RESUMEN
To mount an immune response, T lymphocytes must successfully search for foreign material bound to the surface of antigen-presenting cells. How T cells optimize their chances of encountering and responding to these antigens is unknown. T cell motility in tissues resembles a random or Levy walk and is regulated in part by external factors including chemokines and lymph-node topology, but motility parameters such as speed and propensity to turn may also be cell intrinsic. Here we found that the unconventional myosin 1g (Myo1g) motor generates membrane tension, enforces cell-intrinsic meandering search, and enhances T-DC interactions during lymph-node surveillance. Increased turning and meandering motility, as opposed to ballistic motility, is enhanced by Myo1g. Myo1g acts as a "turning motor" and generates a form of cellular "flânerie." Modeling and antigen challenges show that these intrinsically programmed elements of motility search are critical for the detection of rare cognate antigen-presenting cells.
Asunto(s)
Vigilancia Inmunológica , Miosinas/metabolismo , Linfocitos T/citología , Animales , Células Presentadoras de Antígenos/inmunología , Células Presentadoras de Antígenos/metabolismo , Membrana Celular/metabolismo , Movimiento Celular , Ganglios Linfáticos/inmunología , Ratones , Antígenos de Histocompatibilidad Menor , Miosinas/genética , Receptores de Antígenos de Linfocitos T/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismoRESUMEN
Parasites have been associated with possible anticancer activity, including Trypanosoma cruzi, which has been linked to inhibiting the growth of solid tumors. To better understand this antitumor effect, we investigated the association of anti-T. cruzi antibodies with B cells of the acute lymphoblastic leukemia (ALL) SUPB15 cell line. The antibodies were generated in rabbits. IgGs were purified by affinity chromatography. Two procedures (flow cytometry (CF) and Western blot(WB)) were employed to recognize anti-T. cruzi antibodies on SUPB15 cells. We also used CF to determine whether the anti-T. cruzi antibodies could suppress SUPB15 cells. The anti-T. cruzi antibodies recognized 35.5% of the surface antigens of SUPB15. The complement-dependent cytotoxicity (CDC) results demonstrate the cross-suppression of anti-T. cruzi antibodies on up to 8.4% of SUPB15 cells. For the WB analysis, a band at 100 kDa with high intensity was sequenced using mass spectrometry, identifying the protein as nucleolin. This protein may play a role in the antitumor effect on T. cruzi. The anti-T. cruzi antibodies represent promising polyclonal antibodies that have the effect of tumor-suppressive cross-linking on cancer cells, which should be further investigated.
Asunto(s)
Anticuerpos Antiprotozoarios , Leucemia-Linfoma Linfoblástico de Células Precursoras , Trypanosoma cruzi , Trypanosoma cruzi/inmunología , Leucemia-Linfoma Linfoblástico de Células Precursoras/inmunología , Humanos , Línea Celular Tumoral , Animales , Conejos , Anticuerpos Antiprotozoarios/inmunología , Proteínas de Unión al ARN/inmunología , Proteínas de Unión al ARN/metabolismo , Nucleolina , Fosfoproteínas/inmunología , Fosfoproteínas/metabolismoRESUMEN
Epithelial barrier impairment is a hallmark of several pathologic processes in the gut, including inflammatory bowel diseases. Several intracellular signals prevent apoptosis in intestinal epithelial cells. Herein, we show that in colonocytes, rictor/mammalian target of rapamycin complex 2 (mTORC2) signaling is a prosurvival stimulus. Mechanistically, mTORC2 activates Akt, which, in turn, inhibits apoptosis by phosphorylating B-cell lymphoma 2 (BCL2) associated agonist of cell death (Bad) and preventing caspase-3 activation. Nevertheless, during inflammation, rictor/mTORC2 signaling declines and Akt activity is reduced. Consequently, active caspase-3 increases in surface colonocytes undergoing apoptosis/anoikis and causes epithelial barrier breakdown. Likewise, Rictor ablation in intestinal epithelial cells interrupts mTORC2/Akt signaling and increases apoptosis/anoikis of surface colonocytes without affecting the crypt architecture. The increase in epithelial permeability induced by Rictor ablation produces a mild inflammatory response in the colonic mucosa, but minimally affects the development/establishment of colitis. The data identify a previously unknown mechanism by which rictor/mTORC2 signaling regulates apoptosis/anoikis in intestinal epithelial cells during colitis and clarify its role in the maintenance of the intestinal epithelial barrier.
Asunto(s)
Apoptosis/fisiología , Colitis/patología , Células Epiteliales/metabolismo , Mucosa Intestinal/patología , Proteína Asociada al mTOR Insensible a la Rapamicina/metabolismo , Animales , Colitis/metabolismo , Células Epiteliales/patología , Mucosa Intestinal/metabolismo , Ratones , Transducción de Señal/fisiologíaRESUMEN
Myosin 1g (Myo1g) is a mechanoenzyme associated with actin filaments, expressed exclusively in hematopoietic cells, and involved in various cellular functions, including cell migration, adhesion, and membrane trafficking. Despite the importance of Myo1g in distinct functions, there is currently no monoclonal antibody (mAb) against Myo1g. mAbs are helpful tools for the detection of specific antigens in tumor cells and other tissues. The development of mAbs against targeted dysregulated molecules in cancer cells remains a crucial tool for aiding in the diagnosis and the treatment of patients. Using hybridoma technology, we generated a panel of hybridomas specific for Myo1g. ELISA, immunofluorescence, and Western blot assay results revealed the recognition of Myo1g by these novel monoclonal antibodies in normal and transformed T and B cells. Here, we report the development and application of new monoclonal antibodies against Myo1g for their potential use to detect its overexpression in acute lymphoblastic leukemia (ALL) patients.
Asunto(s)
Anticuerpos Monoclonales , Leucemia Linfoide , Miosinas , Anticuerpos Monoclonales/metabolismo , Niño , Ensayo de Inmunoadsorción Enzimática , Humanos , Hibridomas/metabolismo , Leucemia Linfoide/genética , Leucemia Linfoide/metabolismo , Antígenos de Histocompatibilidad Menor/metabolismo , Miosinas/genética , Miosinas/metabolismoRESUMEN
Yin-Yang transcription factor 1 (YY1) is involved in tumor progression, metastasis and has been shown to be elevated in different cancers, including leukemia. The regulatory mechanism underlying YY1 expression in leukemia is still not understood. Bioinformatics analysis reveal three Hypoxia-inducible factor 1-alpha (HIF-1α) putative binding sites in the YY1 promoter region. The regulation of YY1 by HIF-1α in leukemia was analyzed. Mutation of the putative YY1 binding sites in a reporter system containing the HIF-1α promoter region and CHIP analysis confirmed that these sites are important for YY1 regulation. Leukemia cell lines showed that both proteins HIF-1α and YY1 are co-expressed under hypoxia. In addition, the expression of mRNA of YY1 was increased after 3 h of hypoxia conditions and affect several target genes expression. In contrast, chemical inhibition of HIF-1α induces downregulation of YY1 and sensitizes cells to chemotherapeutic drugs. The clinical implications of HIF-1α in the regulation of YY1 were investigated by evaluation of expression of HIF-1α and YY1 in 108 peripheral blood samples and by RT-PCR in 46 bone marrow samples of patients with pediatric acute lymphoblastic leukemia (ALL). We found that the expression of HIF-1α positively correlates with YY1 expression in those patients. This is consistent with bioinformatic analyses of several databases. Our findings demonstrate for the first time that YY1 can be transcriptionally regulated by HIF-1α, and a correlation between HIF-1α expression and YY1 was found in ALL clinical samples. Hence, HIF-1α and YY1 may be possible therapeutic target and/or biomarkers of ALL.
Asunto(s)
Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Factor de Transcripción YY1/metabolismo , Adolescente , Línea Celular Tumoral , Niño , Preescolar , Regulación Neoplásica de la Expresión Génica , Humanos , Lactante , Recién NacidoRESUMEN
Acute lymphoblastic leukemia (ALL) is the most common pediatric cancer, constituting 80% of all acute leukemias in minors. Despite the increase in the success of therapies, disease-free survival is over 80% in most cases. For the remaining 20% of patients, new strategies are needed to allow us to know and select those at greatest risk of relapse. We evaluated by immunohistochemistry the expression of the transcription factor YY1 and found that it is overexpressed in peripheral blood leukemia cells of pediatric patients with ALL with Pro-B and T phenotype compared to control samples. Over expression of YY1 was associated with a significantly lower chance of survival. We also evaluated by RT-PCR in bone marrow samples from ALL pediatric patients the association of YY1 expression with the percentage of blasts. High levels of YY1 were present in samples with higher percent of blasts in these patients. In addition, ALL pediatric patients with a poor response to therapy had higher levels at the nuclear level of YY1 than those who responded well to chemotherapy. In conclusion, our data suggest that YY1 could serve in pediatric ALL as markers of evolution and response for this disease, mainly in patients with pro-B and T immunophenotype. It is also suggested that YY1 is implicated in the expanse of blast in these patients.
Asunto(s)
Regulación Leucémica de la Expresión Génica , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Regulación hacia Arriba , Factor de Transcripción YY1/genética , Adolescente , Niño , Preescolar , Femenino , Humanos , Inmunofenotipificación , Lactante , Masculino , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Pronóstico , Factor de Transcripción YY1/análisisRESUMEN
Amyloid-beta oligomers (AßO) have been proposed as the most potent neurotoxic and inflammation inducers in Alzheimer's disease (AD). AßO contribute to AD pathogenesis by impairing the production of several cytokines and inflammation-related signaling pathways, such as the Janus kinases/signal transducer of transcription factor-3 (JAK/STAT3) pathway. STAT3 modulates glial activation, indirectly regulates Aß deposition, and induces cognitive decline in AD transgenic models. However, in vivo studies using an AßO microinjection rat model have not yet explored STAT3 role. The main purpose of this study was to elucidate if a single microinjection of AßO could promote an increased expression of STAT3 in glial cells favoring neuroinflammation and neurodegeneration. We designed a model of intrahippocampal microinjection and assessed glial activation, cytokines production, STAT3 expression, and neurodegeneration in time. Our results showed robust expression of STAT3 in glial cells (mainly in astrocytes) and neurons, correlating with neuronal death in response to AßO administration. A STAT3 inhibition assay conducted in rat primary hippocampal cultures, suggested that the induction of the transcription factor by AßO in astrocytes leads them to an activation state that may favor neuronal death. Notwithstanding, pharmacological inhibition of the JAK2/STAT3 pathway should be focused on astrocytes because it is also essential in neurons survival. Overall, these findings strongly suggest the participation of STAT3 in the development of neurodegeneration.
Asunto(s)
Péptidos beta-Amiloides/metabolismo , Astrocitos/metabolismo , Gliosis/etiología , Gliosis/metabolismo , Neuronas/metabolismo , Factor de Transcripción STAT3/metabolismo , Enfermedad de Alzheimer/etiología , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Animales , Astrocitos/patología , Biomarcadores , Muerte Celular , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Técnica del Anticuerpo Fluorescente , Gliosis/patología , Hipocampo/metabolismo , Hipocampo/patología , Inmunohistoquímica , Agregado de Proteínas , Agregación Patológica de Proteínas/genética , Agregación Patológica de Proteínas/metabolismo , Multimerización de Proteína , Ratas , Factor de Transcripción STAT3/genéticaRESUMEN
SLK is essential for embryonic development and may play a key role in wound healing, tumor growth, and metastasis. Expression and activation of SLK are increased in kidney development and during recovery from ischemic acute kidney injury. Overexpression of SLK in glomerular epithelial cells/podocytes in vivo induces injury and proteinuria. Conversely, reduced SLK expression leads to abnormalities in cell adhesion, spreading, and motility. Tight regulation of SLK expression thus may be critical for normal renal structure and function. We produced podocyte-specific SLK-knockout mice to address the functional role of SLK in podocytes. Mice with podocyte-specific deletion of SLK showed reduced glomerular SLK expression and activity compared with control. Podocyte-specific deletion of SLK resulted in albuminuria at 4-5 mo of age in male mice and 8-9 mo in female mice, which persisted for up to 13 mo. At 11-12 mo, knockout mice showed ultrastructural changes, including focal foot process effacement and microvillous transformation of podocyte plasma membranes. Mean foot process width was approximately twofold greater in knockout mice compared with control. Podocyte number was reduced by 35% in knockout mice compared with control, and expression of nephrin, synaptopodin, and podocalyxin was reduced in knockout mice by 20-30%. In summary, podocyte-specific deletion of SLK leads to albuminuria, loss of podocytes, and morphological evidence of podocyte injury. Thus, SLK is essential to the maintenance of podocyte integrity as mice age.
Asunto(s)
Albuminuria/enzimología , Glomérulos Renales/enzimología , Podocitos/enzimología , Proteínas Serina-Treonina Quinasas/metabolismo , Factores de Edad , Albuminuria/genética , Albuminuria/fisiopatología , Animales , Adhesión Celular , Células Cultivadas , Colágeno/metabolismo , Femenino , Predisposición Genética a la Enfermedad , Glomérulos Renales/fisiopatología , Glomérulos Renales/ultraestructura , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Fenotipo , Podocitos/ultraestructura , Proteínas Serina-Treonina Quinasas/deficiencia , Proteínas Serina-Treonina Quinasas/genética , Ratas , Proteínas Represoras/metabolismo , Factores Sexuales , Sialoglicoproteínas/genética , Sialoglicoproteínas/metabolismo , Proteínas WT1RESUMEN
Myosin 1g (Myo1g) is a hematopoietic-specific myosin expressed mainly by lymphocytes. Here, we report the localization of Myo1g in B-cell membrane compartments such as lipid rafts, microvilli, and membrane extensions formed during spreading. By using Myo1g-deficient mouse B cells, we detected abnormalities in the adhesion ability and chemokine-induced directed migration of these lymphocytes. We also assessed a role for Myo1g in phagocytosis and exocytosis processes, as these were also irregular in Myo1g-deficient B cells. Taken together, our results show that Myo1g acts as a main regulator of different membrane/cytoskeleton-dependent processes in B lymphocytes.
Asunto(s)
Linfocitos B/inmunología , Linfocitos B/metabolismo , Movimiento Celular/genética , Movimiento Celular/inmunología , Citoesqueleto/metabolismo , Endocitosis/fisiología , Exocitosis/fisiología , Miosinas/genética , Animales , Adhesión Celular/genética , Células Cultivadas , Femenino , Ratones , Ratones Noqueados , Antígenos de Histocompatibilidad Menor , Miosinas/metabolismo , Transporte de ProteínasRESUMEN
ERM (ezrin, radixin moesin) proteins in lymphocytes link cortical actin to plasma membrane, which is regulated in part by ERM protein phosphorylation. To assess whether phosphorylation of ERM proteins regulates lymphocyte migration and membrane tension, we generated transgenic mice whose T-lymphocytes express low levels of ezrin phosphomimetic protein (T567E). In these mice, T-cell number in lymph nodes was reduced by 27%. Lymphocyte migration rate in vitro and in vivo in lymph nodes decreased by 18% to 47%. Lymphocyte membrane tension increased by 71%. Investigations of other possible underlying mechanisms revealed impaired chemokine-induced shape change/lamellipod extension and increased integrin-mediated adhesion. Notably, lymphocyte homing to lymph nodes was decreased by 30%. Unlike most described homing defects, there was not impaired rolling or sticking to lymph node vascular endothelium but rather decreased migration across that endothelium. Moreover, decreased numbers of transgenic T cells in efferent lymph suggested defective egress. These studies confirm the critical role of ERM dephosphorylation in regulating lymphocyte migration and transmigration. Of particular note, they identify phospho-ERM as the first described regulator of lymphocyte membrane tension, whose increase probably contributes to the multiple defects observed in the ezrin T567E transgenic mice.
Asunto(s)
Membrana Celular/patología , Movimiento Celular/fisiología , Proteínas del Citoesqueleto/fisiología , Ganglios Linfáticos/patología , Mutación/genética , Linfocitos T/patología , Migración Transendotelial y Transepitelial/fisiología , Animales , Membrana Celular/metabolismo , Ganglios Linfáticos/metabolismo , Recuento de Linfocitos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Fosfoproteínas/metabolismo , Fosforilación , Linfocitos T/metabolismoRESUMEN
Many cellular processes depend on ERM (ezrin, moesin, and radixin) proteins mediating regulated linkage between plasma membrane and actin cytoskeleton. Although conformational activation of the ERM protein is mediated by the membrane PIP2, the known properties of the two described PIP2-binding sites do not explain activation. To elucidate the structural basis of possible mechanisms, we generated informative moesin mutations and tested three attributes: membrane localization of the expressed moesin, moesin binding to PIP2, and PIP2-induced release of moesin autoinhibition. The results demonstrate for the first time that the POCKET containing inositol 1,4,5-trisphosphate on crystal structure (the "POCKET" Lys-63, Lys-278 residues) mediates all three functions. Furthermore the second described PIP2-binding site (the "PATCH," Lys-253/Lys-254, Lys-262/Lys-263) is also essential for all three functions. In native autoinhibited ERM proteins, the POCKET is a cavity masked by an acidic linker, which we designate the "FLAP." Analysis of three mutant moesin constructs predicted to influence FLAP function demonstrated that the FLAP is a functional autoinhibitory region. Moreover, analysis of the cooperativity and stoichiometry demonstrate that the PATCH and POCKET do not bind PIP2 simultaneously. Based on our data and supporting published data, we propose a model of progressive activation of autoinhibited moesin by a single PIP2 molecule in the membrane. Initial transient binding of PIP2 to the PATCH initiates release of the FLAP, which enables transition of the same PIP2 molecule into the newly exposed POCKET where it binds stably and completes the conformational activation.
Asunto(s)
Citoesqueleto de Actina/metabolismo , Membrana Celular/metabolismo , Proteínas de Microfilamentos/metabolismo , Fosfatidilinositol 4,5-Difosfato/metabolismo , Citoesqueleto de Actina/genética , Sitios de Unión , Membrana Celular/genética , Humanos , Células Jurkat , Proteínas de Microfilamentos/genética , Mutación , Fosfatidilinositol 4,5-Difosfato/genéticaRESUMEN
Myosin 1c (Myo1c) is a member of the unconventional class I myosins of vertebrates, which directly link the plasma membrane with the microfilament cortical web. Although this molecular motor has been implicated in cell functions such as cytoskeleton organization, cell motility, nuclear transcription, and endocytosis, its role in hematopoietic cells is largely unknown. In this study, we show that Myo1c is abundantly expressed in murine B lymphocytes and is preferentially located at the plasma membrane, especially in peripheral processes such as microvilli. We observed that this motor concentrates at the growing membrane protrusions generated during B cell spreading and that it is actively recruited to the immune synapse. Interestingly, Myo1c was detected in lipid rafts of B cells and showed strong colocalization with MHC-II, particularly after cross-linking of these molecules. By transfection of a dominant negative form of Myo1c or specific siRNA, we also detected alterations in the spreading and Ag-presenting ability of these cells. The data suggest that Myo1c is involved in the cytoskeleton dynamics and membrane protein anchoring or sorting in B lymphocytes.
Asunto(s)
Presentación de Antígeno/inmunología , Linfocitos B/inmunología , Citoesqueleto/metabolismo , Sinapsis Inmunológicas/inmunología , Miosinas/inmunología , Animales , Linfocitos B/metabolismo , Separación Celular , Citoesqueleto/inmunología , Femenino , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Inmunoprecipitación , Ratones , Ratones Endogámicos C57BL , Miosinas/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Immunological mechanisms of non-IgE-mediated cow's milk protein allergy (CMPA) are not well understood. Such a circumstance requires attention with the aim of discovering new biomarkers that could lead to better diagnostic assays for early treatment. Here, we sought both to investigate the mechanism that underlies non-IgE-mediated CMPA and to identify cow's milk immunoreactive proteins in a Mexican pediatric patient group (n = 34). Hence, we determined the IgE and IgG1-4 subclass antibody levels against cow's milk proteins (CMP) by ELISA. Then, we performed 2D-Immunoblots using as first antibody immunoglobulins in the patients'serum that bound specifically against CMP together with CMP enrichment by ion-exchange chromatography. Immunoreactive proteins were identified by mass spectrometry-based proteomics. The serological test confirmed absence of specific IgE in the CMPA patients but showed significant increase in antigen-specific IgG1. Additionally, we identified 11 proteins that specifically bound to IgG1. We conclude that the detection of specific IgG1 together with an immunoproteomics approach is highly relevant to the understanding of CMPA's physiopathology and as a possible aid in making a prognosis since current evidence indicates IgG1 occurrence as an early signal of potential risk toward development of IgE-mediated food allergy. SIGNIFICANCE: Allergies are one of the most studied topics in the field of public health and novel protein allergens are found each year. Discovery of new principal and regional allergens has remarkable repercussions in precise molecular diagnostics, prognostics, and more specific immunotherapies. In this context, specific IgE is widely known to mediate physiopathology; however, allergies whose mechanism does not involve this immunoglobulin are poorly understood although their incidence has increased. Therefore, accurate diagnosis and adequate treatment are delayed with significant consequences on the health of pediatric patients. The study of type and subtypes of immunoglobulins associated with the immunoreactivity of cow's milk proteins together with an immunoproteomics approach allows better comprehension of physiopathology, brings the opportunity to discover new potential cow's milk protein allergens and may help in prognosis prediction (IgG1 occurrence as an early signal of possible risk toward development of IgE-mediated food allergy).
Asunto(s)
Hipersensibilidad a los Alimentos , Hipersensibilidad a la Leche , Animales , Femenino , Bovinos , Hipersensibilidad a la Leche/diagnóstico , Inmunoglobulina E , Hipersensibilidad a los Alimentos/diagnóstico , Alérgenos , Proteínas de la Leche , Inmunoglobulina GRESUMEN
BACKGROUND: Myosin 1g (Myo1g) has recently been identified as a potential diagnostic biomarker in childhood acute lymphocytic leukemia (ALL). CASE REPORT: We describe the case of a 1-year-old Mexican female patient. Although initially studied for hepatomegaly, an infectious or genetic etiology was excluded. Liver biopsy showed infiltration by neoplastic B-cell precursors (BCPs), and bone marrow (BM) aspirate showed 14.5% of BCPs. In a joint session of the oncology, hematology, and pathology departments, low-risk (LR) BCP-ALL of hepatic origin with aberrant myeloid markers was diagnosed. Although treatment was initiated, the patient presented early with BM relapse. Modest overexpression of Myo1g was observed from the onset. However, at the end of the steroid window, expression increased significantly and remained elevated during this first relapse to BM. The parents refused hematopoietic stem cell transplantation, but she continued chemotherapy. After a second BM relapse at 5 years of age, the phenotype switched to myeloid. Her parents then opted for palliative care, and the patient died two months later at home. CONCLUSIONS: This case shows the potential use of Myo1g in clinical practice as a high-risk indicator. Myo1g monitoring may reveal a high risk and relapse trend, even when typical parameter values are not altered: Myo1g could be used to classify patients from low to high risk from diagnosis, allowing patients to promptly receive the best treatment and potentially modifying prognosis and survival.
INTRODUCCIÓN: Recientemente se ha identificado a miosina 1g (Myo1g) como un potencial biomarcador de diagnóstico en la leucemia linfoblástica aguda (LLA) infantil. CASO CLÍNICO: Se describe el caso de una paciente mexicana de 1 año de edad. Aunque inicialmente se estudió por hepatomegalia, se descartó una etiología infecciosa o genética. La biopsia hepática mostró infiltración por precursores de células B neoplásicas (PCB) y un aspirado de médula ósea (MO) mostró 14.5% de PCB. En una sesión conjunta de los departamentos de oncología, hematología y patología, se diagnosticó PCB-LLA de bajo riesgo de origen hepático con marcadores mieloides aberrantes. Aunque se inició tratamiento, la paciente presentó tempranamente recaída de MO. Se observó una modesta sobreexpresión de Myo1g. Sin embargo, al final de la ventana de esteroides, la expresión aumentó considerablemente y permaneció elevada durante esta primera recaída a MO. El trasplante de células madre hematopoyéticas fue rechazado por los padres, pero se continuó con la quimioterapia. Tras una segunda recaída de MO a los 5 años, el fenotipo cambió a mieloide. Sus padres optaron entonces por cuidados paliativos y la paciente falleció dos meses después en su domicilio. CONCLUSIONES: Este caso muestra el potencial uso de Myo1g como indicador de alto riesgo en la práctica clínica. El seguimiento de Myo1g puede revelar una tendencia de alto riesgo y recaídas, incluso cuando los valores de los parámetros rutinarios son aparentemente normales; Myo1g podría utilizarse para clasificar a los pacientes de bajo a alto riesgo desde el diagnóstico, lo que permitiría que los pacientes reciban el mejor tratamiento de manera oportuna, modificando potencialmente el pronóstico y la supervivencia.
Asunto(s)
Trasplante de Células Madre Hematopoyéticas , Leucemia-Linfoma Linfoblástico de Células Precursoras , Femenino , Humanos , Biomarcadores , Fenotipo , Leucemia-Linfoma Linfoblástico de Células Precursoras/patología , RecurrenciaRESUMEN
PTP1B plays a key role in developing different types of cancer. However, the molecular mechanism underlying this effect is unclear. To identify molecular targets of PTP1B that mediate its role in tumorigenesis, we undertook a SILAC-based phosphoproteomic approach, which allowed us to identify Cdk3 as a novel PTP1B substrate. Substrate trapping experiments and docking studies revealed stable interactions between the PTP1B catalytic domain and Cdk3. In addition, we observed that PTP1B dephosphorylates Cdk3 at tyrosine residue 15 in vitro and interacts with it in human glioblastoma cells. Next, we found that pharmacological inhibition of PTP1B or its depletion with siRNA leads to cell cycle arrest with diminished activity of Cdk3, hypophosphorylation of Rb, and the downregulation of E2F target genes Cdk1, Cyclin A, and Cyclin E1. Finally, we observed that the expression of a constitutively active Cdk3 mutant bypasses the requirement of PTP1B for cell cycle progression and expression of E2F target genes. These data delineate a novel signaling pathway from PTP1B to Cdk3 required for efficient cell cycle progression in an Rb-E2F dependent manner in human GB cells.
Asunto(s)
Glioblastoma , Humanos , Glioblastoma/genética , División Celular , Transducción de Señal , Puntos de Control del Ciclo Celular , Ciclo Celular/fisiología , Proteína Tirosina Fosfatasa no Receptora Tipo 1/genética , Proteína Tirosina Fosfatasa no Receptora Tipo 1/metabolismoRESUMEN
Class I Myosins are a subfamily of motor proteins with ATPase activity and a characteristic structure conserved in all myosins: A N-Terminal Motor Domain, a central Neck and a C terminal Tail domain. Humans have eight genes for these myosins. Class I Myosins have different functions: regulate membrane tension, participate in endocytosis, exocytosis, intracellular trafficking and cell migration. Cell migration is influenced by many cellular components including motor proteins, like myosins. Recently has been reported that changes in myosin expression have an impact on the migration of cancer cells, the formation of infiltrates and metastasis. We propose that class I myosins might be potential markers for future diagnostic, prognostic or even as therapeutic targets in leukemia and other cancers.Abbreviations: Myo1g: Myosin 1g; ALL: Acute Lymphoblastic Leukemia, TH1: Tail Homology 1; TH2: Tail Homology 2; TH3: Tail Homology 3.
Asunto(s)
Miosinas , Neoplasias , Movimiento Celular , HumanosRESUMEN
Mucosal innate immunity functions as the first line of defense against invading pathogens. Members of the IL-1 family are key cytokines upregulated in the inflamed mucosa. Inflammatory cytokines are regulated by limiting their function and availability through their activation and secretion mechanisms. IL-1 cytokines secretion is affected by the lack of a signal peptide on their sequence, which prevents them from accessing the conventional protein secretion pathway; thus, they use unconventional protein secretion pathways. Here we show in mouse macrophages that LPS/ATP stimulation induces cytokine relocalization to the plasma membrane, and conventional secretion blockade using monensin or Brefeldin A triggers no IL-36γ accumulation within the cell. In silico modeling indicates IL-36γ can pass through both the P2X7R and Gasdermin D pores, and both IL-36γ, P2X7R and Gasdermin D mRNA are upregulated in inflammation; further, experimental blockade of these receptors' limits IL-36γ release. Our results demonstrate that IL-36γ is secreted mainly by an unconventional pathway through membrane pores formed by P2X7R and Gasdermin D.
Asunto(s)
Inmunidad Mucosa , Proteínas de Unión a Fosfato/metabolismo , Proteínas Citotóxicas Formadoras de Poros/metabolismo , Receptores Purinérgicos P2X7/metabolismo , Animales , Transporte Biológico , Citocinas/metabolismo , Interleucina-1 , RatonesRESUMEN
Monocytes play important roles in anti-microbial and anti-viral responses and chronic inflammatory diseases. Monocytes' functions are altered by aging. We investigated age-changes in calcium (Ca2+) response to CCL2 and LPS in human monocytes. CCL2 and LPS induced a slow increase of the cytosolic Ca2+ level, with a maximum response at â¼360 s and â¼300 s, respectively, in monocytes of young and older adults. No difference was observed in the magnitude and in the Ca2+ kinetic with both stimuli. Furthermore, store-operated Ca2+ entry and plasma membrane expression of ORAI1 showed no difference between both groups. In summary, monocytes from older adults maintained the capacity to mobilize calcium as their counterparts in young adults suggesting that the mechanisms underlying the dysfunctions in monocytes in aging might not involve alterations in Ca2+ flow through the plasma membrane.
Asunto(s)
Calcio , Lipopolisacáridos , Anciano , Calcio/metabolismo , Señalización del Calcio/fisiología , Quimiocina CCL2 , Humanos , Monocitos/metabolismo , Proteína ORAI1/genética , Proteína ORAI1/metabolismo , Molécula de Interacción Estromal 1/metabolismoRESUMEN
The vulnerability of older adults to bacterial infections has been associated with age-related changes in neutrophils. We analyzed the consequences of aging on calcium (Ca2+) mobilization and TRPM2 and CRAC channels expression in human neutrophils. The percentages of granulocytes, mature neutrophils, and neutrophil precursors were equivalent between young and older adults. However, neutrophil chemotaxis towards IL-8, C5a, or fMLP was lower in older adults of both sexes. Interestingly, a stronger Ca2+ transient followed by an identical Ca2+ influx to IL-8 was observed in older adult females. In addition, the Ca2+ response to LPS was delayed and prolonged in neutrophils of older adult males. There was no significant difference in Ca2+ response to fMLP, C5a, or store-operated Ca2+ entry in the older adults. There were also no differences in the expression of CXCR2, CD88, FPLR1, and TLR4. Interestingly, TRPM2- and ORAI1-mRNA expression was lower in neutrophils of older adults, mainly in females. Both channels were detected intracellularly in the neutrophils. TRPM2 was in late endosomes in young adults and in lysosomes in older adult neutrophils. In summary, defective neutrophil chemotaxis in aging seemed not to stem from alterations in Ca2+ signals; nevertheless, the low TRPM2 and ORAI1 expression may affect other functions.
Asunto(s)
Envejecimiento , Canales de Calcio Activados por la Liberación de Calcio , Señalización del Calcio , Neutrófilos , Factores Sexuales , Canales Catiónicos TRPM , Anciano , Canales de Calcio Activados por la Liberación de Calcio/metabolismo , Femenino , Humanos , Interleucina-8/farmacología , Masculino , Neutrófilos/metabolismo , Canales Catiónicos TRPM/genética , Canales Catiónicos TRPM/metabolismoRESUMEN
Severe acute respiratory syndrome (SARS)-coronavirus (CoV)-2 infection in children and adolescents primarily causes mild or asymptomatic coronavirus disease 2019 (COVID-19), and severe illness is mainly associated with comorbidities. However, the worldwide prevalence of COVID-19 in this population is only 1%-2%. In Mexico, the prevalence of COVID-19 in children has increased to 10%. As serology-based studies are scarce, we analyzed the clinical features and serological response (SARS-CoV-2 structural proteins) of children and adolescents who visited the Hospital Infantil de México Federico Gómez (October 2020-March 2021). The majority were 9-year-old children without comorbidities who were treated as outpatients and had mild-to-moderate illness. Children aged 6-10 years and adolescents aged 11-15 years had the maximum number of symptoms, including those with obesity. Nevertheless, children with comorbidities such as immunosuppression, leukemia, and obesity exhibited the lowest antibody response, whereas those aged 1-5 years with heart disease had the highest levels of antibodies. The SARS-CoV-2 spike receptor-binding domain-localized peptides and M and E proteins had the best antibody response. In conclusion, Mexican children and adolescents with COVID-19 represent a heterogeneous population, and comorbidities play an important role in the antibody response against SARS-CoV-2 infection.