A molecular brake that modulates spliceosome pausing at detained introns contributes to neurodegeneration

Emerging evidence suggests that intron-detaining transcripts (IDTs) are a nucleus-detained and polyadenylated mRNA pool for cell to quickly and effectively respond to environmental stimuli and stress. However, the underlying mechanisms of detained intron (DI) splicing are still largely unknown. Here, we suggest that post-transcriptional DI splicing is paused at the Bact state, an active spliceosome but not catalytically primed, which depends on Smad Nuclear Interacting Protein 1 (SNIP1) and RNPS1 (a serine-rich RNA binding protein) interaction. RNPS1 and Bact components preferentially dock at DIs and the RNPS1 docking is sufficient to trigger spliceosome pausing. Haploinsufficiency of Snip1 attenuates neurodegeneration and globally rescues IDT accumulation caused by a previously reported mutant U2 snRNA, a basal spliceosomal component. Snip1 conditional knockout in the cerebellum decreases DI splicing efficiency and causes neurodegeneration. Therefore, we suggest that SNIP1 and RNPS1 form a molecular brake to promote spliceosome pausing, and that its misregulation contributes to neurodegeneration.

Group Ⅱ introns and the application in biotechnology: a review / 生物工程学报

Group Ⅱ introns are self-splicing ribozymes, which insert directly into target sites in DNA with high frequency through "retrohoming". They specifically and efficiently recognize and splice DNA target sites, endowing themselves with great potential in genetic engineering. This paper reviewed the gene targeting principle of group Ⅱ introns and the application in microbial genetic modification, and then analyzed the limitations of them in multi-functional gene editing and eukaryotes based on the "retrohoming" characteristics and the dependence on high Mg2+ concentration. Finally, we dissected the potential of group Ⅱ introns in the development of novel gene editing tools based on our previous research outcome and the structural characteristics of the introns, hoping to provide a reference for the application of group Ⅱ introns in biotechnology.

A pedigree analysis of a rare RhD 336-1G>A intron variant / 中华医学遗传学杂志

OBJECTIVE@#To explore the molecular mechanism of a case where RhD genotyping did not match serological results.@*METHODS@#The serological results of 8 members from two generations of this family were analyzed. And according to Mendelian law of inheritance, RhD genotyping, zygotic type determination and gene sequencing were performed for the family members.@*RESULTS@#The proband and one of her cousins have the same RhD alleles, both of them have a 336-1G>A intron variant RhD allele and a complete RhD deletion allele. The variant alleles are inherited from two of their parents with blood relationship, while the complete-deleted alleles come from the other. 336-1G>A means that the last base G of the second intron of the RhD gene is mutated to A, which leads to a negative RhD serology and a positive genotype in the proband.@*CONCLUSION@#There was a rare 336-1G> A intron variant gene (RhD * 01N.25) in this family, which was a recessive gene relative to the RhD gene and resulted in RhD phenotype negative.

Retrotransposon insertion polymorphism of the porcine esr gene and its association with production performances of Large White pigs / 生物工程学报

Estrogen receptor (esr) mediates the effects of estrogen on the expression of related genes, thereby regulating the growth and reproduction of mammals. To investigate the effect of retrotransposon insertion polymorphism (RIP) of the porcine esr gene on porcine growth performance, retrotransposon insertion polymorphism of the esr gene were predicted by comparative genomics and bioinformatics, and PCR was used to verify the insertion polymorphisms in different porcine breeds. Finally, the correlation analysis between the genotypes and performance of Large White pigs was conducted. The results showed that four retrotransposon polymorphic sites were identified in the esr1 and esr2 genes, which are esr1-SINE- RIP1 located in intron 2 of the esr1 gene, esr1-LINE-RIP2 and RIP3-esr1- SINE located in intron 5 of the gene, and esr2-LINE-RIP located in intron 1 of the esr2 gene, respectively. Among them, insertion of a 287 bp of SINE into intron 2 of the esr1 gene significantly affected (P<0.05) the live back fat thickness and 100 kg body weight back fat thickness of Large White pigs. Moreover, the live back fat thickness and back fat thickness at 100 kg body weight of homozygous with insertion (SINE+/+) was significantly greater than that of heterozygous with insertion (SINE+/-) and homozygous without insertion (SINE-/-). Therefore, esr1-SINE-RIP1 could be used as a molecular marker to assist the selection of deposition traits in Large White pigs.

Analysis of genetic variants in a case with Rotor syndrome / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis for a patient featuring Rotor syndrome.@*METHODS@#Clinical data of the patient was collected. Whole exome sequencing (WES) based on high-throughput sequencing technology was carried out. Long-interspersed element-1 (LINE-1) insertion in intron 5 of the SLCO1B3 gene was detected by using tri-primer single tube PCR.@*RESULTS@#WES revealed that the patient has carried homozygous c.1738C>T nonsense variants of the SLCO1B1 gene. He was also found to harbor a homozygous insertion of LINE-1 in intron 5 of the SLCO1B3 gene, which has caused skipping of exon 5 or exons 5 to 7 and introduced a stop codon in the SLCO1B3 transcript.@*CONCLUSION@#The homozygous c.1738C>T variant of the SLCO1B1 gene and homozygous insertion of LINE-1 in intron 5 of the SLCO1B3 gene probably underlay the Rotor syndrome in this patient.

Genoma humano: aspectos estructurales / Human genome: structural aspects / Genoma humano: aspectos estruturais

El genoma humano, como el de todos los mamíferos y aves, es un mosaico de isocoros, los que son regiones muy largas de ADN (>>100 kb) que son homogéneas en cuanto a su composición de bases. Los isocoros pueden ser divididos en un pequeño número de familias que cubren un amplio rango de niveles de GC (GC es la relación molar de guanina+citosina en el ADN). En el genoma humano encontramos cinco familias, que (yendo de valores bajos a altos de GC) son L1, L2, H1, H2 y H3. Este tipo de organización tiene importantes consecuencias funcionales, tales como la diferente concentración de genes, su regulación, niveles de transcripción, tasas de recombinación, tiempo de replicación, etc. Además, la existencia de los isocoros lleva a las llamadas "correlaciones composicionales", lo que significa que en la medida en que diferentes secuencias están localizadas en diferentes isocoros, todas sus regiones (exones y sus tres posiciones de los codones, intrones, etc.) cambian su contenido en GC, y como consecuencia, cambian tanto el uso de aminoácidos como de codones sinónimos en cada familia de isocoros. Finalmente, discutimos el origen de estas estructuras en un marco evolutivo.

The human genome, as the genome of all mammals and birds, are mosaic of isochores, which are very long streches (>> 100 kb) of DNA that are homogeneous in base composition. Isochores can be divided in a small number of families that cover a broad range of GC levels (GC is the molar ratio of guanine+cytosine in DNA). In the human genome, we find five families, which are (going from GC- poor to GC- rich) L1, L2, H1, H2 and H3. This organization has important consequences, as is the case of the concentration of genes, their regulation, transcription levels, rate of recombination, time of replication, etc. Furthermore, the existence of isochores has as a consequence the so called "compositional correlations", which means that as long as sequences are placed in different families of isochores, all of their regions (exons and their three codon positions, introns, etc.) change their GC content, and as a consequence, both codon and amino acids usage change in each isochore family. Finally, we discuss the origin of isochores within an evolutioary framework.

O genoma humano, como todos os mamíferos e aves, é um mosaico de isocóricas, que são muito longas regiões de ADN (>> 100 kb) que são homogéneos na sua composição de base. Isóquos podem ser divididos em um pequeno número de famílias que cobrem uma ampla gama de níveis de GC (GC é a razão molar de guanina + citosina no DNA). No genoma humano, encontramos cinco famílias, que (variando de valores baixos a altos de GC) são L1, L2, H1, H2 e H3. Este tipo de organização tem importantes conseqüências funcionais, como a diferente concentração de genes, sua regulação, níveis de transcrição, taxas de recombinação, tempo de replicação, etc. Além disso, a existência de isocóricas portada chamado "correlações de composição", o que significa que, na medida em que diferentes sequências estão localizados em diferentes isocóricas, todas as regiões (exs e três posições de codões, intrs, etc.) mudam seu conteúdo em GC e, como consequência, alteram tanto o uso de aminoácidos quanto de códons sinônimos em cada família de isócoros. Finalmente, discutimos a origem dessas estruturas em uma estrutura evolucionária.

Identification of the MYST3-CREBBP fusion gene in infants with acute myeloid leukemia and hemophagocytosis

ABSTRACT Background: Acute myeloid leukemia presenting the MYST3-CREBBP fusion gene is a rare subgroup associated with hemophagocytosis in early infancy and monocytic differentiation. The aim of this study was to define the relevant molecular cytogenetic characteristics of a unique series of early infancy acute myeloid leukemia cases (≤24 months old), based on the presence of hemophagocytosis by blast cells at diagnosis. Methods: A series of 266 infant cases of acute myeloid leukemia was the reference cohort for the present analysis. Acute myeloid leukemia cases with hemophagocytosis by blast cells were reviewed to investigate the presence of the MYST3-CREBBP fusion gene by fluorescence in situ hybridization (FISH) and reverse transcription polymerase chain reaction. Results: Eleven cases with hemophagocytosis were identified with hemophagocytic lymphohistiocytosis being ruled out. Six cases were classified as myelomonocytic leukemia, three as AML-M7 and two as AML-M2. In five cases, the presence of the MYST3-CREBBP fusion gene identified by molecular cytogenetics was confirmed by fluorescence in situ hybridization. All patients received treatment according to the Berlin-Frankfürt-Münster acute myeloid leukemia protocols and only one out of the five patients with the MYST3-CREBBP fusion gene is still alive. Conclusions: Our findings demonstrate that the presence of hemophagocytosis in acute myeloid leukemia was not exclusively associated to the MYST3-CREBBP fusion gene. Improvements in molecular cytogenetics may help to elucidate more complex chromosomal rearrangements in infants with acute myeloid leukemia and hemophagocytosis.

Mutation c.1190-1delG/N in intron 8 and c.1708G>C/N in exon 12 not reported in the IDUA gene developed a clinical phenotype of Scheie syndrome / Mutación c.1190-1delG/N en el intrón 8 y c.1708G>C/N en el exón 12 no reportada en el gen de IDUA desarrolló un fenotipo clínico de síndrome de Scheie

Mucopolysaccharidoses are a group of lysosomal storage disorders caused by deficiency of enzymes catalyzing the degradation of glycosaminoglycans. Mucopoly-saccharidosis I can present a wide range of phenotypic characteristics with three major recognized clinical entities: Hurler and Scheie syndromes represent phenotypes at the severe and mild ends of the clinical spectrum, respectively, and the Hurler-Scheie syndrome is intermediate in phenotypic expression. These are caused by the deficiency or absence of α-L-iduronidase, essential to the metabolism of both dermatan and heparan sulfate, and it is encoded by the IDUA gene. We report the case of a 34-year-old male patient with enzymatic deficiency of α-L-iduronidase, accumulation of its substrate and a previously unreported mutation in the IDUA gene that developed a phenotype of Scheie syndrome.

Las mucopolisacaridosis son un grupo de trastornos de almacenamiento lisosomal causada por la deficiencia de enzimas que catalizan la degradación de glicosaminoglicanos. La mucopolisacaridosis tipo I puede presentar un amplio rango de características fenotípicas englobadas en tres entidades clínicas reconocidas: los síndromes de Hurler y Scheie representan los fenotipos graves y leves del espectro clínico, respectivamente y el síndrome de Hurler-Scheie intermedio en la expresión fenotípica. Estos son causados por la deficiencia o ausencia de la α-L-iduronidasa esencial para el metabolismo del dermatán y el heparán sulfato y es codificada por el gen IDUA. Se presenta el caso de paciente masculino de 34 años de edad con deficiencia enzimática de α-L-iduronidasa, acumulación de su sustrato y una mutación en el gen IDUA, no reportada previamente, que desarrolló un fenotipo del síndrome de Scheie.

Genetic variations of b-MYH7 in Venezuelan patients with hypertrophic cardiomyopathy / Variaciones genéticas en el gen MYH7 en pacientes venezolanos con miocardiopatía hipertrófica

Hypertrophic cardiomyopathy (HCM) is a cardiac disease, characterized by marked hypertrophy and genetic variability. HCM has been associated with sarcomere protein mutations, being cardiac b-myosin (coded by the MYH7 gene) and myosin binding protein C (coded by the MYBPC3 gene) the most frequently affected proteins. As in Venezuela only the clinical analysis are performed in HCM patients, we decided to search for genetic variations in the MYH7 gene. Coding regions, including the junction exon-intron of the MYH7 gene, were studied in 58 HCM patients, whose samples were collected at the ASCARDIO Hospital (Barquisimeto, Lara state, Venezuela) and 106 control subjects from the ASCARDIO Hospital and the IVIC (Barquisimeto Lara state and Miranda, Venezuela, respectively). The blood samples were analyzed by genomic DNA isolation, followed by polymerase chain reaction and sequence analysis. The screening of the MYH7 gene revealed eight already reported polymorphic variants, as well as two intronic variations in these HCM patients. Neither any missense mutations nor other pathological mutations in the MYH7 gene were found in the HCM patients.

La miocardiopatía hipertrófica (MH) es una enfermedad cardiaca primaria, caracterizada por una marcada hipertrofia y variabilidad genética. MH ha sido asociada con mutaciones en las proteínas del sarcómero, siendo la beta miosina cardiaca, codificada por el gen MYH7 y la proteína de unión a miosina C, codificada por el gen MYBPC3, las principalmente afectadas. En Venezuela únicamente se realiza el diagnóstico clínico de MH, por lo cual el objetivo principal de este trabajo fue realizar el análisis genético en los pacientes, iniciando con el gen MYH7. Para ello, se estudió la región codificante, incluyendo la región de unión exón-intron del gen MYH7 en 58 pacientes provenientes de ASCARDIO (Barquisimeto, estado Lara) y 106 controles provenientes de ASCARDIO e IVIC (estados Lara y Miranda, Venezuela). Se colectaron las muestras de sangre para el aislamiento del ADN genómico, se realizó la técnica de PCR, seguido del análisis de secuencias para la detección de mutaciones en pacientes y controles. Se encontraron 8 polimorfismos previamente reportados, y 2 variaciones intrónicas. No se encontraron mutaciones que involucraran un cambio de aminoácido en ninguno de los exones del gen MYH7 de la beta miosina cardiaca.

Prevalência de dislipidemia em adultos de meia-idade com polimorfismo do gene NOS3 e baixa aptidão cardiorrespiratória / Prevalence of dyslipidemia in middle-aged adults with NOS3 gene polymorphism and low cardiorespiratory fitness

OBJETIVO: Analisar a influência da associação dos polimorfismos do gene da sintase do óxido nítrico endotelial (NOS3) para as posições -786T>C, Glu298Asp e íntron 4b/a e a aptidão cardiorrespiratória sobre as concentrações de nitrito/nitrato, pressão arterial, perfil lipídico e prevalência de doenças cardiometabólicas em adultos. SUJEITOS E MÉTODOS: Noventa e duas pessoas foram divididas de acordo com o genótipo: não polimórficas (NP) e polimórficas (P). Posteriormente, foram subdivididas pela aptidão cardiorrespiratória associada ao genótipo: alta (ANP e AP) ou baixa (BNP e BP). RESULTADOS: Os indivíduos que apresentavam polimorfismo para as posições Glu298Asp+Íntron 4b/a e Glu298Asp+-786T>C e baixa aptidão cardiorrespiratória apresentaram maiores valores de colesterol total e maior prevalência de dislipidemia. CONCLUSÃO: Nossos dados demonstram que os polimorfismos do gene da NOS3 para essas duas associações influenciam os níveis de colesterol plasmático, e essa associação foi mais claramente observada quando os indivíduos apresentavam menor nível de aptidão cardiorrespiratória.

OBJECTIVE: To evaluate the influence of the interaction between endothelial nitric oxide synthase gene (NOS3) polymorphisms at positions -786T>C, Glu298Asp and intron 4b/a, and cardiorespiratory fitness on plasma nitrite/nitrate levels, blood pressure, lipid profile, and prevalence of cardiometabolic disorders. SUBJECTS AND METHODS: Ninety-two volunteers were genotyped for NOS3 polymorphisms at positions (-786T>C and Glu298Asp) and (intron 4b/a) and divided according to the genotype: non-polymorphic (NP) and polymorphic (P). After that, they were subdivided according to the cardiorespiratory fitness associated with genotype: high (HNP and HP) and low (LNP and LP). RESULTS: The subjects with polymorphism for the interactions at positions Glu298Asp + intron 4b/a, and Glu298Asp+-786T>C showed the highest values in total cholesterol, as well as dyslipidemia. CONCLUSION: Our findings show that NOS3 gene polymorphisms at positions -786T>C, Glu298Asp, and intron 4b/a exert negative effects on the lipid profile compared with those who do not carry polymorphisms.

EGFR Polymorphism as a Predictor of Clinical Outcome in Advanced Lung Cancer Patients Treated with EGFR-TKI

PURPOSE: Mutations in the epidermal growth factor receptor (EGFR) have been confirmed as predictors of the efficacy of treatment with EGFR-tyrosine kinase inhibitors (TKIs). We investigated whether polymorphisms of the EGFR gene were associated with clinical outcomes in non-small cell lung cancer (NSCLC) patients treated with EGFR-TKI. MATERIALS AND METHODS: A polymorphic dinucleotide repeat in intron 1 [CA simple sequence repeat in intron 1(CA-SSR1)] in intron 1 and single nucleotide polymorphisms (SNP-216) in the promoter region of the EGFR gene were evaluated in 71 NSCLC patients by restriction fragment length polymorphism and DNA sequencing. The relationship between genetic polymorphisms and clinical outcomes of treatment with EGFR-TKIs was evaluated. RESULTS: SNP-216G/T polymorphisms were associated with the efficacy of EGFR-TKI. The response rate for the SNP-216G/T tended to be higher than that for G/G (62.5% vs. 27.4%, p=0.057). The SNP-216G/T genotype was also associated with longer progression-free survival compared with the GG genotype (16.7 months vs. 5.1 months, p=0.005). However, the length of CA-SSR1 was not associated with the efficacy of EGFR-TKI. CONCLUSION: SNP-216G/T polymorphism was a potential predictor of clinical outcomes in NSCLC patients treated with EGFR-TKI.

Polymorphism of the aryl-hydrocarbon receptor gene in intron 10 of human cancers

Polychlorinated dibenzo-p-dioxins (PCDDs) and related halogenated aromatic hydrocarbons (e.g., PCDFs), often called "dioxins", are ubiquitously present environmental contaminants. Some of them, notably 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), are among the most toxic synthetic compounds known. The biological effects of dioxins are mediated via the aryl hydrocarbon receptor (AhR). Mutations in the AhR transactivation domain are linked to sensitivity to the acute lethality of TCDD. We present here a study of AhR gene polymorphism in normal and cancer human tissues affecting pre-mRNA splicing in the AhR gene-coding transactivation domain region (exon 10, intron 10, exon 11 region), previously shown to be associated with AhR dysfunction. We tested 126 pairs of normal and cancer tissue samples from liver, lung, stomach, kidney, mucous, breast, and pancreas of 49 males and 77 females (45-70 years of age). We used in vitro splicing assay, RT-PCR and sequencing methods. Our results showed that in an in vitro system it is possible to reconstitute cellular pre-mRNA splicing events. Tested cancer tissues did not contain mutations in the AhR transactivation domain region when the DNA sequences were compared with those from normal tissues. There were also no differences in AhR mRNA splice variants between normal and malignant breast tissues and no polymorphisms in the studied regions or cDNA.

TP53 PIN3 polymorphism associated with breast cancer risk in Iranian women.

Background: Breast cancer is one of the most common cancers in Iranian women. The p53 gene plays a principal role in genomic stability, and its function varies according to polymorphisms. The aim of our study was to determine the relationship between the intron3 16bp duplication polymorphism of the p53 gene and breast cancer in Iranian women. Materials and Methods: We performed a case-control study among 145 patients with invasive ductal carcinoma of the breast and 145 controls in Isfahan, Iran. The distribution of the intron3 16bp duplication polymorphism was determined by polymerase chain reaction (PCR). The relationship between clinicopathological data and the PIN3 polymorphism was examined using chi-squared analysis. Results: A significant difference was observed in the polymorphism variants in breast cancer specimens compared with controls (P < .001). Among the cancer patients, 59.9% were below the age of 50 years; and 67.5% of the patients in this group had the intron3 16bp duplication polymorphism. Conclusions: PIN3 Ins 16bp duplication polymorphism is a genetically predisposing factor for breast cancer development in Iranian women and may be causal in patients under the age of 50 years.

Análisis de las inversiones del intrón 1 y 22 y secuenciación del gen F8 para el diagnóstico genético-molecular de hemofilia A en Chile / Introns 1 and 22 inversions and F8 gene sequencing for molecular diagnosis of hemophilia A in Chile

Background: Hemophilia A is an inherited disorder caused by alterations in factor VIII gene (F8) located on the X-chromosome, the intron 22 inversion being the most common mutation. The rest are predominantly point mutations distributed along this large gene of 26 exons. Aim: To implement a molecular diagnostic test to detect mutations in the F8 gene in Chilean patients with Hemophilia A. Material and Methods: To validate the testing methods, we analyzed samples with intron 22 and intron 1 inversion, and with point mutations previously studied, as well as one subject without Hemophilia. We also studied unrelated Chilean patients with Hemophilia A and their female relatives for carrier testing. Intron 22 and intron 1 inversions were studied by long distance polymerase chain reaction (PCR) and point mutations by sequencing the coding and promoter regions of the F8 gene. Results: The results obtained in all samples used for validation were concordant with those obtained previously. In the Chilean patients, the intron 22 inversion and point mutations previously described were observed. In 6 out of 9 patients with mild Hemophilia A we found the same mutation (Arg2159Cys) in exon 23, which has been described as prevalent in mild Hemophilia A. Conclusions: The analysis of intron 22 and intron 1 inversions, as well as of point mutations in the F8 gene will help us to confirm the diagnosis in patients with severe, moderate and mild Hemophilia A, and also it will allow us to perform carrier testing and to provide better genetic counseling.

Long-term follow-up of an 8-year-old boy with insulinoma as the first manifestation of a familial form of multiple endocrine neoplasia type 1 / Seguimento de longo prazo em um menino de 8 anos de idade com insulinoma como primeira manifestação de neoplasia endócrina múltipla tipo 1

Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant hereditary cancer syndrome characterized mostly by parathyroid, enteropancreatic, and anterior pituitary tumors. We present a case of an 8-year-old boy referred because of hypoglycemic attacks. His diagnosis was pancreatic insulinoma. Paternal grandmother died due to repeated gastroduodenal ulcerations and a paternal aunt presented similar manifestations. At a first evaluation, the father presented only gastric ulceration but subsequently developed hyperparathyroidism and lung carcinoid tumor. During almost 15 years of follow-up, three brothers and the index case presented hyperparathyroidism and hyperprolactinemia. Molecular study showed a G to A substitution in intron 4, at nine nucleotides upstream of the splicing acceptor site, causing a splicing mutation. All affected members of the family have the same mutation. Paternal grandmother and aunt were not studied and the mother does not carry any mutation. MEN1 is a rare condition that requires permanent medical assistance. Early clinical and genetic identification of affected individuals is essential for their own surveillance and also for genetic counseling.

A neoplasia endócrina múltipla tipo 1 (NEM1) é uma doença hereditária autossômica dominante, caracterizada principalmente por tumores de paratireoide, enteropancreáticos e adeno-hipofisários. Apresentamos o caso de um menino com 8 anos encaminhado por crises de hipoglicemia. Seu diagnóstico foi insulinoma pancreático. Sua avó paterna faleceu por úlceras gastroduodenais de repetição e a tia paterna tinha as mesmas manifestações. Na primeira avaliação, o pai apresentou apenas úlcera gástrica, porém com a evolução desenvolveu hiperparatireoidismo e tumor carcinoide pulmonar. Durante cerca de 15 anos de seguimento, os três irmãos e o caso índice desenvolveram hiperparatireoidismo e hiperprolactinemia. O estudo molecular mostrou a substituição G por A no intron 4, a nove nucleotídeos do sítio aceptor de splicing, criando um novo sítio de splicing. Todos os membros da família afetados e estudados tinham a mesma mutação. A NEM1 é uma condição rara que requer assistência médica permanente. As identificações clínicas e genéticas precoces são essenciais para o tratamento e aconselhamento genético.

Clinical features and molecular analysis of arginine-vasopressin neurophysin II gene in long-term follow-up patients with idiopathic central diabetes insipidus / Apresentação clínica e análise molecular do gene da arginina-vasopressina neurofisina II de pacientes com diabetes insípido central idiopático com longo seguimento

INTRODUCTION: Central diabetes insipidus (DI) characterized by polyuria, polydipsia and inability to concentrate urine, has different etiologies including genetic, autoimmune, post-traumatic, among other causes. Autosomal dominant central DI presents the clinical feature of a progressive decline of arginine-vasopressin (AVP) secretion. OBJECTIVE: In this study, we characterized the clinical features and sequenced the AVP-NPII gene of seven long-term follow-up patients with idiopathic central DI in an attempt to determine whether a genetic cause would be involved. METHODS: The diagnosis of central DI was established by fluid deprivation test and hyper-tonic saline infusion. For molecular analysis, genomic DNA was extracted and the AVP-NPII gene was amplified by polymerase chain reaction and sequenced. RESULTS: Sequencing analysis revealed a homozygous guanine insertion in the intron 2 (IVS2 +28 InsG) of the AVP-NPII gene in four patients, which represents an alternative gene assembly. No mutation in the code region of the AVP-NPII gene was found. CONCLUSIONS: The homozygous guanine insertion in intron 2 (IVS2 +28 InsG) is unlikely to contribute to the AVP-NPII gene modulation in DI. In addition, the etiology of idiopathic central DI in children may not be apparent even after long-term follow-up, and requires continuous etiological surveillance.

INTRODUÇÃO: O diabetes insípido (DI) central, caracterizado por poliúria, polidipsia e inabilidade em concentrar a urina, apresenta diferentes etiologias, incluindo causas genética, autoimune, pós-traumática, entre outras. O DI central autossômico dominante apresenta a característica clínica de falência progressiva da secreção da arginina-vasopressina (AVP). OBJETIVO: No presente estudo, caracterizou-se a apresentação clínica e sequenciou-se o gene AVP-NPII de sete pacientes com DI central idiopático seguidos de longa data na tentativa de determinar se uma causa genética estava envolvida na etiologia. MÉTODOS: O diagnóstico do DI central foi estabelecido por meio do teste de jejum hídrico e infusão de salina hipertônica. Para a realização da análise molecular, o DNA genômico foi extraído e o gene AVP-NPII foi amplificado pela reação em cadeia da polimerase e, posteriormente, sequenciado. RESULTADOS: A análise do sequenciamento do gene AVP-NPII revelou uma inserção em homozigose de uma guanina no íntron 2 (IVS2 +28 InsG) em quatro pacientes, correspondendo a um arranjo alternativo do gene. Nenhuma mutação da região codificadora do gene AVP-NPII foi encontrada. CONCLUSÕES: A inserção em homozigose de uma guanina no íntron 2 (IVS2 +28 InsG) provavelmente não contribui na modulação do gene AVP-NPII no DI. Adicionalmente, a etiologia do DI central idiopático em crianças pode não se tornar evidente mesmo após um longo período de seguimento, necessitando de contínua vigilância da etiologia.

Caracterização Molecular da Deficiência de Fator VIII em Pacientes Hemofílicos de Minas Gerais

Hemofilia A, uma doença hemorrágica ligada ao cromossomo X, é causada pela deficiência ou disfunção do fator VIII da coagulação (FVIII). A doença resulta de alterações genéticas no gene do fator VIII (F8). Clinicamente, a hemofilia A é caracterizada por hemorragias em diferentes partes do corpo e o tratamento recomendado é a reposição do fator FVIII deficiente. O desenvolvimento de anticorpos inbidores para o FVIII é a principal complicação do tratamento. Estes anticorpos neutralizam a atividade do FVIII e, portanto invalidam a terapia. Os riscos para o desenvolvimento destes anticorpos são multifatoriais e envolvem fatores genéticos e ambientais. O objetivo deste estudo foi determinar as bases moleculares da hemofilia A grave e sua correlação com o desenvolvimento de inibidores em associação com o perfil sorológico par doenças virais. As análises moleculares consistiram da determinação da inversão do íntron 1 e do íntron 22 (inv1 e inv 22), além da determinação de outras alterações genéticas presentes na região codificadora de F8.

Inicialmente, 150 pacientes, com hemofilia A grave (níveis de atividade de fator VIII inferior a 1%) registrados no Hemominas, foram selecionados para este estudo. Nós caracterizamos 50 pacientes, dos quais, 8 tinham inv1, 29 tinham inv22 e os pacientes restantes apresentaram mutações sem sentido, mutações de sentido trocado e mutações de mudança de fase de leitura. Posteriormente, um grupo de 40 pacientes, registrados no hemocentro de Campinas e com diagnóstico molecular (32 tinham inv 22 e os pacientes restantes apresentaram mutações sem sentido, mutações de sentido trocado e mutações de mudança de fase de leitura) foram arrolados em nosso estudo. Informações a cerca de inibidores e perfil sorológico de 90 pacientes foram coletadas a partir de prontuários médicos. Dados moleculares e clínicos foram comparados por meio de análises estatísticas. As análises estatísticas não revelaram uma correlação positiva entre as diferentes categorias de mutação e a presença/ausência de inibidores sob influência do perfil sorológico.

Caracterização molecular da deficiência de fator VIII em pacientes hemofílicos de Minas Gerais / Molecular characterization of factor VIII deficiency in patients with Hemophilia Minas Gerais

Hemofilia A, uma doença hemorrágica ligada ao cromossomo X, é causada pela deficiência ou disfunção do fator VIII da coagulação (FVIII). A doença resulta de alterações genéticas no gene do fator VIII (F8). Clinicamente, a hemofilia A é caracterizada por hemorragias em diferentes partes do corpo e o tratamento recomendado é a reposição do fator FVIII deficiente. O desenvolvimento de anticorpos inbidores para o FVIII é a principal complicação do tratamento. Estes anticorpos neutralizam a atividade do FVIII e, portanto invalidam a terapia. Os riscos para o desenvolvimento destes anticorpos são multifatoriais e envolvem fatores genéticos e ambientais. O objetivo deste estudo foi determinar as bases moleculares da hemofilia A grave e sua correlação com o desenvolvimento de inibidores em associação com o perfil sorológico par doenças virais. As análises moleculares consistiram da determinação da inversão do íntron 1 e do íntron 22 (inv1 e inv 22), além da determinação de outras alterações genéticas presentes na região codificadora de F8.

Inicialmente, 150 pacientes, com hemofilia A grave (níveis de atividade de fator VIII inferior a 1%) registrados no Hemominas, foram selecionados para este estudo. Nós caracterizamos 50 pacientes, dos quais, 8 tinham inv1, 29 tinham inv22 e os pacientes restantes apresentaram mutações sem sentido, mutações de sentido trocado e mutações de mudança de fase de leitura. Posteriormente, um grupo de 40 pacientes, registrados no hemocentro de Campinas e com diagnóstico molecular (32 tinham inv 22 e os pacientes restantes apresentaram mutações sem sentido, mutações de sentido trocado e mutações de mudança de fase de leitura) foram arrolados em nosso estudo. Informações a cerca de inibidores e perfil sorológico de 90 pacientes foram coletadas a partir de prontuários médicos. Dados moleculares e clínicos foram comparados por meio de análises estatísticas. As análises estatísticas não revelaram uma correlação positiva entre as diferentes categorias de mutação e a presença/ausência de inibidores sob influência do perfil sorológico.

Short nucleotide sequences signal spliceosomal binding in nucleic acids.

We have explored the region around the splice sites of the human intron and exons from the exon-intron database (EID) and located a number of short 6-nucleotide and 7-nucleotide sequences that are relatively common in the regions. These short sequences, we expect play an important role in the selection of the appropriate splicing process. We propose that the external signals via short recognition sequences play the deterministic role in the actual splicing process. We have obtained 50 such sequences each from the exon and intron from the beginning and from the ending and noted a number of common features.