RESUMO
Most Mendelian disorders, including neuromuscular disorders, display extensive clinical heterogeneity that cannot be solely explained by primary genetic mutations. This phenotypic variability is largely attributed to the presence of disease modifiers, which can exacerbate or lessen the severity and progression of the disease. LAMA2-deficient congenital muscular dystrophy (LAMA2-CMD) is a fatal degenerative muscle disease resulting from mutations in the LAMA2 gene encoding Laminin-α2. Progressive muscle weakness is predominantly observed in the lower limbs in LAMA2-CMD patients, whereas upper limbs muscles are significantly less affected. However, very little is known about the molecular mechanism underlying differential pathophysiology between specific muscle groups. Here, we demonstrate that the triceps muscles of the dy2j/dy2j mouse model of LAMA2-CMD demonstrate very mild myopathic findings compared with the tibialis anterior (TA) muscles that undergo severe atrophy and fibrosis, suggesting a protective mechanism in the upper limbs of these mice. Comparative gene expression analysis reveals that S-Adenosylmethionine decarboxylase (Amd1) and Spermine oxidase (Smox), two components of polyamine pathway metabolism, are downregulated in the TA but not in the triceps of dy2j/dy2j mice. As a consequence, the level of polyamine metabolites is significantly lower in the TA than triceps. Normalization of either Amd1 or Smox expression in dy2j/dy2j TA ameliorates muscle fibrosis, reduces overactive profibrotic TGF-ß pathway and leads to improved locomotion. In summary, we demonstrate that a deregulated polyamine metabolism is a characteristic feature of severely affected lower limb muscles in LAMA2-CMD. Targeted modulation of this pathway represents a novel therapeutic avenue for this devastating disease.
Assuntos
Laminina/genética , Distrofia Muscular do Cíngulo dos Membros/genética , Distrofia Muscular Animal/genética , Poliaminas/metabolismo , Adenosilmetionina Descarboxilase/genética , Animais , Modelos Animais de Doenças , Regulação da Expressão Gênica , Humanos , Locomoção/genética , Locomoção/fisiologia , Camundongos , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiopatologia , Distrofia Muscular do Cíngulo dos Membros/fisiopatologia , Distrofia Muscular Animal/fisiopatologia , Mutação , Oxirredutases atuantes sobre Doadores de Grupo CH-NH/genética , Transdução de Sinais , Fator de Crescimento Transformador beta/genética , Poliamina OxidaseRESUMO
Our increasing knowledge of how genomic variants affect human health and the falling costs of whole-genome sequencing are driving the development of individualized genomic medicine. This new clinical paradigm uses knowledge of an individual's genomic variants to anticipate, diagnose and manage disease. While individualized genetic medicine offers the promise of transformative change in health care, it forces us to reconsider existing ethical, scientific and clinical paradigms. The potential benefits of pre-symptomatic identification of at-risk individuals, improved diagnostics, individualized therapy, accurate prognosis and avoidance of adverse drug reactions coexist with the potential risks of uninterpretable results, psychological harm, outmoded counseling models and increased health care costs. Here we review the challenges, opportunities and limits of integrating genomic analysis into pediatric clinical practice and describe a model for implementing individualized genomic medicine. Our multidisciplinary team of bioinformaticians, health economists, health services and policy researchers, ethicists, geneticists, genetic counselors and clinicians has designed a 'Genome Clinic' research project that addresses multiple challenges in pediatric genomic medicine--ranging from development of bioinformatics tools for the clinical assessment of genomic variants and the discovery of disease genes to health policy inquiries, assessment of clinical care models, patient preference and the ethics of consent.
Assuntos
Testes Genéticos , Genética Médica , Genoma Humano , Genômica , Pediatria , Medicina de Precisão , Ética Médica , Aconselhamento Genético , Predisposição Genética para Doença , Testes Genéticos/ética , Testes Genéticos/métodos , Testes Genéticos/normas , Genética Médica/ética , Genética Médica/métodos , Genética Médica/normas , Genômica/ética , Genômica/métodos , Genômica/tendências , Humanos , Pediatria/métodos , Pediatria/normas , Medicina de Precisão/ética , Medicina de Precisão/métodos , Medicina de Precisão/normas , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
We describe seven patients with KDM6A (located on Xp11.3 and encodes UTX) mutations, a rare cause of Kabuki syndrome (KS2, MIM 300867) and report, for the first time, germ-line missense and splice-site mutations in the gene. We demonstrate that less than 5% cases of Kabuki syndrome are due to KDM6A mutations. Our work shows that similar to the commoner Type 1 Kabuki syndrome (KS1, MIM 147920) caused by KMT2D (previously called MLL2) mutations, KS2 patients are characterized by hypotonia and feeding difficulties during infancy and poor postnatal growth and short stature. Unlike KS1, developmental delay and learning disability are generally moderate-severe in boys but mild-moderate in girls with KS2. Some girls may have a normal developmental profile. Speech and cognition tend to be more severely affected than motor development. Increased susceptibility to infections, join laxity, heart, dental and ophthalmological anomalies are common. Hypoglycaemia is more common in KS2 than in KS1. Facial dysmorphism with KDM6A mutations is variable and diagnosis on facial gestalt alone may be difficult in some patients. Hypertrichosis, long halluces and large central incisors may be useful clues to an underlying KDM6A mutation in some patients.
Assuntos
Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/genética , Face/anormalidades , Genes Ligados ao Cromossomo X , Doenças Hematológicas/diagnóstico , Doenças Hematológicas/genética , Histona Desmetilases/genética , Mutação , Proteínas Nucleares/genética , Doenças Vestibulares/diagnóstico , Doenças Vestibulares/genética , Substituição de Aminoácidos , Criança , Pré-Escolar , Éxons , Fácies , Feminino , Ordem dos Genes , Estudos de Associação Genética , Humanos , Masculino , Taxa de Mutação , Fenótipo , Reprodutibilidade dos Testes , Análise de Sequência de DNARESUMO
Chronic graft-vs-host disease (cGVHD) myositis is a rare complication of hematopoietic SCT, for which the pathogenesis and optimal therapy are unclear. We performed immunohistochemistry on muscle biopsies from pediatric cGVHD myositis and typical cases of autoimmune dermatomyositis and polymyositis. The immunostaining pattern of cGVHD myositis was distinct from that of typical cases of autoimmunity. There was a high proportion of CD20+ and CD68+ cells, and the best therapeutic response was achieved with rituximab (anti-CD20). These results suggest that cGVHD myositis may be mediated by different leukocytes than similar autoimmune diseases and that treatment may be optimized by targeting the specific cellular infiltrates identified in affected tissue.
Assuntos
Doença Enxerto-Hospedeiro/tratamento farmacológico , Doença Enxerto-Hospedeiro/patologia , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Anticorpos Monoclonais Murinos/uso terapêutico , Antígenos CD/imunologia , Antígenos CD20/imunologia , Antígenos de Diferenciação Mielomonocítica/imunologia , Doenças Autoimunes/imunologia , Linfócitos B/imunologia , Criança , Dermatomiosite/patologia , Dermatomiosite/terapia , Doença Enxerto-Hospedeiro/imunologia , Humanos , Imuno-Histoquímica , Polimiosite/patologia , Polimiosite/terapia , RituximabRESUMO
OBJECTIVE: Neuronal nitric oxide synthase (nNOS), normally expressed at the sarcolemmal membrane, is known to be mislocalized to the sarcoplasm in several forms of muscular dystrophy. Our objectives were to characterize further the range of patients manifesting aberrant nNOS sarcolemmal immunolocalization and to study nNOS localization in animal models of nondystrophic myopathy. METHODS: We carried out a retrospective cross-sectional study. We performed immunofluorescent staining for nNOS on biopsy specimens from 161 patients with acquired and nondystrophin inherited neuromuscular conditions. The localization of sarcolemmal nNOS correlated with mobility and functional status. Muscle specimens from mouse models of steroid-induced and starvation-related atrophy were studied for qualitative and quantitative nNOS expression. RESULTS: Sarcolemmal nNOS staining was abnormal in 42% of patients with inherited myopathic conditions, 25% with acquired myopathic conditions, 57% with neurogenic conditions, and 93% with hypotonia. Interestingly, we found significant associations between mobility status or muscle function and sarcolemmal nNOS expression. Furthermore, mouse models of catabolic stress also demonstrated mislocalization of sarcolemmal nNOS. CONCLUSION: Our analyses indicate that nNOS mislocalization is observed in a broad range of nondystrophic neuromuscular conditions associated with impaired mobility status and catabolic stress. Our findings suggest that the assessment of sarcolemmal localization of nNOS represents an important tool for the evaluation of muscle biopsies of patients with a variety of inherited and acquired forms of neuromuscular disorders.
Assuntos
Músculo Esquelético/metabolismo , Doenças Neuromusculares/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Sarcolema/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Distribuição de Qui-Quadrado , Criança , Pré-Escolar , Estudos Transversais , Imunofluorescência , Humanos , Lactente , Camundongos , Pessoa de Meia-Idade , Distrofia Muscular Animal/genética , Distrofia Muscular Animal/metabolismo , Doenças Neuromusculares/genética , Óxido Nítrico Sintase Tipo I/genética , Estudos Retrospectivos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sarcolema/genética , SciuridaeRESUMO
Cardiomyopathy is a multifactorial disease, and the dystrophin-glycoprotein complex has been implicated in the pathogenesis of both hereditary and acquired forms of the disease. Using mouse models of cardiomyopathy made by ablating genes for components of the sarcoglycan complex, we show that long-term treatment with verapamil, a calcium channel blocker with vasodilator properties, can alleviate the severe cardiomyopathic phenotype, restoring normal serum levels for cardiac troponin I and normal cardiac muscle morphology. Interruption of verapamil treatment leads again to vascular dysfunction and acute myocardial necrosis, indicating that predilection for cardiomyopathy is a continuing process. In contrast, verapamil did not prevent cardiac muscle pathology in dystrophin-deficient mdx mice, which neither show a disruption of the sarcoglycan complex in vascular smooth muscle nor vascular dysfunction. Hence, our data strongly suggest that pharmacological intervention with verapamil merits investigation as a potential therapeutic option not only for patients with sarcoglycan mutations, but also for patients with idiopathic cardiomyopathy associated with myocardial ischemia not related to atherosclerotic coronary artery disease.
Assuntos
Cardiomiopatias/prevenção & controle , Proteínas do Citoesqueleto/deficiência , Glicoproteínas de Membrana/deficiência , Proteínas de Membrana/deficiência , Miocárdio/metabolismo , Proteínas de Neoplasias , Vasodilatadores/uso terapêutico , Verapamil/uso terapêutico , Fatores Etários , Animais , Cardiomiopatias/sangue , Cardiomiopatias/genética , Proteínas de Transporte/genética , Circulação Coronária , Proteínas do Citoesqueleto/genética , Modelos Animais de Doenças , Glicoproteínas de Membrana/genética , Proteínas de Membrana/genética , Camundongos , Miocárdio/patologia , Sarcoglicanas , Troponina/sangueRESUMO
Muscular dystrophies represent a heterogeneous group of disorders, which have been largely classified by clinical phenotype. In the last 10 years, identification of novel skeletal muscle genes including extracellular matrix, sarcolemmal, cytoskeletal, cytosolic, and nuclear membrane proteins has changed the phenotype-based classification and shed new light on the molecular pathogenesis of these disorders. A large number of genes involved in muscular dystrophy encode components of the dystrophin-glycoprotein complex (DGC) which normally links the intracellular cytoskeleton to the extracellular matrix. Mutations in components of this complex are thought to lead to loss of sarcolemmal integrity and render muscle fibers more susceptible to damage. Recent evidence suggests the involvement of vascular smooth muscle DGC in skeletal and cardiac muscle pathology in some forms of sarcoglycan-deficient limb-girdle muscular dystrophy. Intriguingly, two other forms of limb-girdle muscular dystrophy are possibly caused by perturbation of sarcolemma repair mechanisms. The complete clarification of these various pathways will lead to further insights into the pathogenesis of this heterogeneous group of muscle disorders.
Assuntos
Músculos/patologia , Distrofias Musculares/genética , Distrofias Musculares/patologia , HumanosRESUMO
BACKGROUND: In a systematic analysis of inherited forms of cardiomyopathy, we previously identified a family with X-linked dilated cardiomyopathy characterised by a mutation in the rod region of dystrophin. We have now attempted to eludicate the genetic mechanism involved in this disease, as well as the role of dystrophin-associated glycoproteins. METHODS: The affected dystrophin epitope, which lacks binding to the dys-1 antibody, was analysed by single-strand conformation polymorphism analysis, reverse-transcription PCR, and DNA sequencing. Effects on dystrophin-associated glycoproteins were studied by immunohistochemistry and western blotting. FINDINGS: A translation-termination mutation (C4148T) in exon 29 of the dystrophin gene was found in all affected family members. Alternative splicing rescued the reading frame and led to the expression of a dystrophin molecule lacking 50 aminoacids both in cardiac and skeletal muscle. Immunohistochemical analysis of the dystrophin-associated proteins revealed a reduction of beta-sarcoglycan and delta-sarcoglycan in the sarcolemma of cardiac muscle but not skeletal muscle tissue. However, western blotting revealed similar amounts of sarcoglycan subunits in both tissues. INTERPRETATION: The molecular mechanism of this subtype of X-linked cardiomyopathy may be explained by a conformational change in exon-29-deleted dystrophin, resulting in disruption of the sarcoglycan assembly in heart muscle but not skeletal muscle.
Assuntos
Cardiomiopatia Dilatada/genética , Códon sem Sentido/genética , Proteínas do Citoesqueleto/genética , Distrofina/genética , Ligação Genética/genética , Glicoproteínas de Membrana/genética , Cromossomo X/genética , Distroglicanas , Feminino , Humanos , Masculino , Mutação , Linhagem , SarcoglicanasRESUMO
Variabilities of serum total cholesterol and potassium results provided to 11 medical clinics were assessed using an audit sample-split specimen design. This involved collection of 3 tubes of blood from each of 302 patients, with 1 split specimen divided into 3 audit samples: 1 was sent to the original participating laboratory, another to a commercial referral laboratory, and the third to an academic referee laboratory. Two methods were used to assess variability of test results. Method 1 was based on result pairs corresponding to the split specimen and its corresponding audit sample. Method 2 was based on audit sample results only. The 2 methods provided comparable results for total cholesterol; the estimated coefficient of variation was 1.0% to 3.7%. However, method 1 consistently provided higher estimates of variability for potassium; the estimated SD was 0.096 to 0.168 mmol/L for method 1, while it was 0.035 to 0.090 mmol/L for method 2. Method 1 is more practical, but method 2 can provide a more accurate assessment of analytic variability.
Assuntos
Colesterol/sangue , Técnicas de Laboratório Clínico/normas , Potássio/sangue , Humanos , Laboratórios Hospitalares , Consultórios Médicos , Reprodutibilidade dos TestesRESUMO
Limb-girdle muscular dystrophy type 2E (LGMD 2E) is caused by mutations in the beta-sarcoglycan gene, which is expressed in skeletal, cardiac, and smooth muscle. beta-sarcoglycan-deficient (Sgcb-null) mice developed severe muscular dystrophy and cardiomyopathy with focal areas of necrosis. The sarcoglycan-sarcospan and dystroglycan complexes were disrupted in skeletal, cardiac, and smooth muscle membranes. epsilon-sarcoglycan was also reduced in membrane preparations of striated and smooth muscle. Loss of the sarcoglycan-sarcospan complex in vascular smooth muscle resulted in vascular irregularities in heart, diaphragm, and kidneys. Further biochemical characterization suggested the presence of a distinct epsilon-sarcoglycan complex in skeletal muscle that was disrupted in Sgcb-null mice. Thus, perturbation of vascular function together with disruption of the epsilon-sarcoglycan-containing complex represents a novel mechanism in the pathogenesis of LGMD 2E.
Assuntos
Proteínas do Citoesqueleto/genética , Glicoproteínas de Membrana/genética , Músculo Esquelético/patologia , Distrofia Muscular Animal/genética , Distrofia Muscular Animal/patologia , Miocárdio/patologia , Animais , Cardiomiopatias/genética , Cardiomiopatias/patologia , Proteínas do Citoesqueleto/deficiência , Proteínas do Citoesqueleto/fisiologia , Distroglicanas , Distrofina/genética , Pulmão/patologia , Glicoproteínas de Membrana/deficiência , Glicoproteínas de Membrana/fisiologia , Camundongos , Camundongos Knockout , Microssomos/patologia , NecroseRESUMO
We rescreened Papanicolaou smear slides from 40,245 women, which had been examined by 81 cytology screeners, scored the screeners' work performance, and compared these scores with the results of the screeners' performance on glass slide and computer-based proficiency tests. All diagnoses (i.e., from the proficiency tests, the original slides, and the rescreened slides) were classified in the 4 diagnostic categories specified in the Clinical Laboratory Improvement Amendments. The rescreening scores were standardized to account for different distributions of abnormalities in the proficiency tests and rescreened slides. We compared a standardized score with the proficiency test scores. Of the cases, 91% were categorized as normal, benign, or reactive changes when rescreened, and 98% of these agreed with the original diagnosis. Sixteen percent of low-grade and 15% of high-grade intraepithelial lesions were classified as normal. The rank correlation between the rescreening scores and both proficiency tests was 0.24 using a scoring scheme for cytotechnologists. The correlation between the rescreening and proficiency testing scores indicates that performance on a 10-slide test gives some indication of the true performance of screeners. The computer-based test shows promise as an alternative to the glass slide test but needs further development and validation.
Assuntos
Pessoal de Laboratório Médico/normas , Teste de Papanicolaou , Esfregaço Vaginal/normas , Reações Falso-Negativas , Feminino , Humanos , Competência Profissional , Controle de Qualidade , Displasia do Colo do Útero/diagnósticoRESUMO
To investigate mechanisms in the pathogenesis of cardiomyopathy associated with mutations of the dystrophin-glycoprotein complex, we analyzed genetically engineered mice deficient for either alpha-sarcoglycan (Sgca) or delta-sarcoglycan (Sgcd). We found that only Sgcd null mice developed cardiomyopathy with focal areas of necrosis as the histological hallmark in cardiac and skeletal muscle. Absence of the sarcoglycan-sarcospan (SG-SSPN) complex in skeletal and cardiac membranes was observed in both animal models. Loss of vascular smooth muscle SG-SSPN complex was only detected in Sgcd null mice and associated with irregularities of the coronary vasculature. Administration of a vascular smooth muscle relaxant prevented onset of myocardial necrosis. Our data indicate that disruption of the SG-SSPN complex in vascular smooth muscle perturbs vascular function, which initiates cardiomyopathy and exacerbates muscular dystrophy.
Assuntos
Cardiomiopatia Dilatada/genética , Proteínas de Transporte/fisiologia , Proteínas do Citoesqueleto/fisiologia , Glicoproteínas de Membrana/fisiologia , Proteínas de Membrana/fisiologia , Músculo Liso Vascular/metabolismo , Distrofia Muscular Animal/genética , Proteínas de Neoplasias , Animais , Cardiomiopatia Dilatada/metabolismo , Cardiomiopatia Dilatada/patologia , Vasos Coronários/patologia , Proteínas do Citoesqueleto/deficiência , Proteínas do Citoesqueleto/genética , Substâncias Macromoleculares , Glicoproteínas de Membrana/deficiência , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Knockout , Músculo Liso Vascular/patologia , Distrofia Muscular Animal/metabolismo , Distrofia Muscular Animal/patologia , Miocárdio/patologia , Necrose , Condicionamento Físico Animal/efeitos adversos , SarcoglicanasRESUMO
The integrins are a large family of heterodimeric transmembrane cellular receptors which mediate the association between the extracellular matrix (ECM) and cytoskeletal proteins. The alpha7beta1 integrin is a major laminin binding integrin in skeletal and cardiac muscle and is thought to be involved in myogenic differentiation and migration processes. The main binding partners of the alpha7 integrin are laminin-1 (alpha1-beta1-gamma1), laminin-2 (alpha2-beta1-gamma1) and laminin-4 (alpha2-beta2-gamma1). Targeted deletion of the gene for the alpha7 integrin subunit (ITGA7) in mice leads to a novel form of muscular dystrophy. In the present study we have investigated the expression of two alternative splice variants, the alpha7B and beta1D integrin subunits, in normal human skeletal muscle, as well as in various forms of muscular dystrophy. In normal human skeletal muscle the expression of the alpha7 integrin subunit appeared to be developmentally regulated: it was first detected at 2 years of age. In contrast, the beta1D integrin could be detected in immature and mature muscle in the sarcolemma of normal fetal skeletal muscle at 18 weeks gestation. The expression of alpha7B integrin was significantly reduced at the sarcolemma in six patients with laminin alpha2 chain deficient congenital muscular dystrophy (CMD) (age >2 years). However, this reduction was not correlated with the amount of laminin alpha2 chain expressed. In contrast, the expression of the laminin alpha2 chain was not altered in the skeletal muscle of the alpha7 knock-out mice. These data argue in favor that there is not a tight correlation between the expression of the alpha7 integrin subunit and that of the laminin alpha2 chain in either human or murine dystrophic muscle. Interestingly, in dystrophinopathies (Duchenne and Becker muscular dystrophy; DMD/BMD) expression of alpha7B was upregulated irrespective of the level of dystrophin expression as shown by a strong sarcolemmal staining pattern even in young boys (age <2 years). The expression of the beta1D integrin subunit was not altered in any of our patients with different types of muscular dystrophy. In contrast, sarcolemmal expression of beta1D integrin was significantly reduced in the alpha7 integrin knock-out mice, whereas the expression of the components of the DGC was not altered. The secondary loss of alpha7B in laminin alpha2 chain deficiency defines a biochemical change in the composition of the plasma membrane resulting from a primary protein deficiency in the basal lamina. These findings, in addition to the occurrence of a muscular dystrophy in alpha7 deficient mice, implies that the alpha7B integrin is an important laminin receptor within the plasma membrane which plays a significant role in skeletal muscle function and stability.
Assuntos
Antígenos CD/genética , Cadeias alfa de Integrinas , Laminina/deficiência , Laminina/genética , Músculo Esquelético/fisiopatologia , Distrofias Musculares/genética , Adolescente , Adulto , Envelhecimento , Sequência de Aminoácidos , Animais , Anticorpos , Antígenos CD/fisiologia , Criança , Pré-Escolar , Proteínas do Citoesqueleto/genética , Distroglicanas , Distrofina/genética , Desenvolvimento Embrionário e Fetal , Feto , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Lactente , Recém-Nascido , Integrinas/genética , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Desenvolvimento Muscular , Músculo Esquelético/embriologia , Músculo Esquelético/crescimento & desenvolvimento , Distrofias Musculares/congênito , Isoformas de Proteínas/genética , SarcoglicanasRESUMO
The congenital muscular dystrophies (CMDs) are a heterogeneous group of disorders. Among these, the laminin alpha 2 chain 'merosin' deficient CMD is caused by mutations of the LAMA2 gene on chr 6q2 and Fukuyama CMD is linked to chr 9q31. We report a 7-year-old boy who was born to consanguineous healthy parents. His motor and mental development were slow. Creatine kinase (CK) was elevated (2.100 U/l), and the muscle biopsy was dystrophic. He sat unsupported at 12 months and took his first steps at 3 years of age. At 6 years of age he could walk up to 500 m. He was mentally retarded and spoke single words only. At 1 year, MR imaging of the brain showed abnormal increased periventricular T2-signal, consistent with dysmyelination as well as pontocerebellar hypoplasia and several cerebellar cysts. The pattern of gyration was normal. Follow-up at 4 years showed normalization of the previously abnormal periventricular T2-signal. Immunohistochemical analysis of the skeletal muscle showed normal expression of laminin alpha 2 for a C-terminal antibody and antibodies to the 300 and 150 kDa fragments, as well as of laminins alpha 5, beta 1, beta 2 and gamma 1. The boy has two healthy younger brothers. Linkage analysis excluded the candidate loci on chromosomes 6q2 and 9q31. As such, the patient's data are suggestive of a new form of laminin alpha 2 positive CMD characterized by transient brain dysmyelination, pontocerebellar hypoplasia and mental retardation.
Assuntos
Cerebelo/anormalidades , Deficiência Intelectual/metabolismo , Laminina/análise , Distrofias Musculares/metabolismo , Bainha de Mielina/fisiologia , Ponte/anormalidades , Biópsia , Criança , Ligação Genética , Marcadores Genéticos , Haplótipos , Humanos , Deficiência Intelectual/patologia , Imageamento por Ressonância Magnética , Masculino , Distrofias Musculares/congênito , Linhagem , Fatores de TempoRESUMO
To evaluate a split-specimen design to identify problems in the testing process in hospital and physician office laboratories, we examined the testing for serum total cholesterol (n = 646) and potassium (n = 732) at 11 medical clinics evaluating 30-199 patients (mean, 125). Clinic personnel collected three tubes of blood from each patient. One specimen was processed routinely, the second was sent to a referral laboratory (RL), and the third specimen was sent to a holding facility for storage. The corresponding stored sample was retrieved and divided into three audit samples randomly and when result difference for the first two specimens exceeded critical values; one audit sample was sent to the original participant, the second to the RL, and the third to a referee laboratory. When three criteria were used, the result discrepancy rates were 2.5-8.7% for potassium and 1.5-4.6% for cholesterol. The split-specimen design could be implemented and evaluated as a monitoring system for a portion of the testing process.
Assuntos
Laboratórios/organização & administração , Manejo de Espécimes , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Colesterol/sangue , Estudos de Avaliação como Assunto , Humanos , Laboratórios/normas , Pessoa de Meia-Idade , Potássio/sangue , Garantia da Qualidade dos Cuidados de Saúde , Valores de ReferênciaRESUMO
Chediak-Higashi syndrome (CHS) is a hereditary, biphasic immunodeficiency syndrome which usually leads to early death, during the first decade. The second phase is characterized by a lymphoproliferative syndrome with histiocytic infiltrations in various tissues. Recently the gene has been identified on chromosome 1q43. In the patient presented here, a mutation within codon 3197 was found, resulting in a frame-shift. Additionally, Duchenne muscular dystrophy (DMD) was diagnosed by immunostaining of the muscle. Unusual for both CHS and DMD muscle weakness and hypotonia became evident during the first months of life. Compared to typical DMD cases we found an increased histiocytic infiltration in the muscle. The underlying muscular dystrophy probably predisposes to the affection of muscle in the second phase of CHS. This patient is presented as an example of modification of the phenotype by a second genetic disease.
Assuntos
Síndrome de Chediak-Higashi/patologia , Distrofias Musculares/patologia , Síndrome de Chediak-Higashi/complicações , Síndrome de Chediak-Higashi/genética , Distrofina/análise , Distrofina/genética , Evolução Fatal , Mutação da Fase de Leitura , Humanos , Imuno-Histoquímica , Lactente , Recém-Nascido , Masculino , Debilidade Muscular/complicações , Debilidade Muscular/patologia , Músculos/metabolismo , Músculos/patologia , Músculos/ultraestrutura , Distrofias Musculares/complicações , Distrofias Musculares/genética , Proteínas/genética , Proteínas de Transporte VesicularRESUMO
OBJECTIVE: To characterize the expression of distinct fragments of laminin alpha2 chain in patients with partial laminin alpha2 chain deficiency and variable clinical severity. BACKGROUND: Deficiency of laminin alpha2 chain caused by mutations of the LAMA2 gene on chromosome 6q2 account for approximately 50% of cases of congenital muscular dystrophy (CMD) in white patients. The complete absence of laminin alpha2 is usually associated with a severe phenotype affecting skeletal muscle and the peripheral and central nervous systems. METHODS: Quantitative assessment of immunofluorescence to study the expression of C- and N-terminal portions of laminin alpha2 chain in five patients with partial laminin alpha2 chain deficiency and variable phenotype. All five patients showed abnormal T2 signal on brain MRI. RESULTS: Immunohistochemistry of muscle specimens showed preserved or minimally reduced expression of the C-terminal region of the laminin alpha2 chain (67 to 74%), but a marked reduction of the N-terminal region in four patients (13 to 19%). One patient with a mild phenotype had a partial reduction (45%) of the C-terminal and the N-terminal (51%) portions of the laminin alpha2 chain. Two patients were unable to walk or sit, although the C-terminal portion of the laminin alpha2 chain was expressed at significant levels (67 to 74%). In contrast, two patients with a similar expression of the C-terminus (67 to 70%) had a milder phenotype and became ambulatory. It was impossible to predict the phenotypes in these four patients with a strong expression of the C-terminus and with low levels of the N-terminus based on the amount of protein expressed. In addition, the laminin beta2 chain was moderately reduced (54 to 75%) in all patients with laminin alpha2 chain deficiency. A strong correlation between the amount of the C-terminus but not for the N-terminus and laminin beta2 reduction could be observed. CONCLUSIONS: N-terminal antibodies to the laminin alpha2 chain provide a more precise immunohistochemical detection of partially laminin alpha2 chain-deficient CMD. The secondary reduction of laminin beta2 chain may better define laminin alpha2 chain-deficient CMD. More data are needed to predict which portions of C-terminus and midrod region of the laminin alpha2 chain result in a semifunctional protein and a milder phenotype.
Assuntos
Laminina/genética , Distrofias Musculares/congênito , Distrofias Musculares/genética , Adulto , Anticorpos , Membrana Basal/química , Membrana Basal/imunologia , Pré-Escolar , Epitopos/análise , Imunofluorescência , Expressão Gênica , Humanos , Técnicas In Vitro , Lactente , Recém-Nascido , Laminina/química , Laminina/imunologia , Fenótipo , Estrutura Terciária de ProteínaRESUMO
OBJECTIVE: The purpose of this study was to assess the feasibility of using a prototype split-specimen design to assess integrity of a portion of the total testing process in medical clinics and laboratories. DESIGN: Two or three tubes of venous blood were collected from 177 patients for analysis of one of three analytes (serum potassium, serum total cholesterol, and whole-blood hemoglobin). Patients were seen at one of the nine clinics participating in this study. In all cases, one tube of blood from each patient was sent to a commercial referral laboratory, and the other tube(s) forwarded to the laboratory that routinely tested specimens for the clinic (participating laboratory) for analysis. Each participating laboratory removed a preanalysis and sometimes a post-analysis aliquot from each specimen and forwarded these to the referral laboratory for analysis. SETTING: The study was conducted in six physician office laboratories (three serving 1 to 4 [mean, 2.7] internists and three serving 3 to 24 [mean, 12] family physicians) and three hospital laboratories (serving hospitals with 100 to more than 700 beds). PATIENTS: Study patients were voluntary participants and provided informed consent. Patient age ranged from 18 to 80 years, and for all the laboratory test was specifically ordered for clinical reasons. Patients who were unable or unwilling to provide informed consent, those for whom testing would require that they provide more than 100 mL of blood, those whose blood was being collected by fingerstick, and those with results that were part of a laboratory test profile were excluded. MAIN OUTCOME MEASURES: Two main outcome measures were assessed: (1) percent differences between split-specimen results exceeding the maximum allowable imprecision level, which was based on published biological variation data (defined as one-half of the intraindividual percent coefficient of variation), for each analyte (result discrepancies); and (2) all "problems" (defined as departures from standard operating procedures) that could be documented by retrospective review of all relevant medical and laboratory records. RESULTS: The rate of result discrepancies was 1 in 20 (5%) for patients in whom hemoglobin was analyzed, 12 in 57 (21%) for patients in whom potassium was analyzed, and 1 in 60 (2%) for patients in whom total cholesterol was analyzed. Results of samples obtained during the aliquoting and storage phases of the total testing process were subject to study-induced problems and were generally not useful in tracing problems to specific stages of the testing process. A total of 28 problems (involving 26 patients) were documented, but only 6 problems were due to routine testing processes. CONCLUSIONS: The feasibility and limitations of a split-specimen design to detect result discrepancies were demonstrated. Most documented problems (22 of 28, or 79%) were study induced. To assess integrity of the total testing process, such problems need to be avoided.
Assuntos
Técnicas de Laboratório Clínico/normas , Laboratórios Hospitalares/normas , Manejo de Espécimes/normas , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Análise Química do Sangue/normas , Colesterol/sangue , Estudos de Viabilidade , Hemoglobinas/análise , Humanos , Pessoa de Meia-Idade , Potássio/sangue , Reprodutibilidade dos TestesRESUMO
We studied the distribution of laminin beta 2 chain in the skeletal muscle basement membrane of 16 patients with congenital muscular dystrophy (CMD) by immunohistochemistry. A dramatic reduction in the laminin beta 2 staining was observed in four patients with classical merosin-negative CMD. A moderate reduction of laminin beta 2 labelling was observed in four patients with partial merosin deficiency and two patients with merosin-positive CMD. Two patients with merosin-positive CMD had no apparent changes in the expression of laminin beta 2. In three patients and one fetus diagnosed as Walker-Warburg syndrome (WWS) the laminin beta 2 pattern was similar to normal controls. We conclude that a primary deficiency in the laminin alpha 2 chain may lead to a vast or moderate reduction in the laminin beta 2 chain in the skeletal muscle membrane.
Assuntos
Laminina/biossíntese , Distrofias Musculares/congênito , Distrofias Musculares/metabolismo , Adolescente , Criança , Pré-Escolar , Feminino , Técnica Direta de Fluorescência para Anticorpo , Humanos , Imuno-Histoquímica , Lactente , Laminina/metabolismo , Masculino , Distrofias Musculares/patologia , GravidezRESUMO
A newborn male infant born to consanguineous parents was found to have diffuse polymicrogyria associated with an unusual pattern of congenital anomalies including microbrachycephaly, turricephaly, blepharophimosis, microstomia with maxillary retrusion and mandibular prognathism, micropenis with cryptorchidism, camptodactyly and adducted thumbs, and a progeroid appearance. The combination of manifestations in our patient represents a unique form of polymicrogyria with congenital anomalies, probably representing a new syndrome.