmented while RNF8 ablation drastically reduced canonical NF-κB activation by Tax. Activation of the non-canonical NF-κB pathway by Tax, however, is unaffected by the loss of RNF8. Using purified components, we further demonstrated biochemically that Tax greatly stimulated RNF8 and Ubc13:Uev1A/Uev2 to assemble long K63-pUb chains. Finally, co-transfection of Tax with increasing amounts of RNF8 greatly induced K63-pUb assembly in a dose-dependent manner. Thus, Tax targets RNF8 and Ubc13:Uev1A/Uev2 to promote the assembly of K63-pUb chains, which signal the activation of TAK1 and multiple downstream kinases including IKK and JNK. Because of the roles RNF8 and K63-pUb chains play in DNA damage repair and cytokinesis, this mechanism may also explain the genomic instability of HTLV-1-transformed T cells and ATL cells.


Asunto(s)
Transformación Celular Neoplásica/metabolismo , Productos del Gen tax/metabolismo , Transducción de Señal/fisiología , Línea Celular , Proteínas de Unión al ADN/metabolismo , Técnica del Anticuerpo Fluorescente , Técnicas de Silenciamiento del Gen , Humanos , Quinasa I-kappa B/metabolismo , Immunoblotting , Quinasas Quinasa Quinasa PAM/metabolismo , Reacción en Cadena de la Polimerasa , Transfección , Ubiquitina-Proteína Ligasas/metabolismo
6.
J Thromb Haemost ; 2024 Sep 02.
Artículo en Inglés | MEDLINE | ID: mdl-39233012

RESUMEN

BACKGROUND: Almost half of severe hemophilia A (HA) cases are caused by an intron 22 inversion (Int22Inv) mutation, which truncates the 26-exon F8 messenger RNA (mRNA) after exon 22. Another F8 transcript, F8B, is initiated from within F8-intron-22. F8B mRNA consists of a short exon spliced to exons 23 to 26 and is expressed in multiple human cell types. It has been hypothesized that Int22Inv patients have self-tolerance to partial factor (F)VIII proteins expressed from these 2 transcripts. FVIII is expressed in endothelial cells, primarily in the liver and lungs. Several studies have reported FVIII expression in other cell types, although this has been controversial. OBJECTIVES: To determine if partial FVIII proteins are expressed from intron 22-inverted and/or F8B mRNA and if FVIII is expressed in nonendothelial cells. METHODS: A panel of FVIII-specific antibodies was validated and employed to label FVIII in cells and tissues and for immunoprecipitation followed by western blots and mass spectrometry proteomics analysis. RESULTS: Immunofluorescent staining localized FVIII to endothelial cells in liver sections from non-HA but not HA-Int22Inv dogs. Neither FVIII nor FVIIIB was detected in human peripheral blood mononuclear cells, B cell or T cell lines, or cell lines expanded from peripheral blood mononuclear cells, whereas FVIII antigen and activity were readily detected in primary nonhemophilic liver sinusoidal endothelial cells. CONCLUSION: If FVIII is expressed in nonendothelial cells or if partial FVIII proteins are expressed in HA-Int22Inv, the concentrations are below the detection limits of these sensitive assays. Our results argue against promotion of immune tolerance through expression of partial FVIII proteins in Int-22Inv patients.

7.
J Biol Chem ; 286(36): 31092-104, 2011 Sep 09.
Artículo en Inglés | MEDLINE | ID: mdl-21724848

RESUMEN

Human T-cell leukemia virus type 1 (HTLV-1) has two late domain (LD) motifs, PPPY and PTAP, which are important for viral budding. Mutations in the PPPY motif are more deleterious for viral release than changes in the PTAP motif. Several reports have shown that the interaction of PPPY with the WW domains of a Nedd4 (neuronal precursor cell-expressed developmentally down-regulated-4) family ubiquitin ligase (UL) is a critical event in virus release. We tested nine members of the Nedd4 family ULs and found that ITCH is the main contributor to HTLV-1 budding. ITCH overexpression strongly inhibited release and infectivity of wild-type (wt) HTLV-1, but rescued the release of infectious virions with certain mutations in the PPPY motif. Electron microscopy showed either fewer or misshapen virus particles when wt HTLV-1 was produced in the presence of overexpressed ITCH, whereas mutants with changes in the PPPY motif yielded normal looking particles at wt level. The other ULs had significantly weaker or no effects on HTLV-1 release and infectivity except for SMURF-1, which caused enhanced release of wt and all PPPY(-) mutant particles. These particles were poorly infectious and showed abnormal morphology by electron microscopy. Budding and infectivity defects due to overexpression of ITCH and SMURF-1 were correlated with higher than normal ubiquitination of Gag. Only silencing of ITCH, but not of WWP1, WWP2, and Nedd4, resulted in a reduction of HTLV-1 budding from 293T cells. The binding efficiencies between the HTLV-1 LD and WW domains of different ULs as measured by mammalian two-hybrid interaction did not correlate with the strength of their effect on HTLV-1 budding.


Asunto(s)
Virus Linfotrópico T Tipo 1 Humano/metabolismo , Proteínas Represoras/fisiología , Ubiquitina-Proteína Ligasas/fisiología , Liberación del Virus , Secuencias de Aminoácidos/genética , Línea Celular , Complejos de Clasificación Endosomal Requeridos para el Transporte , Productos del Gen gag/metabolismo , Virus Linfotrópico T Tipo 1 Humano/genética , Humanos , Microscopía Electrónica , Ubiquitina-Proteína Ligasas Nedd4 , Ubiquitinación
8.
Front Immunol ; 13: 878989, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35651609

RESUMEN

Background: Atopic dermatitis (AD) affects up to 25% of children and 10% of adults in Western countries. When severe or recurrent infections and exceedingly elevated serum IgE levels occur in AD patients, an inborn error of immunity (IEI) may be suspected. The International Union of Immunological Societies classification lists variants in different genes responsible for so-called Hyper-IgE syndromes. Diagnosing an underlying IEI may influence treatment strategies. Methods: Clinical and diagnostic workup of family members are presented including a detailed immunological description and histology of the carcinoma. Functional testing of the novel variant in CARD11 underlying 'CARD11-associated atopy with dominant interference of NF-kB signaling' (CADINS) was performed. Results: We report on an 18-year-old patient with a long-standing history of infections, accompanied by hypogammaglobulinemia, intermittent agranulocytosis, atopy, eosinophilia and colitis. The working diagnosis of common variable immunodeficiency was revised when a novel heterozygous CARD11 variant [c.223C>T; p.(Arg75Trp)] was identified. Functional studies confirmed this variant to have a dominant negative (DN) effect, as previously described in patients with CADINS. Five other family members were affected by severe atopy associated with the above variant, but not hypogammaglobulinemia. Malignancies occurred in two generations: an HPV-positive squamous cell carcinoma and a cutaneous T-cell lymphoma. So far, one patient is under treatment with dupilumab, which has shown marked benefit in controlling severe eczema. Conclusion: The phenotypic spectrum associated with heterozygous CARD11 DN mutations is broad. Partial T-cell deficiency, diminished IFN-γ cytokine and increased IL-4 production, were identified as disease-causing mechanisms. Malignant disease associated with germline CARD11 DN variants has only been reported sporadically. HPV vaccination in teenage years, and cytology screening analogous with routine cervical swabs may be recommended. Treatment with dupilumab, a monoclonal antibody blocking interleukin-4- and interleukin-13 signaling, may be of benefit in controlling severe and extended AD for some patients as reported for STAT3 loss-of-function.


Asunto(s)
Carcinoma , Dermatitis Atópica , Síndrome de Job , Infecciones por Papillomavirus , Adolescente , Adulto , Niño , Dermatitis Atópica/tratamiento farmacológico , Dermatitis Atópica/genética , Humanos , Inmunoglobulina E , Síndrome de Job/diagnóstico , Síndrome de Job/genética
9.
Cell Mol Immunol ; 18(1): 194-205, 2021 01.
Artículo en Inglés | MEDLINE | ID: mdl-31659245

RESUMEN

The adaptive immune response relies on specific apoptotic programs to maintain homeostasis. Conventional effector T cell (Tcon) expansion is constrained by both forkhead box P3 (FOXP3)+-regulatory T cells (Tregs) and restimulation-induced cell death (RICD), a propriocidal apoptosis pathway triggered by repeated stimulation through the T-cell receptor (TCR). Constitutive FOXP3 expression protects Tregs from RICD by suppressing SLAM-associated protein (SAP), a key adaptor protein that amplifies TCR signaling strength. The role of transient FOXP3 induction in activated human CD4 and CD8 Tcons remains unresolved, but its expression is inversely correlated with acquired RICD sensitivity. Here, we describe a novel role for FOXP3 in protecting human Tcons from premature RICD during expansion. Unlike FOXP3-mediated protection from RICD in Tregs, FOXP3 protects Tcons through a distinct mechanism requiring de novo transcription that does not require SAP suppression. Transcriptome profiling and functional analyses of expanding Tcons revealed that FOXP3 enhances expression of the SLAM family receptor CD48, which in turn sustains basal autophagy and suppresses pro-apoptotic p53 signaling. Both CD48 and FOXP3 expression reduced p53 accumulation upon TCR restimulation. Furthermore, silencing FOXP3 expression or blocking CD48 decreased the mitochondrial membrane potential in expanding Tcons with a concomitant reduction in basal autophagy. Our findings suggest that FOXP3 governs a distinct transcriptional program in early-stage effector Tcons that maintains RICD resistance via CD48-dependent protective autophagy and p53 suppression.


Asunto(s)
Antígeno CD48/metabolismo , Muerte Celular , Factores de Transcripción Forkhead/metabolismo , Activación de Linfocitos , Receptores de Antígenos de Linfocitos T/metabolismo , Proteína Asociada a la Molécula de Señalización de la Activación Linfocitaria/metabolismo , Linfocitos T Reguladores/inmunología , Apoptosis , Autofagia , Antígeno CD48/genética , Factores de Transcripción Forkhead/genética , Humanos , Receptores de Antígenos de Linfocitos T/genética , Transducción de Señal , Proteína Asociada a la Molécula de Señalización de la Activación Linfocitaria/genética , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo
10.
Front Immunol ; 9: 2078, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30283440

RESUMEN

The caspase recruitment domain family member 11 (CARD11 or CARMA1)-B cell CLL/lymphoma 10 (BCL10)-MALT1 paracaspase (MALT1) [CBM] signalosome complex serves as a molecular bridge between cell surface antigen receptor signaling and the activation of the NF-κB, JNK, and mTORC1 signaling axes. This positions the CBM complex as a critical regulator of lymphocyte activation, proliferation, survival, and metabolism. Inborn errors in each of the CBM components have now been linked to a diverse group of human primary immunodeficiency diseases termed "CBM-opathies." Clinical manifestations range from severe combined immunodeficiency to selective B cell lymphocytosis, atopic disease, and specific humoral defects. This surprisingly broad spectrum of phenotypes underscores the importance of "tuning" CBM signaling to preserve immune homeostasis. Here, we review the distinct clinical and immunological phenotypes associated with human CBM complex mutations and introduce new avenues for targeted therapeutic intervention.


Asunto(s)
Proteína 10 de la LLC-Linfoma de Células B/inmunología , Proteínas Adaptadoras de Señalización CARD/inmunología , Guanilato Ciclasa/inmunología , Proteína 1 de la Translocación del Linfoma del Tejido Linfático Asociado a Mucosas/inmunología , Inmunodeficiencia Combinada Grave/inmunología , Transducción de Señal/inmunología , Proteína 10 de la LLC-Linfoma de Células B/genética , Proteína 10 de la LLC-Linfoma de Células B/metabolismo , Proteínas Adaptadoras de Señalización CARD/genética , Proteínas Adaptadoras de Señalización CARD/metabolismo , Guanilato Ciclasa/genética , Guanilato Ciclasa/metabolismo , Homeostasis/genética , Homeostasis/inmunología , Humanos , Activación de Linfocitos/genética , Activación de Linfocitos/inmunología , Proteína 1 de la Translocación del Linfoma del Tejido Linfático Asociado a Mucosas/genética , Proteína 1 de la Translocación del Linfoma del Tejido Linfático Asociado a Mucosas/metabolismo , Mutación/inmunología , Unión Proteica , Inmunodeficiencia Combinada Grave/genética , Inmunodeficiencia Combinada Grave/metabolismo , Transducción de Señal/genética
11.
Front Immunol ; 9: 2944, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30619304

RESUMEN

CARD11 is a lymphocyte-specific scaffold molecule required for proper activation of B- and T-cells in response to antigen. Germline gain-of-function (GOF) mutations in the CARD11 gene cause a unique B cell lymphoproliferative disorder known as B cell Expansion with NF-κB and T cell Anergy (BENTA). In contrast, patients carrying loss-of-function (LOF), dominant negative (DN) CARD11 mutations present with severe atopic disease. Interestingly, both GOF and DN CARD11 variants cause primary immunodeficiency, with recurrent bacterial and viral infections, likely resulting from impaired adaptive immune responses. This report describes a unique four-generation family harboring a novel heterozygous germline indel mutation in CARD11 (c.701-713delinsT), leading to one altered amino acid and a deletion of 4 others (p.His234_Lys238delinsLeu). Strikingly, affected members exhibit both moderate B cell lymphocytosis and atopic dermatitis/allergies. Ectopic expression of this CARD11 variant stimulated constitutive NF-κB activity in T cell lines, similar to other BENTA patient mutations. However, unlike other GOF mutants, this variant significantly impeded the ability of wild-type CARD11 to induce NF-κB activation following antigen receptor ligation. Patient lymphocytes display marked intrinsic defects in B cell differentiation and reduced T cell responsiveness in vitro. Collectively, these data imply that a single heterozygous CARD11 mutation can convey both GOF and DN signaling effects, manifesting in a blended BENTA phenotype with atopic features. Our findings further emphasize the importance of balanced CARD11 signaling for normal immune responses.


Asunto(s)
Proteínas Adaptadoras de Señalización CARD/genética , Mutación con Ganancia de Función , Mutación de Línea Germinal , Guanilato Ciclasa/genética , Síndromes de Inmunodeficiencia/genética , Trastornos Linfoproliferativos/genética , Linfocitos B/inmunología , Linfocitos B/metabolismo , Secuencia de Bases , Proteínas Adaptadoras de Señalización CARD/metabolismo , Salud de la Familia , Femenino , Guanilato Ciclasa/metabolismo , Humanos , Síndromes de Inmunodeficiencia/metabolismo , Síndromes de Inmunodeficiencia/patología , Lactante , Trastornos Linfoproliferativos/patología , Masculino , FN-kappa B/metabolismo , Linaje , Linfocitos T/inmunología , Linfocitos T/metabolismo
12.
Oncotarget ; 8(69): 113583-113597, 2017 Dec 26.
Artículo en Inglés | MEDLINE | ID: mdl-29371931

RESUMEN

Previously, we have demonstrated that progesterone and calcitriol synergistically inhibit growth of endometrial and ovarian cancer by enhancing apoptosis and causing cell cycle arrest. Metastasis is the main reason of mortality in cancer patients. Activation of ADP-Ribosylation Factor 6 (ARF6), Neural Precursor cell expressed Developmentally Downregulated 9 (NEDD9), and Membrane-Type-1 Matrix Metalloproteinase (MT1-MMP) have been implicated in promoting tumor growth and metastasis. We examined the effects of progesterone, calcitriol and progesterone-calcitriol combination on metastasis promoting proteins in endometrial cancer. Expression of ARF6, NEDD9, and MT1-MMP was enhanced in advanced-stage endometrial tumors and in cancer cell lines compared to normal tissues and immortalized EM-E6/E7-TERT endometrial epithelial cells. Knockdown of these proteins significantly inhibited the invasiveness of the cancer cells. The expression levels of all three proteins was reduced with progesterone and progesterone-calcitriol combination treatment, whereas calcitriol alone showed no effect on their expression but moderately decreased MT1-MMP activity. Fluorescence microscopy showed membrane expression of MT1-MMP in vehicle and calcitriol-treated endometrial cancer cells. However, progesterone and calcitriol-progesterone combination treatment revealed MT1-MMP in the cytoplasm. Furthermore, progesterone and calcitriol reduced the activity of MT1-MMP, MMP-9, and MMP-2. In addition, invadopodia regulatory proteins were attenuated in both progesterone and progesterone-calcitriol combination treated cells as well as in MT1-MMP knockdown cells. Thus, targeting the aberrant MT1-MMP signaling with progesterone-calcitriol may be a novel approach to impede MT1-MMP mediated cancer dissemination and may have therapeutic benefits for endometrial cancer patients.

13.
Nat Genet ; 49(8): 1192-1201, 2017 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-28628108

RESUMEN

Few monogenic causes for severe manifestations of common allergic diseases have been identified. Through next-generation sequencing on a cohort of patients with severe atopic dermatitis with and without comorbid infections, we found eight individuals, from four families, with novel heterozygous mutations in CARD11, which encodes a scaffolding protein involved in lymphocyte receptor signaling. Disease improved over time in most patients. Transfection of mutant CARD11 expression constructs into T cell lines demonstrated both loss-of-function and dominant-interfering activity upon antigen receptor-induced activation of nuclear factor-κB and mammalian target of rapamycin complex 1 (mTORC1). Patient T cells had similar defects, as well as low production of the cytokine interferon-γ (IFN-γ). The mTORC1 and IFN-γ production defects were partially rescued by supplementation with glutamine, which requires CARD11 for import into T cells. Our findings indicate that a single hypomorphic mutation in CARD11 can cause potentially correctable cellular defects that lead to atopic dermatitis.


Asunto(s)
Proteínas Adaptadoras de Señalización CARD/genética , Dermatitis Atópica/genética , Mutación de Línea Germinal , Guanilato Ciclasa/genética , Sistema de Transporte de Aminoácidos ASC/metabolismo , Estudios de Cohortes , Análisis Mutacional de ADN , Dermatitis Atópica/inmunología , Femenino , Genes Dominantes , Glutamina/metabolismo , Humanos , Células Jurkat , Activación de Linfocitos , Masculino , Diana Mecanicista del Complejo 1 de la Rapamicina , Antígenos de Histocompatibilidad Menor/metabolismo , Complejos Multiproteicos/metabolismo , FN-kappa B/metabolismo , Linaje , Linfocitos T/inmunología , Linfocitos T/metabolismo , Serina-Treonina Quinasas TOR/metabolismo
15.
Oncotarget ; 7(43): 69733-69748, 2016 Oct 25.
Artículo en Inglés | MEDLINE | ID: mdl-27626172

RESUMEN

Nestin, an intermediate filament protein and a stem cell marker is expressed in several tumors. Until recently, little was known about the expression levels and the role of Nestin in endometrial cancer. Compared to the immortalized endometrial epithelial cell line EM-E6/E7-TERT, endometrial cancer cell lines express high to moderate levels of Nestin. Furthermore, endometrial tumors and tumor cell lines have a cancer stem-like cell subpopulation expressing CD133. Among the cancer lines, AN3CA and KLE cells exhibited both a significantly higher number of CD133+ cells and expressed Nestin at higher levels than Ishikawa cells. Knockdown of Nestin in AN3CA and KLE increased cells in G0/G1 phase of the cell cycle, whereas overexpression in Ishikawa decreased cells in G0/G1 phase and increased cells in S-phase. Nestin knockdown cells showed increased p21, p27, and PNCA levels and decreased expression of cyclin-D1 and D3. In contrast, Nestin overexpression revealed an inverse expression pattern of cell cycle regulatory proteins. Nestin knockdown inhibited cancer cell growth and invasive potential by downregulating TGF-ß signaling components, MMP-2, MMP-9, vimentin, SNAIL, SLUG, Twist, N-cadherin, and upregulating the epithelial cell marker E-cadherin whereas the opposite was observed with Nestin overexpressing Ishikawa cells. Nestin knockdown also inhibited, while overexpression promoted invadopodia formation and pFAK expression. Knockdown of Nestin significantly reduced tumor volume in vivo. Finally, progesterone inhibited Nestin expression in endometrial cancer cells. These results suggest that Nestin can be a therapeutic target for cancer treatment.


Asunto(s)
Neoplasias Endometriales/patología , Nestina/fisiología , Transducción de Señal/fisiología , Factor de Crecimiento Transformador beta/fisiología , Cadherinas/análisis , Línea Celular Tumoral , Regulación hacia Abajo , Transición Epitelial-Mesenquimal , Femenino , Puntos de Control de la Fase G1 del Ciclo Celular , Humanos , Invasividad Neoplásica , Progesterona/farmacología
16.
PLoS One ; 8(1): e53138, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-23308151

RESUMEN

Virus transmission can occur either by a cell-free mode through the extracellular space or by cell-to-cell transmission involving direct cell-to-cell contact. The factors that determine whether a virus spreads by either pathway are poorly understood. Here, we assessed the relative contribution of cell-free and cell-to-cell transmission to the spreading of the human immunodeficiency virus (HIV). We demonstrate that HIV can spread by a cell-free pathway if all the steps of the viral replication cycle are efficiently supported in highly permissive cells. However, when the cell-free path was systematically hindered at various steps, HIV transmission became contact-dependent. Cell-to-cell transmission overcame barriers introduced in the donor cell at the level of gene expression and surface retention by the restriction factor tetherin. Moreover, neutralizing antibodies that efficiently inhibit cell-free HIV were less effective against cell-to-cell transmitted virus. HIV cell-to-cell transmission also efficiently infected target T cells that were relatively poorly susceptible to cell-free HIV. Importantly, we demonstrate that the donor and target cell types influence critically the extent by which cell-to-cell transmission can overcome each barrier. Mechanistically, cell-to-cell transmission promoted HIV spread to more cells and infected target cells with a higher proviral content than observed for cell-free virus. Our data demonstrate that the frequently observed contact-dependent spread of HIV is the result of specific features in donor and target cell types, thus offering an explanation for conflicting reports on the extent of cell-to-cell transmission of HIV.


Asunto(s)
Comunicación Celular , Infecciones por VIH/transmisión , VIH/patogenicidad , Anticuerpos Neutralizantes/inmunología , Células Cultivadas , Técnicas de Cocultivo , Regulación Viral de la Expresión Génica , Células HEK293 , VIH/fisiología , Infecciones por VIH/inmunología , Infecciones por VIH/virología , Humanos , Células Jurkat , Linfocitos T/virología , Internalización del Virus
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