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A 24-day-old male Japanese Black calf presented with weight-bearing lameness in the left hind limb and a swollen pelvis. Ultrasonography revealed the accumulation of fluidity materials with a mixture of hyper- and hypo-echogenicity, enclosed by 5-10-mm-thick capsular structures. On the ventral-dorsal radiograph of the pelvis, irregular and radiolucent osseous changes were evident in the femoral head and acetabulum within the swollen hip joint, with soft-tissue density. Computed tomography (CT) confirmed the relationship between the bony and soft tissue lesions, which was suspected following ultrasonography and radiography, and provided additional findings, such as intra-articular accumulation of gas and the degree of osteolytic changes. Based on the imaging findings and cytology of the arthrocentesis specimen, the patient was diagnosed with hip arthritis and osteomyelitis of the femoral head. Additionally, the severity of the lesions supported our decision to perform a femoral head ostectomy. The postoperative radiographic and CT findings of the treated pelvis were helpful in evaluating the efficacy of this technique. Following treatment, the animal could walk and showed normal development, although it was three postoperative months before weight-bearing lameness improved. To the best of our knowledge, this is the first report to describe the combined use of ultrasonography, radiography, and CT in the diagnosis, preoperative planning, and evaluation of the postoperative effects of bovine hip arthritis. Additionally, this report details the therapeutic efficacy of femoral head ostectomy for bovine hip arthritis, a technique that has not been reported previously.
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PURPOSE: Although morphological renal abnormalities in children with febrile urinary tract infection (fUTI) have been showed a predictive factor for recurrent infection, there are no available data on recurrence regarding sonographic renal enlargement at first fUTI episode, especially focusing on whether renal enlargement is temporary or not. MATERIALS AND METHODS: This cohort study reviewed the medical records of children who underwent renal ultrasound during their first fUTI during 2005-2013 and who were aged <15 years at diagnosis. We defined a kidney as temporary enlarged when the kidney length was ≥2 standard deviation above normal renal length for that age on sonography or a difference of ≥1 cm in sonographic length between the right and left kidneys, following normal renal length after antibiotic treatment. RESULTS: A total of 132 children were enrolled, of whom 11 had sonographic temporary temporal renal enlargement during their first fUTI. After completing antibiotic therapy for a first fUTI episode, 20 (15%) children had fUTI recurrence. The clinical characteristics at first episode of fUTI were not significantly different between renal enlargement and nonrenal enlargement groups. Children with temporary renal enlargement at a first fUTI episode had significantly lower fUTI recurrence-free survival proportion than those with nonrenal enlargement according to the Kaplan-Meier method (p = 0.003) Conclusion: Identification of temporary temporal renal enlargement as a predictor of recurrent fUTI may help identify children with a first episode of fUTI who will be warned of close monitoring.
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Nefropatias , Infecções Urinárias , Refluxo Vesicoureteral , Antibacterianos/uso terapêutico , Criança , Estudos de Coortes , Humanos , Reinfecção , Estudos Retrospectivos , Infecções Urinárias/complicações , Infecções Urinárias/diagnósticoRESUMO
BACKGROUND: Continuous negative extra-thoracic pressure (CNEP) can prevent children with apnea developing severe respiratory infection with endotracheal intubation. Little is known about children with mild acute respiratory disease, especially with a focus on clinical respiratory symptoms. METHODS: We conducted a prospective, observational study between July 2014 and July 2017 to evaluate the safety of a modified setting of CNEP in hospitalized children with symptoms of chest-wall retraction or nasal alar breathing without the requirement for immediate intubation therapy in a single center. A modified setting of CNEP was defined as 4 h of treatment comprising 3 consecutive hours of CNEP followed by 1 h of rest. RESULTS: We studied 19 hospitalized children with retraction or nasal breathing but no possible state of endotracheal intubation. The median age at admission was 0.9 years and the duration of CNEP was 6 days. No sedative drugs were used. The percentage of children with retraction or nasal breathing after 24 h from initiation of CNEP was significantly decreased compared with that just before CNEP (68% vs 100%, P = 0.02). Logistic regression showed no statistical evidence of contributing factors for pulmonary symptoms. No patients were transferred to receive intubation, but one boy reinitiated respiratory support within 6 months after discharge. No children had adverse events of upper airway obstruction, skin injury, interfering with access, hypothermia, discomfort from fitting a cuirass, and neck excoriation. CONCLUSIONS: Our results suggest that a modified setting of CNEP management can be tolerated and continued without concern of adverse events.
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Apneia , Intubação Intratraqueal , Criança , Humanos , Pulmão , Masculino , Estudos ProspectivosRESUMO
Very-long-chain acyl-CoA dehydrogenase (VLCAD) deficiency, a condition in which the body is unable to break down long-chain fatty acids properly, is the most common fatty acid oxidation disorder in Japan. Tandem mass spectrometry has been used in newborn screening (NBS), allowing the detection of patients with VLCAD deficiency even before symptoms manifest. However, tandem mass spectrometry has a high false positive rate. We investigated the clinical characteristics of patients with false positive results for tetradecenoyl acylcarnitine (C14:1). This case-control study used data collected between the 1st of January 2014 and the 31st of March 2019. The case group was defined as patients having levels of both C14:1 and C14:1/C2 ratio higher than cut-off levels in the first newborn mass screening, who were eventually diagnosed as false positives by attending doctors at Kobe University Hospital, Palmore Hospital, or Kakogawa Central City Hospital in Japan. The control group comprised 100 patients randomly selected from the three facilities. The false positive group included 17 cases, and the control group contained 300 patients. The demographics of each group did not show any significant differences in sex, body weight at birth, Cesarean section rate, complete breastfeeding rate, or the number of feedings per day. However, the change in body weight at the sampling day of NBS in the false positive and control groups was -10.2%, and - 4.6%, respectively, showing a statistically significant difference (p < 0.01). In addition, body weight gain at the one-month medical checkup was 38.9 g/day in the false positive group and 44.1 g/day in the control group (p < 0.05). An elevation of C14:1 carnitine has been reported in situations involving the catalysis of fatty acid. Therefore, patients with severe body weight loss might be associated with poor sucking or poor milk supply, which might cause a false positive elevation of C14:1 and C14:1/C2. In suspected VLCAD deficiency, attending doctors should pay attention to body weight changes recorded during newborn mass screening.
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Screening is a fundamental strategy for early detection, treatment, and prevention of progression of oral disease and those at high risk for oral disease. While numerous screening tools exist, questionnaires, and saliva tests are often suitable for screening. The questionnaire based on the PRECEDE-PROCEED model was developed, validated, and elucidated on the structural interrelationship between these two methods. In the current investigation, 311 adults had this questionnaire and saliva testing administered simultaneously during an occupational health checkup. The questionnaire was validated by classical test theory, item response theory, and path analysis. Through structural equation modeling, it was found that self-care guidance may be an important role of the family dentist. In addition, self-awareness of oral symptoms was significantly related to saliva test results. However, self-administered questionnaires and saliva tests together may provide more information than either of them alone for the detection, treatment, and prevention of progression of oral disease. Thus, simultaneous application of self-administered questionnaires and saliva tests is recommended during oral health checkups for adults.
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Doenças da Boca , Saúde Bucal , Adulto , Humanos , Exame Físico , Saliva , Inquéritos e QuestionáriosRESUMO
OBJECTIVES: This study aimed to verify the screening performance of our clinical prediction rule for neurological sequelae due to acute encephalopathy (NSAE-CPR), which previously identified the following three variables as predictive of poor outcomes: (1) refractory status epilepticus; (2) consciousness disturbance and/or hemiplegia at 6 hours from onset and (3) aspartate aminotransferase >90 IU/L within 6 hours of onset. DESIGN: Medical community-based multicentre retrospective cohort study. SETTING: Six regional hospitals in Harima and one tertiary centre in Kobe, Japan, from 2008 to 2012. PARTICIPANTS: We enrolled a total of 1612 patients aged <16 years who met the diagnostic criteria for an initial diagnosis of complex febrile seizure. Patients with a history of neurological disease and those included in the derivation cohort were excluded. PRIMARY OUTCOME MEASURES: Univariate and multivariate analyses were performed to determine the association between each of the three predictor variables and poor AE outcome (Pediatric Cerebral Performance Category score ≥2). Receiver operating characteristic curve (ROC) analysis was also performed to assess the screening performance of the NSAE-CPR. RESULTS: The ROC analysis identified at least one of the three predictive variables as an optimal cut-off point, with an area under the curve of 0.915 (95% CI 0.825 to 1.000). The sensitivity, specificity, positive and negative predictive values, positive and negative likelihood ratios and Matthews correlation coefficient were 0.867, 0.954, 0.149, 0.999, 18.704, 0.140 and 0.349, respectively. CONCLUSIONS: Our findings indicate that the NSAE-CPR can be used for the screening and identification of patients with poor outcomes due to acute encephalopathy within 6 hours of onset.
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Encefalopatias , Regras de Decisão Clínica , Transtornos da Consciência , Hemiplegia , Estado Epiléptico , Adulto , Idoso , Encefalopatias/complicações , Transtornos da Consciência/etiologia , Hemiplegia/etiologia , Humanos , Pessoa de Meia-Idade , Prognóstico , Estudos Retrospectivos , Fatores de Risco , Estado Epiléptico/etiologia , Adulto JovemRESUMO
While cyclopropanes have been explored as synthetically valuable building blocks, their transformation without conjugated substituents or directly substituted heteroatoms remains challenging. The current study describes the iridium-catalysed ring-opening hydrosilylation of cyclopropanes. A nitrogen-based directing group was found to control the reactivity of iridium active species as well as the regiochemistry of carbon-carbon bond cleavage and hydrosilylation.
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We aimed to determine the difference in tumor volume associated with the reconstruction model in positron-emission tomography (PET). To reduce the influence of the reconstruction model, we suggested a method to measure the tumor volume using the relative threshold method with a fixed threshold based on peak standardized uptake value (SUVpeak). The efficacy of our method was verified using 18F-2-fluoro-2-deoxy-D-glucose PET/computed tomography images of 20 patients with lung cancer. The tumor volume was determined using the relative threshold method with a fixed threshold based on the SUVpeak. The PET data were reconstructed using the ordered-subset expectation maximization (OSEM) model, the OSEM + time-of-flight (TOF) model, and the OSEM + TOF + point-spread function (PSF) model. The volume differences associated with the reconstruction algorithm (%VD) were compared. For comparison, the tumor volume was measured using the relative threshold method based on the maximum SUV (SUVmax). For the OSEM and TOF models, the mean %VD values were -0.06 ± 8.07 and -2.04 ± 4.23% for the fixed 40% threshold according to the SUVmax and the SUVpeak, respectively. The effect of our method in this case seemed to be minor. For the OSEM and PSF models, the mean %VD values were -20.41 ± 14.47 and -13.87 ± 6.59% for the fixed 40% threshold according to the SUVmax and SUVpeak, respectively. Our new method enabled the measurement of tumor volume with a fixed threshold and reduced the influence of the changes in tumor volume associated with the reconstruction model.
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Processamento de Imagem Assistida por Computador/normas , Carga Tumoral , Adulto , Idoso , Idoso de 80 Anos ou mais , Transporte Biológico , Feminino , Fluordesoxiglucose F18/metabolismo , Humanos , Neoplasias Pulmonares/diagnóstico por imagem , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Imagens de Fantasmas , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Padrões de ReferênciaRESUMO
Use of a rhodium catalyst with (R)-(S)-BPPFA ligand allows efficient synthesis of sila[n]helicenes via dehydrogenative silylation of C-H bonds. By selecting the proper ligands, the current method provides the ability to prepare unsymmetrical sila[n]helicene derivatives without any oxidants. The resulting sila[6]helicene is a rare example of a five-membered ring-fused [6]helicene, which was isolated as a single pure enantiomer without substituents on the terminal benzene rings.
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AIM: To study how lymph node metastasis (LNM) risk is stratified in undifferentiated-type early gastric cancer (undiff-EGC) dependent on combinations of risk factors. METHODS: Five hundred and sixty-seven cases with undiff-EGC undergoing gastrectomy with lymphadenectomy were examined retrospectively. Using clinicopathological factors of patient age, location, size, an endoscopic macroscopic tumor form, ulceration, depth, histology, lymphatic involvement (LI) and venous involvement (VI), LNM risk was examined and stratified by conventional statistical analysis and data-mining analysis. RESULTS: LNM was positive in 44 of 567 cases (7.8%). Univariate analysis revealed > 2 cm, protrusion, submucosal (sm), mixed type, LI and VI as significant prognostic factors and > 2 cm and LI-positive were independent factors by multivariate analysis. In preoperatively evaluable factors excluding LVI, sm and > 2 cm were independent factors. According to the depth and size, cases were categorized into the low-risk group [m and ≤ 2 cm, 0% (LNM incidence)], the moderate-risk group (m and > 2 cm, 5.6%; and sm and ≤ 2 cm, 6.0%), and the high-risk group (sm and > 2 cm, 19.3%). On the other hand, LNM occurred in 1.4% in all LI-negative cases, greatly lower than 28.2% in all LI-positive cases, and LNM incidence was low in LI-negative cases even in the moderate- and high-risk groups. CONCLUSION: LNM-related factors in undiff-EGC were depth and size preoperatively while those were LI and size postoperatively. Among these factors, LI was the most significantly correlated factor.
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Diferenciação Celular , Linfonodos/patologia , Neoplasias Gástricas/patologia , Algoritmos , Distribuição de Qui-Quadrado , Mineração de Dados , Árvores de Decisões , Detecção Precoce de Câncer , Feminino , Gastrectomia , Humanos , Japão , Modelos Logísticos , Excisão de Linfonodo , Linfonodos/cirurgia , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Razão de Chances , Prognóstico , Estudos Retrospectivos , Medição de Risco , Fatores de Risco , Neoplasias Gástricas/cirurgiaRESUMO
A microwave-enhanced plasma generation technique was combined with laser-induced ignition to improve ignition characteristics. A locally intensified microwave field was formed near the laser-induced breakdown plasma. As the plasma absorbed the microwaves, the plasma emission intensity increased. The plasma lifetime could be controlled by changing the microwave oscillation duration. Furthermore, the microwave-enhanced laser-induced breakdown plasma improved the minimum ignition energy of the methane/air pre-mixture with just a small amount of absorbed microwave energy.
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We describe a boy with Fisher syndrome. He presented the typical symptoms of Fisher syndrome, including external ophthalmoplegia, abnormality of convergence, and areflexia, after an episode of Campylobacter enterocolitis. Atypically, however, anti-GA1 antibody was detected in his serum, though anti-GQ1b and anti-GT1a antibodies were not. In addition, the tau protein level in his cerebrospinal fluid was elevated. Generally, Fisher syndrome is a self-limiting disease and has a good prognosis. In our patient, however, mild diplopia and areflexia persisted 6 months after their onset. Here, we report on the first Fisher syndrome patient with anti-GA1 antibody in the serum and elevated tau protein in the cerebrospinal fluid.
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Autoanticorpos/biossíntese , Doenças Autoimunes do Sistema Nervoso/líquido cefalorraquidiano , Gangliosídeos/imunologia , Síndrome de Miller Fisher/líquido cefalorraquidiano , Proteínas tau/biossíntese , Proteínas tau/líquido cefalorraquidiano , Autoanticorpos/sangue , Doenças Autoimunes do Sistema Nervoso/imunologia , Doenças Autoimunes do Sistema Nervoso/microbiologia , Infecções por Campylobacter/líquido cefalorraquidiano , Infecções por Campylobacter/imunologia , Criança , Humanos , Masculino , Síndrome de Miller Fisher/imunologia , Síndrome de Miller Fisher/microbiologia , Regulação para Cima/imunologia , Proteínas tau/sangueRESUMO
We report three familial cases of periodic fever with aphthous stomatitis, pharyngitis, and cervical adenitis syndrome, including a pair of monozygotic twins and their mother. It suggests that periodic fever with aphthous stomatitis, pharyngitis, and cervical adenitis syndrome may have a certain monogenetic background.
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Doenças Hereditárias Autoinflamatórias/genética , Linfadenite/genética , Faringite/genética , Estomatite Aftosa/genética , Adulto , Pré-Escolar , Feminino , Doenças Hereditárias Autoinflamatórias/complicações , Humanos , Lactente , Linfadenite/complicações , Pescoço , Faringite/complicações , Estomatite Aftosa/complicaçõesRESUMO
To evaluate the efficacy of topiramate (TPM) for the treatment of children with epilepsies, we introduced TPM to 45 patients whose epilepsy began in childhood and whose ages ranged from 4 months to 30 years old (mean age: 11 years 7 months). Thirteen of these patients had been diagnosed with generalized epilepsy (GE) (1 cryptogenic, 12 symptomatic), 30 with localization-related epilepsy (LRE) (7 idiopathic, 23 symptomatic), and 2 with unclassified epilepsy [1 case of severe myoclonic epilepsy in infancy (SMEI), 1 case of epilepsy with continuous spikes and waves during slow sleep (CSWS)]. The initial dose of TPM was 1.97 +/- 0.45 mg/kg/day, followed by a slow titration to the maximum dose of 7.32 +/- 1.32 mg/kg/day. After a mean treatment period of 13.5 months (range 4-20 months), the rate of reduction in seizure frequency by more than 50% [50% responder rate (50% RR)] and the rate of complete remission (seizure-free) were 53.8% and 23.1%, respectively, in patients with GE, and 73.3% and 23.3%, respectively, in patients with LRE. TPM was significantly effective against many seizure types including tonic, clonic, complex partial, myoclonic, and atypical absence seizures. Adverse effects included sleepiness in 13 cases (28.9%), weight loss in 6 cases (13.3%), and metabolic acidosis in 2 cases (4.4%); all of these effects were both mild and transient. In conclusion, TPM is effective and safe for the treatment of pediatric epilepsies.
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Anticonvulsivantes/administração & dosagem , Epilepsia/tratamento farmacológico , Frutose/análogos & derivados , Adolescente , Adulto , Fatores Etários , Criança , Pré-Escolar , Quimioterapia Combinada , Tolerância a Medicamentos , Epilepsia/classificação , Feminino , Frutose/administração & dosagem , Humanos , Lactente , Masculino , Topiramato , Resultado do Tratamento , Adulto JovemRESUMO
We report a patient with transient advanced atrioventricular (AV) block induced by obstructive sleep apnea (OSA). This 54-year-old man was diagnosed as having severe OSA and AV block with ventricular asystole for more than 6 s during overnight polysomnography, which occurred just from the onset of OSA before oxygen desaturation had occurred. An electrophysiological study revealed normal AV conduction system function and normal His-Purkinje system function. The resolution of OSA with continuous positive airway pressure therapy improved the advanced AV block. Therefore, the bradyarrhythmia was determined to be an OSA-induced AV block that occurred before oxygen desaturation.
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Bloqueio Atrioventricular/etiologia , Apneia Obstrutiva do Sono/complicações , Humanos , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de DoençaRESUMO
BACKGROUND: Congenital generalized lipodystrophy (CGL), Berardinelli-Seip syndrome, is a rare autosomal recessive disorder characterized by the generalized absence of adipose tissue at birth, severe insulin resistance early in life, hypertriglyceridemia, hepatomegaly, and the development of diabetes mellitus during puberty. Recently, two genes, BSCL2 and AGPAT2, were identified as causative genes for CGL. It has been reported that patients with BSCL mutations present with more severe clinical findings than those with AGPAT2 mutations. However, the clinical course of CGL caused by BSCL2 mutations in infancy has not been fully elucidated. METHODS: Two Japanese infantile patients with CGL from independent families were examined and underwent an oral glucose tolerance test. Insulin resistance and insulin secretion were estimated using the homeostasis model assessment for insulin resistance and the insulinogenic index, respectively. Sequence analysis of the entire coding region of BSCL2 and AGPAT2 was performed. RESULTS: Both CGL patients presented with normal glycemic profiles after oral glucose tolerance tests; however, the values from the homeostasis model assessment of insulin resistance were elevated and well above the cut-off point for diagnosis of infant insulin resistance in both patients. One patient possessed a known homozygous nonsense mutation in exon 8 (c.823C>T) of BSCL2; the other had a novel homozygous missense mutation in exon 5 (c.560A>G) of BSCL2. CONCLUSION: Japanese CGL patients with BSCL2 mutations presented with severe insulin resistance, even during infancy, prior to the development of diabetes mellitus.
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Códon sem Sentido/genética , Subunidades gama da Proteína de Ligação ao GTP/genética , Resistência à Insulina/genética , Lipodistrofia Generalizada Congênita/genética , Mutação de Sentido Incorreto/genética , Aberrações Cromossômicas , Consanguinidade , Análise Mutacional de DNA , Diabetes Mellitus Tipo 1/sangue , Diabetes Mellitus Tipo 1/diagnóstico , Diabetes Mellitus Tipo 1/genética , Feminino , Seguimentos , Genes Recessivos/genética , Teste de Tolerância a Glucose , Hepatomegalia/diagnóstico , Hepatomegalia/genética , Humanos , Lactente , Recém-Nascido , Japão , Lipodistrofia Generalizada Congênita/sangue , Lipodistrofia Generalizada Congênita/diagnóstico , Masculino , Reação em Cadeia da Polimerase , Tomografia Computadorizada por Raios X , Triglicerídeos/sangueRESUMO
Mutations in the dystrophin gene result in the most common inherited muscle disease, Duchenne muscular dystrophy (DMD). Duplications spanning one or more exons have been found to be the second most common disease-causing mutation in the dystrophin gene. Although the duplicated exons are commonly thought to be arranged in tandem, rare noncontiguous exon duplications have been disclosed without clarifying their location or orientation. Here we present the first report that details the exact locations and orientations of noncontiguous duplications in the dystrophin gene. Multiplex ligation-dependent probe amplification analysis of the dystrophin gene of a Japanese boy with DMD revealed that his genomic DNA contained duplications of exons from two separate fragments of the gene: one from exon 45 to exon 48 and the other from exon 55 to exon 63. To clarify the locations and orientations of the duplicated exons, reverse transcription-nested PCR analysis of dystrophin mRNA was conducted. Interestingly, the extra copies of exons 45-48 and exons 55-63 were found to be properly oriented between exons 48 and 49 and exons 63 and 64, respectively. These results indicated that two tandem duplication events occurred in the dystrophin gene of this patient and should contribute to the understanding of the duplication mechanisms that contribute to the development of DMD.
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Distrofina/genética , Duplicação Gênica , Distrofia Muscular de Duchenne/genética , Mapeamento Cromossômico , Cromossomos Humanos X , Primers do DNA , Éxons , Amplificação de Genes , Humanos , Recém-Nascido , Masculino , RNA Mensageiro/genéticaRESUMO
Tetrasomy 9p is a rare clinical syndrome and about 30% of known cases exhibit chromosome mosaicism. The cases with tetrasomy 9p mosaicism have been reported to show the various phenotypes. On the other hand, Klinefelter syndrome is well recognized chromosomal abnormality caused by an additional X chromosome in males (47,XXY), and the characteristic clinical findings include tall stature, immaturity of external genitalia, testicular dysfunction. Here, we report a 10-year-old male with tetrasomy of 9p mosaicism, whose phenotypic feature is mimicking Klinefelter syndrome. He was referred to our hospital for inconspicuous penis. He showed tall height (+2.5 SD). Endocrinological examination revealed the poor testosterone response to human chorionic gonadotropin administration, which indicated the testicular hypofunction, whereas MRI revealed concealed penis as a cause of inconspicuous penis. Because of the phenotype mimicking Klinefelter syndrome, karyotype of his blood lymphocytes was analyzed, and an additional marker chromosome was detected in 6% of the investigated metaphases. Fluorescence in situ hybridization analysis revealed that the marker chromosome was an isochromosome 9p, which resulted in tetrasomy 9p. Chromosome analysis of buccal smear also showed mosaicism for two karyotypes: 5% of cells had the isochromosome of 9p, and the other cells showed normal. This case is the second case with tetrasomy 9p mosaicism mimicking Klinefelter syndrome phenotype in the world. Our case, together with previously reported cases with the same association, indicates the possibility of testicular hypofunction and urogenital anomalies induced by overexpression of some genes on chromosome 9p.
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Aneuploidia , Gonadotropina Coriônica/fisiologia , Cromossomos Humanos Par 9/genética , Síndrome de Klinefelter/genética , Mosaicismo , Fenótipo , Testosterona/biossíntese , Criança , Diagnóstico Diferencial , Humanos , Cariotipagem , Síndrome de Klinefelter/diagnóstico , Síndrome de Klinefelter/patologia , MasculinoRESUMO
The dystrophin gene, which is mutated in Duchenne and Becker muscular dystrophy, is characterized by its extremely large introns. Seven cryptic exons from the intronic sequences of the dystrophin gene have been shown to be inserted into the processed mRNA. In this study, we have cloned seven novel cryptic exons embedded in dystrophin introns that were amplified from dystrophin mRNA isolated from lymphocytes. All of these sequences, which ranged in size from 27 to 151 bp, were found to be cryptic exons because they were completely homologous to intronic sequences (introns 1, 18, 29, 63, 67, and 77), and possessed consensus sequences for branch points, splice acceptor sites, and splice donor sites. Compared with the 77 authentic dystrophin exons, the 14 cryptic exons were characterized by (1) lower Shapiro's splicing probability scores for the splice donor and acceptor sites; (2) smaller and larger densities of splicing enhancer and silencer motifs, respectively; (3) a longer distance between the putative branch site and the splice acceptor site; and (4) with one exception, the introduction of premature stop codons into their respective transcripts. These characteristics indicated that the cryptic exons were weaker than the authentic exons. Our results suggested that a mutation deep within an intron that changed these parameters could cause dystrophinopathy. The cryptic exons identified provide areas that should be examined for the detection of mutations in the dystrophin gene, and they may help us to understand the roles of large dystrophin introns.
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Distrofina/genética , Éxons/genética , Processamento Alternativo/genética , Sequência de Bases , Células Cultivadas , Humanos , Íntrons/genética , Dados de Sequência Molecular , Distrofia Muscular de Duchenne/genética , RNA Mensageiro/análise , RNA Mensageiro/biossínteseRESUMO
BACKGROUND: Myostatin is a negative regulator of skeletal muscle growth. Truncating mutations in the myostatin gene have been reported to result in gross muscle hypertrophy. Duchenne muscular dystrophy (DMD), the most common lethal muscle wasting disease, is a result of an absence of muscle dystrophin. Although this disorder causes a rather uniform pattern of muscle wasting, afflicted patients display phenotypic variability. We hypothesized that genetic variation in myostatin is a modifier of the DMD phenotype. METHODS: We analyzed 102 Japanese DMD patients for mutations in the myostatin gene. RESULTS: Two polymorphisms that are commonly observed in Western countries, p.55A>T and p.153K>R, were not observed in these Japanese patients. An uncommon polymorphism of p.164E>K was uncovered in four cases; each patient was found to be heterozygous for this polymorphism, which had the highest frequency of the polymorphism observed in the Japanese patients. Remarkably, two patients were found to be heterozygous for one of two novel missense mutations (p.95D>H and p.156L>I). One DMD patient carrying a novel missense mutation of p.95D>H was not phenotypically different from the non-carriers. The other DMD patient was found to carry both a novel mutation (p.156L>I) and a known polymorphism (p.164E>K) in one allele, although his phenotype was not significantly modified. Any nucleotide change creating a target site for micro RNAs was not disclosed in the 3' untranslated region. CONCLUSION: Our results indicate that heterozygous missense mutations including two novel mutations did not produce an apparent increase in muscle strength in Japanese DMD cases, even in a patient carrying two missense mutations.