The use of buccal cells for rapid diagnosis of myotonic dystrophy type 1

2010 ◽  
Vol 1 (3) ◽  
Author(s):  
Ian Holt ◽  
Ros Quinlivan ◽  
Jillian Couto ◽  
Darren Monckton ◽  
Glenn Morris
Keyword(s):  
Myotonic Dystrophy ◽  
In Situ Hybridisation ◽  
Rapid Test ◽  
Buccal Cells ◽  
Autosomal Dominant Disorder ◽  
Nuclear Foci ◽  
Cug Repeats ◽  
Positive Results

AbstractType 1 myotonic dystrophy (DM1) is an autosomal dominant disorder caused by a CTG repeat expansion. RNA containing expanded CUG repeats does not leave the nucleus, but accumulates in discrete nuclear foci which sequester the human muscleblind-like (MBNL) proteins. We have examined buccal cells from 15 adult DM1 patients and 7 control non-DM patients to determine whether nuclear foci can be detected by either immunostaining for MBNL1 protein or fluorescent in situ hybridisation (FISH) for the CUG repeat RNA. Both methods detected nuclear foci in all three early-onset patients, but only in a minority of 12 less severe DM1 patients. There were no false-positive results in the 7 controls. Although the method does not reliably identify all DM1 patients, it may prove useful as a rapid test for severe congenital DM1 in floppy babies.

scholarly journals miR-322/-503 rescues myoblast defects in myotonic dystrophy type 1 cell model by targeting CUG repeats

2020 ◽  
Vol 11 (10) ◽  
Author(s):  
Xiaopeng Shen ◽  
Feng Xu ◽  
Meng Li ◽  
Shen Wu ◽  
Jingyi Zhang ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Myotonic Dystrophy ◽  
Cell Model ◽  
C2c12 Cells ◽  
Myoblast Differentiation ◽  
C2c12 Myoblast ◽  
Myotonic Dystrophy Type 1 ◽  
Myotonic Dystrophy Type ◽  
Cug Repeats

Abstract Myotonic dystrophy type 1 (DM1) is the most common type of adult muscular dystrophy caused by the expanded triple-nucleotides (CUG) repeats. Myoblast in DM1 displayed many defects, including defective myoblast differentiation, ribonuclear foci, and aberrant alternative splicing. Despite many were revealed to function in DM1, microRNAs that regulated DM1 via directly targeting the expanded CUG repeats were rarely reported. Here we discovered that miR-322/-503 rescued myoblast defects in DM1 cell model by targeting the expanded CUG repeats. First, we studied the function of miR-322/-503 in normal C2C12 myoblast cells. Downregulation of miR-322/-503 significantly hindered the myoblast differentiation, while miR-322/-503 overexpression promoted the process. Next, we examined the role of miR-322/-503 in the DM1 C2C12 cell model. miR-322/-503 was downregulated in the differentiation of DM1 C2C12 cells. When we introduced ectopic miR-322/-503 expression into DM1 C2C12 cells, myoblast defects were almost fully rescued, marked by significant improvements of myoblast differentiation and repressions of ribonuclear foci formation and aberrant alternative splicing. Then we investigated the downstream mechanism of miR-322/-503 in DM1. Agreeing with our previous work, Celf1 was proven to be miR-322/-503′s target. Celf1 knockdown partially reproduced miR-322/-503′s function in rescuing DM1 C2C12 differentiation but was unable to repress ribonuclear foci, suggesting other targets of miR-322/-503 existed in the DM1 C2C12 cells. As the seed regions of miR-322 and miR-503 were complementary to the CUG repeats, we hypothesized that the CUG repeats were the target of miR-322/-503. Through expression tests, reporter assays, and colocalization staining, miR-322/-503 was proved to directly and specifically target the expanded CUG repeats in the DM1 cell model rather than the shorter ones in normal cells. Those results implied a potential therapeutic function of miR-322/-503 on DM1, which needed further investigations in the future.

Booroola BMPR1B mutation alters early follicular development and oocyte ultrastructure in sheep

2012 ◽  
Vol 24 (2) ◽  
pp. 353 ◽  
Author(s):  
Karen L. Reader ◽  
Lisa J. Haydon ◽  
Roger P. Littlejohn ◽  
Jennifer L. Juengel ◽  
Kenneth P. McNatty
Keyword(s):  
Smooth Endoplasmic Reticulum ◽  
In Situ Hybridisation ◽  
Follicular Development ◽  
Cell Types ◽  
Protein Receptor ◽  
Morphogenetic Protein ◽  
Ultrastructural Characteristics ◽  
Type 3

Booroola ewes homozygous (BB) for a mutation in the bone morphogenetic protein receptor-1b (BMPR1B) gene exhibit higher ovulation rates, have larger diameter oocytes at earlier stages of follicular development (i.e. Type 3) and smaller diameter follicles at ovulation than wild-type (++) sheep. However, it is not known when BMPR1B is first expressed in the developing ovary or the cell types involved. In addition, the effects of the BMPR1B mutation on primordial (Type 1) follicles or during growth to the Type 3 stage are unknown. In the present study, BB and ++ fetal ovaries at Days 30–135 of gestation were screened by in situ hybridisation for BMPR1B mRNA. Ovaries from BB and ++ lambs were examined by microscopy to measure follicular and oocyte ultrastructural characteristics in Type 1–3 follicles. BMPR1B mRNA was observed in ovaries from Day 35 of gestation and was evident in oocytes of newly forming and fully formed Type 1 follicles. In BB animals, the Type 1 follicles had larger mean follicular and oocyte diameters, a greater volume of mitochondria, smooth endoplasmic reticulum and ribosomes and a greater surface area of junctions with the granulosa cells compared with ++ animals. It is concluded that the BMPR1B mutation alters follicular development from the onset of follicular formation.

scholarly journals Use of RNA Fluorescence In Situ Hybridization in the Prenatal Molecular Diagnosis of Myotonic Dystrophy Type I

2006 ◽  
Vol 52 (2) ◽  
pp. 319-322 ◽  
Author(s):  
Emanuela Bonifazi ◽  
Francesca Gullotta ◽  
Laura Vallo ◽  
Raniero Iraci ◽  
Anna Maria Nardone ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Myotonic Dystrophy ◽  
Somatic Mosaicism ◽  
Type I ◽  
Myotonic Dystrophy Type ◽  
Rna Fish ◽  
Ctg Expansion ◽  
Nuclear Foci

Abstract Background: Myotonic dystrophy type 1 (DM1; OMIM #160900) is an autosomal-dominant genetic disorder with multisystemic clinical features associated with a CTG expansion in the 3′ untranslated region of the DMPK gene on chromosome 19q13.3. A long-PCR protocol to detect the DM1 expansion is rapid, sensitive, and accurate, but interpretative limitations can occur when the expansion size exceeds the PCR amplification range and in cases of somatic mosaicism. Methods: To overcome these problems, we used RNA fluorescence in situ hybridization (RNA-FISH) to study cultured cells derived from chorionic villus samples (CVS) with the DM1 mutation. The RNA-FISH method is designed to detect the distinctive DM1 cellular phenotype, characterized by the presence of nuclei with focal ribonuclear inclusions (foci) containing the DMPK expanded transcripts. We analyzed 6 CVS from DM1-predicted pregnancies and 6 CVS from DM1-negative pregnancies. Results: In 4 DM1-predicted fetuses with a CTG expansion >200 CTG, varying numbers of ribonuclear inclusions were clearly visible in all cells. One case with a somatic mosaicism for the DMPK mutation showed 15% of cells with no nuclear foci. No nuclear signals were detected in all controls examined (n = 6) and in 1 DM1-positive sample with a CTG expansion <100 copies. Conclusion: Nuclear foci, and therefore the DM1 mutation they are caused by, can be detected efficiently on interphase nuclei of trophoblast cells with RNA-FISH when the CTG expansion is >200 copies.

Role of RNA and MBNL1 Nuclear Foci in Pathomolecular Mechanism in Myotonic Dystrophy Type 1 and Type 2 (PD6.003)

Neurology ◽  
2012 ◽  
Vol 78 (Meeting Abstracts 1) ◽  
pp. PD6.003-PD6.003
Author(s):  
G. Meola ◽  
V. Renna ◽  
E. Bugiardini ◽  
M. Giagnacovo ◽  
C. Pellicciari ◽  
...  
Keyword(s):  
Myotonic Dystrophy ◽  
Myotonic Dystrophy Type 1 ◽  
Myotonic Dystrophy Type ◽  
Nuclear Foci

scholarly journals Staufen1 impairs stress granule formation in skeletal muscle cells from myotonic dystrophy type 1 patients

2016 ◽  
Vol 27 (11) ◽  
pp. 1728-1739 ◽  
Author(s):  
Aymeric Ravel-Chapuis ◽  
Amanda Klein Gunnewiek ◽  
Guy Bélanger ◽  
Tara E. Crawford Parks ◽  
Jocelyn Côté ◽  
...  
Keyword(s):  
Myotonic Dystrophy ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Muscle Cells ◽  
Cellular Stress Response ◽  
Granule Formation ◽  
A Cell ◽  
Cell Type Specific ◽  
Cug Repeats

Myotonic dystrophy (DM1) is caused by an expansion of CUG repeats (CUGexp) in the DMPK mRNA 3′UTR. CUGexp-containing mRNAs become toxic to cells by misregulating RNA-binding proteins. Here we investigated the consequence of this RNA toxicity on the cellular stress response. We report that cell stress efficiently triggers formation of stress granules (SGs) in proliferating, quiescent, and differentiated muscle cells, as shown by the appearance of distinct cytoplasmic TIA-1– and DDX3-containing foci. We show that Staufen1 is also dynamically recruited into these granules. Moreover, we discovered that DM1 myoblasts fail to properly form SGs in response to arsenite. This blockage was not observed in DM1 fibroblasts, demonstrating a cell type–specific defect. DM1 myoblasts display increased expression and sequestration of toxic CUGexp mRNAs compared with fibroblasts. Of importance, down-regulation of Staufen1 in DM1 myoblasts rescues SG formation. Together our data show that Staufen1 participates in the inhibition of SG formation in DM1 myoblasts. These results reveal that DM1 muscle cells fail to properly respond to stress, thereby likely contributing to the complex pathogenesis of DM1.

Myotonic dystrophy: a cause of acute breathlessness not to be missed

2020 ◽  
pp. practneurol-2020-002573
Author(s):  
Caroline Kramarz ◽  
Abigail Turner ◽  
Vafa Alakbarzade ◽  
Brendan Mclean
Keyword(s):  
Respiratory Failure ◽  
Myotonic Dystrophy ◽  
Neuromuscular Disorder ◽  
Autosomal Dominant Disorder ◽  
Multisystem Disease ◽  
Phenotypic Spectrum ◽  
Life Threatening ◽  
Recurrent Type

Myotonic dystrophy type 1 (DM1), the most common muscular dystrophy in adults, is an autosomal dominant disorder with a wide phenotypic spectrum ranging from oligosymptomatic forms to a life-threatening, multisystem disease. People with DM1 overall have a reduced life expectancy, mainly due to respiratory or cardiac causes. There is no cure but prompt, appropriate symptom management is essential to limit disease-related complications. We present a case of DM1, unrecognised when the patient presented with recurrent type 2 respiratory failure, and initially misdiagnosed as Guillain-Barré syndrome. This misdiagnosis subsequently led to unnecessary investigation and treatment before further detailed neurological examination and collateral family history gave the diagnosis. This case highlights the importance of considering a chronic neuromuscular disorder in patients presenting with acute respiratory failure and an unusual pattern of weakness.

Fetal-type gastrointestinal adenocarcinoma: a morphologically distinct entity with unfavourable prognosis

2017 ◽  
Vol 71 (3) ◽  
pp. 221-227 ◽  
Author(s):  
Kshitij Arora ◽  
Munita Bal ◽  
Angela Shih ◽  
Andrea Moy ◽  
Lawerence Zukerberg ◽  
...  
Keyword(s):  
In Situ Hybridisation ◽  
Morphological Evidence ◽  
Genetic Profile ◽  
Type I ◽  
Type Iii ◽  
Arginase 1 ◽  
Gastric Adenocarcinomas ◽  
Elevated Expression

AimsThis multi-institutional study and a re-evaluation of the TCGA cohort explores the morphological spectrum, genetics and outcome of GI (gastrointestinal) hepatoid tumours, tumours expressing alpha-fetoprotein (AFP) and fetal-type (FT) GI adenocarcinomas.Methods44 tumours with evidence of hepatocellular differentiation were evaluated for morphology as well as by immunohistochemistry for AFP, HepPar1, glypican-3 and arginase-1 and by in situ hybridisation for albumin. Three categories were defined: type I (hepatoid: morphological evidence of hepatocellular differentiation), type II (FT GI adenocarcinoma: tubular profiles and subnuclear vacuolisation, resembling fetal intestine) and type III: positive for at least two hepatocyte-specific markers but lacking morphological evidence of hepatocellular differentiation. GI adenocarcinomas in the TCGA cohort were also evaluated (n=829).Results18 cases were classified as type I, 19 as FT GI adenocarcinomas and 7 as type III (resembling conventional gastrointestinal carcinomas). Serum AFP was elevated in 92% of cases. 93% of tumours were positive for glypican-3, 90% for albumin and 89% for AFP. Arginase-1 was restricted to 35% of type 1 tumours. TCGA gastric tumours with elevated AFP expression showed morphological features of FT GI adenocarcinoma (70%) and were exclusively MSI stable. TCGA gastric adenocarcinomas with high AFP expression showed inferior survival on univariate and multivariate analysis.ConclusionsFT GI adenocarcinomas show a distinctive morphological and immunohistochemical profile. Gastric adenocarcinomas with elevated expression of AFP morphologically resemble FT GI adenocarcinomas, demonstrate aggressive behaviour, independent of grade and stage, and a distinct genetic profile.

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