RESUMEN
Mucopolysaccharidosis type II (MPS II) is an X-linked inherited disease caused by pathogenic variants in the IDS gene, leading to deficiency of the lysosomal enzyme iduronate-2-sulfatase and consequent widespread storage of glycosaminoglycans, leading to several clinical consequences, with progressive manifestations which most times includes cognitive decline. MPS II has wide allelic and clinical heterogeneity and a complex genotype-phenotype correlation. We evaluated data from 501 Brazilian patients diagnosed with MPS II from 1982 to 2020. We genotyped 280 of these patients (55.9%), which were assigned to 206 different families. Point mutations were present in 70% of our patients, being missense variants the most frequent. We correlated the IDS pathogenic variants identified with the phenotype (neuronophatic or non-neuronopathic). Except for two half-brothers, there was no discordance in the genotype-phenotype correlation among family members, nor among MPS II patients from different families with the same single base-pair substitution variant. Mothers were carriers in 82.0% of the cases. This comprehensive study of the molecular profile of the MPS II cases in Brazil sheds light on the genotype-phenotype correlation and helps the better understanding of the disease and the prediction of its clinical course, enabling the provision of a more refined genetic counseling to the affected families.
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Mucopolisacaridosis II , Brasil , Genotipo , Humanos , Masculino , Mucopolisacaridosis II/genética , Mutación , FenotipoRESUMEN
The linear ubiquitin chain assembly complex (LUBAC) is the only known E3 ubiquitin ligase which catalyses the generation of linear ubiquitin linkages de novo LUBAC is a crucial component of various immune receptor signalling pathways. Here, we show that LUBAC forms part of the TRAIL-R-associated complex I as well as of the cytoplasmic TRAIL-induced complex II In both of these complexes, HOIP limits caspase-8 activity and, consequently, apoptosis whilst being itself cleaved in a caspase-8-dependent manner. Yet, by limiting the formation of a RIPK1/RIPK3/MLKL-containing complex, LUBAC also restricts TRAIL-induced necroptosis. We identify RIPK1 and caspase-8 as linearly ubiquitinated targets of LUBAC following TRAIL stimulation. Contrary to its role in preventing TRAIL-induced RIPK1-independent apoptosis, HOIP presence, but not its activity, is required for preventing necroptosis. By promoting recruitment of the IKK complex to complex I, LUBAC also promotes TRAIL-induced activation of NF-κB and, consequently, the production of cytokines, downstream of FADD, caspase-8 and cIAP1/2. Hence, LUBAC controls the TRAIL signalling outcome from complex I and II, two platforms which both trigger cell death and gene activation.
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Muerte Celular , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Activación Transcripcional , Ubiquitina-Proteína Ligasas/metabolismo , Línea Celular , HumanosRESUMEN
BACKGROUND: Minocycline (MIN) is a tetracycline with antioxidant, anti-inflammatory, and neuroprotective properties. Given the likely involvement of inflammation and oxidative stress (IOS) in schizophrenia, MIN has been proposed as a potential adjuvant treatment in this pathology. We tested an early therapeutic window, during adolescence, as prevention of the schizophrenia-related deficits in the maternal immune stimulation (MIS) animal model. METHODS: On gestational day 15, Poly I:C or vehicle was injected in pregnant Wistar rats. A total 93 male offspring received MIN (30 mg/kg) or saline from postnatal day (PND) 35-49. At PND70, rats were submitted to the prepulse inhibition test. FDG-PET and T2-weighted MRI brain studies were performed at adulthood. IOS markers were evaluated in frozen brain tissue. RESULTS: MIN treatment did not prevent prepulse inhibition test behavioral deficits in MIS offspring. However, MIN prevented morphometric abnormalities in the third ventricle but not in the hippocampus. Additionally, MIN reduced brain metabolism in cerebellum and increased it in nucleus accumbens. Finally, MIN reduced the expression of iNOS (prefrontal cortex, caudate-putamen) and increased the levels of KEAP1 (prefrontal cortex), HO1 and NQO1 (amygdala, hippocampus), and HO1 (caudate-putamen). CONCLUSIONS: MIN treatment during adolescence partially counteracts volumetric abnormalities and IOS deficits in the MIS model, likely via iNOS and Nrf2-ARE pathways, also increasing the expression of cytoprotective enzymes. However, MIN treatment during this peripubertal stage does not prevent sensorimotor gating deficits. Therefore, even though it does not prevent all the MIS-derived abnormalities evaluated, our results suggest the potential utility of early treatment with MIN in other schizophrenia domains.
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Antiinflamatorios/farmacología , Antioxidantes/farmacología , Encefalopatías Metabólicas/tratamiento farmacológico , Minociclina/farmacología , Malformaciones del Sistema Nervioso/patología , Trastornos del Neurodesarrollo/tratamiento farmacológico , Estrés Oxidativo/efectos de los fármacos , Efectos Tardíos de la Exposición Prenatal/tratamiento farmacológico , Inhibición Prepulso/efectos de los fármacos , Esquizofrenia/tratamiento farmacológico , Animales , Antiinflamatorios/administración & dosificación , Antioxidantes/administración & dosificación , Conducta Animal/efectos de los fármacos , Encefalopatías Metabólicas/etiología , Modelos Animales de Enfermedad , Femenino , Imagen por Resonancia Magnética , Masculino , Minociclina/administración & dosificación , Malformaciones del Sistema Nervioso/diagnóstico por imagen , Malformaciones del Sistema Nervioso/etiología , Trastornos del Neurodesarrollo/inducido químicamente , Trastornos del Neurodesarrollo/inmunología , Tomografía de Emisión de Positrones , Embarazo , Efectos Tardíos de la Exposición Prenatal/inducido químicamente , Efectos Tardíos de la Exposición Prenatal/inmunología , Ratas , Ratas Wistar , Esquizofrenia/inducido químicamente , Esquizofrenia/inmunologíaRESUMEN
BACKGROUND AND PURPOSE: Minocycline is a broad-spectrum antibiotic, effective as a chronic treatment for recurrent bacterial infections. Beyond its antibiotic action, minocycline also has important anti-inflammatory, antioxidant and antiapoptotic properties. Its efficacy has therefore been evaluated in many neurodegenerative and psychiatric diseases that have an inflammatory basis. Our aim was to review preclinical and clinical studies performed in neurological and psychiatric diseases whose treatment involved the use of minocycline and thereby to discern the possible beneficial effect of minocycline in these disorders. METHODS: Completed and ongoing preclinical studies and clinical trials of minocycline for both neurodegenerative diseases and psychiatric disorders, published from January 1995 to January 2020, were identified through searching relevant databases (https://www.ncbi.nlm.nih.gov/pubmed/, https://clinicaltrials.gov/). A total of 74 preclinical studies and 44 clinical trials and open-label studies were selected. RESULTS: The results of the nearly 20 years of research identified are diverse. While minocycline mostly proved to be effective in animal models, clinical results showed divergent outcomes, with positive results in some studies counterbalanced by a number of cases with no significant improvements. Specific data for each disease are further individually described in this review. CONCLUSIONS: Despite minocycline demonstrating antioxidant and anti-inflammatory effects, discrepancies between preclinical and clinical data indicate that we should be cautious in analyzing the outcomes. Improving and standardizing protocols and refining animal models could help us to determine if minocycline really is a useful drug in the treatment of these pathologies.
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Trastornos Mentales , Enfermedades Neurodegenerativas , Animales , Antibacterianos/uso terapéutico , Antiinflamatorios/uso terapéutico , Humanos , Trastornos Mentales/tratamiento farmacológico , Minociclina/uso terapéutico , Enfermedades Neurodegenerativas/tratamiento farmacológicoRESUMEN
Bi-allelic loss-of-function variants in genes that encode subunits of the adaptor protein complex 4 (AP-4) lead to prototypical yet poorly understood forms of childhood-onset and complex hereditary spastic paraplegia: SPG47 (AP4B1), SPG50 (AP4M1), SPG51 (AP4E1) and SPG52 (AP4S1). Here, we report a detailed cross-sectional analysis of clinical, imaging and molecular data of 156 patients from 101 families. Enrolled patients were of diverse ethnic backgrounds and covered a wide age range (1.0-49.3 years). While the mean age at symptom onset was 0.8 ± 0.6 years [standard deviation (SD), range 0.2-5.0], the mean age at diagnosis was 10.2 ± 8.5 years (SD, range 0.1-46.3). We define a set of core features: early-onset developmental delay with delayed motor milestones and significant speech delay (50% non-verbal); intellectual disability in the moderate to severe range; mild hypotonia in infancy followed by spastic diplegia (mean age: 8.4 ± 5.1 years, SD) and later tetraplegia (mean age: 16.1 ± 9.8 years, SD); postnatal microcephaly (83%); foot deformities (69%); and epilepsy (66%) that is intractable in a subset. At last follow-up, 36% ambulated with assistance (mean age: 8.9 ± 6.4 years, SD) and 54% were wheelchair-dependent (mean age: 13.4 ± 9.8 years, SD). Episodes of stereotypic laughing, possibly consistent with a pseudobulbar affect, were found in 56% of patients. Key features on neuroimaging include a thin corpus callosum (90%), ventriculomegaly (65%) often with colpocephaly, and periventricular white-matter signal abnormalities (68%). Iron deposition and polymicrogyria were found in a subset of patients. AP4B1-associated SPG47 and AP4M1-associated SPG50 accounted for the majority of cases. About two-thirds of patients were born to consanguineous parents, and 82% carried homozygous variants. Over 70 unique variants were present, the majority of which are frameshift or nonsense mutations. To track disease progression across the age spectrum, we defined the relationship between disease severity as measured by several rating scales and disease duration. We found that the presence of epilepsy, which manifested before the age of 3 years in the majority of patients, was associated with worse motor outcomes. Exploring genotype-phenotype correlations, we found that disease severity and major phenotypes were equally distributed among the four subtypes, establishing that SPG47, SPG50, SPG51 and SPG52 share a common phenotype, an 'AP-4 deficiency syndrome'. By delineating the core clinical, imaging, and molecular features of AP-4-associated hereditary spastic paraplegia across the age spectrum our results will facilitate early diagnosis, enable counselling and anticipatory guidance of affected families and help define endpoints for future interventional trials.
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Complejo 4 de Proteína Adaptadora/genética , Cuerpo Calloso/diagnóstico por imagen , Imagen por Resonancia Magnética/tendencias , Paraplejía Espástica Hereditaria/diagnóstico por imagen , Paraplejía Espástica Hereditaria/genética , Adolescente , Adulto , Niño , Preescolar , Estudios de Cohortes , Estudios Transversales , Femenino , Humanos , Lactante , Imagen por Resonancia Magnética/métodos , Masculino , Persona de Mediana Edad , Sistema de Registros , Adulto JovenRESUMEN
Amyloidosis, caused by a mutation in the transthyretin (TTR) gene, is the most common hereditary type disease. More than 120 mutations have been described, with extensive phenotypic heterogeneity. Val30Met (p.Val50Met) is the most frequent mutation, and patients exhibit polyneuropathy, possibly including cardiac, renal, gastrointestinal, and/or ocular involvement. Val122Ile (p.Val142Ile) is the mutation associated with cardiomyopathy, and few cases have been reported in Brazil. Most individuals are heterozygous for one pathogenic mutation. Herein, we report a compound heterozygote with two pathogenic mutations (Val30Met/ Val122Ile), and a family history of a deceased brother with amyloidosis, who also carried the same TTR gene mutations. The patient presented with neuropathic, cardiac, and renal impairment and a faster disease progression. Cases of the double mutation have been linked to changes in disease presentation. The concomitance of two pathogenic mutations may have contributed to more exuberant manifestations and faster disease progression.
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Amiloidosis Familiar/genética , Cardiomiopatías/genética , Polineuropatías/genética , Prealbúmina/genética , Amiloidosis Familiar/fisiopatología , Brasil , Cardiomiopatías/fisiopatología , Progresión de la Enfermedad , Heterocigoto , Humanos , Masculino , Persona de Mediana Edad , Mutación , Linaje , Polineuropatías/fisiopatologíaRESUMEN
Alteration of the KPTN gene, responsible for the coding of kaptin (a protein involved in actin cytoskeletal dynamics), causes a syndrome characterized by macrocephaly, neurodevelopmental delay and epileptic seizures. We report the first Brazilian case of KPTN gene variation, previously described in nine subjects from four interlinked families from an Amish community in Ohio, two Estonian siblings and a 9-year-old boy from Kansas City. We report a case of KPTN-related syndrome in a 5-year-old child which presented macrocephaly, muscular hypotonia, and global development delay. The neurological examination revealed below-expected performance in coordination and balance tests, dyspraxia, and hand-mouth synkinesia. Expressive language was characterized by phono-articulatory imprecision, abundance of phonological processes and morphosyntactic immaturity. Neuropsychological assessment revealed intellectual disability with impairment of verbal and executive functions. Exome sequencing was performed. Analysis revealed a homozygous 2-nucleotide duplication c.597_598dup p.(Ser200Ilefs*55) in the KPTN gene, which is predicted to lead to a translational frameshift and formation of a premature stop codon. The phenotypic profile is similar to the cases described in the other families. Presence of macrocephaly and delayed development indicate the possibility of KPTN gene variation. Genetic testing should be carried out at an early stage in order to reach a timely diagnosis.
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Discapacidades del Desarrollo/patología , Homocigoto , Discapacidad Intelectual/patología , Megalencefalia/patología , Proteínas de Microfilamentos/genética , Hipotonía Muscular/patología , Mutación , Brasil , Preescolar , Discapacidades del Desarrollo/genética , Femenino , Humanos , Discapacidad Intelectual/genética , Megalencefalia/genética , Hipotonía Muscular/genética , Fenotipo , SíndromeRESUMEN
Sarcomas are rare and heterogeneous cancers classically associated with a poor outcome. Sarcomas are 1% of the cancer but recent estimations indicate that sarcomas account for 2% of the estimated cancer-related deaths. Traditional treatment with surgery, radiotherapy, and chemotherapy has improved the outcome for some types of sarcomas. However, novel therapeutic strategies to treat sarcomas are necessary. TNF-related apoptosis-inducing ligand (TRAIL) is a death ligand initially described as capable of inducing apoptosis on tumor cell while sparing normal cells. Only few clinical trials have used TRAIL-based treatments in sarcoma, but they show only low or moderate efficacy of TRAIL. Consequently, novel TRAIL formulations with an improved TRAIL bioactivity are necessary. Our group has developed a novel TRAIL formulation based on tethering this death ligand on a lipid nanoparticle surface (LUV-TRAIL) resembling the physiological secretion of TRAIL as a trasmembrane protein inserted into the membrane of exosomes. We have already demonstrated that LUV-TRAIL shows an improved cytotoxic activity when compared to soluble recombinant TRAIL both in hematological malignancies and epithelial-derived cancers. In the present study, we have tested LUV-TRAIL in several human sarcoma tumor cell lines with different sensitivity to soluble recombinant TRAIL, finding that LUV-TRAIL was more efficient than soluble recombinant TRAIL. Moreover, combined treatment of LUV-TRAIL with distinct drugs proved to be especially effective, sensitizing even more resistant cell lines to TRAIL.
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Apoptosis , Lípidos , Nanopartículas , Sarcoma/metabolismo , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Caspasas/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Humanos , Proteínas Recombinantes , Ligando Inductor de Apoptosis Relacionado con TNF/farmacologíaRESUMEN
PURPOSE: Non-small cell lung cancer (NSCLC) is one the types of cancer with higher prevalence and mortality. Apo2-Ligand/TRAIL is a TNF family member able to induce apoptosis in tumor cells but not in normal cells. It has been tested in clinical trials against different types of human cancer including NSCLC. However, results of clinical trials have shown a limited efficacy of TRAIL-based therapies. Recently we have demonstrated that artificial lipid nanoparticles coated with bioactive Apo2L/TRAIL (LUV-TRAIL) greatly improved TRAIL cytotoxic ability being capable of killing chemoresistant hematological cancer cells. In the present work we have extended the study to NSCLC. METHODS/PATIENTS: LUV-TRAIL-induced cytotoxicity was assessed on different NSCLC cell lines with different sensitivity to soluble TRAIL and on primary human tumor cells from three patients suffering from NSCLC cancer. We also tested LUV-TRAIL-cytotoxic ability in combination with several anti-tumor agents. RESULTS: LUV-TRAIL exhibited a greater cytotoxic effect compared to soluble TRAIL both in A549 cells and primary human NSCLC cells. LUV-TRAIL-induced cell death was dependent on caspase-8 and caspase-3 activation. Moreover, combination of LUV-TRAIL with other anti-tumor agents such as flavopiridol, and SNS-032 clearly enhanced LUV-TRAIL-induced cytotoxicity against NSCLC cancer cells. CONCLUSION: The novel formulation of TRAIL based on displaying it on the surface of lipid nanoparticles greatly increases its anti-tumor activity and has clinical potential in cancer treatment.
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Antineoplásicos/farmacología , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Neoplasias Pulmonares/metabolismo , Nanopartículas/química , Ligando Inductor de Apoptosis Relacionado con TNF/farmacología , Antineoplásicos/química , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Caspasa 3/metabolismo , Caspasa 8/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica , Humanos , Liposomas , Neoplasias Pulmonares/tratamiento farmacológico , Proteínas Recombinantes/química , Proteínas Recombinantes/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF/químicaRESUMEN
The ability of cells to mount an interferon response to virus infections depends on intracellular nucleic acid sensing pattern recognition receptors (PRRs). RIG-I is an intracellular PRR that binds short double-stranded viral RNAs to trigger MAVS-dependent signalling. The RIG-I/MAVS signalling complex requires the coordinated activity of multiple kinases and E3 ubiquitin ligases to activate the transcription factors that drive type I and type III interferon production from infected cells. The linear ubiquitin chain assembly complex (LUBAC) regulates the activity of multiple receptor signalling pathways in both ligase-dependent and -independent ways. Here, we show that the three proteins that constitute LUBAC have separate functions in regulating RIG-I signalling. Both HOIP, the E3 ligase capable of generating M1-ubiquitin chains, and LUBAC accessory protein HOIL-1 are required for viral RNA sensing by RIG-I. The third LUBAC component, SHARPIN, is not required for RIG-I signalling. These data cement the role of LUBAC as a positive regulator of RIG-I signalling and as an important component of antiviral innate immune responses.
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Virus ARN , Ubiquitina-Proteína Ligasas , Ubiquitinación , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitina/metabolismo , Transducción de Señal , Proteína 58 DEAD Box/genética , Virus ARN/metabolismoRESUMEN
The dysregulated immune response and inflammation resulting in severe COVID-19 are still incompletely understood. Having recently determined that aberrant death-ligand-induced cell death can cause lethal inflammation, we hypothesized that this process might also cause or contribute to inflammatory disease and lung failure following SARS-CoV-2 infection. To test this hypothesis, we developed a novel mouse-adapted SARS-CoV-2 model (MA20) that recapitulates key pathological features of COVID-19. Concomitantly with occurrence of cell death and inflammation, FasL expression was significantly increased on inflammatory monocytic macrophages and NK cells in the lungs of MA20-infected mice. Importantly, therapeutic FasL inhibition markedly increased survival of both, young and old MA20-infected mice coincident with substantially reduced cell death and inflammation in their lungs. Intriguingly, FasL was also increased in the bronchoalveolar lavage fluid of critically-ill COVID-19 patients. Together, these results identify FasL as a crucial host factor driving the immuno-pathology that underlies COVID-19 severity and lethality, and imply that patients with severe COVID-19 may significantly benefit from therapeutic inhibition of FasL.
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COVID-19 , Modelos Animales de Enfermedad , Proteína Ligando Fas , SARS-CoV-2 , Animales , Ratones , Líquido del Lavado Bronquioalveolar , COVID-19/patología , COVID-19/inmunología , COVID-19/metabolismo , COVID-19/virología , COVID-19/mortalidad , Proteína Ligando Fas/metabolismo , Inflamación/patología , Inflamación/metabolismo , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Pulmón/patología , Pulmón/virología , Pulmón/metabolismo , Macrófagos/metabolismo , Macrófagos/patología , Ratones Endogámicos C57BLRESUMEN
Prenatal infections and cannabis use during adolescence are well-recognized risk factors for schizophrenia. As inflammation and oxidative stress (OS) contribute to this disorder, anti-inflammatory drugs have been proposed as potential therapies. This study aimed to evaluate the association between delta-9-tetrahydrocannabinol (THC) and schizophrenia-like abnormalities in a maternal immune activation (MIA) model. Additionally, we assessed the preventive effect of cannabidiol (CBD), a non-psychotropic/anti-inflammatory cannabinoid. THC and/or CBD were administered to Saline- and MIA-offspring during periadolescence. At adulthood, THC-exposed MIA-offspring showed significant improvements in sensorimotor gating deficits. Structural and metabolic brain changes were evaluated by magnetic resonance imaging, revealing cortical shrinkage in Saline- and enlargement in MIA-offspring after THC-exposure. Additionally, MIA-offspring displayed enlarged ventricles and decreased hippocampus, which were partially reverted by both cannabinoids. CBD prevented THC-induced reduction in the corpus callosum, despite affecting white matter structure. Post-mortem studies revealed detrimental effects of THC, including increased inflammation and oxidative stress. CBD partially reverted these pro-inflammatory alterations and modulated THC's effects on the endocannabinoid system. In conclusion, contrary to expectations, THC exhibited greater behavioural and morphometric benefits, despite promoting a pro-inflammatory state that CBD partially reverted. Further research is needed to elucidate the underlying mechanisms involved in the observed benefits of THC.
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Cannabidiol , Cannabinoides , Cannabis , Esquizofrenia , Humanos , Embarazo , Femenino , Adulto , Cannabidiol/farmacología , Cannabidiol/uso terapéutico , Esquizofrenia/tratamiento farmacológico , Esquizofrenia/metabolismo , Dronabinol/farmacología , Poli I-C , Inflamación , AntiinflamatoriosRESUMEN
Schizophrenia is a chronic neurodevelopmental disorder with an inflammatory/prooxidant component. N-acetylcysteine (NAC) has been evaluated in schizophrenia as an adjuvant to antipsychotics, but its role as a preventive strategy has not been sufficiently explored. We aimed to evaluate the potential of NAC administration in two-time windows before the onset of symptoms in a schizophrenia-like maternal immune stimulation (MIS) rat model. Pregnant Wistar rats were injected with Poly I:C or Saline on gestational day (GD) 15. Three different preventive approaches were evaluated: 1) NAC treatment during periadolescence in the offspring (from postnatal day [PND] 35 to 49); 2) NAC treatment during pregnancy after MIS challenge until delivery (GD15-21); and 3) NAC treatment throughout all pregnancy (GD1-21). At postnatal day (PND) 70, prepulse inhibition (PPI) and anxiety levels were evaluated. In vivo magnetic resonance (MR) imaging was acquired on PND100 to assess structural changes in gray and white matter, and brain metabolite concentrations. Additionally, inflammation and oxidative stress (IOS) markers were measured ex vivo in selected brain regions. MIS offspring showed behavioral, neuroanatomical, and biochemical alterations. Interestingly, NAC treatment during periadolescence prevented PPI deficits and partially counteracted some biochemical imbalances. Moreover, NAC treatments during pregnancy not only replicated the beneficial outcomes reported by the treatment in periadolescence, but also prevented some neuroanatomical deficits, including reductions in hippocampal and corpus callosum volumes. This study suggests that early reduction of inflammation and prooxidation could help prevent the onset of schizophrenia-like symptoms, supporting the importance of anti-IOS compounds in ameliorating this disorder.
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Acetilcisteína , Esquizofrenia , Femenino , Embarazo , Ratas , Animales , Ratas Wistar , Acetilcisteína/farmacología , Esquizofrenia/tratamiento farmacológico , Esquizofrenia/prevención & control , Poli I-C , InflamaciónRESUMEN
Autosomal dominant leukodystrophy (ADLD) is an adult onset demyelinating disorder that is caused by duplications of the lamin B1 (LMNB1) gene. However, as only a few cases have been analyzed in detail, the mechanisms underlying LMNB1 duplications are unclear. We report the detailed molecular analysis of the largest collection of ADLD families studied, to date. We have identified the minimal duplicated region necessary for the disease, defined all the duplication junctions at the nucleotide level and identified the first inverted LMNB1 duplication. We have demonstrated that the duplications are not recurrent; patients with identical duplications share the same haplotype, likely inherited from a common founder and that the duplications originated from intrachromosomal events. The duplication junction sequences indicated that nonhomologous end joining or replication-based mechanisms such fork stalling and template switching or microhomology-mediated break induced repair are likely to be involved. LMNB1 expression was increased in patients' fibroblasts both at mRNA and protein levels and the three LMNB1 alleles in ADLD patients show equal expression, suggesting that regulatory regions are maintained within the rearranged segment. These results have allowed us to elucidate duplication mechanisms and provide insights into allele-specific LMNB1 expression levels.
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Duplicación de Gen , Lamina Tipo B/genética , Enfermedad de Pelizaeus-Merzbacher/genética , Adulto , Secuencia de Bases , Puntos de Rotura del Cromosoma , Hibridación Genómica Comparativa , ADN/química , ADN/genética , Humanos , Lamina Tipo B/metabolismo , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Enfermedad de Pelizaeus-Merzbacher/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
Human Apo2-ligand/TRAIL is a member of the TNF cytokine superfamily capable of inducing apoptosis on tumor cells while sparing normal cells. Besides its antitumor activity, Apo2L/TRAIL is also implicated in immune regulation. Apo2L/TRAIL is stored inside activated T cells in cytoplasmic multivesicular bodies and is physiologically released to the extracellular medium inserted in the internal membrane vesicles, known as exosomes. In this study we have generated artificial lipid vesicles coated with bioactive Apo2L/TRAIL, which resemble natural exosomes, to analyze their apoptosis-inducing ability on cell lines from hematological tumors. We have tethered Apo2L/TRAIL to lipid vesicles by using a novel Ni(2+)-(N-5-amino-1-carboxylpentyl)-iminodiacetic acid, NTA)-containing liposomal system. This lipidic framework (LUVs-Apo2L/TRAIL) greatly improves Apo2L/TRAIL activity, decreasing by around 14-fold the LC50 on the T-cell leukemia Jurkat. This increase in bioactivity correlated with the greater ability of LUVs-Apo2L/TRAIL to induce caspase-3 activation and is probably due to the increase in local concentration of Apo2L/TRAIL, improving its receptor cross-linking efficiency. More important, liposome-bound Apo2L/TRAIL overcame the resistance to soluble recombinant Apo2L/TRAIL exhibited by tumor cell mutants overexpressing Bcl-xL or by a Bax and Bak-defective Jurkat cell mutant (Jurkat-shBak) and are also effective against other hematologic tumor cells. Jurkat-Bcl-xL and Jurkat-shBak cells are resistant to most chemotherapeutic drugs currently used in cancer treatment, and their sensitivity to LUVs-Apo2L/TRAIL could have potential clinical applications.
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Neoplasias Hematológicas/tratamiento farmacológico , Liposomas/química , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Células Cultivadas , Resistencia a Antineoplásicos , Citometría de Flujo , Humanos , Leucocitos Mononucleares , Liposomas/administración & dosificación , Ligando Inductor de Apoptosis Relacionado con TNF/química , Ligando Inductor de Apoptosis Relacionado con TNF/farmacologíaRESUMEN
BACKGROUND: An observational retrospective study has been conducted, including 52 patients (37 male and 15 female), ranging from 22 to 65 years old, who underwent an orthotopic liver transplantation (OLT) at the Hospital Universitario Central de Asturias (HUCA) between 2007 and 2010. METHODS: The main objective was to evaluate the post-OLT critical complication prognosis usefulness of the precursors of three new biomarkers: mid-regional proadrenomedullin (MR-proADM), carboxy-terminal-proendothelin-1 (CT-ProET-1) and mid-regional proatrial natriuretic peptide (MR-ProANP). As all of them are blood pressure mediators, stress-associated physiological phenomena are expected to affect their expression and secretion, mainly those related to blood circulation. Therefore, as a second goal, the biological variability of the biomarkers has been studied in a set of OLT patients without complications during the first postoperative week. The knowledge of the reference change value of the new biomarkers will be interesting for their correct interpretation in future investigations. The prognostic value of the new biomarkers was also compared to that of procalcitonin (PCT). RESULTS: It has been shown that the basal concentration of the biomarkers is higher in patients that undergo OLT than in the normal population, correlating with the severity of the pathology. The intra-individual biological variation of these biomarkers is similar to other biochemical parameters, the reference change value for OLT patients being 90% for CT-proET-1, 112% for MR-proADM and 127% for MR-proANP. CONCLUSIONS: Multivariate analysis showed that MR-proADM was the best biomarker for the prognosis of severe complications.
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Adrenomedulina/sangre , Factor Natriurético Atrial/sangre , Endotelina-1/sangre , Trasplante de Hígado/métodos , Fragmentos de Péptidos/sangre , Adulto , Anciano , Biomarcadores/sangre , Femenino , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Estudios Retrospectivos , Adulto JovenRESUMEN
Cancer resistance to treatments is a challenge that researchers constantly seek to overcome. For instance, TNF-related apoptosis-inducing ligand (TRAIL) is a potential good prospect as an anti-cancer therapy, as it attacks tumor cells but not normal cells. However, treatments based in soluble TRAIL provided incomplete clinical results and diverse formulations have been developed to improve its bioactivity. In previous works, we generated a new TRAIL formulation based in its attachment to the surface of unilamellar nanoliposomes (LUV-TRAIL). This formulation greatly increased apoptosis in a wide selection of tumor cell types, albeit a few of them remained resistant. On the other hand, it has been described that a metabolic shift in cancer cells can also alter its sensitivity to other treatments. In this work, we sought to increase the sensitivity of several tumor cell types resistant to LUV-TRAIL by previous exposure to the metabolic drug dichloroacetate (DCA), which forces oxidative phosphorylation. Results showed that DCA + LUV-TRAIL had a synergistic effect on both lung adenocarcinoma A549, colorectal HT29, and breast cancer MCF7 cells. Despite DCA inducing intracellular changes in a cell-type specific way, the increase in cell death by apoptosis was clearly correlated with an increase in death receptor 5 (DR5) surface expression in all cell lines. Therefore, DCA-induced metabolic shift emerges as a suitable option to overcome TRAIL resistance in cancer cells.
RESUMEN
Approximately 30 sex chromosome discordant chimera cases have been reported to date. In particular, there are few reported cases of chimerism involving coexisting normal and abnormal lineages that each carries a distinct sex chromosome complement. To our knowledge, this is the first case of sexual chimerism with a simultaneous chromosomal aneuploidy involving chromosome 8. This report represents the data from 11 years of follow-up.
RESUMEN
An electrochemical biosensor for creatinine determination in a drop of whole human blood was developed and applied to the determination of creatinine in real clinical samples. It is based on the modification of a dual carbon working electrode with a combination of three enzymes: creatinine amidohydrolase (CNN), creatine amidinohydrolase (CRN) and sarcosine oxidase (SOX). Electrochemical transduction is performed using horseradish peroxidase (HRP) and potassium hexacyanoferrate(II) as mediator. A drop of human blood is enough to carry out the measurements by differential chronoamperometry where one carbon electrode detects creatine and the other both creatine and creatinine. The integrated differential signal obtained in the biosensor is linear with the concentration of creatinine in blood in the range 0.5-15 mg/dL and the enzyme-modified electrodes are stable for at least 3 months at 4 °C. The biosensor was lined to a reference method based on Isotope Dilution Mass Spectrometry (IDMS) with 50 real human blood samples and the results compared with those obtained by alternative routine techniques based on Jaffé method and an enzymatic method (Cobas 8000 Roche®, Crep2 Roche®). There were no significant differences between the creatinine concentrations found by the routine techniques and the developed biosensor.