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1.
Hum Mol Genet ; 25(3): 571-83, 2016 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-26647307

RESUMO

Megacystis Microcolon Intestinal Hypoperistalsis Syndrome (MMIHS) is a rare congenital disorder, in which heterozygous missense variants in the Enteric Smooth Muscle actin γ-2 (ACTG2) gene have been recently identified. To investigate the mechanism by which ACTG2 variants lead to MMIHS, we screened a cohort of eleven MMIHS patients, eight sporadic and three familial cases, and performed immunohistochemistry, molecular modeling and molecular dynamics (MD) simulations, and in vitro assays. In all sporadic cases, a heterozygous missense variant in ACTG2 was identified. ACTG2 expression was detected in all intestinal layers where smooth muscle cells are present in different stages of human development. No histopathological abnormalities were found in the patients. Using molecular modeling and MD simulations, we predicted that ACTG2 variants lead to significant changes to the protein function. This was confirmed by in vitro studies, which showed that the identified variants not only impair ACTG2 polymerization, but also contribute to reduced cell contractility. Taken together, our results confirm the involvement of ACTG2 in sporadic MMIHS, and bring new insights to MMIHS pathogenesis.


Assuntos
Anormalidades Múltiplas/genética , Actinas/genética , Colo/anormalidades , Mucosa Intestinal/metabolismo , Pseudo-Obstrução Intestinal/genética , Contração Muscular/genética , Músculo Liso/metabolismo , Mutação de Sentido Incorreto , Bexiga Urinária/anormalidades , Anormalidades Múltiplas/metabolismo , Anormalidades Múltiplas/patologia , Actinas/química , Actinas/metabolismo , Colo/metabolismo , Colo/patologia , Evolução Fatal , Feminino , Expressão Gênica , Heterozigoto , Humanos , Recém-Nascido , Pseudo-Obstrução Intestinal/metabolismo , Pseudo-Obstrução Intestinal/patologia , Intestinos/patologia , Masculino , Simulação de Dinâmica Molecular , Músculo Liso/patologia , Linhagem , Multimerização Proteica , Bexiga Urinária/metabolismo , Bexiga Urinária/patologia , Adulto Jovem
2.
Biochim Biophys Acta ; 1852(11): 2352-61, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26282049

RESUMO

Congenital Short Bowel Syndrome (CSBS) is a rare gastrointestinal disorder in which the mean length of the small intestine is substantially reduced when compared to its normal counterpart. Families with several affected members have been described and CSBS has been suggested to have a genetic basis. Recently, our group found mutations in CLMP as the cause of the recessive form of CSBS, and mutations in FLNA as the cause of the X-linked form of the disease. These findings have improved the quality of genetic counselling for CSBS patients and made prenatal diagnostics possible. Moreover, they provided a reliable starting point to further investigate the pathogenesis of CSBS, and to better understand the development of the small intestine. In this review, we present our current knowledge on CSBS and discuss hypotheses on how the recent genetic findings can help understand the cause of CSBS.

3.
Gastroenterology ; 142(3): 453-462.e3, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22155368

RESUMO

BACKGROUND & AIMS: Short-bowel syndrome usually results from surgical resection of the small intestine for diseases such as intestinal atresias, volvulus, and necrotizing enterocolitis. Patients with congenital short-bowel syndrome (CSBS) are born with a substantial shortening of the small intestine, to a mean length of 50 cm, compared with a normal length at birth of 190-280 cm. They also are born with intestinal malrotation. Because CSBS occurs in many consanguineous families, it is considered to be an autosomal-recessive disorder. We aimed to identify and characterize the genetic factor causing CSBS. METHODS: We performed homozygosity mapping using 610,000 K single-nucleotide polymorphism arrays to analyze the genomes of 5 patients with CSBS. After identifying a gene causing the disease, we determined its expression pattern in human embryos. We also overexpressed forms of the gene product that were and were not associated with CSBS in Chinese Hamster Ovary and T84 cells and generated a zebrafish model of the disease. RESULTS: We identified loss-of-function mutations in Coxsackie- and adenovirus receptor-like membrane protein (CLMP) in CSBS patients. CLMP is a tight-junction-associated protein that is expressed in the intestine of human embryos throughout development. Mutations in CLMP prevented its normal localization to the cell membrane. Knock-down experiments in zebrafish resulted in general developmental defects, including shortening of the intestine and the absence of goblet cells. Because goblet cells are characteristic for the midintestine in zebrafish, which resembles the small intestine in human beings, the zebrafish model mimics CSBS. CONCLUSIONS: Loss-of-function mutations in CLMP cause CSBS in human beings, likely by interfering with tight-junction formation, which disrupts intestinal development. Furthermore, we developed a zebrafish model of CSBS.


Assuntos
Intestino Delgado/anormalidades , Mutação de Sentido Incorreto , Receptores Virais/genética , Síndrome do Intestino Curto/genética , Adolescente , Adulto , Animais , Células CHO , Criança , Pré-Escolar , Proteína de Membrana Semelhante a Receptor de Coxsackie e Adenovirus , Cricetinae , Cricetulus , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Predisposição Genética para Doença , Heterozigoto , Homozigoto , Humanos , Lactente , Recém-Nascido , Intestino Delgado/metabolismo , Masculino , Morfogênese , Fenótipo , Polimorfismo de Nucleotídeo Único , Receptores Virais/metabolismo , Síndrome do Intestino Curto/embriologia , Síndrome do Intestino Curto/metabolismo , Síndrome do Intestino Curto/patologia , Transfecção , Adulto Jovem , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
4.
Genet Med ; 15(4): 310-3, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23037936

RESUMO

PURPOSE: Autosomal recessive congenital short bowel syndrome is caused by mutations in CLMP. No mutations were found in the affected males of a family with presumed X-linked congenital short bowel syndrome or in an isolated male patient. Our aim was to identify the disease-causing mutation in these patients. METHODS: We performed mutation analysis of the second exon of FLNA in the two surviving affected males of the presumed X-linked family and in the isolated patient. RESULTS: We identified a novel 2-base-pair deletion in the second exon of FLNA in all these male patients. The deletion is located between two nearby methionines at the N-terminus of filamin A. Previous studies showed that translation of FLNA occurs from both methionines, resulting in two isoforms of the protein. We hypothesized that the longer isoform is no longer translated due to the mutation and that this mutation is therefore not lethal for males in utero. CONCLUSION: Our findings emphasize that congenital short bowel syndrome can be the presenting symptom in male patients with mutations in FLNA.


Assuntos
Filaminas/genética , Mutação , Síndrome do Intestino Curto/diagnóstico , Síndrome do Intestino Curto/genética , Adolescente , Adulto , Sequência de Bases , Éxons , Doenças Genéticas Ligadas ao Cromossomo X/diagnóstico , Doenças Genéticas Ligadas ao Cromossomo X/genética , Humanos , Masculino , Linhagem , Fenótipo , Deleção de Sequência , Adulto Jovem
5.
Eur J Hum Genet ; 24(11): 1627-1629, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27352967

RESUMO

Congenital short bowel syndrome (CSBS) is an intestinal pediatric disorder, where patients are born with a dramatic shortened small intestine. Pathogenic variants in CLMP were recently identified to cause an autosomal recessive form of the disease. However, due to the rare nature of CSBS, only a small number of patients have been reported to date with variants in this gene. In this report, we describe novel inherited variants in CLMP in three CSBS patients derived from two unrelated families, confirming CLMP as the major gene involved in the development of the recessive form of CSBS.


Assuntos
Proteína de Membrana Semelhante a Receptor de Coxsackie e Adenovirus/genética , Mutação , Síndrome do Intestino Curto/genética , Animais , Células CHO , Cricetinae , Cricetulus , Feminino , Genes Recessivos , Humanos , Recém-Nascido , Linhagem , Síndrome do Intestino Curto/diagnóstico
6.
PLoS One ; 8(2): e54649, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23460781

RESUMO

Loss-of-function mutations in CLMP have been found in patients with Congenital Short Bowel Syndrome (CSBS), suggesting that its encoded protein plays a major role in intestinal development. CLMP is a membrane protein that co-localizes with tight junction proteins, but its function is largely unknown. We expressed wild-type (WT)-CLMP and a mutant-CLMP (associated with CSBS) in human intestinal epithelial T84 cells that, as we show here, do not produce endogenous CLMP. We investigated the effects of WT-CLMP and mutant-CLMP proteins on key cellular processes that are important for intestinal epithelial development, including migration, proliferation, viability and transepithelial resistance. Our data showed that expression of WT-CLMP or mutant-CLMP does not affect any of these processes. Moreover, our aggregation assays in CHO cells show that CLMP does not act as a strong adhesion molecule. Thus, our data suggest that, in the in vitro model systems we used, the key processes involved in intestinal epithelial development appear to be unaffected by WT-CLMP or mutant-CLMP. Further research is needed to determine the role of CLMP in the development of the intestine.


Assuntos
Proteína de Membrana Semelhante a Receptor de Coxsackie e Adenovirus/metabolismo , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Epitélio/crescimento & desenvolvimento , Intestinos/crescimento & desenvolvimento , Animais , Células CHO , Moléculas de Adesão Celular/metabolismo , Agregação Celular , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Cricetinae , Cricetulus , Impedância Elétrica , Epitélio/metabolismo , Humanos , Proteínas Mutantes/metabolismo , Transdução Genética
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