Muscular dystrophies and myopathies: the spectrum of mutated genes in the Czech Republic.

Inherited neuromuscular disorder (NMD) is a wide term covering different genetic disorders affecting muscles, nerves, and neuromuscular junctions. Genetic and clinical heterogeneity is the main drawback in a routine gene-by-gene diagnostics. We present Czech NMD patients with a genetic cause identified using targeted next-generation sequencing (NGS) and the spectrum of these causes. Overall 167 unrelated patients presenting NMD falling into categories of muscular dystrophies, congenital muscular dystrophies, congenital myopathies, distal myopathies, and other myopathies were tested by targeted NGS of 42 known NMD-related genes. Pathogenic or probably pathogenic sequence changes were identified in 79 patients (47.3%). In total, 37 novel and 51 known disease-causing variants were detected in 23 genes. In addition, variants of uncertain significance were suspected in 7 cases (4.2%), and in 81 cases (48.5%) sequence changes associated with NMD were not found. Our results strongly indicate that for molecular diagnostics of heterogeneous disorders such as NMDs, targeted panel testing has a high-clinical yield and should therefore be the preferred first-tier approach. Further, we show that in the genetic diagnostic practice of NMDs, it is necessary to take into account different types of inheritance including the occurrence of an autosomal recessive disorder in two generations of one family.

Profile measurements of the electron temperature on the ASDEX Upgrade, COMPASS, and ISTTOK tokamak using Thomson scattering, triple, and ball-pen probes.

The ball-pen probe (BPP) technique is used successfully to make profile measurements of the electron temperature on the ASDEX Upgrade (Axially Symmetric Divertor Experiment), COMPASS (COMPact ASSembly), and ISTTOK (Instituto Superior Tecnico TOKamak) tokamak. The electron temperature is provided by a combination of the BPP potential (ΦBPP) and the floating potential (Vfl) of the Langmuir probe (LP), which is compared with the Thomson scattering diagnostic on ASDEX Upgrade and COMPASS. Excellent agreement between the two diagnostics is obtained for circular and diverted plasmas and different heating mechanisms (Ohmic, NBI, ECRH) in deuterium discharges with the same formula Te = (ΦBPP - Vfl)/2.2. The comparative measurements of the electron temperature using BPP/LP and triple probe (TP) techniques on the ISTTOK tokamak show good agreement of averaged values only inside the separatrix. It was also found that the TP provides the electron temperature with significantly higher standard deviation than BPP/LP. However, the resulting values of both techniques are well in the phase with the maximum of cross-correlation function being 0.8.

High frequency of SH3TC2 mutations in Czech HMSN I patients.

Charcot-Marie-Tooth (CMT) neuropathy type 4C (CMT4C) is an autosomal recessive (AR), demyelinating neuropathy with early spine deformities caused by mutations in the SH3TC2 gene. To determine the spectrum of SH3TC2 mutations in the Czech population, the entire coding region of SH3TC2 was sequenced in 60 unrelated Czech patients. The prevalent mutation was shown to be the p.Arg954Stop. Therefore, 412 additional patients referred for CMT testing were tested for the presence of p.Arg954Stop only. Of 60 patients in whom the SH3TC2 gene was sequenced, at least one mutation was detected in 13 (21.7%) patients and biallelic pathogenic mutations were detected in 7 (11.6%) patients. Of the 412 patients tested for p.Arg954Stop, the mutation was found in 8 patients (1.94%), 6 were homozygous and 2 were heterozygous. The second causative mutation was detected by sequencing in one of the patients but not in the other. Nine novel sequence variants were detected. Their pathogenicity was further tested in silico and in control samples. Mutations in the SH3TC2 gene are a frequent cause of demyelinating hereditary neuropathy among Czech patients. In total, at least one mutation was found in 21 unrelated patients. CMT4C seems to be the most frequent type of AR CMT and one of the most frequent of all CMT types. Mutation p.Arg954Stop is highly prevalent in the Czech population. Patients with demyelinating neuropathy along with non-dominant mode of inheritance and negative for CMT1A/hereditary neuropathy with liability to pressure palsy should be tested for the presence of the p.Arg954Stop mutation or other mutations in the SH3TC2 gene.

Glaucoma drainage implants in the treatment of refractory glaucoma in pediatric patients.

PURPOSE: The aim of this study is to report the clinical course, effectiveness, and safety of glaucoma drainage implants (Molteno and Baerveldt devices) in primary and secondary childhood glaucomas refractory to conventional surgical treatments and medical therapy. METHODS: This retrospective study included 76 children (76 eyes) younger than 18 years who underwent glaucoma drainage device (GDD) implantation in our clinic between 1990 and 2004. The mean age at time of surgery was 6.9+/-5.3 years (range: 4 months to 17.5 years). Intraocular pressure (IOP), visual acuity, corneal diameter, axial length, intraoperative and postoperative complications, and number of glaucoma medications were evaluated. Criteria for success were defined as IOP between 7 and 22 mmHg with or without glaucoma medications, no further glaucoma surgery, the absence of visually threatening complications, and no loss of light perception. Results were compared for children with primary and secondary glaucomas. The mean follow-up was 7.1+/-6.5 years (range: 1.6 to 15.2 years). RESULTS: Mean preoperative and postoperative IOP was 33.6+/-11.4 mmHg and 17.1+/-6.5 mmHg (p<0.001), respectively. Kaplan-Meier survival analysis showed cumulative probability of success: 93% at 6 months, 91% at 1 year, 82% at 2 years, 76% at 3 years, 71% at 4 years, 67% at 5 years, and 65% at 6 years. There was no difference between patients with primary (n=31 eyes) and secondary glaucoma (n=45 eyes) in terms of cumulative success (p=0.186), final IOP, number of medications, or length of follow-up. On average, the GDI surgery was successful for a mean period of 6.7 years. Fourteen eyes of 76 (18.4%) failed: 10 eyes with uncontrolled IOP, 2 eyes with retinal detachment, and 2 eyes with no light perception. Statistical regression model did not show influence of gender and previous surgery. Lower age at the time of surgery was found to be associated with higher probability of treatment failure. CONCLUSIONS: Molteno and Baerveldt glaucoma drainage implants surgery seems to be safe and effective treatment for primary and secondary pediatric glaucoma refractory to the initial surgical procedure and medical therapy.

Charcot-Marie-Tooth neuropathy type 1A combined with Duchenne muscular dystrophy.

We report a 24-year-old male with an unusual combination of two inherited neuromuscular disorders--Charcot-Marie-Tooth (CMT) disease type 1A and Duchenne muscular dystrophy (DMD). A phenotypic presentation of this patient included features of both these disorders. Nerve conduction studies revealed demyelinating peripheral neuropathy. Electromyography showed a profound myogenic pattern. The serum creatine kinase level was highly elevated. Muscle biopsy revealed a dystrophic picture with deficient dystrophin immunostaining. CMT1A duplication on chromosome 17p11.2 was found. The frame-shift mutation c.3609-3612delTAAAinsCTT (p.K1204LfsX11) was detected in the dystrophin gene by analysing mRNA isolated from the muscle tissue. The patient inherited both these mutations from his mother. The combination of CMT1A and DMD has not been reported as yet.

GDAP1 mutations in Czech families with early-onset CMT.

Mutations in the ganglioside-induced differentiation associated protein-1 gene (GDAP1) cause autosomal recessive (AR) demyelinating or axonal Charcot-Marie-Tooth neuropathy (CMT). In order to establish the spectrum and frequency of GDAP1 mutations in Czech population, we sequenced GDAP1 in 74 Czech patients from 69 unrelated families with early-onset demyelinating or axonal CMT compatible with AR inheritance. We identified three isolated patients with GDAP1 mutations in both alleles. In one additional sporadic and one familial case, the second pathogenic mutation remained unknown. Overall, we detected two different mutations, a novel R191X nonsense and a L239F missense mutation. L239F previously described in a German-Italian family is a prevalent mutation in Czech population and we give evidence for its common ancestral origin. All Czech GDAP1 patients developed involvement of all four limbs evident by the end of second decade, except for one isolated patient showing very slow disease progression. All patients displayed axonal type of neuropathy.

Risk factors for critical illness polyneuromyopathy.

Although numerous clinical, laboratory, and pharmacological variables have been reported as significant risk factors for critical illness polyneuromyopathy (CIPM), there is still no consensus on the aetiology of this condition. Objectives of the study were to assess the clinical and electrophysiological incidence and risk factors for CIPM.A cohort of critically ill patients was observed prospectively for a one-month period and the association between neuromuscular involvement and various potential risk factors was evaluated. Sixty one critically ill patients completed the follow-up (30 women, 31 men, median age 59 years).CIPM development was detected clinically in 17 patients (27.9 %) and electrophysiologically in 35 patients (57.4 %). CIPM was significantly associated with the presence and duration of systemic inflammatory response syndrome and the severity of multiple, respiratory, central nervous, and cardiovascular organ failures. The median duration of mechanical ventilation was significantly longer in patients with CIPM than in those without (16 vs 3 days, p<0.001). Independent predictors of CIPM obtainable within the 1(st) week of critical illness were the admission sequential organ failure assessment score (odds ratio [OR], 1.15; 95% confidence interval [CI], 1.02-1.36), the 1(st) week total sequential organ failure assessment scores (OR, 1.14; 95 % CI, 1.06-1.46) and the 1(st) week duration of systemic inflammatory response syndrome (OR, 1.05; 95% CI, 1.01-1.15). They were able to correctly predict the development of CIPM at the end of the 1(st) week in about 80% of critically ill cases.In conclusion, the presence and duration of systemic inflammatory response syndrome and the severity of multiple and several organ failures are associated with increased risk of the development of CIPM.

[A unique case of congenital muscular dystrophy]. / Unikátní prípad kongenitální svalové dystrofie.

The congenital muscular dystrophies (CMD, MDC) represent a heterogeneous group of autosomal recessive disorders manifesting in infancy by muscle weakness and hypotonia. Approximately 40% of patients with CMD have a primary deficiency of the laminin alpha 3. chain of merosin (laminin-2) due to mutations in LAMA2 gene. Laminin-2 bound to alpha-dystroglycan forms a link between actin--associated cytoskeletal proteins and the components of extracellular matrix. Disruption of this axis is responsible for several forms of muscular dystrophy. A unique case of congenital muscular dystrophy simulating a juvenile polymyositis in a muscle biopsy is presented. A profound reduction of alpha-dystroglycan and less pronounced secondary deficiency of alpha 2-laminin were found. All known forms of CMD were excluded, and the disorder was diagnosed as so far undescribed form of CMD. The mutation in a gene encoding the protein, that seems to play a role in a glycosylation of alpha-dystroglycan, is presumed.

Silicone rubber-hydrogel composites as polymeric biomaterials. IV. Silicone matrix-hydrogel filler interaction and mechanical properties.

Composite materials consisting of a silicone rubber matrix and particulate synthetic hydrogels were prepared and their mechanical properties were studied. The influence of the size, shape, aggregation of hydrogel particles, chemical reactions of polymer phases on tensile properties and tear strength of the composite materials were investigated. The relations between the properties, structure and chemical composition of polymer phases of the composite materials and their mechanical properties are discussed.

Silicone rubber-hydrogel composites as polymeric biomaterials. III. An investigation of phase distribution by scanning electron microscopy.

The structure of silicone rubber-hydrogel composite materials was investigated by scanning electron microscopy (SEM) and light microscopy. The polymer phases in these materials composed of the polysiloxane matrix and very small particles of lightly cross-linked poly(2-hydroxyethylmethacrylate) or poly(2-hydroxyethylmethacrylate-co-methacrylic acid) were visualized using both methods. The distribution of polymer phases was studied by SEM of fracture surfaces of the materials. The results are discussed in relation to the transport properties of the materials.

Silicone rubber-hydrogel composites as polymeric biomaterials. I. Biological properties of the silicone rubber-p(HEMA) composite.

A composite material was prepared consisting of silicone rubber matrix and particulate lightly cross-linked poly(2-hydroxyethyl methacrylate) (p(HEMA] hydrogel. The material resembling common silicone rubber is hydrophilic and swells in water like hydrogels. The effects of the implanted composite on tissues of the living organism were tested in rats by methods assessing local acute and chronic inflammatory reactions and calcification by means of radioactive indicators and by histological examination. Results of a 6 month implant study indicated no difference in reactions of the animal body on the silicone rubber-p(HEMA) composite and a non-toxic, non-irritant pure solid p(HEMA) control.

Silicone rubber-hydrogel composites as polymeric biomaterials. II. Hydrophilicity and permeability to water-soluble low-molecular-weight compounds.

The surface and transport properties of water-swollen silicone rubber-hydrogel composites were investigated. Surface wettability of these materials, composed of a polysiloxane matrix and the hydrogel phase consisting of very fine particles of lightly cross-linked poly(2-hydroxyethylmethacrylate), increased markedly with increasing content of the hydrogel phase. For composite materials containing a lightly cross-linked 2-hydroxyethylmethacrylate (HEMA)-methacrylic acid (MAA) copolymer and polymethacrylic acid (PMAA) as the hydrogel phase, permeability to water-soluble organic compounds and drugs were measured. The permeability varied within a broad range depending on the composition and content of the hydrogel phase. High permeation rates could be obtained while still retaining relatively fair mechanical properties. Relationships between the composition of silicone rubber-hydrogel composites, their structure and the permeation coefficients of the individual permeates are discussed.

The role of sulfation in the metabolic activation of N-hydroxy-4'-fluoro-4-acetylaminobiphenyl.

The role of sulfation in the metabolic activation of the liver carcinogen N-hydroxy-4'-fluoro-4-acetylaminobiphenyl (N-OH-FAABP) in male rat liver was investigated. N-OH-FAABP was a substrate for sulfotransferases in vitro and sulfation was inhibited by the sulfotransferase inhibitors pentachlorophenol (PCP) and 2,6-dichloro-4-nitrophenol (DCNP). The main metabolite of N-OH-FAABP excreted in bile in vivo, and in the isolated perfused liver, was identified as the N-O-glucuronide conjugate. Inhibition of sulfation in vivo by PCP or DCNP, or in vitro by omission of inorganic sulfate, resulted in a large increase in the excretion of the N-O-glucuronide conjugate. It was estimated that at least 21% of the dose was sulfated in control animals. Inhibition of sulfation in vivo by PCP or DCNP prevented the covalent binding of N-OH-FAABP to liver (and kidney) macromolecules by 70% and 20% respectively. HPLC analysis of the fluorobiphenyl DNA and RNA adducts showed that the formation of both N-acetylated and deacetylated (deoxy)-guanosine adducts was prevented. Furthermore, omission of inorganic sulfate in the isolated perfused liver prevented the formation of all fluorobiphenyl DNA adducts by 70-80%. It is concluded that two sulfotransferase-dependent pathways exist for the metabolic activation of N-OH-FAABP in male rat liver: (i) direct sulfation of the hydroxamic acid, resulting, upon decomposition of the FAABP-N-sulfate ester, in the formation of N-acetylated DNA adducts and (ii) deacetylation followed by sulfation of the hydroxylamine to FABP-N-sulfate, leading to the formation of deacetylation DNA adducts.

In vivo degradation of polymers. II. Change of mechanical properties and cross-link density in silicone rubber pacemaker lead insulations during long-term implantation in the human body.

The results of mechanical testing, microscopic examinations and swelling of the explanted clinically used silicone rubber pacemaker lead insulations are presented. An experimental set of 100 lead insulations implanted for times ranging from 3 d to 11 yr were evaluated for change of mechanical properties and cross-link density during clinical use. The results show that the silicone rubber tubing suffers a gradual structural change due to its exposure to the physiological environment of the human body. A spiral deformation pattern due to the permanent set of the silicone rubber tubing in contact with the electrode leading wire was observed.

In vivo degradation of polymers. I. Change of mechanical properties in polyethylene pacemaker lead insulations during long-term implantation in the human body.

The results of mechanical testing and microscopic examination of explanted clinically-used polyethylene pacemaker lead insulations are presented. An experimental set of 98 lead insulations implanted for times ranging from 7 d to 11 yr was evaluated for changes in mechanical properties during clinical use. The results showed that the polyethylene tubing suffered a gradual structural change, due to its exposure to the physiological environment of the human body. The decline of the mechanical properties with implantation time is described by empirical formulae. Abrasive deterioration was also observed and is discussed.

Biostability of medical elastomers: a review.

Biostability of synthetic elastomers used for manufacturing artificial replacements of human organs or their parts is a critical property of such materials as it determines the long-term function of a specific biomedical device. This paper presents a critical review of the present knowledge of biostability of elastomeric biomedical materials used as artifacts functioning under the simultaneous effects of dynamic flexing and contact with body fluids. The main topics discussed are silicone rubber, elastomers for artificial blood pumps, and the methodology of model fatigue-life testing.