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1.
Drug Metab Dispos ; 50(5): 646-654, 2022 05.
Artículo en Inglés | MEDLINE | ID: mdl-35221288

RESUMEN

Drug-induced liver injury (DILI) remains one of the major concerns for healthcare providers and patients. Unfortunately, it is difficult to predict and prevent DILI in the clinic because detailed mechanisms of DILI are largely unknown. Many risk factors have been identified for both "intrinsic" and "idiosyncratic" DILI, suggesting that cofactors are an important aspect in understanding DILI. This review outlines the cofactors that potentiate DILI and categorizes them into two types: (1) the specific cofactors that target metabolic enzymes, transporters, antioxidation defense, immune response, and liver regeneration; and (2) the general cofactors that include inflammation, age, gender, comorbidity, gut microbiota, and lifestyle. The underlying mechanisms by which cofactors potentiate DILI are also discussed. SIGNIFICANCE STATEMENT: This review summarizes the risk factors for DILI, which can be used to predict and prevent DILI in the clinic. This work also highlights the gaps in the DILI field and provides future perspectives on the roles of cofactors in DILI.


Asunto(s)
Enfermedad Hepática Inducida por Sustancias y Drogas , Enfermedad Hepática Inducida por Sustancias y Drogas/etiología , Humanos , Inflamación , Hígado , Factores de Riesgo
2.
J Biol Chem ; 292(15): 6148-6162, 2017 04 14.
Artículo en Inglés | MEDLINE | ID: mdl-28228480

RESUMEN

The discovery and application of CRISPR/Cas9 technology for genome editing has greatly accelerated targeted mutagenesis in a variety of organisms. CRISPR/Cas9-mediated site-specific cleavage is typically exploited for the generation of insertions or deletions (indels) after aberrant dsDNA repair via the endogenous non-homology end-joining (NHEJ) pathway or, alternatively, for enhancing homology-directed repair to facilitate the generation of a specific mutation (or "knock-in"). However, there is a need for efficient cellular assays that can measure Cas9/guide RNA activity. Reliable methods for enriching and identifying desired mutants are also lacking. Here we describe a method using the Piggybac transposon for stable genomic integration of an H2B-GFP reporter or a hygromycin resistance gene for assaying Cas9 target cleavage and homology-directed repair. The H2B-GFP fusion protein provides increased stability and an obvious pattern of nuclear localization. This method, called SRIRACCHA (i.e. a stable, but reversible, integrated reporter for assaying CRISPR/Cas-stimulated HDR activity), enables the enrichment of mutants via selection of GFP-positive or hygromycin-resistant mammalian cells (immortalized or non-immortalized) as a surrogate for the modification of the endogenous target site. Currently available hyperactive Piggybac transposase mutants allow both delivery and removal of the surrogate reporters, with minimal risk of generating undesirable mutations. This assay permits rapid screening for efficient guide RNAs and the accelerated identification of mutant clones and is applicable to many cell types. We foresee the utility of this approach in contexts in which the maintenance of genomic integrity is essential, for example, when engineering cells for therapeutic purposes.


Asunto(s)
Sistemas CRISPR-Cas , Eliminación de Gen , Marcación de Gen/métodos , Vectores Genéticos/genética , Animales , Línea Celular Tumoral , Ratones
3.
Stem Cells ; 34(11): 2772-2783, 2016 11.
Artículo en Inglés | MEDLINE | ID: mdl-27350140

RESUMEN

In this study, we identify a novel and essential role for the Krüppel-like zinc finger transcription factor GLI-similar 3 (GLIS3) in the regulation of postnatal spermatogenesis. We show that GLIS3 is expressed in gonocytes, spermatogonial stem cells (SSCs) and spermatogonial progenitors (SPCs), but not in differentiated spermatogonia and later stages of spermatogenesis or in somatic cells. Spermatogenesis is greatly impaired in GLIS3 knockout mice. Loss of GLIS3 function causes a moderate reduction in the number of gonocytes, but greatly affects the generation of SSCs/SPCs, and as a consequence the development of spermatocytes. Gene expression profiling demonstrated that the expression of genes associated with undifferentiated spermatogonia was dramatically decreased in GLIS3-deficient mice and that the cytoplasmic-to-nuclear translocation of FOXO1, which marks the gonocyte-to-SSC transition and is necessary for SSC self-renewal, is inhibited. These observations suggest that GLIS3 promotes the gonocyte-to-SSC transition and is a critical regulator of the dynamics of early postnatal spermatogenesis. Stem Cells 2016;34:2772-2783.


Asunto(s)
Proteínas Represoras/genética , Espermatocitos/metabolismo , Espermatogénesis/genética , Espermatogonias/metabolismo , Células Madre/metabolismo , Testículo/metabolismo , Transactivadores/genética , Animales , Diferenciación Celular , Proteínas de Unión al ADN , Proteína Forkhead Box O1/genética , Proteína Forkhead Box O1/metabolismo , Perfilación de la Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Transporte de Proteínas , Proteínas Represoras/deficiencia , Espermatocitos/citología , Espermatogonias/citología , Células Madre/citología , Testículo/citología , Transactivadores/deficiencia
4.
NMR Biomed ; 28(5): 546-54, 2015 May.
Artículo en Inglés | MEDLINE | ID: mdl-25810360

RESUMEN

Polycystic kidney disease (PKD) is a life-threatening disease that leads to a grotesque enlargement of the kidney and significant loss of function. Several imaging studies with MRI have demonstrated that cyst size in polycystic kidneys can determine disease severity and progression. In the present study, we found that, although kidney volume and cyst volume decreased with drug treatment, renal function did not improve with treatment. Here, we applied dynamic contrast-enhanced MRI to study PKD in a Glis3 (GLI-similar 3)-deficient mouse model. Cysts from this model have a wide range of sizes and develop at an early age. To capture this crucial stage and assess cysts in detail, we imaged during early development (3-17 weeks) and applied high spatiotemporal resolution MRI (125 × 125 × 125 cubic microns every 7.7 s). A drug treatment with rapamycin (also known as sirolimus) was applied to determine whether disease progression could be halted. The effect and synergy (interaction) of aging and treatment were evaluated using an analysis of variance (ANOVA). Structural measurements, including kidney volume, cyst volume and cyst-to-kidney volume ratio, changed significantly with age. Drug treatment significantly decreased these metrics. Functional measurements of time-to-peak (TTP) mean and TTP variance were determined. TTP mean did not change with age, whereas TTP variance increased with age. Treatment with rapamycin generally did not affect these functional metrics. Synergistic effects of treatment and age were not found for any measurements. Together, the size and volume ratio of cysts decreased with drug treatment, whereas renal function remained the same. The quantification of renal structure and function with MRI can comprehensively assess the pathophysiology of PKD and response to treatment.


Asunto(s)
Imagenología Tridimensional/métodos , Imagen por Resonancia Magnética/métodos , Enfermedades Renales Poliquísticas/tratamiento farmacológico , Enfermedades Renales Poliquísticas/patología , Proteínas Represoras/genética , Sirolimus/uso terapéutico , Transactivadores/genética , Animales , Proteínas de Unión al ADN , Interpretación de Imagen Asistida por Computador/métodos , Inmunosupresores/uso terapéutico , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Resultado del Tratamiento
5.
J Perinatol ; 44(9): 1285-1290, 2024 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-38822149

RESUMEN

OBJECTIVE: We aim to describe neonatal respiratory outcomes following previable preterm premature rupture of membranes(PPROM) when gentle ventilation is utilized. We also report maternal morbidity and mortality. STUDY DESIGN: This is a retrospective single-center cohort study of infants delivered between 2016 and 2020 that included infants born at ≥23 weeks without major congenital anomaly after a pregnancy complicated with PPROM before 23 weeks gestation. Statistical analysis utilized unpaired Student's t-test or Mann-Whitney U-test when appropriate. RESULTS: 35 infants from 33 pregnancies were included. 91.4% of infants survived until discharge and 12.1% developed Bronchopulmonary Dysplasia (BPD). Those who developed BPD had significantly lower amniotic fluid levels prior to delivery (p < 0.05). There was no significant maternal morbidity or mortality in this cohort. CONCLUSION: This cohort had high survival and low rates of respiratory morbidities. This suggests the use of gentle ventilation might be the optimal strategy for patients born after previable PPROM.


Asunto(s)
Displasia Broncopulmonar , Rotura Prematura de Membranas Fetales , Humanos , Rotura Prematura de Membranas Fetales/terapia , Recién Nacido , Estudios Retrospectivos , Femenino , Embarazo , Displasia Broncopulmonar/etiología , Masculino , Respiración Artificial/efectos adversos , Edad Gestacional , Adulto , Recien Nacido Prematuro
6.
Proc Natl Acad Sci U S A ; 107(5): 2213-8, 2010 Feb 02.
Artículo en Inglés | MEDLINE | ID: mdl-20133865

RESUMEN

There are a number of challenges associated with designing nanoparticles for medical applications. We define two challenges here: (i) conventional targeting against up-regulated cell surface antigens is limited by heterogeneity in expression, and (ii) previous studies suggest that the optimal size of nanoparticles designed for systemic delivery is approximately 50-150 nm, yet this size range confers a high surface area-to-volume ratio, which results in fast diffusive drug release. Here, we achieve spatial control by biopanning a phage library to discover materials that target abundant vascular antigens exposed in disease. Next, we achieve temporal control by designing 60-nm hybrid nanoparticles with a lipid shell interface surrounding a polymer core, which is loaded with slow-eluting conjugates of paclitaxel for controlled ester hydrolysis and drug release over approximately 12 days. The nanoparticles inhibited human aortic smooth muscle cell proliferation in vitro and showed greater in vivo vascular retention during percutaneous angioplasty over nontargeted controls. This nanoparticle technology may potentially be used toward the treatment of injured vasculature, a clinical problem of primary importance.


Asunto(s)
Vasos Sanguíneos/efectos de los fármacos , Vasos Sanguíneos/lesiones , Sistemas de Liberación de Medicamentos , Nanopartículas/administración & dosificación , Secuencia de Aminoácidos , Animales , Antígenos/genética , Antígenos/metabolismo , Ingeniería Biomédica , Células Cultivadas , Preparaciones de Acción Retardada/administración & dosificación , Humanos , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/lesiones , Nanomedicina , Nanopartículas/química , Paclitaxel/administración & dosificación , Biblioteca de Péptidos , Ratas , Ratas Sprague-Dawley
7.
Exp Cell Res ; 316(4): 507-16, 2010 Feb 15.
Artículo en Inglés | MEDLINE | ID: mdl-20025870

RESUMEN

In this study, we demonstrate that protein kinase C (PKC) activators, including phorbol-12-myristate-13-acetate (PMA), 1,2-dioctanoyl-sn-glycerol (DOG), and platelet-derived growth factor alpha are potent inducers of angiopoietin-like protein 4 (ANGPTL4) expression in several normal lung cell types and carcinoma cell lines. In human airway smooth muscle (HASM) cells induction of ANGPTL4 expression is observed as early as 2 h after the addition of PMA. PMA also increases the level of ANGPTL4 protein released in the medium. PKC inhibitors Ro31-8820 and Gö6983 greatly inhibit the induction of ANGPTL4 mRNA by PMA suggesting that this up-regulation involves activation of PKC. Knockdown of several PKCs by corresponding siRNAs suggest a role for PKCalpha. PMA does not activate MAPK p38 and p38 inhibitors have little effect on the induction of ANGPTL4 indicating that p38 is not involved in the regulation of ANGPTL4 by PMA. In contrast, treatment of HASM by PMA induces phosphorylation and activation of Ra, MEK1/2, ERK1/2, JNK, Elk-1, and c-Jun. The Ras inhibitor manumycin A, the MEK1/2 inhibitor U0126, and the JNK inhibitor SP600125, greatly reduce the increase in ANGPTL4 expression by PMA. Knockdown of MEK1/2 and JNK1/2 expression by corresponding siRNAs inhibits the induction of ANGPTL4. Our observations suggest that the induction of ANGPTL4 by PMA in HASM involves the activation of PKC, ERK, and JNK pathways. This induction may play a role in tissue remodeling during lung injury and be implicated in several lung pathologies.


Asunto(s)
Angiopoyetinas/metabolismo , Pulmón/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Proteína Quinasa C/metabolismo , Acetato de Tetradecanoilforbol/análogos & derivados , Proteína 4 Similar a la Angiopoyetina , Angiopoyetinas/genética , Northern Blotting , Western Blotting , Línea Celular , Línea Celular Tumoral , Humanos , Pulmón/citología , Pulmón/efectos de los fármacos , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Acetato de Tetradecanoilforbol/farmacología
8.
Sci Rep ; 11(1): 10799, 2021 05 24.
Artículo en Inglés | MEDLINE | ID: mdl-34031475

RESUMEN

The impact of prematurity on human development and neonatal diseases, such as bronchopulmonary dysplasia, has been widely reported. However, little is known about the effects of prematurity on the programs of stem cell self-renewal and differentiation of the upper respiratory epithelium, which is key for adaptation to neonatal life. We developed a minimally invasive methodology for isolation of neonatal basal cells from nasopharyngeal (NP) aspirates and performed functional analysis in organotypic cultures to address this issue. We show that preterm NP progenitors have a markedly distinct molecular signature of abnormal proliferation and mitochondria quality control compared to term progenitors. Preterm progenitors had lower oxygen consumption at baseline and were unable to ramp up consumption to the levels of term cells when challenged. Although they formed a mucociliary epithelium, ciliary function tended to decline in premature cells as they differentiated, compared to term cells. Together, these differences suggested increased sensitivity of preterm progenitors to environmental stressors under non-homeostatic conditions.


Asunto(s)
Displasia Broncopulmonar/patología , Perfilación de la Expresión Génica/métodos , Nasofaringe/citología , Oxígeno/metabolismo , Células Madre/citología , Displasia Broncopulmonar/genética , Técnicas de Cultivo de Célula , Diferenciación Celular , Células Cultivadas , Regulación de la Expresión Génica , Humanos , Recién Nacido , Recien Nacido Prematuro , Nasofaringe/metabolismo , Análisis de Secuencia de ARN , Células Madre/metabolismo
9.
JAMA Pediatr ; 175(2): 157-167, 2021 02 01.
Artículo en Inglés | MEDLINE | ID: mdl-33044493

RESUMEN

Importance: Limited data on vertical and perinatal transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and health outcomes of neonates born to mothers with symptomatic or asymptomatic coronavirus disease 2019 (COVID-19) are available. Studies are needed to inform evidence-based infection prevention and control (IP&C) policies. Objective: To describe the outcomes of neonates born to mothers with perinatal SARS-CoV-2 infection and the IP&C practices associated with these outcomes. Design, Setting, and Participants: This retrospective cohort analysis reviewed the medical records for maternal and newborn data for all 101 neonates born to 100 mothers positive for or with suspected SARS-CoV-2 infection from March 13 to April 24, 2020. Testing for SARS-CoV-2 was performed using Cobas (Roche Diagnostics) or Xpert Xpress (Cepheid) assays. Newborns were admitted to well-baby nurseries (WBNs) (82 infants) and neonatal intensive care units (NICUs) (19 infants) in 2 affiliate hospitals at a large academic medical center in New York, New York. Newborns from the WBNs roomed-in with their mothers, who were required to wear masks. Direct breastfeeding after appropriate hygiene was encouraged. Exposures: Perinatal exposure to maternal asymptomatic/mild vs severe/critical COVID-19. Main Outcomes and Measures: The primary outcome was newborn SARS-CoV-2 testing results. Maternal COVID-19 status was classified as asymptomatic/mildly symptomatic vs severe/critical. Newborn characteristics and clinical courses were compared across maternal COVID-19 severity. Results: In total, 141 tests were obtained from 101 newborns (54 girls [53.5%]) on 0 to 25 days of life (DOL-0 to DOL-25) (median, DOL-1; interquartile range [IQR], DOL-1 to DOL-3). Two newborns had indeterminate test results, indicative of low viral load (2.0%; 95% CI, 0.2%-7.0%); 1 newborn never underwent retesting but remained well on follow-up, and the other had negative results on retesting. Maternal severe/critical COVID-19 was associated with newborns born approximately 1 week earlier (median gestational age, 37.9 [IQR, 37.1-38.4] vs 39.1 [IQR, 38.3-40.2] weeks; P = .02) and at increased risk of requiring phototherapy (3 of 10 [30.0%] vs 6 of 91 [7.0%]; P = .04) compared with newborns of mothers with asymptomatic/mild COVID-19. Fifty-five newborns were followed up in a new COVID-19 Newborn Follow-up Clinic at DOL-3 to DOL-10 and remained well. Twenty of these newborns plus 3 newborns followed up elsewhere had 32 nonroutine encounters documented at DOL-3 to DOL-25, and none had evidence of SARS-CoV-2 infection, including 6 with negative retesting results. Conclusions and Relevance: No clinical evidence of vertical transmission was identified in 101 newborns of mothers positive for or with suspected SARS-CoV-2 infection, despite most newborns rooming-in and direct breastfeeding practices.


Asunto(s)
Prueba de COVID-19/estadística & datos numéricos , COVID-19/transmisión , Transmisión Vertical de Enfermedad Infecciosa/estadística & datos numéricos , Complicaciones Infecciosas del Embarazo/epidemiología , Resultado del Embarazo/epidemiología , COVID-19/epidemiología , Femenino , Humanos , Recién Nacido , Masculino , Ciudad de Nueva York , Evaluación de Resultado en la Atención de Salud , Embarazo , Estudios Retrospectivos , SARS-CoV-2/aislamiento & purificación , Adulto Joven
10.
Cerebellum ; 9(3): 310-23, 2010 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-20393820

RESUMEN

Previously, deficiency in the expression of the nuclear orphan receptor TAK1 was found to be associated with delayed cerebellar granule cell migration and Purkinje cell maturation with a permanent deficit in foliation of lobules VI­VII, suggesting a role for TAK1 in cerebellum development. In this study, we confirm that TAK1-deficient (TAK1(−/−)) mice have a smaller cerebellum and exhibit a disruption of lobules VI­VII. We extended these studies and show that at postnatal day 7, TAK1(−/−) mice exhibit a delay in monolayer maturation of dysmorphic calbindin 28K-positive Purkinje cells. The astrocyte-specific glutamate transporter (GLAST) was expressed within Bergmann fibers and internal granule cell layer at significantly lower levels in the cerebellum of TAK1(−/−) mice. At PND21, Golgi-positive Purkinje cells in TAK1(−/−) mice displayed a smaller soma (18%) and shorter distance to first branch point (35%). Neuronal death was not observed in TAK1(−/−) mice at PND21; however, activated microglia were present in the cerebellum, suggestive of earlier cell death. These structural deficits in the cerebellum were not sufficient to alter motor strength, coordination, or activity levels; however, deficits in acoustic startle response, prepulse startle inhibition, and social interactions were observed. Reactions to a novel environment were inhibited in a light/dark chamber, open-field, and home-cage running wheel. TAK1(−/−) mice displayed a plateau in performance on the running wheel, suggesting a deficit in learning to coordinate performance on a motor task. These data indicate that TAK1 is an important transcriptional modulator of cerebellar development and neurodevelopmentally regulated behavior.


Asunto(s)
Conducta Animal/fisiología , Cerebelo/patología , Quinasas Quinasa Quinasa PAM/deficiencia , Neuroglía/patología , Animales , Cerebelo/metabolismo , Transportador 1 de Aminoácidos Excitadores/metabolismo , Aprendizaje/fisiología , Quinasas Quinasa Quinasa PAM/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Actividad Motora/fisiología , Neuroglía/metabolismo , Reflejo de Sobresalto/fisiología , Conducta Social
11.
Semin Perinatol ; 44(7): 151320, 2020 11.
Artículo en Inglés | MEDLINE | ID: mdl-33071033

RESUMEN

During the early months of the COVID-19 pandemic, infection prevention and control (IP&C) for women in labor and mothers and newborns during delivery and receiving post-partum care was quite challenging for staff, patients, and support persons due to a relative lack of evidence-based practices, high rates of community transmission, and shortages of personal protective equipment (PPE). We present our IP&C policies and procedures for the obstetrical population developed from mid-March to mid-May 2020 when New York City served as the epicenter of the pandemic in the U.S. For patients, we describe screening for COVID-19, testing for SARS-CoV-2, and clearing patients from COVID-19 precautions. For staff, we address self-monitoring for symptoms, PPE in different clinical scenarios, and reducing staff exposures to SARS-CoV-2. For visitors/support persons, we address limiting them in labor and delivery, the postpartum units, and the NICU to promote staff and patient safety. We describe management of SARS-CoV-2-positive mothers and their newborns in both the well-baby nursery and in the neonatal ICU. Notably, in the well-baby nursery we do not separate SARS-CoV-2-positive mothers from their newborns, but emphasize maternal mask use and social distancing by placing newborns in isolates and asking mothers to remain 6 feet away unless feeding or changing their newborn. We also encourage direct breastfeeding and do not advocate early bathing. Newborns of SARS-CoV-2-positive mothers are considered persons under investigation (PUIs) until 14 days of life, the duration of the incubation period for SARS-CoV-2. We share two models of community-based care for PUI neonates. Finally, we provide our strategies for enhancing communication and education during the early months of the pandemic.


Asunto(s)
COVID-19/prevención & control , Salas de Parto , Control de Infecciones/organización & administración , Unidades de Cuidado Intensivo Neonatal , Salas Cuna en Hospital , Política Organizacional , COVID-19/diagnóstico , COVID-19/terapia , COVID-19/transmisión , Humanos , Control de Infecciones/métodos , Máscaras , Tamizaje Masivo , Equipo de Protección Personal , Distanciamiento Físico , SARS-CoV-2 , Visitas a Pacientes
12.
Semin Perinatol ; 44(7): 151286, 2020 11.
Artículo en Inglés | MEDLINE | ID: mdl-32826081

RESUMEN

As the COVID-19 pandemic continues to spread worldwide, it is crucial that we determine populations that are at-risk and develop appropriate clinical care policies to protect them. While several respiratory illnesses are known to seriously impact pregnant women and newborns, preliminary data on the novel SARS-CoV-2 Coronavirus suggest that these groups are no more at-risk than the general population. Here, we review the available literature on newborns born to infected mothers and show that newborns of mothers with positive/suspected SARS-CoV-2 infection rarely acquire the disease or show adverse clinical outcomes. With this evidence in mind, it appears that strict postnatal care policies, including separating mothers and newborns, discouraging breastfeeding, and performing early bathing, may be more likely to adversely impact newborns than they are to reduce the low risk of maternal transmission of SARS-CoV-2 or the even lower risk of severe COVID-19 disease in otherwise healthy newborns.


Asunto(s)
Baños , Lactancia Materna , COVID-19/epidemiología , Transmisión Vertical de Enfermedad Infecciosa/estadística & datos numéricos , Aislamiento de Pacientes , Complicaciones Infecciosas del Embarazo , Femenino , Humanos , Recién Nacido , Política Organizacional , Atención Posnatal , Embarazo , Alojamiento Conjunto , SARS-CoV-2
13.
Cancer Res ; 67(16): 7929-36, 2007 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-17699800

RESUMEN

Farnesol (FOH) and other isoprenoid alcohols induce apoptosis in various carcinoma cells and inhibit tumorigenesis in several in vivo models. However, the mechanisms by which they mediate their effects are not yet fully understood. In this study, we show that FOH is an effective inducer of apoptosis in several lung carcinoma cells, including H460. This induction is associated with activation of several caspases and cleavage of poly(ADP-ribose) polymerase (PARP). To obtain insight into the mechanism involved in FOH-induced apoptosis, we compared the gene expression profiles of FOH-treated and control H460 cells by microarray analysis. This analysis revealed that many genes implicated in endoplasmic reticulum (ER) stress signaling, including ATF3, DDIT3, HERPUD1, HSPA5, XBP1, PDIA4, and PHLDA1, were highly up-regulated within 4 h of FOH treatment, suggesting that FOH-induced apoptosis involves an ER stress response. This was supported by observations showing that treatment with FOH induces splicing of XBP1 mRNA and phosphorylation of eIF2alpha. FOH induces activation of several mitogen-activated protein kinase (MAPK) pathways, including p38, MAPK/extracellular signal-regulated kinase (ERK) kinase (MEK)-ERK, and c-jun NH2-terminal kinase (JNK). Inhibition of MEK1/2 by U0126 inhibited the induction of ER stress response genes. In addition, knockdown of the MEK1/2 and JNK1/2 expression by short interfering RNA (siRNA) effectively inhibited the cleavage of caspase-3 and PARP and apoptosis induced by FOH. However, only MEK1/2 siRNAs inhibited the induction of ER stress-related genes, XBP1 mRNA splicing, and eIF2alpha phosphorylation. Our results show that FOH-induced apoptosis is coupled to ER stress and that activation of MEK1/2 is an early upstream event in the FOH-induced ER stress signaling cascade.


Asunto(s)
Apoptosis/efectos de los fármacos , Retículo Endoplásmico/efectos de los fármacos , Farnesol/farmacología , Neoplasias Pulmonares/tratamiento farmacológico , Línea Celular Tumoral , Retículo Endoplásmico/metabolismo , Chaperón BiP del Retículo Endoplásmico , Activación Enzimática/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Neoplasias Pulmonares/enzimología , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Proteínas Quinasas Activadas por Mitógenos/genética , Proteínas Quinasas Activadas por Mitógenos/metabolismo , ARN Interferente Pequeño/genética
14.
Cancer Res ; 67(14): 6647-56, 2007 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-17621610

RESUMEN

In this study, we examine the potential role of receptor-associated protein 80 (RAP80), a nuclear protein containing two ubiquitin-interacting motifs (UIM), in DNA damage response and double-strand break (DSB) repair. We show that following ionizing radiation and treatment with DNA-damaging agents, RAP80 translocates to discrete nuclear foci that colocalize with those of gamma-H2AX. The UIMs and the region of amino acids 204 to 304 are critical for the relocalization of RAP80 to ionizing radiation-induced foci (IRIF). These observations suggest that RAP80 becomes part of a DNA repair complex at the sites of IRIF. We also show that RAP80 forms a complex with the tumor repressor BRCA1 and that this interaction is mediated through the BRCA1 COOH-terminal repeats of BRCA1. The UIMs are not required for the interaction of RAP80 with BRCA1. Knockdown of RAP80 in HEK293 cells significantly reduced DSB-induced homology-directed recombination (HDR). Moreover, inhibition of RAP80 expression by small interfering RNA increased radiosensitivity, whereas increased radioresistance was observed in human breast cancer MCF-7 cells with overexpression of RAP80. Taken together, our data suggest that RAP80 plays an important role in DNA damage response signaling and HDR-mediated DSB repair. We further show that RAP80 can function as a substrate of the ataxia-telangiectasia mutated protein kinase in vitro, which phosphorylates RAP80 at Ser(205) and Ser(402). We show that this phosphorylation is not required for the migration of RAP80 to IRIF.


Asunto(s)
Proteína BRCA1/metabolismo , Proteínas Portadoras/fisiología , Reparación del ADN , Proteínas Nucleares/fisiología , Ubiquitina/química , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Proteínas de la Ataxia Telangiectasia Mutada , Proteínas Portadoras/metabolismo , Proteínas de Ciclo Celular/fisiología , Línea Celular Tumoral , Daño del ADN , Proteínas de Unión al ADN/fisiología , Chaperonas de Histonas , Humanos , Datos de Secuencia Molecular , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinasas/fisiología , Estructura Terciaria de Proteína , Transporte de Proteínas , Homología de Secuencia de Aminoácido , Transducción de Señal , Proteínas Supresoras de Tumor/fisiología
15.
J Clin Invest ; 127(12): 4326-4337, 2017 12 01.
Artículo en Inglés | MEDLINE | ID: mdl-29083325

RESUMEN

Deficiency in Krüppel-like zinc finger transcription factor GLI-similar 3 (GLIS3) in humans is associated with the development of congenital hypothyroidism. However, the functions of GLIS3 in the thyroid gland and the mechanism by which GLIS3 dysfunction causes hypothyroidism are unknown. In the current study, we demonstrate that GLIS3 acts downstream of thyroid-stimulating hormone (TSH) and TSH receptor (TSHR) and is indispensable for TSH/TSHR-mediated proliferation of thyroid follicular cells and biosynthesis of thyroid hormone. Using ChIP-Seq and promoter analysis, we demonstrate that GLIS3 is critical for the transcriptional activation of several genes required for thyroid hormone biosynthesis, including the iodide transporters Nis and Pds, both of which showed enhanced GLIS3 binding at their promoters. The repression of cell proliferation of GLIS3-deficient thyroid follicular cells was due to the inhibition of TSH-mediated activation of the mTOR complex 1/ribosomal protein S6 (mTORC1/RPS6) pathway as well as the reduced expression of several cell division-related genes regulated directly by GLIS3. Consequently, GLIS3 deficiency in a murine model prevented the development of goiter as well as the induction of inflammatory and fibrotic genes during chronic elevation of circulating TSH. Our study identifies GLIS3 as a key regulator of TSH/TSHR-mediated thyroid hormone biosynthesis and proliferation of thyroid follicular cells and uncovers a mechanism by which GLIS3 deficiency causes neonatal hypothyroidism and prevents goiter development.


Asunto(s)
Proliferación Celular , Receptores de Tirotropina/metabolismo , Proteínas Represoras/metabolismo , Glándula Tiroides/metabolismo , Hormonas Tiroideas/biosíntesis , Tirotropina/metabolismo , Transactivadores/metabolismo , Animales , Proteínas de Unión al ADN , Bocio/genética , Bocio/metabolismo , Bocio/prevención & control , Diana Mecanicista del Complejo 1 de la Rapamicina/genética , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Ratones , Ratones Noqueados , Regiones Promotoras Genéticas , Receptores de Tirotropina/genética , Proteínas Represoras/genética , Proteínas Ribosómicas/genética , Proteínas Ribosómicas/metabolismo , Transportadores de Sulfato/genética , Transportadores de Sulfato/metabolismo , Simportadores/genética , Simportadores/metabolismo , Glándula Tiroides/citología , Hormonas Tiroideas/genética , Tirotropina/genética , Transactivadores/genética
16.
Biochem Pharmacol ; 97(3): 256-68, 2015 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-26275811

RESUMEN

In this study, we demonstrate that treatment of T lymphoblastic leukemic Molt4 cells with farnesol activates the apoptosome via the intrinsic pathway of apoptosis. This induction was associated with changes in the level of intracellular potassium and calcium, the dissipation of the mitochondrial and plasma membrane potential, release of cytochrome c, activation of several caspases, and PARP cleavage. The induction of apoptosis by farnesol was inhibited by the addition of the pan-caspase inhibitor Z-VAD-fmk and by the exogenous expression of the anti-apoptotic protein Bcl2. Analysis of the gene expression profiles by microarray analysis revealed that farnesol increased the expression of several genes related to the unfolded protein response (UPR), including CHOP and CHAC1. This induction was associated with the activation of the PERK-eIF2α-ATF3/4 cascade, but not the XBP-1 branch of the UPR. Although farnesol induced activation of the ERK1/2, p38, and JNK pathways, inhibition of these MAPKs had little effect on farnesol-induced apoptosis or the induction of UPR-related genes. Our data indicate that the induction of apoptosis in leukemic cells by farnesol is mediated through a pathway that involves activation of the apoptosome via the intrinsic pathway and induction of the PERK-eIF2α-ATF3/4 cascade in a manner that is independent of the farnesol-induced activation of MAPKs.


Asunto(s)
Factor de Transcripción Activador 3/metabolismo , Factor de Transcripción Activador 4/metabolismo , Anticarcinógenos/farmacología , Apoptosis/efectos de los fármacos , Farnesol/farmacología , Factor de Transcripción CHOP/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Humanos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patología , Proteínas Proto-Oncogénicas c-bcl-2/genética , Especies Reactivas de Oxígeno/metabolismo , Transcriptoma/efectos de los fármacos , Transfección
17.
PLoS One ; 8(3): e58417, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-23505504

RESUMEN

CD44 is a multifunctional membrane receptor implicated in the regulation of several biological processes, including inflammation. CD44 expression is elevated in liver and white adipose tissue (WAT) during obesity suggesting a possible regulatory role for CD44 in metabolic syndrome. To study this hypothesis, we examined the effect of the loss of CD44 expression on the development of various features of metabolic syndrome using CD44 null mice. Our study demonstrates that CD44-deficient mice (CD44KO) exhibit a significantly reduced susceptibility to the development of high fat-diet (HFD)-induced hepatic steatosis, WAT-associated inflammation, and insulin resistance. The decreased expression of genes involved in fatty acid synthesis and transport (Fasn and Cd36), de novo triglyceride synthesis (Mogat1), and triglyceride accumulation (Cidea, Cidec) appears in part responsible for the reduced hepatic lipid accumulation in CD44KO(HFD) mice. In addition, the expression of various inflammatory and cell matrix genes, including several chemokines and its receptors, osteopontin, and several matrix metalloproteinases and collagen genes was greatly diminished in CD44KO(HFD) liver consistent with reduced inflammation and fibrogenesis. In contrast, lipid accumulation was significantly increased in CD44KO(HFD) WAT, whereas inflammation as indicated by the reduced infiltration of macrophages and expression of macrophage marker genes, was significantly diminished in WAT of CD44KO(HFD) mice compared to WT(HFD) mice. CD44KO(HFD) mice remained considerably more insulin sensitive and glucose tolerant than WT(HFD) mice and exhibited lower blood insulin levels. Our study indicates that CD44 plays a critical role in regulating several aspects of metabolic syndrome and may provide a new therapeutic target in the management of insulin resistance.


Asunto(s)
Dieta Alta en Grasa , Hígado Graso/genética , Receptores de Hialuranos/genética , Resistencia a la Insulina/genética , Paniculitis/genética , Adiposidad/genética , Animales , Peso Corporal/genética , Complejo CD3/metabolismo , Activación Enzimática , Hígado Graso/metabolismo , Hígado Graso/prevención & control , Perfilación de la Expresión Génica , Receptores de Hialuranos/metabolismo , Inflamación/genética , Inflamación/metabolismo , Hígado/enzimología , Hígado/lesiones , Linfocitos/inmunología , Linfocitos/metabolismo , Masculino , Ratones , Ratones Noqueados , Paniculitis/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo
18.
Int J Epidemiol ; 40(1): 65-101, 2011 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-21097954

RESUMEN

BACKGROUND: Despite the current obesity epidemic, maternal underweight remains a common occurrence with potential adverse perinatal outcomes. Our objective was to determine the relationship between maternal underweight and preterm birth (PTB) and low birth weight (LBW) in singleton pregnancies in developing and developed countries. METHODS: We followed the MOOSE consensus statement. We searched MEDLINE and EMBASE from their inceptions. We included studies that assessed the effect of maternal underweight compared with normal weight according to body mass index in singleton gestations on our two primary outcomes: PTB (<37 weeks) and LBW (<2500 g). Two assessors independently reviewed citations, extracted data and assessed quality. RESULTS: A total of 78 studies were included involving 1 025 794 women. The overall risk of PTB was increased in the cohort studies of underweight women [adjusted relative risk (RR) 1.29, 95% confidence interval (CI) 1.15-1.46], as were the risks of spontaneous PTB (adjusted RR 1.32, 95% CI 1.10-1.57) and induced PTB (adjusted RR 1.21, 95% CI 1.07-1.36). Underweight women had an increased risk of an LBW infant (adjusted RR 1.64, 95% CI 1.38-1.94). In developed countries, underweight women had an increased risk of PTB (RR 1.22, 95% CI 1.15-1.30) but not in developing countries (RR 0.99, 95% CI 0.67-1.45). In both developed and developing countries, underweight women were at increased risk of having an LBW infant (RR 1.48, 95% CI 1.29-1.68, and RR 1.52, 95% CI 1.25-1.85, respectively). CONCLUSIONS: In this systematic review and meta-analyses, we determined that singletons born to underweight women have higher risks of PTB (overall, spontaneous and induced) and LBW than those born to women with normal weight.


Asunto(s)
Recién Nacido de Bajo Peso , Madres , Nacimiento Prematuro/epidemiología , Delgadez , Índice de Masa Corporal , Femenino , Humanos , Recién Nacido , Embarazo , Resultado del Embarazo , Factores de Riesgo
19.
Diabetes ; 60(1): 177-88, 2011 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-20864514

RESUMEN

OBJECTIVE: The nuclear receptor TAK1/TR4/NR2C2 is expressed in several tissues that are important in the control of energy homeostasis. In this study, we investigate whether TAK1 functions as a regulator of lipid and energy homeostasis and has a role in metabolic syndrome. RESEARCH DESIGN AND METHODS: We generated TAK1-deficient (TAK1⁻(/)⁻) mice to study the function of TAK1 in the development of metabolic syndrome in aged mice and mice fed a high-fat diet (HFD). (Immuno)histochemical, biochemical, and gene expression profile analyses were performed to determine the effect of the loss of TAK1 expression on lipid homeostasis in liver and adipose tissues. In addition, insulin sensitivity, energy expenditure, and adipose-associated inflammation were compared in wild-type (WT) and TAK1⁻(/)⁻ mice fed a HFD. RESULTS: TAK1-deficient (TAK1⁻(/)⁻) mice are resistant to the development of age- and HFD-induced metabolic syndrome. Histo- and biochemical analyses showed significantly lower hepatic triglyceride levels and reduced lipid accumulation in adipose tissue in TAK1⁻(/)⁻ mice compared with WT mice. Gene expression profiling analysis revealed that the expression of several genes encoding proteins involved in lipid uptake and triglyceride synthesis and storage, including Cidea, Cidec, Mogat1, and CD36, was greatly decreased in the liver and primary hepatocytes of TAK1⁻(/)⁻ mice. Restoration of TAK1 expression in TAK1⁻(/)⁻ hepatocytes induced expression of several lipogenic genes. Moreover, TAK1⁻(/)⁻ mice exhibited reduced infiltration of inflammatory cells and expression of inflammatory genes in white adipose tissue, and were resistant to the development of glucose intolerance and insulin resistance. TAK1⁻(/)⁻ mice consume more oxygen and produce more carbon dioxide than WT mice, suggesting increased energy expenditure. CONCLUSIONS: Our data reveal that TAK1 plays a critical role in the regulation of energy and lipid homeostasis, and promotes the development of metabolic syndrome. TAK1 may provide a new therapeutic target in the management of obesity, diabetes, and liver steatosis.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/deficiencia , Hígado Graso/prevención & control , Inflamación/prevención & control , Obesidad/complicaciones , Receptores de Esteroides/deficiencia , Receptores de Hormona Tiroidea/deficiencia , Tejido Adiposo/anatomía & histología , Tejido Adiposo/patología , Animales , Grasas de la Dieta , Epidídimo , Hígado Graso/patología , Citometría de Flujo , Inflamación/patología , Resistencia a la Insulina , Masculino , Síndrome Metabólico/genética , Síndrome Metabólico/patología , Síndrome Metabólico/prevención & control , Ratones , Ratones Noqueados , Tamaño de los Órganos , ARN/genética , ARN/aislamiento & purificación , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
20.
Biomaterials ; 30(8): 1627-34, 2009 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-19111339

RESUMEN

Current approaches to encapsulate and deliver therapeutic compounds have focused on developing liposomal and biodegradable polymeric nanoparticles (NPs), resulting in clinically approved therapeutics such as Doxil/Caelyx and Genexol-PM, respectively. Our group recently reported the development of biodegradable core-shell NP systems that combined the beneficial properties of liposomal and polymeric NPs for controlled drug delivery. Herein we report the parameters that alter the biological and physicochemical characteristics, stability, drug release properties and cytotoxicity of these core-shell NPs. We further define scalable processes for the formulation of these NPs in a reproducible manner. These core-shell NPs consist of (i) a poly(D,L-lactide-co-glycolide) hydrophobic core, (ii) a soybean lecithin monolayer, and (iii) a poly(ethylene glycol) shell, and were synthesized by a modified nanoprecipitation method combined with self-assembly. Preparation of the NPs showed that various formulation parameters such as the lipid/polymer mass ratio and lipid/lipid-PEG molar ratio controlled NP physical stability and size. We encapsulated a model chemotherapy drug, docetaxel, in the NPs and showed that the amount of lipid coverage affected its drug release kinetics. Next, we demonstrated a potentially scalable process for the formulation, purification, and storage of NPs. Finally, we tested the cytotoxicity using MTT assays on two model human cell lines, HeLa and HepG2, and demonstrated the biocompatibility of these particles in vitro. Our data suggest that the PLGA-lecithin-PEG core-shell NPs may be a useful new controlled release drug delivery system.


Asunto(s)
Sistemas de Liberación de Medicamentos/métodos , Ácido Láctico/síntesis química , Lecitinas/síntesis química , Nanopartículas/química , Polietilenglicoles/síntesis química , Ácido Poliglicólico/síntesis química , Taxoides/administración & dosificación , Muerte Celular/efectos de los fármacos , Química Farmacéutica , Preparaciones de Acción Retardada/farmacología , Docetaxel , Estabilidad de Medicamentos , Células HeLa , Humanos , Cinética , Ácido Láctico/química , Ácido Láctico/farmacología , Lecitinas/química , Lecitinas/farmacología , Lípidos/química , Tamaño de la Partícula , Polietilenglicoles/química , Polietilenglicoles/farmacología , Ácido Poliglicólico/química , Ácido Poliglicólico/farmacología , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Taxoides/farmacología
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