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1.
Immunity ; 36(5): 769-81, 2012 May 25.
Artículo en Inglés | MEDLINE | ID: mdl-22608497

RESUMEN

The coordination of nutrient and energy availability with cell growth and division is essential for proper immune cell development and function. By using a chemical mutagenesis strategy in mice, we identified a pedigree that has a complete block in B cell development at the pre-B cell stage resulting from a deletion in the Fnip1 gene. Enforced expression of an immunoglobulin transgene failed to rescue B cell development. Whereas essential pre-B cell signaling molecules were activated normally in Fnip1-null pre-B cells, the metabolic regulators AMPK and mTOR were dysregulated, resulting in excessive cell growth and enhanced sensitivity to apoptosis in response to metabolic stress (pre-B cell receptor crosslinking, oncogene activation). These results indicate that Folliculin-interacting protein 1 (Fnip1) is vital for B cell development and metabolic homeostasis and reveal a metabolic checkpoint that may ensure that pre-B cells have sufficient metabolic capacity to support division, while limiting lymphomagenesis caused by deregulated growth.


Asunto(s)
Linfocitos B/citología , Linfocitos B/metabolismo , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Diferenciación Celular/genética , Estrona/genética , Estrona/metabolismo , Quinasas de la Proteína-Quinasa Activada por el AMP , Animales , Apoptosis/genética , División Celular/genética , Hematopoyesis/genética , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas Pesadas de Inmunoglobulina/metabolismo , Ratones , Ratones Transgénicos , Células Precursoras de Linfocitos B/metabolismo , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo , Transducción de Señal/genética , Serina-Treonina Quinasas TOR/genética , Serina-Treonina Quinasas TOR/metabolismo
2.
Am J Obstet Gynecol ; 225(1): 89.e1-89.e16, 2021 07.
Artículo en Inglés | MEDLINE | ID: mdl-33412130

RESUMEN

BACKGROUND: Intra-amniotic infection or inflammation is common in early preterm birth and associated with substantial neonatal lung morbidity owing to fetal exposure to proinflammatory cytokines and infectious organisms. Amniotic fluid interleukin 8, a proinflammatory cytokine, was previously correlated with the development of neonatal bronchopulmonary dysplasia, but whether amniotic fluid cytokines or placental pathology more accurately predicts neonatal lung pathology and morbidity is unknown. We have used a pregnant nonhuman primate model of group B Streptococcus infection to study the pathogenesis of intra-amniotic infection, bacterial invasion of the amniotic cavity and fetus, and microbial-host interactions. In this nonhuman primate model, we have studied the pathogenesis of group B Streptococcus strains with differing potential for virulence, which has resulted in a spectrum of intra-amniotic infection and fetal lung injury that affords the opportunity to study the inflammatory predictors of fetal lung pathology and injury. OBJECTIVE: This study aimed to determine whether fetal lung injury is best predicted by placental histopathology or the cytokine response in amniotic fluid or maternal plasma. STUDY DESIGN: Chronically catheterized pregnant monkeys (Macaca nemestrina, pigtail macaque) at 116 to 125 days gestation (term at 172 days) received a choriodecidual inoculation of saline (n=5), weakly hemolytic group B Streptococcus strain (n=5, low virulence), or hyperhemolytic group B Streptococcus strain (n=5, high virulence). Adverse pregnancy outcomes were defined as either preterm labor, microbial invasion of the amniotic cavity, or development of the fetal inflammatory response syndrome. Amniotic fluid and maternal and fetal plasma samples were collected after inoculation, and proinflammatory cytokines (tumor necrosis factor alpha, interleukin beta, interleukin 6, interleukin 8) were measured by a multiplex assay. Cesarean delivery was performed at the time of preterm labor or within 1 week of inoculation. Fetal necropsy was performed at the time of delivery. Placental pathology was scored in a blinded fashion by a pediatric pathologist, and fetal lung injury was determined by a semiquantitative score from histopathology evaluating inflammatory infiltrate, necrosis, tissue thickening, or collapse scored by a veterinary pathologist. RESULTS: The principal findings in our study are as follows: (1) adverse pregnancy outcomes occurred more frequently in animals receiving hyperhemolytic group B Streptococcus (80% with preterm labor, 80% with fetal inflammatory response syndrome) than in animals receiving weakly hemolytic group B Streptococcus (40% with preterm labor, 20% with fetal inflammatory response syndrome) and in controls (0% preterm labor, 0% fetal inflammatory response syndrome); (2) despite differences in the rate of adverse pregnancy outcomes and fetal inflammatory response syndrome, fetal lung injury scores were similar between animals receiving the weakly hemolytic group B Streptococcus strains and animals receiving the hyperhemolytic group B Streptococcus strains; (3) fetal lung injury score was significantly correlated with peak amniotic fluid cytokines interleukin 6 and interleukin 8 but not tumor necrosis factor alpha or interleukin 1 beta; and (4) fetal lung scores were poorly correlated with maternal and fetal plasma cytokine levels and placental pathology. CONCLUSION: Amniotic fluid interleukin 6 and interleukin 8 levels were superior predictors of fetal lung injury than placental histopathology or maternal plasma cytokines. This evidence supports a role for amniocentesis in the prediction of neonatal lung morbidity owing to intra-amniotic infection, which cannot be provided by cytokine analysis of maternal plasma or placental histopathology.


Asunto(s)
Líquido Amniótico/química , Citocinas/sangre , Interleucina-6/análisis , Interleucina-8/análisis , Lesión Pulmonar/embriología , Placenta/patología , Líquido Amniótico/microbiología , Animales , Modelos Animales de Enfermedad , Femenino , Inflamación/embriología , Inflamación/microbiología , Pulmón/embriología , Pulmón/microbiología , Pulmón/patología , Lesión Pulmonar/diagnóstico , Lesión Pulmonar/microbiología , Macaca nemestrina , Masculino , Embarazo , Resultado del Embarazo , Infecciones Estreptocócicas/embriología , Streptococcus agalactiae
3.
Proc Natl Acad Sci U S A ; 112(2): 424-9, 2015 Jan 13.
Artículo en Inglés | MEDLINE | ID: mdl-25548157

RESUMEN

Mammalian skeletal muscle is broadly characterized by the presence of two distinct categories of muscle fibers called type I "red" slow twitch and type II "white" fast twitch, which display marked differences in contraction strength, metabolic strategies, and susceptibility to fatigue. The relative representation of each fiber type can have major influences on susceptibility to obesity, diabetes, and muscular dystrophies. However, the molecular factors controlling fiber type specification remain incompletely defined. In this study, we describe the control of fiber type specification and susceptibility to metabolic disease by folliculin interacting protein-1 (Fnip1). Using Fnip1 null mice, we found that loss of Fnip1 increased the representation of type I fibers characterized by increased myoglobin, slow twitch markers [myosin heavy chain 7 (MyH7), succinate dehydrogenase, troponin I 1, troponin C1, troponin T1], capillary density, and mitochondria number. Cultured Fnip1-null muscle fibers had higher oxidative capacity, and isolated Fnip1-null skeletal muscles were more resistant to postcontraction fatigue relative to WT skeletal muscles. Biochemical analyses revealed increased activation of the metabolic sensor AMP kinase (AMPK), and increased expression of the AMPK-target and transcriptional coactivator PGC1α in Fnip1 null skeletal muscle. Genetic disruption of PGC1α rescued normal levels of type I fiber markers MyH7 and myoglobin in Fnip1-null mice. Remarkably, loss of Fnip1 profoundly mitigated muscle damage in a murine model of Duchenne muscular dystrophy. These results indicate that Fnip1 controls skeletal muscle fiber type specification and warrant further study to determine whether inhibition of Fnip1 has therapeutic potential in muscular dystrophy diseases.


Asunto(s)
Proteínas Portadoras/fisiología , Fibras Musculares de Contracción Rápida/patología , Fibras Musculares de Contracción Rápida/fisiología , Fibras Musculares de Contracción Lenta/patología , Fibras Musculares de Contracción Lenta/fisiología , Distrofia Muscular de Duchenne/patología , Distrofia Muscular de Duchenne/fisiopatología , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Proteínas Portadoras/genética , Modelos Animales de Enfermedad , Masculino , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Ratones Endogámicos mdx , Ratones Noqueados , Mitocondrias Musculares/metabolismo , Mitocondrias Musculares/patología , Complejos Multiproteicos/metabolismo , Contracción Muscular/fisiología , Fatiga Muscular/fisiología , Distrofia Muscular de Duchenne/genética , Mioglobina/metabolismo , Cadenas Pesadas de Miosina/metabolismo , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma , Serina-Treonina Quinasas TOR/metabolismo , Factores de Transcripción/deficiencia , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
4.
J Immunol ; 192(1): 300-7, 2014 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-24298015

RESUMEN

Nucleotide-binding oligomerization domain 2 (NOD2) is a cytosolic pathogen recognition receptor that regulates susceptibility to a variety of infections and chronic diseases. Burkholderia pseudomallei, a facultative intracellular bacterium, causes the tropical infection melioidosis. We hypothesized that NOD2 may participate in host defense in melioidosis. We performed a series of in vitro assays and in vivo experiments and analyzed the association of human genetic variation with infection to delineate the contribution of NOD2 to the host response to B. pseudomallei. We found that transfection with NOD2 mediated NF-κB activation induced by B. pseudomallei stimulation of HEK293 cells. After low-dose inoculation with aerosolized B. pseudomallei, Nod2-deficient mice showed impaired clinical responses and permitted greater bacterial replication in the lung and dissemination to the spleen compared with wild-type mice. IL-6 and KC levels were higher in the lungs of Nod2-deficient mice. In a cohort of 1562 Thai subjects, a common genetic polymorphism in the NOD2 region, rs7194886, was associated with melioidosis, and this effect was most pronounced in women. rs7194886 was not associated with differences in cytokine production induced by whole-blood stimulation with the NOD2 ligand, muramyl dipeptide, or B. pseudomallei. To our knowledge, these findings are the first to characterize the role of NOD2 in host defense in mammalian melioidosis.


Asunto(s)
Burkholderia pseudomallei/inmunología , Melioidosis/genética , Melioidosis/inmunología , Proteína Adaptadora de Señalización NOD2/genética , Animales , Línea Celular Tumoral , Citocinas/sangre , Citocinas/metabolismo , Modelos Animales de Enfermedad , Células HEK293 , Humanos , Inmunidad Innata/genética , Interleucina-6/sangre , Interleucina-6/metabolismo , Pulmón/inmunología , Pulmón/metabolismo , Pulmón/microbiología , Melioidosis/metabolismo , Melioidosis/mortalidad , Ratones , Ratones Noqueados , Proteína Adaptadora de Señalización NOD2/deficiencia , Proteína Adaptadora de Señalización NOD2/metabolismo , Polimorfismo de Nucleótido Simple
5.
Infect Immun ; 83(10): 3909-17, 2015 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-26195546

RESUMEN

The mechanisms underlying fetal lung injury remain poorly defined. MicroRNAs (miRNAs) are small noncoding, endogenous RNAs that regulate gene expression and have been implicated in the pathogenesis of lung disease. Using a nonhuman primate model of choriodecidual infection, we sought to determine if differentially expressed miRNAs were associated with acute fetal lung injury. After inoculating 10 chronically catheterized pregnant monkeys (Macaca nemestrina) with either group B streptococcus (GBS) at 1 × 10(6) CFU (n = 5) or saline (n = 5) in the choriodecidual space, we extracted fetal lung mRNA and miRNA and profiled the changes in expression by microarray analysis. We identified 9 differentially expressed miRNAs in GBS-exposed fetal lungs, but of these, only miR-155-5p was validated by quantitative reverse transcription-PCR (P = 0.02). Significantly elevated miR-155-5p expression was also observed when immortalized human fetal airway epithelial (FeAE) cells were exposed to proinflammatory cytokines (interleukin-6 [IL-6] and tumor necrosis factor alpha [TNF-α]). Overexpression of miR-155-5p in FeAE cells in turn increased the production of IL-6 and CXCL10/gamma interferon-induced protein 10, which are implicated in leukocyte recruitment but also in protection from lung injury. Interestingly, while miR-155-5p decreased fibroblast growth factor 9 (FGF9) expression in a luciferase reporter assay, FGF9 levels were actually increased in GBS-exposed fetal lungs in vivo. FGF9 overexpression is associated with abnormal lung development. Thus, upregulation of miR-155-5p may serve as a compensatory mechanism to lessen the increase in FGF9 and prevent aberrant lung development. Understanding the complicated networks regulating lung development in the setting of infection is a key step in identifying how to prevent fetal lung injury leading to bronchopulmonary dysplasia.


Asunto(s)
Enfermedades Fetales/genética , Enfermedades Fetales/microbiología , Pulmón/metabolismo , Infecciones Estreptocócicas/embriología , Infecciones Estreptocócicas/genética , Streptococcus/fisiología , Animales , Modelos Animales de Enfermedad , Femenino , Enfermedades Fetales/metabolismo , Humanos , Interleucina-6/genética , Interleucina-6/metabolismo , Pulmón/crecimiento & desarrollo , Pulmón/microbiología , Macaca nemestrina , Masculino , Embarazo , Infecciones Estreptocócicas/metabolismo , Infecciones Estreptocócicas/microbiología , Streptococcus/genética , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo
6.
J Immunol ; 189(8): 3936-46, 2012 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-22984076

RESUMEN

Adoptive T cell therapy (ACT) for the treatment of established cancers is actively being pursued in clinical trials. However, poor in vivo persistence and maintenance of antitumor activity of transferred T cells remain major problems. TGF-ß is a potent immunosuppressive cytokine that is often expressed at high levels within the tumor microenvironment, potentially limiting T cell-mediated antitumor activity. In this study, we used a model of autochthonous murine prostate cancer to evaluate the effect of cell-intrinsic abrogation of TGF-ß signaling in self/tumor-specific CD8 T cells used in ACT to target the tumor in situ. We found that persistence and antitumor activity of adoptively transferred effector T cells deficient in TGF-ß signaling were significantly improved in the cancerous prostate. However, over time, despite persistence in peripheral lymphoid organs, the numbers of transferred cells in the prostate decreased and the residual prostate-infiltrating T cells were no longer functional. These findings reveal that TGF-ß negatively regulates the accumulation and effector function of transferred self/tumor-specific CD8 T cells and highlight that, when targeting a tumor Ag that is also expressed as a self-protein, additional substantive obstacles are operative within the tumor microenvironment, potentially hampering the success of ACT for solid tumors.


Asunto(s)
Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/patología , Epítopos de Linfocito T/inmunología , Neoplasias de la Próstata/inmunología , Neoplasias de la Próstata/patología , Transducción de Señal/inmunología , Factor de Crecimiento Transformador beta/antagonistas & inhibidores , Factor de Crecimiento Transformador beta/fisiología , Traslado Adoptivo/métodos , Animales , Linfocitos T CD8-positivos/trasplante , Femenino , Masculino , Ratones , Ratones Noqueados , Ratones Transgénicos , Neoplasias de la Próstata/terapia , Miembro 25 de Receptores de Factores de Necrosis Tumoral/genética , Transducción de Señal/genética
7.
bioRxiv ; 2024 Aug 06.
Artículo en Inglés | MEDLINE | ID: mdl-39211124

RESUMEN

Genome-wide association studies have identified SH2B3 as an important non-MHC gene for islet autoimmunity and type 1 diabetes (T1D). In this study, we found a single SH2B3 haplotype significantly associated with increased risk for human T1D, and this haplotype carries the single nucleotide variant rs3184504*T in SH2B3. To better characterize the role of SH2B3 in T1D, we used mouse modeling and found a T cell-intrinsic role for SH2B3 regulating peripheral tolerance. SH2B3 deficiency had minimal effect on TCR signaling or proliferation across antigen doses, yet enhanced cell survival and cytokine signaling including common gamma chain-dependent and interferon-gamma receptor signaling. SH2B3 deficient CD8+T cells showed augmented STAT5-MYC and effector-related gene expression partially reversed with blocking autocrine IL-2 in culture. Using the RIP-mOVA model, we found CD8+ T cells lacking SH2B3 promoted early islet destruction and diabetes without requiring CD4+ T cell help. SH2B3-deficient cells demonstrated increased survival post-transfer compared to control cells despite a similar proliferation profile in the same host. Next, we created a spontaneous NOD .Sh2b3 -/- mouse model and found markedly increased incidence and accelerated T1D across sexes. Collectively, these studies identify SH2B3 as a critical mediator of peripheral T cell tolerance limiting the T cell response to self-antigens. Article Highlights: The rs3184504 polymorphism, encoding a hypomorphic variant of the negative regulator SH2B3, strongly associates with T1D.SH2B3 deficiency results in hypersensitivity to cytokines, including IL-2, in murine CD4+ and CD8+ T cells.SH2B3 deficient CD8+ T cells exhibit a comparable transcriptome to wild-type CD8+ T cells at baseline, but upon antigen stimulation SH2B3 deficient cells upregulate genes characteristic of enhanced JAK/STAT signaling and effector functions.We found a T-cell intrinsic role of SH2B3 leading to severe islet destruction in an adoptive transfer murine T1D model, while global SH2B3 deficiency accelerated spontaneous NOD diabetes across sexes.

8.
J Urol ; 190(3): 1090-5, 2013 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-23583535

RESUMEN

PURPOSE: Focused ultrasound has the potential to expel small stones or residual stone fragments from the kidney, or move obstructing stones to a nonobstructing location. We evaluated the efficacy and safety of ultrasonic propulsion in a live porcine model. MATERIALS AND METHODS: Calcium oxalate monohydrate kidney stones and laboratory model stones (2 to 8 mm) were ureteroscopically implanted in the renal pelvicalyceal system of 12 kidneys in a total of 8 domestic swine. Transcutaneous ultrasonic propulsion was performed using an HDI C5-2 imaging transducer (ATL/Philips, Bothell, Washington) and the Verasonics® diagnostic ultrasound platform. Successful stone relocation was defined as stone movement from the calyx to the renal pelvis, ureteropelvic junction or proximal ureter. Efficacy and procedure time was determined. Three blinded experts evaluated histological injury to the kidney in the control, sham treatment and treatment arms. RESULTS: All 26 stones were observed to move during treatment and 17 (65%) were relocated successfully to the renal pelvis (3), ureteropelvic junction (2) or ureter (12). Average ± SD successful procedure time was 14 ± 8 minutes and a mean of 23 ± 16 ultrasound bursts, each about 1 second in duration, were required. There was no evidence of gross or histological injury to the renal parenchyma in kidneys exposed to 20 bursts (1 second in duration at 33-second intervals) at the same output (2,400 W/cm(2)) used to push stones. CONCLUSIONS: Noninvasive transcutaneous ultrasonic propulsion is a safe, effective and time efficient means to relocate calyceal stones to the renal pelvis, ureteropelvic junction or ureter. This technology holds promise as a useful adjunct to surgical management for renal calculi.


Asunto(s)
Cálculos Renales/terapia , Terapia por Ultrasonido/instrumentación , Terapia por Ultrasonido/métodos , Animales , Oxalato de Calcio/química , Modelos Animales de Enfermedad , Diseño de Equipo , Seguridad de Equipos , Femenino , Inmunohistoquímica , Cálculos Renales/diagnóstico por imagen , Cálculos Renales/patología , Litotricia/métodos , Porcinos , Resultado del Tratamiento , Ultrasonografía
9.
Int J Exp Pathol ; 93(6): 421-8, 2012 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-23136994

RESUMEN

Melioidosis is a tropical disease caused by ingestion, percutaneous inoculation or inhalation of the Gram-negative soil saprophyte Burkholderia pseudomallei. We developed a reproducible experimental murine model of pneumonic melioidosis induced by inhalation of aerosolized B. pseudomallei 1026b. In a series of experiments performed to bracket the lethal dose, we found that C57BL/6 mice were modestly more resistant than BALB/c mice (median lethal dose 334 CFU/lung vs 204 CFU/lung). We further characterized infection and pulmonary inflammation in C57BL/6 mice infected with a sublethal dose. We observed pulmonary replication and dissemination of bacteria to distant organs in the first days after infection, followed by bacterial containment by day 4 and no evidence of recrudescent infection for up to 2 months. We measured a robust host inflammatory response notable for a neutrophilic bronchoalveolar lavage fluid profile, elevated cytokines and chemokines in the lung and serum and scattered foci of neutrophilic infiltrates in the alveoli and in a perivascular distribution on histological analysis. We previously noted a similar pattern of inflammation in mice infected with aerosolized B. thailandensis. This report builds on the limited literature describing experimental murine pneumonic melioidosis induced by aerosol and characterizes pulmonary infection and resultant inflammation in C57BL/6 mice infected with aerosolized B. pseudomallei. This model has utility for the study of bacterial and host factors that contribute to the virulence of melioidosis.


Asunto(s)
Burkholderia pseudomallei/fisiología , Melioidosis/microbiología , Neumonía Bacteriana/microbiología , Aerosoles , Animales , Biomarcadores/metabolismo , Líquido del Lavado Bronquioalveolar/citología , Burkholderia pseudomallei/patogenicidad , Citocinas/metabolismo , Modelos Animales de Enfermedad , Exposición por Inhalación/efectos adversos , Longevidad , Pulmón/metabolismo , Pulmón/microbiología , Pulmón/patología , Melioidosis/metabolismo , Melioidosis/mortalidad , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Neutrófilos/patología , Neumonía Bacteriana/metabolismo , Neumonía Bacteriana/mortalidad , Organismos Libres de Patógenos Específicos , Tasa de Supervivencia
10.
JCI Insight ; 7(9)2022 05 09.
Artículo en Inglés | MEDLINE | ID: mdl-35531955

RESUMEN

Hematopoietic protein-1 (Hem-1) is a member of the actin-regulatory WASp family verprolin homolog (WAVE) complex. Loss-of-function variants in the NCKAP1L gene encoding Hem-1 were recently discovered to result in primary immunodeficiency disease (PID) in children, characterized by poor specific Ab responses, increased autoantibodies, and high mortality. However, the mechanisms of how Hem-1 deficiency results in PID are unclear. In this study, we utilized constitutive and B cell-specific Nckap1l-KO mice to dissect the importance of Hem-1 in B cell development and functions. B cell-specific disruption of Hem-1 resulted in reduced numbers of recirculating follicular (FO), marginal zone (MZ), and B1 B cells. B cell migration in response to CXCL12 and -13 were reduced. T-independent Ab responses were nearly abolished, resulting in failed protective immunity to Streptococcus pneumoniae challenge. In contrast, T-dependent IgM and IgG2c, memory B cell, and plasma cell responses were more robust relative to WT control mice. B cell-specific Hem-1-deficient mice had increased autoantibodies against multiple autoantigens, and this correlated with hyperresponsive BCR signaling and increased representation of CD11c+T-bet+ age-associated B cell (ABC cells) - alterations associated with autoimmune diseases. These results suggest that dysfunctional B cells may be part of a mechanism explaining why loss-of-function Hem-1 variants result in recurring infections and autoimmunity.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales , Autoanticuerpos , Enfermedades Autoinmunes , Linfocitos B , Inmunidad Humoral , Actinas , Proteínas Adaptadoras Transductoras de Señales/inmunología , Animales , Linfocitos B/inmunología , Ratones , Ratones Noqueados
11.
Leukemia ; 36(1): 42-57, 2022 01.
Artículo en Inglés | MEDLINE | ID: mdl-34193976

RESUMEN

Philadelphia chromosome-like acute lymphoblastic leukemia (Ph-like ALL) is a high-risk subtype of B-ALL often associated with genetic variants that alter cytokine receptor signaling, including mutations in the interleukin-7 receptor (IL7R). To investigate whether IL7R variants are leukemia-initiating, we built mouse models expressing activated Il7r (aIL7R). B-cell intrinsic aIL7R mice developed spontaneous B-ALL, demonstrating sufficiency of Il7r activating mutations in leukemogenesis. Concomitant introduction of a knock-out allele in the associated adapter protein Lnk (encoded by Sh2b3) or a dominant-negative variant of the transcription factor Ikaros (Ikzf1) increased disease penetrance. The resulting murine leukemias displayed monoclonality and recurrent somatic Kras mutations and efficiently engrafted into immunocompetent mice. Phosphoproteomic analyses of aIL7R leukemic cells revealed constitutive Stat5 signaling and B cell receptor (BCR)-like signaling despite the absence of surface pre-BCR. Finally, in vitro treatment of aIL7R leukemic B-cells with Jak, mTOR, or Syk inhibitors blocked growth, confirming that each pathway is active in this mouse model of IL7R-driven B-ALL.


Asunto(s)
Regulación Leucémica de la Expresión Génica , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patología , Receptores de Interleucina-7/metabolismo , Animales , Apoptosis , Proliferación Celular , Humanos , Ratones , Ratones Endogámicos NOD , Ratones SCID , Leucemia-Linfoma Linfoblástico de Células Precursoras B/etiología , Leucemia-Linfoma Linfoblástico de Células Precursoras B/metabolismo , Receptores de Interleucina-7/genética , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto
12.
J Exp Med ; 218(4)2021 04 05.
Artículo en Inglés | MEDLINE | ID: mdl-33600594

RESUMEN

Hematopoietic protein-1 (Hem-1) is a hematopoietic cell-specific actin-regulatory protein. Loss-of-function (LOF) variants in the NCKAP1L gene encoding Hem-1 have recently been found to result in primary immunodeficiency disease (PID) in humans, characterized by recurring respiratory infections, asthma, and high mortality. However, the mechanisms of how Hem-1 variants result in PID are not known. In this study, we generated constitutive and myeloid cell-specific Nckap1l-KO mice to dissect the importance of Hem-1 in lung immunity. We found that Hem-1-deficient mice accumulated excessive surfactant and cell debris in airways (pulmonary alveolar proteinosis) due to impaired development of alveolar macrophages (AMs) and reduced expression of the AM differentiation factor Pparg. Residual Hem-1-deficient AMs shifted to a proinflammatory phenotype, and Hem-1-deficient neutrophils and monocytes failed to migrate normally. Myeloid cell-specific Hem-1-deficient mice exhibited increased morbidity following influenza A virus or Streptococcus pneumoniae challenge. These results provide potential mechanisms for how LOF variants in Hem-1 result in recurring respiratory diseases.


Asunto(s)
Actinas/metabolismo , Proteínas Adaptadoras Transductoras de Señales/fisiología , Diferenciación Celular/genética , Macrófagos Alveolares/inmunología , Proteinosis Alveolar Pulmonar/inmunología , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Diferenciación Celular/inmunología , Movimiento Celular/genética , Movimiento Celular/inmunología , Células Cultivadas , Modelos Animales de Enfermedad , Femenino , Pulmón/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/inmunología , Neutrófilos/inmunología , PPAR gamma/metabolismo , Fagocitosis/genética , Fagocitosis/inmunología , Proteinosis Alveolar Pulmonar/metabolismo , Transducción de Señal/genética , Transducción de Señal/inmunología
13.
Mamm Genome ; 21(3-4): 130-42, 2010 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-20111866

RESUMEN

Members of the CHD protein family play key roles in gene regulation through ATP-dependent chromatin remodeling. This is facilitated by chromodomains that bind histone tails, and by the SWI2/SNF2-like ATPase/helicase domain that remodels chromatin by moving histones. Chd6 is ubiquitously expressed in both mouse and human, with the highest levels of expression in the brain. The Chd6 gene contains 37 exons, of which exons 12-19 encode the highly conserved ATPase domain. To determine the biological role of Chd6, we generated mouse lines with a deletion of exon 12. Chd6 without exon 12 is expressed at normal levels in mice, and Chd6 Exon 12 -/- mice are viable, fertile, and exhibit no obvious morphological or pathological phenotype. Chd6 Exon 12 -/- mice lack coordination as revealed by sensorimotor analysis. Further behavioral testing revealed that the coordination impairment was not due to muscle weakness or bradykinesia. Histological analysis of brain morphology revealed no differences between Chd6 Exon 12 -/- mice and wild-type (WT) controls. The location of CHD6 on human chromosome 20q12 is overlapped by the linkage map regions of several human ataxias, including autosomal recessive infantile cerebellar ataxia (SCAR6), a nonprogressive cerebrospinal ataxia. The genomic location, expression pattern, and ataxic phenotype of Chd6 Exon 12 -/- mice indicate that mutations within CHD6 may be responsible for one of these ataxias.


Asunto(s)
ADN Helicasas/metabolismo , Exones/genética , Actividad Motora/genética , Eliminación de Secuencia/genética , Animales , Conducta Animal/fisiología , Regulación de la Expresión Génica , Humanos , Ratones , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Fenotipo , Filogenia , Equilibrio Postural/genética , Transducción de Señal/genética
14.
Am J Physiol Lung Cell Mol Physiol ; 297(6): L1112-9, 2009 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-19801452

RESUMEN

Pseudomonas aeruginosa is a leading cause of hospital-acquired pneumonia and an important pathogen in patients with chronic lung disease, such as cystic fibrosis and bronchiectasis. The contribution of Toll-like receptor 5 (TLR5) to the innate immune response to this organism is incompletely understood. We exposed wild-type and TLR5-deficient (Tlr5(-/-)) mice to aerosolized P. aeruginosa at low and high inocula and assessed bacterial clearance, lung inflammation, and cytokine production 4 and 24 h after infection. Bacterial clearance was impaired in Tlr5(-/-) mice after low-inoculum, but not high-inoculum, infection. Early bronchoalveolar accumulation of neutrophils was reduced in Tlr5(-/-) mice after low- and high-dose infection. Cytokine responses, including markedly impaired monocyte chemoattractant protein-1 production 4 h after low- and high-inoculum challenge, were selectively altered in Tlr5(-/-) mice. In contrast, there was no impairment of bacterial clearance, neutrophil recruitment, or monocyte chemoattractant protein-1 production in Tlr5(-/-) mice after infection with a nonflagellated isotypic strain of P. aeruginosa. Thus TLR5-mediated recognition of flagellin is involved in activating pulmonary defenses against P. aeruginosa and contributes to antibacterial resistance in a manner that is partially inoculum dependent. These data are the first to demonstrate a unique role for TLR5 in the innate immune response to P. aeruginosa lung infection.


Asunto(s)
Inmunidad Innata/inmunología , Neumonía/inmunología , Neumonía/microbiología , Infecciones por Pseudomonas/inmunología , Infecciones por Pseudomonas/microbiología , Pseudomonas aeruginosa/inmunología , Receptor Toll-Like 5/inmunología , Animales , Líquido del Lavado Bronquioalveolar/citología , Recuento de Células , Quimiocina CCL2/inmunología , Citocinas/metabolismo , Femenino , Flagelos/inmunología , Inflamación/inmunología , Inflamación/patología , Pulmón/inmunología , Pulmón/microbiología , Pulmón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Neutrófilos/citología , Neutrófilos/inmunología , Neumonía/patología , Infecciones por Pseudomonas/patología , Receptor Toll-Like 5/deficiencia , Factor de Necrosis Tumoral alfa/inmunología
15.
Toxicol Pathol ; 37(6): 799-804, 2009 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-19773593

RESUMEN

The systemic inflammatory response syndrome (SIRS) is a clinicopathological manifestation of overexuberant acute-phase inflammation caused by infectious or noninfectious etiologies. The systemic release of pro-inflammatory cytokines, chemokines, and lipid and vasoactive mediators induces endothelial damage and microvascular thrombosis, potentially culminating in disseminated intravascular coagulation (DIC), acute respiratory distress syndrome (ARDS), and multiple organ dysfunction (MOD) or failure (MOF). We present five cases in the pig-tailed macaque and olive baboon where SIRS resulted in MOF, ARDS, DIC, and the Waterhouse-Friderichsen syndrome; each with gross and histological elements manifested as edema, deposition of fibrin, hemorrhage, and thrombosis. In the described cases, SIRS was the end-common pathway for multiple risk factors that parallel those documented in humans: major surgery, obstetric complications, and infection. The diagnosis of SIRS should be considered when evaluating nonhuman primate (NHP) cases of MOF manifesting with histological evidence of vascular leakage. Experimental manipulation of NHP models may be complicated by SIRS and accompanying rapid clinical decompensation. Such adverse events may compromise toxicological studies and should be avoided when possible.


Asunto(s)
Lesión Pulmonar Aguda/veterinaria , Coagulación Intravascular Diseminada/veterinaria , Insuficiencia Multiorgánica/veterinaria , Síndrome de Respuesta Inflamatoria Sistémica/veterinaria , Lesión Pulmonar Aguda/sangre , Lesión Pulmonar Aguda/patología , Animales , Citocinas/metabolismo , Coagulación Intravascular Diseminada/sangre , Coagulación Intravascular Diseminada/patología , Femenino , Macaca nemestrina , Masculino , Insuficiencia Multiorgánica/sangre , Insuficiencia Multiorgánica/patología , Papio anubis , Factores de Riesgo , Síndrome de Respuesta Inflamatoria Sistémica/sangre , Síndrome de Respuesta Inflamatoria Sistémica/patología
16.
Infect Immun ; 76(12): 5862-72, 2008 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-18852243

RESUMEN

Pneumonia caused by Staphylococcus aureus is a growing concern in the health care community. We hypothesized that characterization of the early innate immune response to bacteria in the lungs would provide insight into the mechanisms used by the host to protect itself from infection. An adult mouse model of Staphylococcus aureus pneumonia was utilized to define the early events in the innate immune response and to assess the changes in the airway proteome during the first 6 h of pneumonia. S. aureus actively replicated in the lungs of mice inoculated intranasally under anesthesia to cause significant morbidity and mortality. By 6 h postinoculation, the release of proinflammatory cytokines caused effective recruitment of neutrophils to the airway. Neutrophil influx, loss of alveolar architecture, and consolidated pneumonia were observed histologically 6 h postinoculation. Bronchoalveolar lavage fluids from mice inoculated with phosphate-buffered saline (PBS) or S. aureus were depleted of overabundant proteins and subjected to strong cation exchange fractionation followed by liquid chromatography and tandem mass spectrometry to identify the proteins present in the airway. No significant changes in response to PBS inoculation or 30 min following S. aureus inoculation were observed. However, a dramatic increase in extracellular proteins was observed 6 h postinoculation with S. aureus, with the increase dominated by inflammatory and coagulation proteins. The data presented here provide a comprehensive evaluation of the rapid and vigorous innate immune response mounted in the host airway during the earliest stages of S. aureus pneumonia.


Asunto(s)
Neumonía Estafilocócica/inmunología , Proteoma/inmunología , Infecciones Estafilocócicas/inmunología , Animales , Western Blotting , Líquido del Lavado Bronquioalveolar/química , Líquido del Lavado Bronquioalveolar/citología , Cromatografía Liquida , Citocinas/análisis , Citocinas/inmunología , Femenino , Pulmón/microbiología , Pulmón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Infiltración Neutrófila/inmunología , Neumonía Estafilocócica/microbiología , Neumonía Estafilocócica/patología , Infecciones Estafilocócicas/patología , Staphylococcus aureus
17.
Mol Cell Biol ; 25(16): 6990-7004, 2005 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-16055712

RESUMEN

The corepressor mSin3A is the core component of a chromatin-modifying complex that is recruited by multiple gene-specific transcriptional repressors. In order to understand the role of mSin3A during development, we generated constitutive germ line as well as conditional msin3A deletions. msin3A deletion in the developing mouse embryo results in lethality at the postimplantation stage, demonstrating that it is an essential gene. Blastocysts derived from preimplantation msin3A null embryos and mouse embryo fibroblasts (MEFs) lacking msin3A display a significant reduction in cell division. msin3A null MEFs also show mislocalization of the heterochromatin protein, HP1alpha, without alterations in global histone acetylation. Heterozygous msin3A(+/-) mice with a systemic twofold decrease in mSin3A protein develop splenomegaly as well as kidney disease indicative of a disruption of lymphocyte homeostasis. Conditional deletion of msin3A from developing T cells results in reduced thymic cellularity and a fivefold decrease in the number of cytotoxic (CD8) T cells, while helper (CD4) T cells are unaffected. We show that CD8 development is dependent on mSin3A at a step downstream of T-cell receptor signaling and that loss of mSin3A specifically decreases survival of double-positive and CD8 T cells. Thus, msin3A is a pleiotropic gene which, in addition to its role in cell cycle progression, is required for the development and homeostasis of cells in the lymphoid lineage.


Asunto(s)
Cromatina/metabolismo , Proteínas Represoras/fisiología , Linfocitos T/citología , Animales , Apoptosis , Blastocisto , Western Blotting , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD8-positivos/metabolismo , Ciclo Celular , Diferenciación Celular , Linaje de la Célula , Proliferación Celular , Células Cultivadas , Cromatina/química , Homólogo de la Proteína Chromobox 5 , Proteínas Cromosómicas no Histona/metabolismo , Exones , Fibroblastos/citología , Fibroblastos/metabolismo , Citometría de Flujo , Eliminación de Gen , Regulación del Desarrollo de la Expresión Génica , Genotipo , Glomerulonefritis Membranosa , Heterocromatina/metabolismo , Heterocigoto , Ratones , Ratones Transgénicos , Modelos Biológicos , Modelos Genéticos , Recombinación Genética , Complejo Correpresor Histona Desacetilasa y Sin3 , Esplenomegalia , Linfocitos T/metabolismo , Linfocitos T Citotóxicos/citología , Timo/citología , Factores de Tiempo
18.
PLoS One ; 13(6): e0197973, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-29897930

RESUMEN

Birt-Hogg-Dube' Syndrome (BHDS) is a rare genetic disorder in humans characterized by skin hamartomas, lung cysts, pneumothorax, and increased risk of renal tumors. BHDS is caused by mutations in the BHD gene, which encodes for Folliculin, a cytoplasmic adapter protein that binds to Folliculin interacting proteins-1 and -2 (Fnip1, Fnip2) as well as the master energy sensor AMP kinase (AMPK). Whereas kidney-specific deletion of the Bhd gene in mice is known to result in polycystic kidney disease (PKD) and renal cell carcinoma, the roles of Fnip1 in renal cell development and function are unclear. In this study, we utilized mice with constitutive deletion of the Fnip1 gene to show that the loss of Fnip1 is sufficient to result in renal cyst formation, which was characterized by decreased AMPK activation, increased mTOR activation, and metabolic hyperactivation. Using RNAseq, we found that Fnip1 disruption resulted in many cellular and molecular changes previously implicated in the development of PKD in humans, including alterations in the expression of ion and amino acid transporters, increased cell adhesion, and increased inflammation. Loss of Fnip1 synergized with Tsc1 loss to hyperactivate mTOR, increase Erk activation, and greatly accelerate the development of PKD. Our results collectively define roles for Fnip1 in regulating kidney development and function, and provide a model for how loss of Fnip1 contributes to PKD and perhaps renal cell carcinoma.


Asunto(s)
Proteínas Portadoras/genética , Quistes/genética , Eliminación de Gen , Riñón/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Transcripción Genética/genética , Proteína 1 del Complejo de la Esclerosis Tuberosa/genética , Animales , Proteínas Portadoras/metabolismo , Quistes/patología , Activación Enzimática/genética , Células Epiteliales/metabolismo , Células Epiteliales/patología , Perfilación de la Expresión Génica , Genotipo , Riñón/crecimiento & desarrollo , Riñón/patología , Ratones , Tamaño de los Órganos/genética , Fosforilación Oxidativa , Proteína 1 del Complejo de la Esclerosis Tuberosa/deficiencia
19.
Cancer Discov ; 8(6): 750-763, 2018 06.
Artículo en Inglés | MEDLINE | ID: mdl-29563103

RESUMEN

Chimeric antigen receptor (CAR) T-cell immunotherapy has revolutionized the treatment of refractory leukemias and lymphomas, but is associated with significant toxicities, namely cytokine release syndrome (CRS) and neurotoxicity. A major barrier to developing therapeutics to prevent CAR T cell-mediated neurotoxicity is the lack of clinically relevant models. Accordingly, we developed a rhesus macaque (RM) model of neurotoxicity via adoptive transfer of autologous CD20-specific CAR T cells. Following cyclophosphamide lymphodepletion, CD20 CAR T cells expand to 272 to 4,450 cells/µL after 7 to 8 days and elicit CRS and neurotoxicity. Toxicities are associated with elevated serum IL6, IL8, IL1RA, MIG, and I-TAC levels, and disproportionately high cerebrospinal fluid (CSF) IL6, IL2, GM-CSF, and VEGF levels. During neurotoxicity, both CD20 CAR and non-CAR T cells accumulate in the CSF and in the brain parenchyma. This RM model demonstrates that CAR T cell-mediated neurotoxicity is associated with proinflammatory CSF cytokines and a pan-T cell encephalitis.Significance: We provide the first immunologically relevant, nonhuman primate model of B cell-directed CAR T-cell therapy-mediated CRS and neurotoxicity. We demonstrate CAR and non-CAR T-cell infiltration in the CSF and in the brain during neurotoxicity resulting in pan-encephalitis, accompanied by increased levels of proinflammatory cytokines in the CSF. Cancer Discov; 8(6); 750-63. ©2018 AACR.This article is highlighted in the In This Issue feature, p. 663.


Asunto(s)
Antígenos CD20/inmunología , Ciclofosfamida/administración & dosificación , Inmunoterapia Adoptiva/efectos adversos , Síndromes de Neurotoxicidad/inmunología , Receptores de Antígenos de Linfocitos T/inmunología , Animales , Línea Celular Tumoral , Ciclofosfamida/efectos adversos , Modelos Animales de Enfermedad , Humanos , Células K562 , Macaca mulatta , Síndromes de Neurotoxicidad/etiología , Trasplante Autólogo
20.
Physiol Rep ; 5(21)2017 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-29142002

RESUMEN

Staphylococcus aureus is an important cause of acute bacterial pneumonia. Toll-like receptor 2 (TLR2) recognizes multiple components of the bacterial cell wall and activates innate immune responses to gram-positive bacteria. We hypothesized that TLR2 would have an important role in pulmonary host defense against S. aureus TLR null (TLR2-/-) mice and wild type (WT) C57BL/6 controls were challenged with aerosolized S. aureus at a range of inocula for kinetic studies of cytokine and antimicrobial peptide expression, lung inflammation, bacterial killing by alveolar macrophages, and bacterial clearance. Survival was measured after intranasal infection. Pulmonary induction of most pro-inflammatory cytokines was significantly blunted in TLR2-/- mice 4 and 24 h after infection in comparison with WT controls. Bronchoalveolar concentrations of cathelicidin-related antimicrobial peptide also were reduced in TLR2-/- mice. Lung inflammation, measured by enumeration of bronchoalveolar neutrophils and scoring of histological sections, was significantly blunted in TLR2-/- mice. Phagocytosis of S. aureus by alveolar macrophages in vivo after low-dose infection was unimpaired, but viability of ingested bacteria was significantly greater in TLR2-/- mice. Bacterial clearance from the lungs was slightly impaired in TLR2-/- mice after low-dose infection only; bacterial elimination from the lungs was slightly accelerated in the TLR2-/- mice after high-dose infection. Survival after high-dose intranasal challenge was 50-60% in both groups. TLR2 has a significant role in early innate immune responses to S. aureus in the lungs but is not required for bacterial clearance and survival from S. aureus pneumonia.


Asunto(s)
Neumonía Estafilocócica/inmunología , Staphylococcus aureus , Receptor Toll-Like 2/inmunología , Animales , Péptidos Catiónicos Antimicrobianos/metabolismo , Carga Bacteriana , Líquido del Lavado Bronquioalveolar/inmunología , Recuento de Colonia Microbiana , Citocinas/biosíntesis , Femenino , Inmunidad Innata , Pulmón/microbiología , Macrófagos Alveolares/inmunología , Macrófagos Alveolares/microbiología , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Fagocitosis/inmunología , Neumonía Estafilocócica/microbiología , Neumonía Estafilocócica/patología , Staphylococcus aureus/crecimiento & desarrollo , Receptor Toll-Like 2/deficiencia , Catelicidinas
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