Clinical features and MYO5B mutations of a family affected by microvillus inclusion disease / 中国当代儿科杂志

Microvillus inclusion disease (MVID) is an autosomal recessive disorder caused by biallelic mutations in the MYO5B or STX3 gene. Refractory diarrhea and malabsorption are the main clinical manifestations. The aim of this study was to investigate the clinical features and MYO5B gene mutations of an infant with MVID. A 21-day-old female infant was referred to the hospital with the complaint of diarrhea for 20 days. On physical examination, growth retardation of the body weight and length was found along with moderately jaundiced skin and sclera. Breath sounds were clear in the two lungs and the heart sounds were normal. The abdomen was distended and the veins in the abdominal wall were observed. The liver and spleen were not palpable. Biochemical analysis revealed raised serum total bile acids, bilirubin, transaminases and γ-glutamyl transpeptidase while decreased levels of serum sodium, chloride, phosphate and magnesium. Blood gas analysis indicated metabolic acidosis. The preliminary diagnosis was congenital diarrhea, and thus parenteral nutrition was given along with other symptomatic and supportive measures. However, diarrhea, metabolic acidosis and electrolyte disturbance were intractable, and the cholestatic indices, including transaminases, γ-glutamyl transpeptidase, bilirubin and total bile acids, remained at increased levels. One month later, the patient was discharged and then lost contact. On genetic analysis, the infant was proved to be a compound heterozygote of the c.310+2Tdup and c.1966C>T(p.R656C) variants of the gene MYO5B, with c.310+2Tdup being a novel splice-site mutation. MVID was thus definitely diagnosed.

Phenotypic and genetic analysis of a family affected with microvillus inclusion disease / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To explore the clinical features and mutations of MYO5B gene in a family affected with microvillus inclusion disease.</p><p><b>METHODS</b>Clinical data of an infant affected with microvillus inclusion disease was collected. Genomic DNA was extracted from peripheral blood samples from the patient and her parents. PCR amplification and Sanger sequencing were performed to analyze all the exons and their flanking sequences of the MYO5B gene.</p><p><b>RESULTS</b>The patient presented with complicated manifestations including respiratory distress syndrome, dehydration, acidosis, bowel dilatation, liver and kidney dysfunction, and severe and intractable diarrhea. A compound mutation of the MYO5B gene, i.e., IVS37-1G>C/c.2729_2731delC (p.R911Afs916X), was discovered in the patient. The former was a splice-site mutation inherited from the mother, while the latter was a frameshift mutation inherited from the father. Both were not reported previously.</p><p><b>CONCLUSION</b>Based on the clinical and molecular evidence, the patient was diagnosed with microvillus inclusion disease. Above finding has expanded the mutation spectrum of the MYO5B gene, which can provide valuable information for genetic counseling for the family.</p>

Myosin Va is developmentally regulated and expressed in the human cerebellum from birth to old age

Myosin Va functions as a processive, actin-based motor molecule highly enriched in the nervous system, which transports and/or tethers organelles, vesicles, and mRNA and protein translation machinery. Mutation of myosin Va leads to Griscelli disease that is associated with severe neurological deficits and a short life span. Despite playing a critical role in development, the expression of myosin Va in the central nervous system throughout the human life span has not been reported. To address this issue, the cerebellar expression of myosin Va from newborns to elderly humans was studied by immunohistochemistry using an affinity-purified anti-myosin Va antibody. Myosin Va was expressed at all ages from the 10th postnatal day to the 98th year of life, in molecular, Purkinje and granular cerebellar layers. Cerebellar myosin Va expression did not differ essentially in localization or intensity from childhood to old age, except during the postnatal developmental period. Structures resembling granules and climbing fibers in Purkinje cells were deeply stained. In dentate neurons, long processes were deeply stained by anti-myosin Va, as were punctate nuclear structures. During the first postnatal year, myosin Va was differentially expressed in the external granular layer (EGL). In the EGL, proliferating prospective granule cells were not stained by anti-myosin Va antibody. In contrast, premigratory granule cells in the EGL stained moderately. Granule cells exhibiting a migratory profile in the molecular layer were also moderately stained. In conclusion, neuronal myosin Va is developmentally regulated, and appears to be required for cerebellar function from early postnatal life to senescence.

Supplementation with 0.1% and 2% vitamin e in diabetic rats: analysis of myenteric neurons immunostained for myosin-V and nNOS in the jejunum / Suplementação com vitamina E 0,1% e 2% em ratos diabéticos: análise de neurônios mioentéricos imunomarcados para miosina-V e nNOS no jejuno

CONTEXT: Diabetes mellitus is a disease characterized by hyperglycemia that, when allowed to progress long-term untreated, develops vascular and neurological complications, which are responsible for the development of alterations in the enteric nervous system in diabetic patients. In the gastrointestinal tract, diabetes mellitus promotes motor and sensory changes, and in the reflex function of this system, causing gastroparesis, diarrhea, constipation, megacolon, slow gastrointestinal transit, gastric stasis and dilation with decreased or increased peristaltic contractions. Several studies have shown that oxidative stress is the main responsible for the vascular and neurological complications affecting the enteric nervous system of diabetics. OBJECTIVE: The effects of 0.1% and 2% vitamin E on myosin-V- and nNOS-immunoreactive neurons in the jejunum of diabetic rats were investigated. METHODS: Thirty rats were divided into the groups: normoglycemic, normoglycemic treated with 0.1% vitamin E, normoglycemic treated with 2% vitamin E, diabetic, diabetic treated with 0.1% vitamin E, and diabetic treated with 2% vitamin E. The neuronal density and areas of neuron cell bodies were determined. RESULTS: Diabetes (diabetic group) significantly reduced the number of myosin-V-immunoreactive neurons compared with the normoglycemic group. The diabetic treated with 0.1% vitamin E and diabetic treated with 2% vitamin E groups did not exhibit a greater density than the D group (P>0.05). Nitrergic density did not change with diabetes (P>0.05). The areas of myosin-V- and nNOS-immunoreactive neurons significantly increased in the normoglycemic treated with 2% vitamin E and diabetic groups compared with the normoglycemic group. CONCLUSION: Supplementation with 2% vitamin E had a neurotrophic effect only in the area of myosin-V-immunoreactive neurons compared with the diabetic group.

CONTEXTO: O diabetes mellitus (DM) é uma doença caracterizada pela hiperglicemia que a longo prazo, quando não tratada, desenvolve complicações vasculares e neurológicas, responsáveis pelo desenvolvimento das alterações no sistema nervoso entérico de pacientes diabéticos. Em nível gastrointestinal o DM provoca modificações motoras, sensoriais e na função reflexa desse sistema, podendo ocasionar gastroparesia, diarreia, constipação, megacólon, lentidão do trânsito gastrointestinal, estase e dilatação gástrica com diminuição ou aumento de contrações peristálticas. Diversos estudos têm evidenciado que o estresse oxidativo é o principal responsável pelas complicações vasculares e neurológicas que atingem o sistema nervoso entérico de diabéticos. OBJETIVO: O efeito da vitamina E 0,1% e 2 sobre a miosina-V e nNOS imunorreativas em neurônios do jejuno de ratos diabéticos foram investigados. MÉTODOS: Trinta ratos foram divididos em grupos: normoglicêmicos (NU), normoglicêmicos tratados com vitamina E 0,1% (NE1), normoglicêmicos tratados com vitamina E 2% (NE2), diabético (UD), diabéticos tratados com vitamina E 0,1% (DE1), e diabéticos tratados com vitamina E 2% (DE2). A densidade neuronal e áreas de corpos celulares de neurônios foram determinadas. RESULTADOS: Diabetes (UD grupo) reduziu significativamente o número de neurônios miosina-V imunorreativos quando comparado com o grupo UN. Os grupos DE1 e DE2 não exibem uma maior densidade do que o grupo D (P>0,05). Densidade nitrérgicos não se alterou com diabetes (P>0,05). As áreas dos neurônios miosina-V e nNOS imunorreativos aumentaram significativamente nos grupos NE2 e UD comparados com o grupo UN. CONCLUSÃO: A suplementação com vitamina E 2% teve um efeito neurotrófico apenas na área da miosina-V imunorreativos neurônios em comparação com o grupo UD.

Analysis of myosin-V immunoreactive myenteric neurons from arthritic rats / Análise dos neurônios mioentéricos miosina-V imunoreativos de ratos artríticos

CONTEXT: The inflammatory response itself and the consequent oxidative stress are able to promote neurodegeneration. So, it is possible that enteric nervous system is affected by inflammatory diseases threatening quality of life of patients. However, gastrointestinal symptoms of arthritis are usually attributed to anti-inflammatory drugs rather than neural damage. OBJECTIVE: To confirm if the general population of myenteric neurons from the ileum and jejunum of rats is affected by arthritis. METHODS: Twenty Holtzmann rats, 58-day-old male, were used and divided in four groups: control group (C30), arthritic group (Art30), older control group (C60) and older arthritic group (Art60). At 58 days old, the animals in groups Art30 and Art60 received an injection of the complete Freund's adjuvant in order to induce arthritis. The whole-mount preparations of ileum and jejunum were processed for myosin-V immunohistochemistry. Quantitative and morphometric analyses were performed. RESULTS: Groups Art30 and Art60 presented, respectively, a reduction of 2 percent and 6 percent in intestinal area when compared to their control groups. No significant differences were observed in general neuronal density among the four groups (P>0.05). Group C60 presented a reduction of 14.4 percent and 10.9 percent in mean neuronal cell body area when compared to group C30 (P<0.05), for the ileum and jejunum, respectively. The other groups had a similar mean neuronal cell body area (P>0.05). CONCLUSION: Arthritis does not promote quantitative or morphological damages in general myenteric population. However, studies in progress have revealed some significant alterations in myenteric neurons subpopulations (nitrergic and VIP-ergic neurons).

CONTEXTO: A resposta inflamatória e o estresse oxidativo acentuados em decorrência da artrite reumatóide são capazes de promover neurodegeneração. Nessas condições, é possível que o sistema nervoso entérico seja afetado, diminuindo a qualidade de vida dos pacientes. No entanto, os sintomas da artrite no trato gastrointestinal são geralmente associados ao uso de medicamentos anti-inflamatórios do que a um possível dano neural. OBJETIVO: Verificar se a população geral de neurônios mioentéricos do íleo e do jejuno de ratos artríticos é afetada pela artrite. MÉTODOS: Foram utilizados 20 ratos Holtzmann, inicialmente com 58 dias de idade, divididos em 4 grupos: controle com 88 dias (C30); artrítico com 88 dias (Art30); controle com 118 dias (C60) e artrítico com 118 dias (Art60). Os animais dos grupos Art30 e Art60 receberam aos 58 dias de idade o adjuvante completo de Freund para indução da artrite. Os preparados totais de íleo e jejuno foram submetidos a imunoistoquímica para a proteína miosina-V. Realizou-se análises quantitativas e morfométricas dos neurônios. RESULTADOS: Os animais Art30 e Art60 apresentaram, respectivamente, redução de 2 por cento e 6 por cento na área intestinal em relação aos seus controles. Não foram observadas diferenças na densidade neuronal geral entre os quatro grupos (P>0,05). O grupo C60 apresentou redução de 14,4 por cento e 10,9 por cento na área média do corpo celular neuronal em relação ao grupo C30 (P<0,05). Os demais grupos apresentaram área média de corpo celular semelhante (P>0,05). CONCLUSÃO: A artrite não provocou alterações quantitativas ou morfológicas na população mioentérica geral, entretanto, estudos em andamento revelam alterações significativas em subpopulações de neurônios mioentéricos (nitrérgicos e VIP-érgicos).

Differential patterns of myosin Va expression during the ontogenesis of the rat hippocampus

Myosin Va is an actin-based, processive molecular motor protein highly enriched in the nervous tissue of vertebrates. It has been associated with processes of cellular motility, which include organelle transport and neurite outgrowth. The in vivo expression of myosin Va protein in the developing nervous system of mammals has not yet been reported. We describe here the immunolocalization of myosin Va in the developing rat hippocampus. Coronal sections of the embryonic and postnatal rat hippocampus were probed with an affinity-purified, polyclonal anti-myosin Va antibody. Myosin Va was localized in the cytoplasm of granule cells in the dentate gyrus and of pyramidal cells in Ammon's horn formation. Myosin Va expression changed during development, being higher in differentiating rather than already differentiated granule and pyramidal cells. Some of these cells presented a typical migratory profile, while others resembled neurons that were in the process of differentiation. Myosin Va was also transiently expressed in fibers present in the fimbria. Myosin Va was not detected in germinative matrices of the hippocampus proper or of the dentate gyrus. In conclusion, myosin Va expression in both granule and pyramidal cells showed both position and time dependency during hippocampal development, indicating that this motor protein is under developmental regulation.

Comparative analysis of two immunohistochemical methods for antigen retrieval in the optical lobe of the honeybee Apis mellifera: Myosin-v assay

The present study compared two heating methods currently used for antigen retrieval (AR) immunostaining: the microwave oven and the steam cooker. Myosin-V, a molecular motor involved in vesicle transport, was used as a neuronal marker in honeybee Apis mellifera brains fixed in formalin. Overall, the steam cooker showed the most satisfactory AR results. At 100 ºC, tissue morphology was maintained and revealed epitope recovery, while evaporation of the AR solution was markedly reduced; this is important for stabilizing the sodium citrate molarity of the AR buffer and reducing background effects. Standardization of heat-mediated AR of formalin-fixed and paraffin-embedded tissue sections results in more reliable immunostaining of the honeybee brain.

Intramanchette transport during primate spermiogenesis: expression of dynein, myosin Va, motor recruiter myosin Va, VIIa-Rab27a/b interacting protein, and Rab27b in the manchette during human and monkey spermiogenesis / 亚洲男科学杂志(英文版)

<p><b>AIM</b>To show whether molecular motor dynein on a microtubule track, molecular motor myosin Va, motor recruiter myosin Va, VIIa-Rab27a/b interacting protein (MyRIP), and vesicle receptor Rab27b on an F-actin track were present during human and monkey spermiogenesis involving intramanchette transport (IMT).</p><p><b>METHODS</b>Spermiogenic cells were obtained from three men with obstructive azoospermia and normal adult cynomolgus monkey (Macaca fascicularis). Immunocytochemical detection and reverse transcription-polymerase chain reaction (RT-PCR) analysis of the proteins were carried out. Samples were analyzed by light microscope.</p><p><b>RESULTS</b>Using RT-PCR, we found that dynein, myosin Va, MyRIP and Rab27b were expressed in monkey testis. These proteins were localized to the manchette, as shown by immunofluorescence, particularly during human and monkey spermiogenesis.</p><p><b>CONCLUSION</b>We speculate that during primate spermiogenesis, those proteins that compose microtubule-based and actin-based vesicle transport systems are actually present in the manchette and might possibly be involved in intramanchette transport.</p>

Effect of age on the myosin-V immunoreactive myenteric neurons of rats ileum

Alterations in the gastrointestinal neuromuscular function related to age have been demonstrated in human and animal models. This study analyzes the effects of the aging process on the area of the neuronal cell bodies of the myenteric plexus in the antimesenteric and intermediate regions of the ileal circumference of Wistar, 12 month-old in comparison 3 month-old animals. The ileum was removed and whole-mount preparations immunostained by the antibody anti-myosin-V were processed. The morphometric analyses were performed using a computerized image analysis system, with a subsequent distribution of neurons by size in intervals of 100 micro2. The cellular body morphometry revealed a significant increase in the size of the myosin-V- immunoreactive myenteric neurons from 12 month-old animals when compared with 3 month –old animals. However, significant differences between the regions were not observed; these observations were not age-dependent. The implications of these results in relation to the increase of the body weight, size of the small intestine, general organization of the myenteric plexus, staining method of neurons and the possible factors involved in the regulation and/or control of the volume of neronal cells due to aging, are discussed.

Immunoelectronic localization of myosin-Va in rat cerebellum

Myosin V is an unconventional type of actin-binding myosin that participates in cytoplasmic organelle transport. Although this unconventional myosin has been extensively studied, its subcellular localization in the mammalian cerebellum has not been determined. In this work, we used an antibody against the tail domain of the myosin-Va heavy chain and a secondary antibody labeled with protein A-gold (15 nm) to study the subcellular distribution of this protein. Myosin-Va was found in the cytoplasm, where it was associated with a filament (probably actin). This protein was also detected in the plasma membrane of axons and dendrites in the molecular layer in rat cerebellum.