Fast myosin immunostaining of skeletal muscle transversal sections obtained from a healthy patient (A), DM1 patients (B-D) and DM2 patients (E-G).

<p>Type 2 fibers (fast positive fibers) are stained in brown. Muscle from DM1-E1 (B) and DM2-PS (E) patients show a normal histological muscle pattern similar to those observed in control muscle section (A). Muscle from DM1-E2 (C) and DM1-CDM (D) patients show a high fiber size variability with both type 1 (unstained fibers; white arrows) and type 2 (black arrows) atrophic fibers, fast positive nuclear clumps (arrowheads) and a preferential type 1 fiber central nucleation (asterisks). Muscle from DM2-PDM and DM2-PROMM patients also show high fiber size variability with very small type 2 fibers (black arrows), type 2 nuclear clumps (arrowheads) and a preferential type 2 fiber central nucleation (asterisks).</p

Clinical data on DM patients used in this study.

a<p>Medical Research Council, scale for muscle strength; scale (0–5 grade) on 15 muscles at both sides in the upper and lower limbs for a total of 150 maximum score.</p>b<p>Electrocardiogram, included first-degree atrio-ventricular block, incomplete or complete bundle-branch block.</p>c<p>Muscle Impairment Rating Scale, stage of the disease for DM1 patients <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0083777#pone.0083777-Mathieu1" target="_blank">[73]</a>.</p

Results of QRT-PCR experiments to quantify the expression level of the <i>CLCN1</i> mRNA in biceps brachii muscle samples from DM2 patients (DM2-PS (n = 4), DM2-PDM (n = 5) and DM2-PROMM (n = 5)) and controls.

<p>Each experiment has been performed in triplicate and the relative amount of the <i>CLCN1</i> transcripts has been determined using the β₂-microglobulin as endogenous control gene. Bars represent standard deviation.</p

Metahistograms have been obtained from the analysis of muscle fiber diameters in DM1 patients (A-C) and in DM2 patients (D-F).

<p>The results are based on sections immunostained for MHC fast or slow myosin. Tables show the relative atrophy or hypertrophy factors in each subphenotype considered. Data relative to each DM1 and DM2 phenotypic groups have been obtained by pooling the findings of each patient: DM1-E1 (n = 3), DM1-E2 (n = 5), DM1-CDM (n = 3), DM2-PS (n = 4), DM2-PDM (n = 5) and DM2-PROMM (n = 5).</p

Genome Wide Identification of Aberrant Alternative Splicing Events in Myotonic Dystrophy Type 2

<div><p>Myotonic dystrophy type 2 (DM2) is a genetic, autosomal dominant disease due to expansion of tetraplet (CCTG) repetitions in the first intron of the ZNF9/CNBP gene. DM2 is a multisystemic disorder affecting the skeletal muscle, the heart, the eye and the endocrine system. According to the proposed pathological mechanism, the expanded tetraplets have an RNA toxic effect, disrupting the splicing of many mRNAs. Thus, the identification of aberrantly spliced transcripts is instrumental for our understanding of the molecular mechanisms underpinning the disease. The aim of this study was the identification of new aberrant alternative splicing events in DM2 patients. By genome wide analysis of 10 DM2 patients and 10 controls (CTR), we identified 273 alternative spliced exons in 218 genes. While many aberrant splicing events were already identified in the past, most were new. A subset of these events was validated by qPCR assays in 19 DM2 and 15 CTR subjects. To gain insight into the molecular pathways involving the identified aberrantly spliced genes, we performed a bioinformatics analysis with Ingenuity system. This analysis indicated a deregulation of development, cell survival, metabolism, calcium signaling and contractility. In conclusion, our genome wide analysis provided a database of aberrant splicing events in the skeletal muscle of DM2 patients. The affected genes are involved in numerous pathways and networks important for muscle physio-pathology, suggesting that the identified variants may contribute to DM2 pathogenesis.</p></div

Needle-like instruments for steering through solid organs: A review of the scientific and patent literature

High accuracy and precision in reaching target locations inside the human body is necessary for the success of percutaneous procedures, such as tissue sample removal (biopsy), brachytherapy, and localized drug delivery. Flexible steerable needles may allow the surgeon to reach targets deep inside solid organs while avoiding sensitive structures (e.g. blood vessels). This article provides a systematic classification of possible mechanical solutions for three-dimensional steering through solid organs. A scientific and patent literature search of steerable instrument designs was conducted using Scopus and Web of Science Derwent Innovations Index patent database, respectively. First, we distinguished between mechanisms in which deflection is induced by the pre-defined shape of the instrument versus mechanisms in which an actuator changes the deflection angle of the instrument on demand. Second, we distinguished between mechanisms deflecting in one versus two planes. The combination of deflection method and number of deflection planes led to eight logically derived mechanical solutions for three-dimensional steering, of which one was dismissed because it was considered meaningless. Next, we classified the instrument designs retrieved from the scientific and patent literature into the identified solutions. We found papers and patents describing instrument designs for six of the seven solutions. We did not find papers or patents describing instruments that steer in one-plane on-demand via an actuator and in a perpendicular plane with a pre-defined deflection angle via a bevel tip or a pre-curved configuration.Accepted Author ManuscriptMedical Instruments & Bio-Inspired Technolog

Increased PHKA1 exon 19 inclusion in DM2 patients.

<p>A) EAA analysis identified an AS event on exon 19 of PHKA1 mRNA transcript ENST00000339490. The AS area is enlarged and exon 19, more frequently included in DM2 patients, is highlighted in solid gray. B) The box plot shows the increased expression of Affymetrix probe set 4012322 recognizing exon 19, in DM2 patients compared to CTR (n = 10, **** p<0.0001). Values are normalized for the levels of the whole transcript. The splice index (SI) is indicated. C) Validation qPCR assays were performed using the specific primer pair indicated as black arrowhead in panel A. Results are shown as fold change (DM2 = 19, CTR = 15; ** p<0.01).</p

Increased NDUFV3 exon 3 skipping in DM2 patients.

<p>A) EAA analysis identified an AS event on exon 3 of NDUFV3 mRNA transcript ENST00000354250. The AS area is enlarged and exon 3, less frequently included in DM2 patients, is highlighted in solid gray. B) The box plot shows the decreased expression of Affymetrix probe set 3922937 recognizing exon 3, in DM2 patients compared to CTR (n = 10, **** p<0.0001). Values are normalized for the levels of the whole transcript. The splice index (SI) is indicated. C) Validation qPCR assays were performed using the specific primer pair indicated as black arrowhead in panel A. Results are shown as fold change (DM2 = 19, CTR = 15; ** p<0.01).</p

Increased LAMC2 exon 23 inclusion in DM2 patients.

<p>A) EAA analysis identified an AS event on exon 23 of LAMC2 mRNA transcript ENST00000264144. The AS area is enlarged and exon 23, more frequently included in DM2 patients, is highlighted in solid gray. B) The box plot shows the increased expression of Affymetrix probe set 2371184, recognizing exon 23, in DM2 patients compared to CTR (n = 10, * p<0.05). Values are normalized for the levels of the whole transcript. The splice index (SI) is indicated. C) Validation qPCR assays were performed using the specific primer pair indicated as black arrowhead in panel A. Results are shown as fold change (DM2 = 19, CTR = 15; *p<0.05).</p

Increased ZMND11 exon 2 inclusion in DM2 patients.

<p>A) EAA analysis identified an AS event on exon 2 of ZMND11 mRNA transcript ENST00000381584. The AS area is enlarged and exon 2, more frequently included in DM2 patients, is highlighted in solid gray. B) The box plot shows the increased expression of Affymetrix probe set 3231406 recognizing exon 2, in DM2 patients compared to CTR (n = 10, * p<0.05). Values are normalized for the levels of the whole transcript. The splice index (SI) is indicated. C) Validation qPCR assays were performed using the specific primer pair indicated as black arrowhead in panel A. Results are shown as fold change (DM2 = 19, CTR = 15; * p<0.05).</p