RESUMEN
BACKGROUND AND AIMS: Mutations in the disheveled, Egl-10 and pleckstrin domain-containing protein 5 (DEPDC5) gene have emerged as an important cause of various familial focal epilepsy syndromes. However, the significance of DEPDC5 mutations in patients with sporadic focal epilepsy has yet to be characterized. MATERIALS AND METHODS: We studied a kindred of familial focal epilepsy with variable foci using whole-exome sequencing. We subsequently studied a cohort of 293 patients with focal epilepsy and sequenced all exons of DEPDC5 using targeted resequencing. RESULTS: We reported a Taiwanese family with a novel splice site mutation which affected mRNA splicing and activated the downstream mammalian target of rapamycin (mTOR) pathway. Among patients with focal epilepsies, the majority (220/293) of these patients had sporadic focal epilepsy without malformation of cortical development. Two (0.9%) of these patients had probably pathogenic mutations in the DEPDC5 gene. DISCUSSION AND CONCLUSIONS: Our finding suggests that DEPDC5 is not only the most common gene for familial focal epilepsy but also could be a significant gene for sporadic focal epilepsy. Since focal epilepsies account for more than 60% of all epilepsies, the effect of mTORC1 inhibitor on patients with focal epilepsy due to DEPDC5 mutations will be an important future direction of research.
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Epilepsias Parciales/genética , Predisposición Genética a la Enfermedad , Proteínas Represoras/genética , Serina-Treonina Quinasas TOR/genética , Adolescente , Niño , Preescolar , Epilepsias Parciales/patología , Femenino , Proteínas Activadoras de GTPasa , Humanos , Masculino , Diana Mecanicista del Complejo 1 de la Rapamicina/genética , Mutación , Linaje , Empalme del ARN/genética , Secuenciación del ExomaRESUMEN
BACKGROUND AND PURPOSE: Investigation of the relationship between mitochondrial DNA (mtDNA) variants and Parkinson disease (PD) remains an issue awaiting more supportive evidence. Moreover, an affirming cellular model study is also lacking. METHODS: The index mtDNA variants and their defining mitochondrial haplogroup were determined in 725 PD patients and 744 non-PD controls. Full-length mtDNA sequences were also conducted in 110 cases harboring various haplogroups. Cybrid cellular models, composed by fusion of mitochondria-depleted rho-zero cells and donor mitochondria, were used for a rotenone-induced PD simulation study. RESULTS: Multivariate logistic regression analysis revealed that subjects harboring the mitochondrial haplogroup B5 have resistance against PD (odds ratio 0.50, 95% confidence interval 0.32-0.78; P = 0.002). Furthermore, a composite mtDNA variant group consisting of A10398G and G8584A at the coding region was found to have resistance against PD (odds ratio 0.50, 95% confidence interval 0.33-0.78; P = 0.001). In cellular studies, B4 and B5 cybrids were selected according to their higher resistance to rotenone, in comparison with cybrids harboring other haplogroups. The B5 cybrid, containing G8584A/A10398G variants, showed more resistance to rotenone than the B4 cybrid not harboring these variants. This is supported by findings of low reactive oxygen species generation and a low apoptosis rate in the B5 cybrid, whereas a higher expression of autophagy was observed in the B4 cybrid particularly under medium dosage and longer treatment time with rotenone. CONCLUSIONS: Our studies, offering positive results from clinical investigations and cybrid experiments, provide data supporting the role of variant mtDNA in the risk of PD.
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ADN Mitocondrial/genética , Variación Genética , Enfermedad de Parkinson/genética , Anciano , Femenino , Haplotipos , Humanos , Masculino , Persona de Mediana Edad , Factores de RiesgoRESUMEN
Viral envelope proteins play important roles in viral infection and assembly. The grouper iridovirus ORF 64L (GIV-64L) was predicted to encode an envelope protein and was conserved in all sequenced Ranaviruses. In this study, the complete nucleotide sequence of the GIV-64L gene (1215 bp) was cloned into the isopropyl ß-D-1-thiogalactopyranoside (IPTG) induction prokaryotic expression vector pET23a. The approximately 50.2 kDa recombinant GIV-64L-His protein was induced, purified and used as an immunogen to immunize BALB/c mice. Three monoclonal antibodies (mAbs), all IgG1 class antibodies against GIV-64L protein, were produced by enzyme-linked immunosorbent assay. Reverse transcription polymerase chain reaction analyses revealed GIV-64L to be a late gene when expressed in grouper kidney cells during GIV infection with cycloheximide (an inhibitor of protein synthesis) or cytosine arabinoside (an inhibitor of DNA synthesis) present. Finally, one of the established mAbs, GIV-64L-mAb-17, was used in Western blotting and an immunofluorescence assay, which showed that GIV-64L protein was expressed at 24 h post-infection and localized only in the cytoplasm in GIV-infected cells, packed into a whole virus particle. The presently characterized GIV-64L mAbs should have widespread applications in GIV immunodiagnostics and other research, and these results should offer important insights into the pathogenesis of GIV.
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Anticuerpos Monoclonales/metabolismo , Infecciones por Virus ADN/virología , Enfermedades de los Peces/virología , Iridovirus/genética , Proteínas del Envoltorio Viral/genética , Animales , Anticuerpos Monoclonales/genética , Línea Celular , Infecciones por Virus ADN/diagnóstico , Infecciones por Virus ADN/patología , Enfermedades de los Peces/diagnóstico , Enfermedades de los Peces/patología , Peces , Regulación Viral de la Expresión Génica , Iridovirus/clasificación , Ratones , Ratones Endogámicos BALB C , Filogenia , Transporte de Proteínas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Alineación de Secuencia , Factores de Tiempo , Proteínas del Envoltorio Viral/metabolismoRESUMEN
The role of high mobility group box 1 (HMGB1) has been demonstrated in stroke and coronary artery disease but not in peripheral arterial occlusive disease (PAOD). The pathogenesis of HMGB1 in acute and chronic vascular injury is also not well understood. We hypothesized that HMGB1 induces inflammatory markers in diabetic PAOD patients. We studied 36 diabetic patients, including 29 patients with PAOD, who had undergone amputation for diabetic foot and 7 nondiabetic patients who had undergone amputation after traumatic injury. Expression of HMGB1 and inflammatory markers were quantified using immunohistochemical staining. Mitochondrial DNA copy number was quantified using real-time polymerase chain reaction. Compared with that in the traumatic amputation group, HMGB1 expression in vessels was significantly higher in the diabetes and diabetic PAOD groups. In all subjects, arterial stenosis grade was positively correlated with the expression levels of HMGB1, 8-hydroxyguanosine, malondialdehyde, vascular cell adhesion molecule 1, and inflammatory markers CD3, and CD68 in both the intima and the media of vessels. Furthermore, HMGB1 expression level was positively correlated with 8-hydroxyguanosine, vascular cell adhesion molecule 1, nuclear factor-kB, CD3, and CD68 expression. Within the PAOD subgroup, subjects with HMGB1 expression had higher expression of the autophagy marker LC3A/B and higher mitochondrial DNA copy number. HMGB1 may be an inflammatory mediator with roles in oxidative damage and proinflammatory and inflammatory processes in diabetic atherogenesis. Moreover, it may have dual effects by compensating for increased mitochondrial DNA copy number and increased autophagy marker expression.
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Aterosclerosis/metabolismo , Diabetes Mellitus Tipo 2/complicaciones , Pie Diabético/metabolismo , Proteína HMGB1/metabolismo , Amputación Quirúrgica , Arteriopatías Oclusivas/genética , Arteriopatías Oclusivas/metabolismo , Aterosclerosis/genética , Biomarcadores , Pie Diabético/genética , Pie Diabético/cirugía , Expresión Génica , Proteína HMGB1/genética , Humanos , Inflamación , Estrés Oxidativo , Enfermedad Arterial Periférica/genética , Enfermedad Arterial Periférica/metabolismoRESUMEN
Grouper iridovirus (GIV) is one of the most important viral pathogens in grouper, particularly at the fry and fingerling stages. The study of GIV pathogenicity has been hampered by the lack of proper immunological reagents to study the expression and function of viral proteins in the infected cells. In this study, two mouse monoclonal antibodies (mAbs) against GIV 55L and 97L proteins were produced. Enzyme-linked immunosorbent assay (ELISA) and Western blotting were used to screen these hybridomas, resulting in the identification of two high-affinity mAbs named GIV55L-mAb-2 and GIV97L-mAb-3, respectively. Both mAbs belong to the IgG1 isotype and were effective in detecting their respective target viral protein. Reverse-transcription polymerase chain reaction (RT-PCR) and Western blot analyses of GIV-infected GK cells revealed that GIV 97L is an immediate early gene, whereas GIV 55L a late one. The localization of 55L and 97L in GIV-infected cells was further characterized by immunofluorescence microscopy with the mAbs. The 55L protein mainly aggregated in the cytoplasm while 97L distributed in both the nucleus and cytoplasm of the infected cells. These studies demonstrate the validity of the two mAbs as immunodiagnostic and research reagents.
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Anticuerpos Monoclonales/metabolismo , Anticuerpos Antivirales/metabolismo , Infecciones por Virus ADN/veterinaria , Enfermedades de los Peces/metabolismo , Iridovirus/metabolismo , Proteínas Virales/metabolismo , Secuencia de Aminoácidos , Animales , Western Blotting , Línea Celular , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Infecciones por Virus ADN/metabolismo , Ensayo de Inmunoadsorción Enzimática , Perfilación de la Expresión Génica , Hibridomas , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , Alineación de Secuencia , Proteínas Virales/química , Proteínas Virales/genéticaRESUMEN
Grouper iridovirus (GIV) belongs to the Ranavirus genus and is one of the most important viral pathogens in grouper, particularly at the fry and fingerling stages. In this study, we identified and characterized the GIV-2L gene, which encodes a protein of unknown function. GIV-2L is 1242 bp in length, with a predicted protein mass of 46.2 kDa. It displayed significant identity only with members of the Ranavirus and Iridovirus genera. We produced mouse monoclonal antibodies against the GIV-2L protein by immunizing mice with GIV-2L-His-tag recombinant protein. By inhibiting de novo protein and DNA synthesis in GIV-infected cells, we showed that GIV-2L was a late gene during the viral replication. Finally, immunofluorescence microscopy revealed that GIV-2L protein accumulated in both the nucleus and cytoplasm of infected cells. These results offer important insights into the pathogenesis of GIV.
RESUMEN
Hyperglycemia-induced reactive oxygen species production can cause diabetes and its complications, including atherosclerosis. The role of mitochondrial DNA variants and mitochondrial copy number in the pathogenesis of diabetic atherogenesis is not well understood. We examined 36 diabetic patients who had undergone amputation for diabetic foot and seven non-diabetic patients who had undergone amputation after traumatic injury. Mitochondrial DNA was extracted and used for sequencing. Single nucleotide polymorphisms (SNPs) relative to the Cambridge reference sequence were analyzed. Mitochondrial DNA copy number was quantified by real-time PCR. Twenty-one novel variants were detected in 29 diabetic patients with arterial stenosis; six of the variants were heteroplasmic, and most occurred in highly evolutionarily conserved residues. These variants were more prevalent in patients with arterial stenosis than in those without stenosis. The novel variants included four in complex I (ND1: C3477A/C, A3523A/G; ND5: C13028A/C, C13060A/C), one in complex IV (COX1: T6090A/T), and one in rRNA (12srRNA: G857G/T). Compared with non-diabetic patients, the diabetic patients had significantly less mitochondrial DNA. Furthermore, among diabetic patients with arterial stenosis, there was a significant positive correlation between mitochondrial DNA copy number and the number of total SNPs. In conclusion, we identified six novel heteroplasmic mitochondrial DNA variants among diabetic patients with arterial stenosis, and we found that diabetic atherogenesis is associated with decreased amounts of mitochondrial DNA.
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Aterosclerosis/genética , Variaciones en el Número de Copia de ADN/genética , ADN Mitocondrial/genética , Complicaciones de la Diabetes/genética , Secuencia de Aminoácidos , Secuencia de Bases , Secuencia Conservada/genética , Análisis Mutacional de ADN , Complejo I de Transporte de Electrón/química , Complejo I de Transporte de Electrón/genética , Complejo IV de Transporte de Electrones/química , Complejo IV de Transporte de Electrones/genética , Humanos , Mitocondrias/genética , Proteínas Mitocondriales/química , Proteínas Mitocondriales/genética , Datos de Secuencia Molecular , Polimorfismo de Nucleótido Simple/genéticaRESUMEN
This study aimed to investigate the effects of eight metals on the anaerobic digestion of the organic fraction of municipal solid waste (OFMSW) in bioreactors. Anaerobic bioreactors containing 200 mL MSW mixed completely with 200 m L sludge seeding. Ca and K (0, 1000, 2000 and 6,000 mg L(-1)) and Cr, Ni, Zn, Co, Mo and W (0, 5, 50 and 100 mg L(-1)) of various dose were added to anaerobic bioreactors to examine their anaerobic digestion performance. Results showed that except K and Zn, Ca (~728 to ~1,461 mg L(-1)), Cr (~0.0022 to ~0.0212 mg L(-1)), Ni (~0.801 to ~5.362 mg L(-1)), Co (~0.148 to ~0.580 mg L(-1)), Mo (~0.044 to ~52.94 mg L(-1)) and W (~0.658 to ~40.39 mg L(-1)) had the potential to enhance the biogas production. On the other hand, except Mo and W, inhibitory concentrations IC(50) of Ca, K, Cr, Ni, Zn and Co were found to be ~3252, ~2097, ~0.124, ~7.239, ~0.482, ~8.625 mg L(-1), respectively. Eight spiked metals showed that they were adsorbed by MSW to a different extent resulting in different liquid metals levels and potential stimulation and inhibition on MSW anaerobic digestion. These results were discussed and compared to results from literature.
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Metales/metabolismo , Eliminación de Residuos/métodos , Adsorción , Anaerobiosis , Biocombustibles , Reactores Biológicos , Metales/química , Metales Pesados/metabolismo , Aguas del AlcantarilladoRESUMEN
BACKGROUND: A T-to-C transition at mitochondrial DNA (mtDNA) nucleotide position 16189 can generate a variable length polycytosine tract (poly-C). This tract variance has been associated with disease. A suggested pathogenesis is that it interferes with the replication process of mtDNA, which in turn decreases the mtDNA copy number and generates disease. METHODS: In this study, 837 healthy adults' blood samples were collected and determined for their mtDNA D-loop sequence. The mtDNA copy number in the leucocytes and serum levels of oxidative thiobarbituric acid reactive substance (TBARS) and antioxidative thiols were measured. All subjects were then categorised into three groups: wild type or variant mtDNA with presence of an interrupted/uninterrupted poly-C at 16180-16195 segment. RESULTS: A step-wise multiple linear regression analysis identified factors affecting expression of mtDNA copy number including TBARS, thiols, age, body mass index and the mtDNA poly-C variant. Subjects harbouring a variant uninterrupted poly-C showed lowest mean (SD) mtDNA copy number (330 (178)), whereas an increased copy number was noted in subjects harbouring variant, interrupted poly-C (420 (273)) in comparison with wild type (358 (215)). The difference between the three groups and between the uninterrupted poly-C and the composite data from the interrupted poly-C and wild type remained consistent after adjustment for TBARS, thiols, age and body mass index (p=0.001 and p=0.011, respectively). A trend for decreased mtDNA copy number in association with increased number of continuous cytosine within the 16180-16195 segment was noted (p(trend)<0.006). CONCLUSIONS: Our results substantiate a previous suggestion that the mtDNA 16189 variant can cause alteration of mtDNA copy number in human blood cells.
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ADN Mitocondrial/genética , Dosificación de Gen , Variación Genética/genética , Poli C/genética , Adulto , Anciano , ADN Mitocondrial/sangre , ADN Mitocondrial/química , Femenino , Humanos , Leucocitos/metabolismo , Modelos Lineales , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADN , Compuestos de Sulfhidrilo/sangre , Sustancias Reactivas al Ácido Tiobarbitúrico/metabolismoRESUMEN
AIM: To investigate the gender differences associated with a thinner intima-media thickness (IMT) of the common carotid artery (CCA) in women. MATERIALS AND METHODS: In a sample of 218 consecutive healthy volunteers comprising 110 men and 108 women, the IMT of the CCA was measured using B-mode ultrasonography. Blood pressure, fasting blood sugar, body mass index (BMI), blood lipid profile, homocysteine, folic acid, uric acid, high sensitive C-reactive protein, and thiobarbituric acid reactive substances (TBARS) levels were measured and compared with each other in both genders. RESULTS: The IMT of the CCA was significantly thinner in women than in men (p=0.012). Blood pressure, fasting plasma glucose, BMI, low-density lipoprotein cholesterol, triglycerides, homocysteine, uric acid, and TBARS were significantly (p<0.05) lower, folic acid and high-density lipoprotein cholesterol (HDL-C) were significantly (p<0.0001) higher in women compared with men. Multivariable logistic regression analysis revealed that higher serum levels of homocysteine, uric acid, and TBARS, and lower serum levels of HDL-C were significantly (p<0.05) associated with male sex. Multiple linear regression analysis further revealed that age, sex, and BMI were independently associated with CCA IMT. CONCLUSIONS: The IMT of the CCA was thinner in women than in men. Traditional vascular risk factors explain only a small amount of variance in multivariate regression models supporting the hypothesis that other behavioural, sex hormone-related or genetic factors, which have not been sufficiently explored so far, may play a role in the gender differences of IMT.
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Arteria Carótida Común , Túnica Media , Adulto , Factores de Edad , Anciano , Biomarcadores/sangre , Glucemia/análisis , Índice de Masa Corporal , Proteína C-Reactiva/análisis , Arteria Carótida Común/anatomía & histología , Arteria Carótida Común/diagnóstico por imagen , HDL-Colesterol/sangre , LDL-Colesterol/sangre , Femenino , Humanos , Masculino , Persona de Mediana Edad , Reproducibilidad de los Resultados , Factores de Riesgo , Factores Sexuales , Triglicéridos/sangre , Túnica Media/anatomía & histología , Túnica Media/diagnóstico por imagen , Ultrasonografía , Adulto JovenRESUMEN
BACKGROUND AND PURPOSE: Although a clear protocol for reduction of recurrent ischaemic stroke (RIS) has been established, few studies have compared the stroke subtype distribution and risk factors between RIS and first-ever stroke (FES). METHODS: This one-year hospital-based study enrolled 587 FES and 475 RIS patients. Patients were categorized into four stroke subtypes according to a modified TOAST stroke subtype classification system. Risk factor profiles were compared between the two major stroke groups and between the corresponding four subtypes to discriminate the significant risk factors for RIS. RESULTS: A multivariate regression analysis identified hypertension (OR, 1.87; 95% CI, 1.34-2.62), diabetes mellitus (DM) (OR, 1.57; 95% CI, 1.22-2.02), low high-density lipoprotein (LHDL) (OR, 1.43; 95% CI, 1.08-1.88) and older age as significant RIS risk factors. The significance of the former three RIS factors was further recognized in its large-vessel subtype. Moreover, metabolic syndrome was significantly more common in the recurrent stroke group (P = 0.01), including its large-vessel subtype (P = 0.04). Progressively increasing odds ratios from 1.49 to 2.02, in accordance with increased number of diagnostic components of metabolic syndrome for recurrent large-vessel ischaemic stroke, were noted. CONCLUSIONS: Metabolic syndrome likely plays a crucial role in the development of RIS, including large-vessel infarction in modern-day Taiwan.
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Infarto Cerebral/etiología , Infarto Cerebral/prevención & control , Síndrome Metabólico/complicaciones , Factores de Edad , Anciano , Infarto Cerebral/clasificación , HDL-Colesterol/sangre , Complicaciones de la Diabetes/epidemiología , Femenino , Humanos , Hipertensión/complicaciones , Hipertensión/epidemiología , Masculino , Recurrencia , Factores de RiesgoRESUMEN
Status epilepticus results in preferential neuronal cell loss in the hippocampus. We evaluated the hypothesis that the repertoire of intracellular events in the vulnerable hippocampal CA3 subfield after induction of experimental temporal lobe status epilepticus entails upregulation of nitric oxide synthase II (NOS II), followed by the release of mitochondrial cytochrome c that triggers the cytosolic caspase-3 cascade, leading to apoptotic cell death. In Sprague-Dawley rats, significant and temporally correlated upregulation of NOS II (3-24h), but not NOS I or II expression, enhanced cytosolic translocation of cytochrome c (days 1 and 3), augmented activated caspase-3 in cytosol (days 1, 3 and 7) and DNA fragmentation (days 1, 3 and 7) was detected bilaterally in the hippocampal CA3 subfield after elicitation of sustained seizure activity by microinjection of kainic acid into the unilateral CA3 subfield. Application bilaterally into the hippocampal CA3 subfield of a selective NOS II inhibitor, S-methylisothiourea, significantly blunted these apoptotic events; a selective NOS I inhibitor, N(omega)-propyl-l-arginine or a potent NOS III inhibitor, N(5)-(1-iminoethyl)-l-ornithine was ineffective. We conclude that upregulation of NOS II contributes to apoptotic cell death in the hippocampal CA3 subfield via a cytochrome c/caspase-3 signaling cascade following the induction of experimental temporal lobe status epilepticus.
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Apoptosis/fisiología , Caspasa 3/fisiología , Citocromos c/fisiología , Epilepsia del Lóbulo Temporal/fisiopatología , Hipocampo/fisiología , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Transducción de Señal/fisiología , Estado Epiléptico/fisiopatología , Regulación hacia Arriba/efectos de los fármacos , Animales , Western Blotting , Citosol/enzimología , Fragmentación del ADN/efectos de los fármacos , Electroencefalografía/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Técnica del Anticuerpo Fluorescente , Lateralidad Funcional/fisiología , Hipocampo/citología , Etiquetado Corte-Fin in Situ , Masculino , Microscopía Confocal , Óxido Nítrico/biosíntesis , Óxido Nítrico/fisiología , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Ratas , Ratas Sprague-DawleyRESUMEN
Solitary lymphangioma of the spleen is an extremely rare, benign vasoformative tumour that typically shows no notable enhancement on imaging studies. Few descriptions of the MRI features have been reported and, to our knowledge, the findings on dynamic gadolinium-enhanced MR images have never been described. We report a case in which MR images showed diffuse and prolonged enhancement of a splenic mass. On histological correlation, splenic lymphangiomas with abundant fibrous stroma may account for this specific enhancing pattern.
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Linfangioma/diagnóstico , Neoplasias del Bazo/diagnóstico , Medios de Contraste , Diagnóstico Diferencial , Femenino , Gadolinio , Humanos , Imagen por Resonancia Magnética/métodos , Persona de Mediana EdadRESUMEN
Spectral analysis of heart rate variability provides a probe to assess the function of the sympathetic and parasympathetic nervous systems. Time-frequency analysis of heart rate variability is useful for investigating autonomic nervous function in patients with syncope or non-sustained ventricular tachycardia, or in anaesthesia, etc. In this paper, we developed an algorithm for continuous and online analysis of heart rate variability. The algorithm was simulated and evaluated in MATLAB, and implemented on the digital signal processor. The electrocardiogram signals from MIT/BIH arrhythmia database and one patient with syncope demonstrate the capability of the proposed method in the continuous and online analysis of heart rate variability.
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Algoritmos , Arritmias Cardíacas/diagnóstico , Arritmias Cardíacas/fisiopatología , Diagnóstico por Computador/métodos , Frecuencia Cardíaca , Procesamiento de Señales Asistido por Computador , Humanos , Sistemas en Línea , Reproducibilidad de los Resultados , Estudios Retrospectivos , Sensibilidad y EspecificidadRESUMEN
As populations age, osteoporosis is becoming an important public health care problem. Urinary level of the cross-linked N-telopeptide of type I collagen has been reported to be a sensitive marker of bone resorption. Recently, we synthesized and characterized 10 overlapping peptides covering the N-telopeptide of alpha-2 type I collagen and reported their relative binding response to anti-type I collagen cross-linked N-telopeptide (NTX) antibodies determined by a competitive-inhibition enzyme-linked immunosorbent assay (ELISA). In this study, we design an assay based on the surface plasmon resonance (SPR) technology to detect binding interaction of each peptide fragment of NTX with the anti-NTX monoclonal antibodies. Anti-NTX monoclonal antibodies were immobilized on the surface of sensor chip by amine-coupling procedure. Serial dilutions of each peptide were prepared and injected separately onto the antibodies-immobilized sensor chip. The real-time association and dissociation interactions of each peptide were detected and reported as sensorgrams. Binding response of each peptide to the monoclonal antibodies was determined, and the SPR results were compared with the ELISA results. We demonstrate that the trends of binding potency of peptide fragments detected by SPR are in good correlation to the results obtained by ELISA, indicating that our developed SPR-based method can be further applied to detect the NTX fragments in urine and to monitor the bone loss in humans. The potent peptide fragments identified by both assays are promising for further preparation of specific monoclonal antibodies in order to develop bioassays for bone loss in humans.
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Colágeno/química , Fragmentos de Péptidos/química , Resonancia por Plasmón de Superficie , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Sitios de Unión de Anticuerpos , Biomarcadores/orina , Colágeno/inmunología , Colágeno Tipo I , Ensayo de Inmunoadsorción Enzimática , Humanos , Osteoporosis/diagnóstico , Osteoporosis/orina , Fragmentos de Péptidos/inmunología , Fragmentos de Péptidos/orina , Valor Predictivo de las PruebasRESUMEN
Investigation of the clinical manifestations of MELAS-specific A3243G mitochondrial DNA (mtDNA) point mutation has suggested that the A3243G mutation of mtDNA can cause certain subtypes of diabetes mellitus (DM) and contributes about 0.15% of the overall incidence of diabetes. However, a relationship between the diabetic syndrome and the proportion of mutant mtDNA in affected tissues remains unclear. In this article, we report the results of our investigation of 14 diabetic and 23 non-diabetic patients who had the A3243G mutant mtDNA. The proportions of mutant mtDNA in different tissues were noted to change variably and neither heteroplasmy of mutant mtDNA in various tissues nor the proportion of mutated mtDNA in a specific tissue showed a correlation with the clinical phenotype of DM. Generation of a diabetic syndrome was not predictable from either the content of mutant mtDNA in leukocytes, hair follicles, or in muscle tissues. Further study showed that muscle tissue has the highest proportion of mutant mtDNA followed by hair follicles and by blood cells. Moreover, we observed that as the patient's age increased, all tissue showed a declining proportion of mutant mtDNA. These findings suggest that age may play a role in the manifestation of diabetes in patients with A3243G mutation of mtDNA.
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Pueblo Asiatico/genética , ADN Mitocondrial/genética , Diabetes Mellitus/genética , Mutación/genética , Adulto , Humanos , Persona de Mediana Edad , Fenotipo , Valores de Referencia , TaiwánAsunto(s)
Bacteriemia/microbiología , Absceso Encefálico/microbiología , Vibriosis/diagnóstico , Antibacterianos , Bacteriemia/diagnóstico , Bacteriemia/tratamiento farmacológico , Absceso Encefálico/diagnóstico , Absceso Encefálico/tratamiento farmacológico , Quimioterapia Combinada/administración & dosificación , Resultado Fatal , Humanos , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Índice de Severidad de la Enfermedad , Tomografía Computarizada por Rayos X , Vibrio cholerae/clasificaciónRESUMEN
Rat pups age of 14 postnatal day (P14) were subjected to lithium-pilocarpine (Li-PC) model of status epilepticus (SE). Control rats (n=6) were given an equivalent volume of saline intraperitoneally. Behavioral testing began on P60 including the Morris water maze, the radial arm maze, and the rotarod test. Brain were then analyzed with cresyl violet stain for histological lesions and evaluated for mossy fiber sprouting with the Timm stain. We observed spatial memory deficits both in the Morris water maze and radial arm maze in Li-PC-treated rat. There was no motor impairment in Li-PC-treated rat by the rotarod test. Two of six Li-PC-treated rats showed cell loss in hippocampal CA1 subfield. The Timm staining pattern was similar in both control and Li-PC-treated rats. Result of this study suggests that Li-PC-induced SE in immature rats cause long-term cognitive deficit and permanent cell loss in hippocampal CA1, but spare motor impairment.
Asunto(s)
Envejecimiento/efectos de los fármacos , Epilepsia del Lóbulo Temporal/patología , Hipocampo/crecimiento & desarrollo , Aprendizaje por Laberinto/efectos de los fármacos , Trastornos de la Memoria/patología , Degeneración Nerviosa/patología , Estado Epiléptico/patología , Envejecimiento/fisiología , Animales , Modelos Animales de Enfermedad , Epilepsia del Lóbulo Temporal/fisiopatología , Femenino , Hipocampo/patología , Hipocampo/fisiopatología , Litio/farmacología , Masculino , Aprendizaje por Laberinto/fisiología , Trastornos de la Memoria/inducido químicamente , Trastornos de la Memoria/fisiopatología , Actividad Motora/efectos de los fármacos , Actividad Motora/fisiología , Trastornos del Movimiento/etiología , Trastornos del Movimiento/patología , Trastornos del Movimiento/fisiopatología , Agonistas Muscarínicos/farmacología , Degeneración Nerviosa/inducido químicamente , Degeneración Nerviosa/fisiopatología , Neuronas/efectos de los fármacos , Neuronas/patología , Pilocarpina/farmacología , Ratas , Ratas Sprague-Dawley , Estado Epiléptico/inducido químicamente , Estado Epiléptico/fisiopatologíaRESUMEN
BACKGROUND AND PURPOSE: The authors attempt to determine whether hemodynamically significant extracranial internal carotid artery (ICA) lesions correlate with the severity of first-ever hemispheric ischemic stroke. METHODS: Carotid duplex was used to evaluate carotid arteries. The National Institutes of Health Stroke Scale was used to describe the severity of the stroke and was stratified as follows: 1-6 = mild, 7-15 = moderate, > 15 = severe. Duplex findings were categorized according to velocity criteria into < 50% stenosis if ICA peak systolic velocity (PSV) (cm/s) < 140 and > 50% stenosis if ICA PSV > 140 or ratio of ICA and common carotid artery in PSV > 2. No detectable flow at ICA was considered occlusion. Stroke subtype was classified according to TOAST criteria. RESULTS: Two hundred nineteen consecutive patients were enrolled, including 127 with mild, 65 with moderate, and 27 with severe stroke. The prevalence of ICA stenosis > 50% in each group was 3.6%, 1.4%, 0.9%, respectively. Two patients in the severe group had total ICA occlusion. The overall prevalence of significant ICA lesions was 6.8%. CONCLUSIONS: There is no positive correlation of stroke severity with the severity of duplex findings, which may be due to low prevalence of significant ICA lesions or other stroke mechanisms. Most of the patients had mild stroke, and the majority had ICA stenosis < 50%. Small-vessel occlusion tended to have mild severity of stroke. Intracranial artery lesions or other factors causing stroke in Taiwanese should be investigated. Given the low incidence of significant extracranial carotid disease in symptomatic Taiwanese stroke patients, routine screening of symptomatic Taiwanese for extracranial carotid artery disease does not provide enough information to determine stroke mechanism, and transcranial Doppler should be added to the screening tests.