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We recently described a subgroup of autopsied COVID-19 subjects (â¼40%), termed 'profibrotic phenotype,' who exhibited clusters of myofibroblasts (Mfbs), which were positive for the collagen-specific chaperone heat shock protein 47 (HSP47+) in situ. This report identifies increased, localized (hot spot restricted) expression of αSMA, COLα1, POSTN and FAP supporting the identity of HSP47+ cells as myofibroblasts and characterizing a profibrotic extracellular matrix (ECM) phenotype. Coupled with increased GRP78 in COVID-19 subjects, these data could reflect induction of the unfolded protein response for mitigation of proteostasis (i.e., protein homeostasis) dysfunction in discrete clusters of cells. ECM shifts in selected COVID-19 subjects occur without significant increases in either global trichrome positive staining or myocardial injury based quantitively on standard H&E scoring. Our findings also suggest distinct mechanism(s) for ECM remodeling in the setting of SARS-CoV-2 infection. The ratio of CD163+/CD68+ cells is increased in hot spots of profibrotic hearts compared with either controls or outside of hot spots in COVID-19 subjects. In sum, matrix remodeling of human COVID-19 hearts in situ is characterized by site-restricted profibrotic mediated (e.g., HSP47+ Mfbs, CD163+ Mφs) modifications in ECM (i.e., COLα1, POSTN, FAP), with a strong correlation between COLα1 and HSP47+cells within hot spots. Given the established associations of viral infection (e.g., human immunodeficiency virus; HIV), myocardial fibrosis and sudden cardiac death, early screening tools (e.g., plasma biomarkers, noninvasive cardiac magnetic resonance imaging) for diagnosis, monitoring and treatment of fibrotic ECM remodeling are warranted for COVID-19 high-risk populations.
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COVID-19 , Miofibroblastos , Humanos , Miofibroblastos/metabolismo , COVID-19/patología , SARS-CoV-2 , Corazón , Proteínas del Choque Térmico HSP47/genética , Proteínas del Choque Térmico HSP47/metabolismo , FibrosisRESUMEN
This study aims to assess the relationship between risk perception, attitude, and avoidance among residents toward an urban incinerator in Taichung, Taiwan. A cross-sectional study was conducted and three schools were enrolled. The case group was composed of 514 residents who live near an incinerator. The control group was composed of 264 people nearly the same age and who have lived in that area basically the same period of time. All participants were interviewed using a structured questionnaire. Results of this study showed that there was no significant difference between the exposure group and the control group in risk perception and attitude regarding the incinerator. However, the exposure group showed a significantly higher desire to move within one year or move sometime in the future than the control group. Therefore, these people should encourage the Environmental Protection Administration (EPA) to do everything it can to make sure that the incinerator operates safely.
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Reacción de Prevención , Exposición a Riesgos Ambientales , Conocimientos, Actitudes y Práctica en Salud , Incineración , Medición de Riesgo , Residuos Sólidos , Adulto , Estudios Transversales , Femenino , Humanos , Masculino , Persona de Mediana Edad , Percepción , TaiwánRESUMEN
Mutations of HSPB5 (also known as CRYAB or αB-crystallin), a bona fide heat shock protein and molecular chaperone encoded by the HSPB5 (crystallin, alpha B) gene, are linked to multisystem disorders featuring variable combinations of cataracts, cardiomyopathy, and skeletal myopathy. This study aimed to investigate the pathological mechanisms involved in an early-onset myofibrillar myopathy manifesting in a child harboring a homozygous recessive mutation in HSPB5, 343delT. To study HSPB5 343delT protein dynamics, we utilize model cell culture systems including induced pluripotent stem cells derived from the 343delT patient (343delT/343delT) along with isogenic, heterozygous, gene-corrected control cells (WT KI/343delT) and BHK21 cells, a cell line lacking endogenous HSPB5 expression. 343delT/343delT and WT KI/343delT-induced pluripotent stem cell-derived skeletal myotubes and cardiomyocytes did not express detectable levels of 343delT protein, contributable to the extreme insolubility of the mutant protein. Overexpression of HSPB5 343delT resulted in insoluble mutant protein aggregates and induction of a cellular stress response. Co-expression of 343delT with WT prevented visible aggregation of 343delT and improved its solubility. Additionally, in vitro refolding of 343delT in the presence of WT rescued its solubility. We demonstrate an interaction between WT and 343delT both in vitro and within cells. These data support a loss-of-function model for the myopathy observed in the patient because the insoluble mutant would be unavailable to perform normal functions of HSPB5, although additional gain-of-function effects of the mutant protein cannot be excluded. Additionally, our data highlight the solubilization of 343delT by WT, concordant with the recessive inheritance of the disease and absence of symptoms in carrier individuals.
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Cardiomiopatías/genética , Cardiomiopatías/metabolismo , Catarata/genética , Catarata/metabolismo , Enfermedades Musculares/genética , Enfermedades Musculares/metabolismo , Cadena B de alfa-Cristalina/genética , Cadena B de alfa-Cristalina/metabolismo , Cardiomiopatías/etiología , Catarata/etiología , Femenino , Homocigoto , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Masculino , Modelos Biológicos , Fibras Musculares Esqueléticas/metabolismo , Enfermedades Musculares/etiología , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Miocitos Cardíacos/metabolismo , Linaje , Agregación Patológica de Proteínas/genética , Agregación Patológica de Proteínas/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Eliminación de Secuencia , Solubilidad , Cadena B de alfa-Cristalina/químicaRESUMEN
BACKGROUND/AIMS: Resistance of leukemia stem cells (LSCs) to chemotherapy in patients with acute myeloid leukemia (AML) causes relapse of disease. Hedgehog (Hh) signaling plays a critical role in the maintenance and differentiation of cancer stem cells. Yet its role in AML remains controversial. The purpose of the present study is to investigate the role of GLI1, the transcriptional activator of Hh signaling, in AML progenitor cells and to explore the anti-AML effects of GLI small-molecule inhibitor GANT61. METHODS: The expression of GLI1 mRNA and protein were examined in AML progenitor cells and normal cells. The proliferation, colony formation, apoptosis and differentiation of AML progenitor cells were also analyzed in the presence of GANT61. RESULTS: Kasumi-1 and KG1a cells, containing more CD34+ cells, expressed higher level of GLI1 compared to U937 and NB4 cells with fewer CD34+ cells. Consistently, a positive correlation between the protein levels of GLI1 and CD34 was validated in the bone marrow mononuclear cells (BMMC) of AML patients tested. GANT61 inhibited the proliferation and colony formation in AML cell lines. Importantly, GANT61 induced apoptosis in CD34+ enriched Kasumi-1 and KG1a cells, whereas it induced differentiation in U937 and NB4 cells. Furthermore, GANT61 enhanced the cytotoxicity of cytarabine (Ara-c) in primary CD34+ AML cells, indicating that inhibition of GLI1 could be a promising strategy to enhance chemosensitivity. CONCLUSIONS: The present findings suggested that Hh signaling was activated in AML progenitor cells. GLI1 acted as a potential target for AML therapy.
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Antígenos CD34/metabolismo , Leucemia Mieloide Aguda/patología , Células Madre Neoplásicas/metabolismo , Proteína con Dedos de Zinc GLI1/antagonistas & inhibidores , Proteína con Dedos de Zinc GLI1/metabolismo , Adolescente , Adulto , Anciano , Apoptosis/efectos de los fármacos , Células de la Médula Ósea/citología , Células de la Médula Ósea/metabolismo , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Femenino , Puntos de Control de la Fase G1 del Ciclo Celular/efectos de los fármacos , Humanos , Leucemia Mieloide Aguda/metabolismo , Masculino , Persona de Mediana Edad , Células Madre Neoplásicas/citología , Piridinas/farmacología , Pirimidinas/farmacología , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Adulto Joven , Proteína con Dedos de Zinc GLI1/genéticaRESUMEN
Human induced pluripotent stem cell-derived cardiomyocyte (iPSC-CM)-based assays are emerging as a promising tool for the in vitro preclinical screening of QT interval-prolonging side effects of drugs in development. A major impediment to the widespread use of human iPSC-CM assays is the low throughput of the currently available electrophysiological tools. To test the precision and applicability of the near-infrared fluorescent voltage-sensitive dye 1-(4-sulfanatobutyl)-4-{ß[2-(di-n-butylamino)-6-naphthyl]butadienyl}quinolinium betaine (di-4-ANBDQBS) for moderate-throughput electrophysiological analyses, we compared simultaneous transmembrane voltage and optical action potential (AP) recordings in human iPSC-CM loaded with di-4-ANBDQBS. Optical AP recordings tracked transmembrane voltage with high precision, generating nearly identical values for AP duration (AP durations at 10%, 50%, and 90% repolarization). Human iPSC-CMs tolerated repeated laser exposure, with stable optical AP parameters recorded over a 30-min study period. Optical AP recordings appropriately tracked changes in repolarization induced by pharmacological manipulation. Finally, di-4-ANBDQBS allowed for moderate-throughput analyses, increasing throughput >10-fold over the traditional patch-clamp technique. We conclude that the voltage-sensitive dye di-4-ANBDQBS allows for high-precision optical AP measurements that markedly increase the throughput for electrophysiological characterization of human iPSC-CMs.
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2-Naftilamina/análogos & derivados , Potenciales de Acción , Colorantes Fluorescentes , Células Madre Pluripotentes Inducidas/fisiología , Miocitos Cardíacos/fisiología , Compuestos de Quinolinio/química , Imagen de Colorante Sensible al Voltaje/métodos , 2-Naftilamina/química , Diferenciación Celular , Humanos , Células Madre Pluripotentes Inducidas/citología , Rayos Infrarrojos , Miocitos Cardíacos/citologíaRESUMEN
2-Arylamino-4-aryl-pyrimidines were found to be potent inhibitors of PAK1 kinase. The synthesis and SAR are described. The incorporation of a bromide at the 5-position of the pyrimidine core and in combination with a 1,2-dimethylpiperazine pendant domain yielded a lead compound with potent PAK1 inhibition and anti-proliferative activity in various colon cancer cell lines.
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Compuestos de Anilina/síntesis química , Inhibidores de Proteínas Quinasas/síntesis química , Pirimidinonas/síntesis química , Quinasas p21 Activadas/antagonistas & inhibidores , Compuestos de Anilina/química , Compuestos de Anilina/toxicidad , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Humanos , Unión Proteica , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/toxicidad , Pirimidinonas/química , Pirimidinonas/toxicidad , Relación Estructura-Actividad , Quinasas p21 Activadas/metabolismoRESUMEN
Doxorubicin (DOX), one of the most effective and widely used anticancer drugs, has the major limitation of cancer treatment-related cardiotoxicity (CTRTOX) in the clinic. Reactive oxygen species (ROS) generation and mitochondrial dysfunction are well-known consequences of DOX-induced injury to cardiomyocytes. This study aimed to explore the mitochondrial functional consequences and associated mechanisms of pretreatment with carvedilol, a ß-blocking agent known to exert protection against DOX toxicity. When disease modeling was performed using cultured rat cardiac muscle cells (H9c2 cells) and human iPSC-derived cardiomyocytes (iPSC-CMs), we found that prophylactic carvedilol mitigated not only the DOX-induced suppression of mitochondrial function but that the mitochondrial functional readout of carvedilol-pretreated cells mimicked the readout of cells overexpressing the major regulator of mitochondrial biogenesis, PGC-1α. Carvedilol pretreatment reduces mitochondrial oxidants, decreases cell death in both H9c2 cells and human iPSC-CM and maintains the cellular 'redox poise' as determined by sustained expression of the redox sensor Keap1 and prevention of DOX-induced Nrf2 nuclear translocation. These results indicate that, in addition to the already known ROS-scavenging effects, carvedilol has a hitherto unrecognized pro-reducing property against the oxidizing conditions induced by DOX treatment, the sequalae of DOX-induced mitochondrial dysfunction and compromised cell viability. The novel findings of our preclinical studies suggest future trial design of carvedilol prophylaxis, such as prescreening for redox state, might be an alternative strategy for preventing oxidative stress writ large in lieu of the current lack of clinical evidence for ROS-scavenging agents.
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The LMNA gene encodes the nuclear envelope proteins Lamins A and C, which comprise a major part of the nuclear lamina, provide mechanical support to the nucleus, and participate in diverse intracellular signaling. LMNA mutations give rise to a collection of diseases called laminopathies, including dilated cardiomyopathy (LMNA-DCM) and muscular dystrophies. Although nuclear deformities are a hallmark of LMNA-DCM, the role of nuclear abnormalities in the pathogenesis of LMNA-DCM remains incompletely understood. Using induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) from LMNA mutant patients and healthy controls, we show that LMNA mutant iPSC-CM nuclei have altered shape or increased size compared to healthy control iPSC-CM nuclei. The LMNA mutation exhibiting the most severe nuclear deformities, R249Q, additionally caused reduced nuclear stiffness and increased nuclear fragility. Importantly, for all cell lines, the degree of nuclear abnormalities corresponded to the degree of Lamin A/C and Lamin B1 mislocalization from the nuclear envelope. The mislocalization was likely due to altered assembly of Lamin A/C. Collectively, these results point to the importance of correct lamin assembly at the nuclear envelope in providing mechanical stability to the nucleus and suggest that defects in nuclear lamina organization may contribute to the nuclear and cellular dysfunction in LMNA-DCM.
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Background Cardiac fibrosis complicates SARS-CoV-2 infections and has been linked to arrhythmic complications in survivors. Accordingly, we sought evidence of increased HSP47 (heat shock protein 47), a stress-inducible chaperone protein that regulates biosynthesis and secretion of procollagen in heart tissue, with the goal of elucidating molecular mechanisms underlying cardiac fibrosis in subjects with this viral infection. Methods and Results Using human autopsy tissue, immunofluorescence, and immunohistochemistry, we quantified Hsp47+ cells and collagen α 1(l) in hearts from people with SARS-CoV-2 infections. Because macrophages are also linked to inflammation, we measured CD163+ cells in the same tissues. We observed irregular groups of spindle-shaped HSP47+ and CD163+ cells as well as increased collagen α 1(I) deposition, each proximate to one another in "hot spots" of ≈40% of hearts after SARS-CoV-2 infection (HSP47+ P<0.05 versus nonfibrotics and P<0.001 versus controls). Because HSP47+ cells are consistent with myofibroblasts, subjects with hot spots are termed "profibrotic." The remaining 60% of subjects dying with COVID-19 without hot spots are referred to as "nonfibrotic." No control subject exhibited hot spots. Conclusions Colocalization of myofibroblasts, M2(CD163+) macrophages, and collagen α 1(l) may be the first evidence of a COVID-19-related "profibrotic phenotype" in human hearts in situ. The potential public health and diagnostic implications of these observations require follow-up to further define mechanisms of viral-mediated cardiac fibrosis.
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COVID-19 , Miofibroblastos , Humanos , Miofibroblastos/metabolismo , SARS-CoV-2 , Colágeno/metabolismo , Proteínas de Choque Térmico/metabolismo , Colágeno Tipo I/metabolismo , Fenotipo , Macrófagos/metabolismo , FibrosisRESUMEN
Mutations in the human LMNA gene cause a collection of diseases called laminopathies, which includes muscular dystrophy and dilated cardiomyopathy. The LMNA gene encodes lamins, filamentous proteins that form a meshwork on the inner side of the nuclear envelope. How mutant lamins cause muscle disease is not well understood, and treatment options are currently limited. To understand the pathological functions of mutant lamins so that therapies can be developed, we generated new Drosophila models and human iPS cell-derived cardiomyocytes. In the Drosophila models, muscle-specific expression of the mutant lamins caused nuclear envelope defects, cytoplasmic protein aggregation, activation of the Nrf2/Keap1 redox pathway, and reductive stress. These defects reduced larval motility and caused death at the pupal stage. Patient-derived cardiomyocytes expressing mutant lamins showed nuclear envelope deformations. The Drosophila models allowed for genetic and pharmacological manipulations at the organismal level. Genetic interventions to increase autophagy, decrease Nrf2/Keap1 signaling, or lower reducing equivalents partially suppressed the lethality caused by mutant lamins. Moreover, treatment of flies with pamoic acid, a compound that inhibits the NADPH-producing malic enzyme, partially suppressed lethality. Taken together, these studies have identified multiple new factors as potential therapeutic targets for LMNA-associated muscular dystrophy.
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Vaccinations are widely credited with reducing death rates from COVID-19, but the underlying host-viral mechanisms/interactions for morbidity and mortality of SARS-CoV-2 infection remain poorly understood. Acute respiratory distress syndrome (ARDS) describes the severe lung injury, which is pathologically associated with alveolar damage, inflammation, non-cardiogenic edema, and hyaline membrane formation. Because proteostatic pathways play central roles in cellular protection, immune modulation, protein degradation, and tissue repair, we examined the pathological features for the unfolded protein response (UPR) using the surrogate biomarker glucose-regulated protein 78 (GRP78) and co-receptor for SARS-CoV-2. At autopsy, immunostaining of COVID-19 lungs showed highly elevated expression of GRP78 in both pneumocytes and macrophages compared with that of non-COVID control lungs. GRP78 expression was detected in both SARS-CoV-2-infected and un-infected pneumocytes as determined by multiplexed immunostaining for nucleocapsid protein. In macrophages, immunohistochemical staining for GRP78 from deceased COVID-19 patients was increased but overlapped with GRP78 expression taken from surgical resections of non-COVID-19 controls. In contrast, the robust in situ GRP78 immunostaining of pneumocytes from COVID-19 autopsies exhibited no overlap and was independent of age, race/ethnicity, and gender compared with that from non-COVID-19 controls. Our findings bring new insights for stress-response pathways involving the proteostatic network implicated for host resilience and suggest that targeting of GRP78 expression with existing therapeutics might afford an alternative therapeutic strategy to modulate host-viral interactions during SARS-CoV-2 infections.
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Células Epiteliales Alveolares/metabolismo , COVID-19/metabolismo , Estrés del Retículo Endoplásmico , Proteínas de Choque Térmico/análisis , Receptores de Coronavirus/análisis , SARS-CoV-2/patogenicidad , Adulto , Anciano , Anciano de 80 o más Años , Células Epiteliales Alveolares/patología , Células Epiteliales Alveolares/virología , Autopsia , COVID-19/mortalidad , COVID-19/patología , COVID-19/virología , Estudios de Casos y Controles , Chaperón BiP del Retículo Endoplásmico , Femenino , Interacciones Huésped-Patógeno , Humanos , Macrófagos Alveolares/metabolismo , Macrófagos Alveolares/virología , Masculino , Persona de Mediana Edad , Proteostasis , Regulación hacia Arriba , Adulto JovenRESUMEN
The genetic architecture of atrial fibrillation (AF) encompasses low impact, common genetic variants and high impact, rare variants. Here, we characterize a high impact AF-susceptibility allele, KCNQ1 R231H, and describe its transcontinental geographic distribution and history. Induced pluripotent stem cell-derived cardiomyocytes procured from risk allele carriers exhibit abbreviated action potential duration, consistent with a gain-of-function effect. Using identity-by-descent (IBD) networks, we estimate the broad- and fine-scale population ancestry of risk allele carriers and their relatives. Analysis of ancestral migration routes reveals ancestors who inhabited Denmark in the 1700s, migrated to the Northeastern United States in the early 1800s, and traveled across the Midwest to arrive in Utah in the late 1800s. IBD/coalescent-based allele dating analysis reveals a relatively recent origin of the AF risk allele (~5000 years). Thus, our approach broadens the scope of study for disease susceptibility alleles to the context of human migration and ancestral origins.
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Fibrilación Atrial/genética , Predisposición Genética a la Enfermedad/genética , Canal de Potasio KCNQ1/genética , Mutación Missense , Polimorfismo de Nucleótido Simple , Potenciales de Acción , Alelos , Dinamarca , Emigrantes e Inmigrantes , Femenino , Genotipo , Geografía , Humanos , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/metabolismo , Masculino , Persona de Mediana Edad , Miocitos Cardíacos/citología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/fisiología , Linaje , Factores de Riesgo , UtahRESUMEN
Mutations in B cell lymphoma 2-associated athanogene 3 (BAG3) are recurrently associated with dilated cardiomyopathy (DCM) and muscular dystrophy. Using isogenic genome-edited human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), we examined how a DCM-causing BAG3 mutation (R477H), as well as complete loss of BAG3 (KO), impacts myofibrillar organization and chaperone networks. Although unchanged at baseline, fiber length and alignment declined markedly in R477H and KO iPSC-CMs following proteasome inhibition. RNA sequencing revealed extensive baseline changes in chaperone- and stress response protein-encoding genes, and protein levels of key BAG3 binding partners were perturbed. Molecular dynamics simulations of the BAG3-HSC70 complex predicted a partial disengagement by the R477H mutation. In line with this, BAG3-R477H bound less HSC70 than BAG3-WT in coimmunoprecipitation assays. Finally, myofibrillar disarray triggered by proteasome inhibition in R477H cells was mitigated by overexpression of the stress response protein heat shock factor 1 (HSF1). These studies reveal the importance of BAG3 in coordinating protein quality control subsystem usage within the cardiomyocyte and suggest that augmenting HSF1 activity might be beneficial as a means to mitigate proteostatic stress in the context of BAG3-associated DCM.
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Proteínas Adaptadoras Transductoras de Señales , Proteínas Reguladoras de la Apoptosis , Cardiomiopatía Dilatada/genética , Células Madre Pluripotentes Inducidas/metabolismo , Miocitos Cardíacos/metabolismo , Proteínas Adaptadoras Transductoras de Señales/química , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Reguladoras de la Apoptosis/química , Proteínas Reguladoras de la Apoptosis/genética , Proteínas Reguladoras de la Apoptosis/metabolismo , Cardiomiopatía Dilatada/metabolismo , Edición Génica , Técnicas de Inactivación de Genes , Proteínas del Choque Térmico HSC70/química , Proteínas del Choque Térmico HSC70/genética , Proteínas del Choque Térmico HSC70/metabolismo , Humanos , Simulación de Dinámica Molecular , Mutación Missense/genéticaRESUMEN
Recently, certain studies have demonstrated in vitro that prostaglandin E2 (PGE2) promotes human cluster of differentiation (CD)34+ cell homing. However, the subtype receptors activated by PGE2 are unknown, as the PGE2 receptor EP1-4 subtypes (EP1-4) are expressed on the membrane of human CD34+ cells. Based on the above, the present study aimed to screen the receptor subtype activity by PGE2 to promote human CD34+ cell homing. It was observed that human CD34+ cells expressed the four PGE2 subreceptors, particularly EP2 and 4. PGE2 increased EP2 and 4 mRNA expression significantly, while EP1 and 3 mRNA exhibited no significant alteration. PGE2, EP2 agonist (EP2A), and EP4A upregulated CXC chemokine receptor 4 mRNA and protein expression in human CD34+ cells, and promoted stromal cellderived factor 1α (SDF1α) expression in bone marrow mesenchymal stem cells (BMMSCs). These phenomena were inhibited by the associated receptor antagonists. PGE2, EP2A, and EP4A facilitated human CD34+ cell migration towards SDF1α and BMMSCs. The results of the present study suggested that PGE2 promoted human CD34+ cell homing through EP2 and 4 receptors in vitro.
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Antígenos CD34/metabolismo , Dinoprostona/metabolismo , Células Madre Hematopoyéticas/metabolismo , Subtipo EP2 de Receptores de Prostaglandina E/metabolismo , Subtipo EP4 de Receptores de Prostaglandina E/metabolismo , Biomarcadores , Movimiento Celular , Células Cultivadas , Quimiocina CXCL12/genética , Quimiocina CXCL12/metabolismo , Dinoprostona/farmacología , Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Trasplante de Células Madre Hematopoyéticas , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/efectos de los fármacos , Humanos , Inmunofenotipificación , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Fenotipo , Receptores CXCR4/genética , Receptores CXCR4/metabolismo , Subtipo EP2 de Receptores de Prostaglandina E/genética , Subtipo EP4 de Receptores de Prostaglandina E/genéticaRESUMEN
BACKGROUND: Hemophagocytic lymphohistiocytosis (HLH) is a rare and potentially fatal condition that can be primary or secondary. Secondary HLH due to Mycobacterium tuberculosis (TB) is uncommon. We report a case of tuberculosis-associated HLH in an umbilical cord blood transplant (UCBT) recipient and discuss its clinical characteristics and challenges. METHODS: Hematologic investigations, bone marrow aspirates, Xpert MTB/RIF test of TB with peripheral blood were performed. Immune modulation with anti-TB therapy was initiated. RESULTS: Subsequent treatment with anti-TB treatment resulted rapid clinical response and disease remission. CONCLUSION: It is important to consider TB as one of the underlying cause of HLH in high-risk patients, particularly those in immunodeficient states. Early diagnosis and treatment can improve the survival rates of patients with tuberculosis-associated HLH.
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Sangre Fetal/trasplante , Linfohistiocitosis Hemofagocítica/complicaciones , Receptores de Trasplantes , Tuberculosis/complicaciones , Adulto , Femenino , Humanos , Linfohistiocitosis Hemofagocítica/sangre , Linfohistiocitosis Hemofagocítica/diagnóstico , Linfohistiocitosis Hemofagocítica/terapia , Mycobacterium tuberculosis/efectos de los fármacos , Mycobacterium tuberculosis/fisiología , Tuberculosis/tratamiento farmacológicoRESUMEN
Genome editing in induced pluripotent stem cells is currently hampered by the laborious and expensive nature of identifying homology-directed repair (HDR)-modified cells. We present an approach where isolation of cells bearing a selectable, HDR-mediated editing event at one locus enriches for HDR-mediated edits at additional loci. This strategy, called co-targeting with selection, improves the probability of isolating cells bearing HDR-mediated variants and accelerates the production of disease models.
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Edición Génica , Marcación de Gen , Genoma Humano , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/metabolismo , Sistemas CRISPR-Cas , Línea Celular , Reparación del ADN por Unión de Extremidades , Técnicas de Sustitución del Gen , Vectores Genéticos , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Células Madre Pluripotentes Inducidas/efectos de los fármacos , Reparación del ADN por RecombinaciónRESUMEN
Pim-1 has emerged as an attractive target for developing therapeutic agents for treating disorders involving abnormal cell growth, especially cancers. Herein we present lead optimization, chemical synthesis and biological evaluation of pyrazolo[1,5-a]pyrimidine compounds as potent and selective inhibitors of Pim-1 starting from a hit from virtual screening. These pyrazolo[1,5-a]pyrimidine compounds strongly inhibited Pim-1 and Flt-3 kinases. Selected compounds suppressed both the phosphorylation of BAD protein in a cell-based assay and 2-dimensional colony formation in a clonogenic cell survival assay at submicromolar potency, suggesting that cellular activity was mediated through inhibition of Pim-1. Moreover, these Pim-1 inhibitors did not show significant hERG inhibition at 30 µM concentration. The lead compound proved to be highly selective against a panel of 119 oncogenic kinases, indicating it had an improved safety profile compared with the first generation Pim-1 inhibitor SGI-1776.
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To determine HLA-A genetic polymorphism in Chinese populations and establish ethnic genetic database, 165 Han and 162 Uygur subjects were investigated with a non-isotopic and sensitive method PCR-SSOP. 22 alleles were identified in Han with the most frequent allele being HLA-A * 1101 (19.7%), followed by * 0201 (12.72%). Also, 22 alleles were identified in Uygur with * 2407 (17.90%) being the most frequent one and the frequencies of following alleles: * 0201, * 0101, * 3301 were higher than 10%. HLA-A * 0203, * 0205, * 0210, * 0302, * 2403 and * 3302 were only detected in Han; meanwhile * 0205, * 0211, * 2301, * 2502, * 68012 and * 6802 were only in Uygur. According to Hardy-Weinberg equilibrium, each allele showed no significant (P > 0.05) deviation between the expected frequency and the observed one. Heterozygosity (H), discrimination power (DP) and probability of paternity exclusion (EPP) of HLA-A locus from Han nationality were computed to be 0.9029, 0.9776 and 0.8592; and those from Uygur as 0.9063, 0.9379 and 0.7885. These results suggest that HLA-A DNA polymorphism and the database of two Chinese populations have useful applications in processing forensic casework (as personal identification, paternity test), tracing population migration and genetic diagnosis.
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Antígenos HLA-A/genética , Reacción en Cadena de la Polimerasa/métodos , Alelos , China , ADN/genética , Sondas de ADN/genética , Frecuencia de los Genes , Genotipo , Humanos , Hibridación de Ácido Nucleico/métodos , Polimorfismo GenéticoRESUMEN
The HLA-Cw loci polymorphism in Uygur population was investigated using the PCR- sequence specific oligonucleotide probe (SSOP) method,and the genetic database on the distribution of gene frequency of the HLA-Cw loci was established. From 146 individuals of Uygur population,18 HLA-Cw alleles were detected. The gene frequency was from 0.0069 to 0.2460. The four most common alleles were HLA-Cw*04(24.60%),07(11.51%),08(10 10%),14(12.02%),and they covered 58.23% of total alleles detected from Uygur population.We have made a survey of HLA-Cw alleles frequencies in a Uygur population,with blank frequency being lowered to 0.0064. The distribution of genotype frequencies met the law of Hardy-Weinberg equilibrium by hi-square test. The frequency data can be used in forensic and paternity tests to estimate the frequency of a DNA profile in the Uygur population,transplant matching and anthropology.
RESUMEN
Advances in induced pluripotent stem cell (iPSC) technology have set the stage for routine derivation of patient- and disease-specific human iPSC-cardiomyocyte (CM) models for preclinical drug screening and personalized medicine approaches. Peripheral blood mononuclear cells (PBMCs) are an advantageous source of somatic cells because they are easily obtained and readily amenable to transduction. Here, we report that the electrophysiological properties and pharmacological responses of PBMC-derived iPSC CM are generally similar to those of iPSC CM derived from other somatic cells, using patch-clamp, calcium transient, and multielectrode array (MEA) analyses. Distinct iPSC lines derived from a single patient display similar electrophysiological features and pharmacological responses. Finally, we demonstrate that human iPSC CMs undergo acute changes in calcium-handling properties and gene expression in response to rapid electrical stimulation, laying the foundation for an in-vitro-tachypacing model system for the study of human tachyarrhythmias.