Characterization of RAN Translation and Antisense Transcription in Primary Cell Cultures of Patients with Myotonic Dystrophy Type 1

Traducció RAN; Transcripció antisentit; Moduladors fenotípicsTraducción RAN; Transcripción antisentido; Moduladores fenotípicosRAN translation; Antisense transcription; Phenotypic modulatorsMyotonic Dystrophy type 1 (DM1) is a muscular dystrophy with a multi-systemic nature. It was one of the first diseases in which repeat associated non-ATG (RAN) translation was described in 2011, but has not been further explored since. In order to enhance our knowledge of RAN translation in DM1, we decided to study the presence of DM1 antisense (DM1-AS) transcripts (the origin of the polyglutamine (polyGln) RAN protein) using RT-PCR and FISH, and that of RAN translation via immunoblotting and immunofluorescence in distinct DM1 primary cell cultures, e.g., myoblasts, skin fibroblasts and lymphoblastoids, from ten patients. DM1-AS transcripts were found in all DM1 cells, with a lower expression in patients compared to controls. Antisense RNA foci were found in the nuclei and cytoplasm of a subset of DM1 cells. The polyGln RAN protein was undetectable in all three cell types with both approaches. Immunoblots revealed a 42 kD polyGln containing protein, which was most likely the TATA-box-binding protein. Immunofluorescence revealed a cytoplasmic aggregate, which co-localized with the Golgi apparatus. Taken together, DM1-AS transcript levels were lower in patients compared to controls and a small portion of the transcripts included the expanded repeat. However, RAN translation was not present in patient-derived DM1 cells, or was in undetectable quantities for the available methods.The research of G. Nogales-Gadea and A. Ramos-Fransi is funded by Instituto de Salud Carlos III (grant numbers PI15/01756 and PI18/00713) and co-financed by Fondos FEDER. G. Nogales-Gadea is supported by a Miguel Servet research contract (ISCIII CD14/00032, ISCIII CPII19/00021, and FEDER) and by a Trampoline Grant #21108 from AFM Telethon. E. Koehorst is funded by the La Caixa Foundation (ID 100010434), fellowship code LCF/BQ/IN18/11660019, cofunded by the European Union’s Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement no. 713673. The research of M. Suelves is funded by Ministerio de Ciencia e Innovación (grant number PID2020-118730RB-I00) and co-financed by Fondos FEDER. J. Núñez-Manchón is funded by Instituto de Salud Carlos III I-PFIS fellowship (grant number IFI20/00022). G. Lucente is supported by a Rio Hortega contract (ISCIII CM16/00016 and FEDER). J. Chojnacki is supported by European Union’s Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement no. 793830. The work of A.P. Gómez-Escribano and R.P. Vázquez-Manrique is funded by the ISCIII (CPII16/00004, PI17/00011 and PI20/00114) and the Fundación Ramón Areces (CIVP19S8119). This work was supported by the CERCA program/ Government of Catalonia. The funding bodies had no role in the design of the study and the collection, analysis, and interpretation of data

Current Incidence and risk factors of fecal incontinence after acute stroke affecting functionally independent people

Background: Previously published retrospective series show a high prevalence of fecal incontinence (FI) in stroke patients. We aimed to analyze in a prospective series the current incidence of FI in acute stroke in functionally independent patients and its evolution over time and the patient characteristics associated with the appearance of FI in acute stroke. Methods: We included consecutive patients with acute stroke admitted in our stroke unit who fulfilled the following inclusion criteria: a first episode of stroke, aged >18 years, with no previous functional dependency [modified Rankin Scale (mRS) = 2] and without previous known FI. FI was assessed by a multidisciplinary trained team using dedicated questionnaires at 72 ± 24 h (acute phase) and at 90 ± 15 days (chronic phase). Demographic, medical history, clinical and stroke features, mortality, and mRS at 7 days were collected. Results: Three hundred fifty-nine (48.3%) of 749 patients (mean age 65.9 ± 10, 64% male, 84.1% ischemic) fulfilled the inclusion criteria and were prospectively included during a 20-month period. FI was identified in 23 patients (6.4%) at 72 ± 24 h and in 7 (1.9%) at 90 days ± 15 days after stroke onset. FI was more frequent in hemorrhagic strokes (18 vs. 5%, p 0.007) and in more severe strokes [median National Institute of Health Stroke Scale (NIHSS) 18 (14–22) vs. 5 (3–13), p < 0.0001]. No differences were found regarding age, sex, vascular risk factors, or other comorbidities, or affected hemisphere. Patients with NIHSS =12 (AUC 0.81, 95% CI 0.71 to 0.89) had a 17-fold increase for the risk of FI (OR 16.9, IC 95% 4.7–60.1) adjusted for covariates.Peer ReviewedPostprint (published version

Three-dimensional imaging in myotonic dystrophy type 1

Altres ajuts: The research of G. Nogales-Gadea, A. Ramos-Fransi, and A. Lucia is funded by Instituto de Salud Carlos III and cofinanced by Fondos FEDER. G. Nogales-Gadea is supported by a Miguel Servet research contract and by a Trampoline Grant #21108 from AFM Telethon. A. Ballester-Lopez is funded by an FI Agaur fellowship and Generalitat de Catalunya. E. Koehorst is funded by the La Caixa Foundation (ID 100010434), fellowship code LCF/BQ/IN18/11660019, cofunded by the European Union's Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement no. 713673. I. Linares-Pardo is funded by CP14/00032 and SGR 1520 (GRC) Generalitat de Catalunya. J. Núñez-Manchón was funded by AFM Telethon Trampoline Grant #21108. G. Lucente was supported by a Rio Hortega contract. J. Chojnacki is supported by European Union's Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant . The funding bodies had no role in the design of the study and collection, analysis, and interpretation of data.We aimed to determine whether 3D imaging reconstruction allows identifying molecular:clinical associations in myotonic dystrophy type 1 (DM1). We obtained myoblasts from 6 patients with DM1 and 6 controls. We measured cytosine-thymine-guanine (CTG) expansion and detected RNA foci and muscleblind like 1 (MBNL1) through 3D reconstruction. We studied dystrophia myotonica protein kinase (DMPK) expression and splicing alterations of MBNL1, insulin receptor, and sarcoplasmic reticulum Ca(2+)-ATPase 1. Three-dimensional analysis showed that RNA foci (nuclear and/or cytoplasmic) were present in 45%-100% of DM1-derived myoblasts we studied (range: 0-6 foci per cell). RNA foci represented <0.6% of the total myoblast nuclear volume. CTG expansion size was associated with the number of RNA foci per myoblast (r = 0.876 [95% confidence interval 0.222-0.986]) as well as with the number of cytoplasmic RNA foci (r = 0.943 [0.559-0.994]). Although MBNL1 colocalized with RNA foci in all DM1 myoblast cell lines, colocalization only accounted for 1% of total MBNL1 expression, with the absence of DM1 alternative splicing patterns. The number of RNA foci was associated with DMPK expression (r = 0.967 [0.079-0.999]). On the other hand, the number of cytoplasmic RNA foci was correlated with the age at disease onset (r = −0.818 [−0.979 to 0.019]). CTG expansion size modulates RNA foci number in myoblasts derived from patients with DM1. MBNL1 sequestration plays only a minor role in the pathobiology of the disease in these cells. Higher number of cytoplasmic RNA foci is related to an early onset of the disease, a finding that should be corroborated in future studies

A DM1 family with interruptions associated with atypical symptoms and late onset but not with a milder phenotype

Carriage of interruptions in CTG repeats of the myotonic dystrophy protein kinase gene has been associated with a broad spectrum of myotonic dystrophy type 1 (DM1) phenotypes, mostly mild. However, the data available on interrupted DM1 patients and their phenotype are scarce. We studied 49 Spanish DM1 patients, whose clinical phenotype was evaluated in depth. Blood DNA was obtained and analyzed through triplet‐primed polymerase chain reaction (PCR), long PCR‐Southern blot, small pool PCR, AciI digestion, and sequencing. Five patients of our registry (10%), belonging to the same family, carried CCG interruptions at the 3’ end of the CTG expansion. Some of them presented atypical traits such as a very late onset of symptoms (&gt;50 years) and a severe axial and proximal weakness requiring walking assistance. They also showed classic DM1 symptoms including cardiac and respiratory dysfunction, which were severe in some of them. Sizes and interrupted allele patterns were determined, and we found a contraction and an expansion in two intergenerational transmissions. Our study contributes to the observation that DM1 patients carrying interruptions present with atypical clinical features that can make DM1 diagnosis difficult, with a later than expected age of onset and a previously unreported aging‐related severe disease manifestation

Demencia frontotemporal y manía: estudio de un caso y revisión de la literatura

Introducción: La demencia frontotemporal es un término descriptivo usado para referirse a la demencia asociada invariablemente con degeneración de los lóbulos frontal y temporal, en ausencia de enfermedad de Alzheimer. Se trata de un grupo heterogéneo de enfermedades neurodegenerativas con diferentes manifestaciones clínicas, componente genético y alteración anatomopatológica. Sus síntomas psiquiátricos son de inicio insidioso, con deterioro de la personalidad y progresiva afectación de la conducta. Generalmente estos cambios se producen antes de encontrar anomalías en las pruebas de neuroimagen, sus características dependen de la localización de las lesiones cerebrales y suelen preceder o eclipsar a las discapacidades cognitivas. La apatía y la abulia son los síntomas iniciales más comunes y se presentan en la mayoría de los pacientes cuando la enfermedad progresa. La desinhibición se manifiesta por diferentes comportamientos sociales inapropiados como: hilaridad inadecuada, exhibicionismo, inquietud, impulsividad, sexualidad y violencia incontrolada, jocosidad inapropiada y humor pueril; pudiendo estar presente en la mitad de los pacientes. Caso clínico: Se presenta un caso clínico representativo, en el que la primera manifestación de una demencia frontotemporal consiste en un cuadro de características maníacas. También se revisa la bibliografía específica publicada hasta la fecha. Conclusiones: Es importante realizar un correcto diagnóstico diferencial entre una demencia incipiente y el debut tardío de un trastorno afectivo bipolar, así como las exploraciones complementarias que pueden ser de utilidad a tal fin

A novel mutation in the valosin-containing-protein gene found in a Spanish family

Sin financiación2.651 JCR (2018) Q2, 99/199 Clinical Neurology; Q3, 160/267 Neurosciences0.990 SJR (2018) Q2, 70/165 Neurology, 126/378 Neurology (clinical)No data IDR 2018UE

A DM1 family with interruptions associated with atypical symptoms and late onset but not with a milder phenotype

Carriage of interruptions in CTG repeats of the myotonic dystrophy protein kinase gene has been associated with a broad spectrum of myotonic dystrophy type 1 (DM1) phenotypes, mostly mild. However, the data available on interrupted DM1 patients and their phenotype are scarce. We studied 49 Spanish DM1 patients, whose clinical phenotype was evaluated in depth. Blood DNA was obtained and analyzed through triplet‐primed polymerase chain reaction (PCR), long PCR‐Southern blot, small pool PCR, AciI digestion, and sequencing. Five patients of our registry (10%), belonging to the same family, carried CCG interruptions at the 3′‐end of the CTG expansion. Some of them presented atypical traits such as very late onset of symptoms ( > 50 years) and a severe axial and proximal weakness requiring walking assistance. They also showed classic DM1 symptoms including cardiac and respiratory dysfunction, which were severe in some of them. Sizes and interrupted allele patterns were determined, and we found a contraction and an expansion in two intergenerational transmissions. Our study contributes to the observation that DM1 patients carrying interruptions present with atypical clinical features that can make DM1 diagnosis difficult, with a later than expected age of onset and a previously unreported aging‐related severe disease manifestation.Sin financiación4.878 JCR (2020) Q1, 44/175 Genetics & Heredity1.981 SJR (2020) Q1, 51/340 GeneticsNo data IDR 2019UE

Preliminary Findings on CTG Expansion Determination in Different Tissues from Patients with Myotonic Dystrophy Type 1

Myotonic Dystrophy type 1 (DM1) is characterized by a high genetic and clinical variability. Determination of the genetic variability in DM1 might help to determine whether there is an association between CTG (Cytosine-Thymine-Guanine) expansion and the clinical manifestations of this condition. We studied the variability of the CTG expansion (progenitor, mode, and longest allele, respectively, and genetic instability) in three tissues (blood, muscle, and tissue) from eight patients with DM1. We also studied the association of genetic data with the patients' clinical characteristics. Although genetic instability was confirmed in all the tissues that we studied, our results suggest that CTG expansion is larger in muscle and skin cells compared with peripheral blood leukocytes. While keeping in mind that more research is needed in larger cohorts, we have provided preliminary evidence suggesting that the estimated progenitor CTG size in muscle could be potentially used as an indicator of age of disease onset and muscle function impairment.Instituto de Salud Carlos III (Grant Numbers: PI15/01756; P18/00713)AFM Telethon (Trampoline grant number #21108)AFM Telethon Trampoline Grant #21108FI Agaur fellowship FI_B 01090“La Caixa” Foundation (ID 100010434), fellowship code LCF/BQ/IN18/11660019, co-funded by the European Union’s Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement n°713673CP14/00032Miguel Servet research contract (ISCIII CD14/00032, CPII19/00021, and FEDER)Rio Hortega contract (ISCIII CM16/00016 and FEDER)Personal honoraria from Shire-Takeda, Amicus, Kyowa-Kirin, and Sanofi-Genzyme4.096 JCR (2020) Q2, 65/175 Genetics & Heredity1.337 SJR (2020) Q2, 99/340 GeneticsNo data IDR 2019UE

Myotilinopathy unmasked by statin treatment: A case report

Sin financiación2.393 JCR (2018) Q3, 115/199 Clinical Neurology, 184/261 Neurosciences0.950 SJR (2018) Q2, 134/378 Neurology (clinical), 80/188 Physiology, 45/108 Physiology (medical); Q3, 59/90 Cellular and Molecular NeuroscienceNo data IDR 2018UE

Three-dimensional imaging in myotonic dystrophy type 1: Linking molecular alterations with disease phenotype

Objective: We aimed to determine whether 3D imaging reconstruction allows identifying molecular:clinical associations in myotonic dystrophy type 1 (DM1). Methods: We obtained myoblasts from 6 patients with DM1 and 6 controls. We measured cytosine-thymine-guanine (CTG) expansion and detected RNA foci and muscleblind like 1 (MBNL1) through 3D reconstruction. We studied dystrophia myotonica protein kinase (DMPK) expression and splicing alterations of MBNL1, insulin receptor, and sarcoplasmic reticulum Ca(2+)-ATPase 1. Results: Three-dimensional analysis showed that RNA foci (nuclear and/or cytoplasmic) were present in 45%-100% of DM1-derived myoblasts we studied (range: 0-6 foci per cell). RNA foci represented <0.6% of the total myoblast nuclear volume. CTG expansion size was associated with the number of RNA foci per myoblast (r = 0.876 [95% confidence interval 0.222-0.986]) as well as with the number of cytoplasmic RNA foci (r = 0.943 [0.559-0.994]). Although MBNL1 colocalized with RNA foci in all DM1 myoblast cell lines, colocalization only accounted for 1% of total MBNL1 expression, with the absence of DM1 alternative splicing patterns. The number of RNA foci was associated with DMPK expression (r = 0.967 [0.079-0.999]). On the other hand, the number of cytoplasmic RNA foci was correlated with the age at disease onset (r = -0.818 [-0.979 to 0.019]). Conclusions: CTG expansion size modulates RNA foci number in myoblasts derived from patients with DM1. MBNL1 sequestration plays only a minor role in the pathobiology of the disease in these cells. Higher number of cytoplasmic RNA foci is related to an early onset of the disease, a finding that should be corroborated in future studies.Sin financiación3.485 JCR (2020) Q2, 96/208 Clinical Neurology1.262 SJR (2020) Q1, 92/373 Neurology (clinical)No data IDR 2019UE