Long-term radiocarbon variation studies in the air and tree rings of Slovakia.

Fifty-five years of radiocarbon variation studies are reviewed with an emphasis on a better understanding of the impacts of the Bohunice nuclear power plant and fossil fuel CO2 on the atmosphere and biosphere of Slovakia. The maximum Δ14C levels in the air up to about 1200‰ were observed during the 1970s at the Zlkovce monitoring station, which after 2005 decreased to <30‰. A relative decrease in the atmospheric Δ14C levels due to increasing levels of fossil CO2 in the atmosphere has also been significant, for example, in Bratislava down to about -330‰, but after 2005 they were only <50‰ below the Jungfraujoch European clean-air level. The tree-ring data, averaging the annual Δ14C levels for several stations in Slovakia, have been in agreement with the atmospheric data, as well as with the newly established clean-air station at Jasná in central Slovakia. Future 14C levels will depend strongly on fossil CO2 levels in the atmosphere, which will change the bomb 14C era to the fossil CO2 era. New investigations of 14C variations in the atmosphere-biosphere-hydrosphere compartments represent a great challenge for radiocarbon science, important for better understanding of environmental processes, climate change, and impacts of human activities on the total environment. This new era of radiocarbon research will also need new developments in radiocarbon analytical technologies, as further progress in accuracy and precision of results (<1‰) will be needed to meet the new radiocarbon challenges.

Pediatric Inflammatory Multisystem Syndrome (PIMS) - Potential role for cytokines such Is IL-6.

COVID-19 is a transmissible respiratory disease caused by coronavirus SARS-CoV-2, which is similar to SARS or MERS. Its increased severity was noted in aged patients usually over 65 years of age. Children and young people have an asymptomatic or mild course of the disease.Unfortunately, the number of children with problems after mild or asymptomatic COVID-19 recovery is increasing and their troubles resemble Kawasaki disease, although the laboratory findings seem to be different. This condition is called pediatric inflammatory multisystem syndrome (PIMS), and it is a new disease seen in children directly influenced by previous SARS-CoV-2 infection. The literature reports that PIMS typically follows 2-4 weeks after SARS-CoV-2 infection. The clinical symptoms of the affected children are extremely complex, ranging from gastrointestinal to cardiovascular problems with frequent skin and mucosal manifestations, and without intensive treatment they can be fatal. The exact causes of PIMS are recently unknown, however, it is explained as hyperactivation of immunity.In this minireview, we summarize data on the prominent role of the IL-6-IL-6R-STAT3 axis in PIMS aetiopathogenesis. Therapeutic manipulation of IL-6 or IL-6 receptor could be an approach to the treatment of children with severe PIMS.

Predictors of exercise capacity in patients with atrial correction of transposition of great arteries.

OBJECTIVES: To examine the relationship of clinical, biochemical and imaging parameters to maximum oxygen uptake in patients after atrial correction of transposition of great arteries. BACKGROUND: Exercise tolerance is a key determinant of quality of life in patients with adult congenital heart disease. It is determined by a large scale of factors often different from general cardiology. METHODS: 86 consecutive patients after Senning correction of TGA were subjected to clinical and echocardiographic examination, Holter monitoring, blood tests of NT-proBNP, MRI of the heart and exercise test. Parameters of these examinations were correlated to VO2 max. RESULTS: The average age of patients was 28±3.5 years. The average systemic right ventricular function determined by MRI was 51.9±7.9 %. The average NT-proBPN was 124.3±23.6 ng/l, VO2 max. 31.7±6.5    ml/kg/min and the heart rate reserve 106±24 /min. Neither systemic right ventricular systolic function nor   NT-proBPN predicted VO2 max., whereas the heart rate reserve did (p=0.003). CONCLUSION: An inability to increase heart rate during exercise noted in a considerable number of patients after atrial switch of TGA caused a decreased exercise tolerance. It is not solely the global systolic function of either ventricle that influences the exercise performance, rather it is the ability to increase heart rate and overall cardiac output appropriately (Tab. 3, Fig. 6, Ref. 28).

Creatine Kinase and Myoglobin Levels as Indicators of Perioperative Muscle Damage during Open- and Mini-Invasive Stabilization of Thoracic and Lumbar Spine Fracture - a Prospective Randomized Study.

PURPOSE OF THE STUDY In this randomized prospective study, we monitored and compared perioperative changes in skeletal muscle enzymes blood levels in open and mini-invasive stabilization of thoracolumbar spine fractures. The established hypothesis was to confirm higher blood levels of muscle enzymes in open stabilization. MATERIAL AND METHODS This study included 38 patients with the mean age of 46.4 years. 19 injuries were managed in an open procedure and 19 procedures were mini-invasive. Venous blood was taken intermittently at short intervals to determine the levels of skeletal muscle enzymes. The catalytic concentration of creatine kinase was determined via an enzymatic UV-test, and the concentration of myoglobin via electro-chemiluminescent immunoassay. Enzyme levels were processed statistically. The Wilcoxon test was used. RESULTS The median increase in the values of both enzymes is higher in the mini-invasive method than in the open method in both the surgery phase for the injury and in the extraction phase. The median increase in the values of both enzymes is higher in both methods for the primary procedure phase compared to the extraction phase. All results are statistically significant at p of <0.05. All tests were calculated using the MATLAB Statistics Toolbox. DISCUSSION A very surprising finding, when testing the hypothesis of the levels increasing mainly in open stabilization, was confirming the opposite. Both enzymes were higher in the mini-invasive approach to stabilising the spine after the injury, but also after the extraction. This contradicts the available literature. However, this can be explained by the methodology of enzyme levels determination in the previously published studies. We believe that this phenomenon can be partially caused by an iatrogenic mini-compartment of muscles in the postoperative period, absence of wound drainage, but also by higher muscle contusion when inserting bolts through the tubes via small incisions, when the tubes penetrate to the entry points relatively violently and the muscles in this area are affected more than in the classical skeletization. CONCLUSIONS Analysis of biochemical changes in open and mini-invasive surgery did not confirm the hypothesis that levels of creatine kinase and myoglobin enzymes increase especially in open stabilization. On the contrary, they were statistically significantly higher in mini-invasive procedures. Key words: creatine kinase, myoglobin, muscle enzymes, spine fracture, spine surgery, miniinvasive surgery.

[Outcomes of Treatment of Calcaneal Fractures Using Locking Compression Plate with or without Augmentation]. / Výsledky lécení zlomeniny patní kosti LCP dlahou s augmentací a bez augmentace.

PURPOSE OF THE STUDY The retrospective study aims to compare the outcomes of augmentation of neutral triangle formed after the calcaneal fracture reduction with osteosynthesis using the locking compression plate with the outcomes of osteosynthesis without augmentation. MATERIAL AND METHODS In the period from 2005 to January 2011, 98 patients with 114 calcaneal fractures were treated by open reduction and internal fixation (ORIF) method with the use of calcaneal plate. The group included 15 women (15.3%) and 83 men (84.7%), with the mean age of 39.2 years (12-62 years). There were 16 cases of bilateral calcaneal fractures, of which 2 in women (12.5%) and 14 in men (87.5%). The fractures were classified based on the Sanders classification as Type I -IV. The patients with Type II and III fracture according to the Sanders classification were indicated for surgical treatment with ORIF using the calcaneal LCP. The defect in the region of neutral triangle of the calcaneus was filled with the injectable hydroxyapatite cement. The cohort of operated patients was evaluated based on the AOFAS (American Orthopaedic Foot and Ankle Society) Clinical Rating System. The bilateral fractures were not evaluated. This group shows a substantially higher occurrence of associated injuries, which causes strong distortion of results. RESULTS The cohort of 82 operated patients with Sanders Type II and III fractures were evaluated. In 20 fractures (24.4%) the defect in the calcaneus body was filled with hydroxyapatite cement. In osteosynthesis without defect augmentation an excellent result was achieved in 21 patients (33.9%). A good result was reported in 24 patients (38.7%), a satisfactory result in 12 patients (19.4%) and a poor result in 5 patients (8.0%). In patients with osteosynthesis of the calcaneus with augmentation, an excellent result was achieved in 6 cases (30%), a good result in 5 cases (25%), a satisfactory result in 7 cases (35%) and a poor result in 2 cases (10%). Early postoperative complications were observed in a total of 13 patients (15.8%). In osteosynthesis without defect augmentation, there were 2 cases of wound dehiscence (3.2%), 3 cases of marginal skin necrosis (4.8%), 4 cases of superficial wound infection (6.4%) and 1 case of deep wound infection (1.6%). In patients with performed augmentation, wound dehiscence was seen in 1 case (5%), superficial wound infection in 1 case (5%) and superficial marginal skin necrosis in 1 case (5%). Deep wound infection was not reported in this group. DISCUSSION A question frequently discussed is the filling of defect in the diaphysis of calcaneus. Brodt et al. state a statistically higher stability of the calcaneus in osteosynthesis with augmentation, but he does not use the locking plate. Longino in his study compares the results of calcaneal osteosynthesis with spongioplasty with a graft from pelvis and without it and does not observe any major differences in the final outcome in his cohort. Elsner evaluates the results in 18 patients, in whom calcium phosphate cement augmentation was used for osteosynthesis. Over the period of three years he did not observe a higher rate of complications. Schildhauer assesses the early load of the calcaneus with tricalcium phosphate cement augmentation. After three weeks of full loading no loss of reduction was found. Thordarson evaluates 11 operated patients with Sanders type II and III calcaneal fracture. The defect of the calcaneus was filled by calcium phosphate cement. In this small group 1 case of serious infection complication and 1 loss of reduction with full load at six weeks after the surgery were observed. In our group no differences were observed between the outcomes of patients with performed or not performed defect augmentation in the neutral triangle site. A higher percentage of complications in patients with filled defect of the calcaneus was not observed either. CONCLUSIONS The operative treatment of displaced intra-articular fractures consisting of open reduction from extended lateral approach and internal calcaneal LCP fixation brings good results. We concluded that there is no statistically significant difference in the results of osteosynthesis with a locking plate alone and combined with augmentation of diaphyseal defect of the calcaneus. Filling of the diaphyseal defect in the calcaneus is not necessary, it neither accelerates the healing, nor brings better treatment outcomes Key words: calcaneal fracture, cement augmentation of defect, locking plate, extended lateral approach.

Stable COX17 Downregulation Leads to Alterations in Mitochondrial Ultrastructure, Decreased Copper Content and Impaired Cytochrome c Oxidase Biogenesis in HEK293 Cells.

Cox17 is an assembly factor that participates in early cytochrome c oxidase (COX, CcO) assembly stages. Cox17 shuttles copper ions from the cytosol to the mitochondria and, together with Sco1 and Sco2, provides copper ions to the Cox1 and Cox2 mitochondrially encoded subunits. In Saccharomyces cerevisiae, Cox17 also modulates mitochondrial membrane architecture due to the interaction of Cox17 with proteins of the MICOS complex (mitochondrial contact site and cristae organizing system). There is currently no data regarding the impact of long-term Cox17 deficiency in human cells. Here, we present construction and characterization of three stable COX17 shRNA-downregulated HEK293 cell lines that have less than 10 % of the residual Cox17 protein level. Cox17-depleted cell lines exhibited decreased intramitochondrial copper content, decreased CcO subunit levels (Cox1, Cox4 and Cox5a) and accumulation of CcO subcomplexes. Similarly to yeast cells, mitochondria in Cox17-downregulated HEK293 cell lines exhibited ultrastructural changes including cristae reduction and mitochondrial swelling. Characterization of the molecular pathogenesis of long-term Cox17 deficiency complements our knowledge of the mitochondrial copper metabolism and assembly of cytochrome c oxidase in human cells.

Effect of shock waves combined with cytostatics on the growths of tumors in vivo.

Based on their field of application, the physical parameters of shock waves differ. Experiments referred to in this article used tandem shock waves generated on the surface of a composite anode. There, individual pores of the anode produce multichannel discharges. The composite anode may have a variety of shapes, which, consequently, influence the arrangement of the entire apparatus and the area of their application. Experiments referred to in this article utilise an anode divided into two parts that generated tandem shock waves. The previously conducted experiments have clearly shown that the effect of a tandem shock wave can be very well localized in the focal area, causing necrosis and apoptosis of the tumor cells, and enhancing the effect of cytostatics. This study investigated the effect of tandem shock waves with concomitantly administered cytostatics. We conducted our experiments on Lewis rats. The rats were injected with syngeneic sarcoma tumor cells intradermally and caudally on both the right and left sides. The highest rate of tumor growth inhibition was observed in the cisplatin-treated group that was subsequently treated with shock waves. The effect of shock waves on cell membranes is well described as they increase their permeability due to sonodynamic effect induced by cavitation. The results of experiments referred to in this article conducted in vivo in experimental animals enable us to note that the shock wave increases the effect of chemotherapy administered.

[Verification of the Necessity for the Open Reduction and Tension Band Wiring of the Displaced Olecranon Fracture in Younger Children]. / Overení nutnosti otevrené repozice a tahové cerkláze u dislokované zlomeniny okovce u mensích detí.

PURPOSE OF THE STUDY Olecranon fractures in skeletally immature patients are rather rare and represent up to 7% of elbow skeletal injuries. Although the majority of olecranon fractures is constituted by undisplaced fractures treated conservatively with good outcomes, a few of them require surgery. The aim of the study was to compare two different approaches of surgical treatment - the open reduction with tension band wiring - cerclage (ORCe) and the closed reduction and percutaneous pinning (CRPP). MATERIAL AND METHODS 37 patients (28 boys, 9 girls) were included in the retrospective multicentric study. The patients were treated at two different institutions (the Clinic of Paediatric Surgery, Orthopaedics and Traumatology, the University Hospital Brno and the Department of Paediatric Surgery and Traumatology, the University Hospital Hradec Králové, Czech Republic). 17 patients underwent the ORCe procedure, while 20 patients were treated using the CRPP method. Different parameters were statistically compared in the groups (demographic data, data concerning the course of the therapy, outcome of the therapy in terms of movements' restrictions and complications such as osteosynthesis failure and infection). RESULTS Both the groups were comparable in terms of demographic data because no statistically significant difference was observed in terms of the age (p = 0.082), the affected site (p = 1.000) and the gender (p = 0.462). Statistically significant difference between these two groups was found in the interval between the implementation and the removal of the osteosynthetic material (p < 0.001) and in the length of cast immobilisation (p = 0.047). The number of patients with movement restriction up to 10° was statistically significantly higher in patients who underwent the CRPP procedure (p = 0.040), but no statistically significant difference was seen between these two groups in terms of movement restriction more than 10° (p = 0.609). One revision surgery was performed in the CRPP group, however with no statistical significance (p = 0.350). DISCUSSION The multicentric study included 37 children, who underwent surgical treatment of a displaced olecranon fracture. Compared to the studies dealing with this topic the number of patients included in this study is relatively high. It advocates the possibility of using the CRPP method as a good alternative to the gold-standard ORCe technique because no difference in terms of the number of revision surgeries and the clinically important movement restriction of more than 10° were seen. It also brings along advantages such as a simple surgical technique, good functional and cosmetic effects, reducing the risk of ischemic insult of growth plate and the possibility of osteosynthetic material removal at an outpatient department with no need for general anaesthesia. The risk of a higher radiation exposure of both the patient and the surgical team should be considered as a disadvantage of the closed method. CONCLUSIONS The mini-invasive CRPP appears to be a good alternative option to the ORCe method for the treatment of isolated olecranon fractures in children offering the advantages such as avoiding extensive open procedure and simple implants removal. Key words:children, olecranon, fracture, tension band wiring - cerclage, percutaneous pinning, elbow.

[Comparison of Two Methods of Minimally Invasive Osteosynthesis for Proximal Radius Fractures in Paediatric Patients]. / Porovnání dvou metod miniinvazivní osteosyntézy u zlomeniny proximálního konce vretenní kosti v detském veku.

PURPOSE OF THE STUDY The presented study was construed as a retrospective multicentric clinical study focused on paediatric skeletal injuries of the proximal radius. As a general rule, the Type I displaced fractures (Judet classification) are treated conservatively, with no reduction. In the case of Type II-IV displacement, the fracture necessitates reduction or is also transfixed by a Kirschner wire (K-wire) or a Prevot nail (P-nail) where subsequent fragment instability occurs. The comparison aimed to ascertain whether there is a statistically significant difference between the two methods. No difference was expected by the authors, therefore a null hypothesis was set. MATERIAL AND METHODS The patients were treated at the Clinic of Paediatric Surgery, Orthopaedics and Traumatology (CPSOT) of the Faculty of Medicine of the Masaryk University and at the Clinic of Orthopaedics and Traumatology of the Musculoskeletal System of the University Hospital in Pilsen in the period from 2006 to 2015. Two methods of closed reduction and minimally- invasive osteosynthesis were evaluated. The first method was the elastic stable intramedullary nailing (ESIN) with a P-nail, the second method was an osteosynthesis using a K-wire. In the clinical part of the study, comparisons were made based on the monitoring of the same parameters - final restriction of movement, time to full weight bearing of the extremity and incidence of serious complications. RESULTS The final group comprised a total of 31 patients, of whom 7 boys and 24 girls aged 3-16 years with the median of 9-10 years. Some restriction of movement following the treatment occurred in a total of seven patients (44%) with the K-wire and in four patients (27%) with the P-nail. When comparing the movement at 5% level of significance using the Chi-Square tests, no significant difference was found (p = 0.446). When evaluating the serious complications at 5% level of significance using the Chi-Square tests, the difference between the two methods of treatment was again insignificant (p = 0.365). When the full weight bearing was compared (median K-wire 8 weeks, median P-nail 10 weeks), a statistically significant difference was obtained at 5% level of significance using the Fischer exact test (p = 0.003). DISCUSSION In these fractures, usually the metaphysis or the physis are involved in the injury, in which case the fractures are classified according to Salter and Harris, with the most frequent occurrence of SHII epiphyseal separation and rare SHIII and SHIV epiphyseal fractures. The radial head fractures are mostly caused by valgus force. Therefore, the individuals with a higher elbow valgosity are more prone to injuries. The girls strongly prevail (77%) also in our study. In general, our results as well as the literature have proven that as the displacement increases, the necessity of closed reduction and osteosynthesis grows. The post-treatment complications in our group were observed in 35.5% of patients, namely most often in the form of limited movement. The literature refers to complications in 26.5-53% of patients. CONCLUSIONS The clinical results clearly show that when comparing the complications after the radial head fracture in children there is no statistically significant difference between the methods of osteosynthesis. This study shall serve as a starting point for the currently ongoing prospective multicentric study evaluating the modified ESIN technique using a pre-bent Kirschner wire. Key words:children, fracture, radius, proximal, osteosynthesis.

Buccal Respiratory Chain Complexes I and IV Quantities in Huntington's Disease Patients.

Alterations in mitochondrial parameters are an important hallmark of Huntington's disease (HD). The ubiquitous expression of mutant huntingtin raises the prospect that mitochondrial disturbances can also be detected and monitored through buccal epithelial cells. In a group of 34 patients with Huntington's disease and a group of 22 age-related healthy volunteers, respiratory complex I and IV protein quantities in buccal epithelial cells were measured using the dipstick immunocapture assay. The protein quantity of respiratory complex I correlates with age (r = 0.427, P = 0.026, FWE-P = 0.156) in the patient group, but not in the group of healthy subjects. Our non-invasive approach allows us to obtain valuable information for the studies of mitochondrial biochemical parameters in patients with neurodegenerative diseases and could also be useful in epidemiological studies.

Changes in transcription pattern lead to a marked decrease in COX, CS and SQR activity after the developmental point of the 22(nd) gestational week.

Tissue differentiation and proliferation throughout fetal development interconnect with changes in the oxidative phosphorylation system (OXPHOS) on the cellular level. Reevaluation of the expression data revealed a significant increase in COX4 and MTATP6 liver transcription levels after the 22(nd) gestational week (GW) which inspired us to characterize its functional impact. Specific activities of cytochrome c oxidase (COX), citrate synthase (CS), succinate-coenzyme Q reductase (SQR) and mtDNA determined by spectrophotometry and RT-PCR were studied in a set of 25 liver and 18 skeletal muscle samples at 13(th) to 29(th) GW. Additionally, liver hematopoiesis (LH) was surveyed by light microscopy. The mtDNA content positively correlated with the gestational age only in the liver. The activities of COX, CS and SQR in both liver and muscle isolated mitochondria significantly decreased after the 22(nd) GW in comparison with earlier GW. A continuous decline of LH, not correlating with the documented OXPHOS-specific activities, was observed from the 14(th) to the 24(th) GW indicating their exclusive reflection of liver tissue processes. Two apparently contradictory processes of increasing mtDNA transcription and decreasing OXPHOS-specific activities seem to be indispensable for rapid postnatal adaptation to high energy demands. The inadequate capacity of mitochondrial energy production may be an important factor in the mortality of children born before the critical developmental point of the 22(nd) GW.

[Clinical Results of Endoscopic Treatment of Greater Trochanteric Pain Syndrome]. / Výsledky endoskopické terapie bolestivého syndromu velkého trochanteru.

PURPOSE OF THE STUDY This retrospective study aims to present short-term clinical outcomes of endoscopic treatment of patients with greater trochanteric pain syndrome (GTPS). MATERIAL AND METHODS The evaluated study population was composed of a total of 19 patients (16 women, 3 men) with the mean age of 47 years (19-63 years). In twelve cases the right hip joint was affected, in the remaining seven cases it was the left side. The retrospective evaluation was carried out only in patients with greater trochanteric pain syndrome caused by independent chronic trochanteric bursitis without the presence of m. gluteus medius tear not responding to at least 3 months of conservative treatment. In patients from the followed-up study population, endoscopic trochanteric bursectomy was performed alone or in combination with iliotibial band release. The clinical results were evaluated preoperatively and with a minimum follow-up period of 1 year after the surgery (mean 16 months). The Visual Analogue Scale (VAS) for assessment of pain and WOMAC (Western Ontario MacMaster) score were used. In both the evaluated criteria (VAS and WOMAC score) preoperative and postoperative results were compared. Moreover, duration of surgery and presence of postoperative complications were assessed. Statistical evaluation of clinical results was carried out by an independent statistician. In order to compare the parameter of WOMAC score and VAS pre- and post-operatively the Mann-Whitney Exact Test was used. The statistical significance was set at 0.05. RESULTS The preoperative VAS score ranged 5-9 (mean 7.6) and the postoperative VAS ranged 0-5 (mean 2.3). The WOMAC score ranged 56.3-69.7 (mean 64.2) preoperatively and 79.8-98.3 (mean 89.7) postoperatively. When both the evaluated parameters of VAS and WOMAC score were compared in time, a statistically significant improvement (p<0.05) was achieved postoperatively. The mean duration of surgical procedure was 68 minutes. Moreover, in peritrochanteric space apart from chronic bursitis also another pathology was found in a total of 14 cases (74%). In six cases (32%) it was a mild degeneration of m. gluteus medius (treated only with debridement) and in eight patients who underwent surgery (42%) the dorsal third of tractus iliotibialis was hypertrophic and protruding into bursa (treated with an incision of the hypertrophied part of the band). No serious neurovascular or thromboembolic complications were recorded. Only minor postoperative complications in a total of 7 patients who underwent surgery (37%) occurred. DISCUSSION There are lots of studies in literature presenting the results of endoscopic treatment of GTPS either using an independent trochanteric bursectomy or its combination with iliotibial band release. In our study we succeeded in achieving similar clinical results as those achieved by the other authors engaged in this area. CONCLUSIONS It has been proven in this retrospective study that the technique of endoscopic trochanteric bursectomy in patients with greater trochanteric pain syndrome yields statistically significant improvement of clinical results with the concurrent minimum incidence of postoperative complications. Key words: greater trochanteric pain syndrome, peritrochanteric space, recalcitrant trochanteric bursitis, hip arthroscopy, endoscopic trochanteric bursectomy, iliotibial band release.

Molecular Diagnostics of Copper-Transporting Protein Mutations Allows Early Onset Individual Therapy of Menkes Disease.

Menkes disease is a severe X-linked recessive disorder caused by a defect in the ATP7A gene, which encodes a membrane copper-transporting ATPase. Deficient activity of the ATP7A protein results in decreased intestinal absorption of copper, low copper level in serum and defective distribution of copper in tissues. The clinical symptoms are caused by decreased activities of copper-dependent enzymes and include neurodegeneration, connective tissue disorders, arterial changes and hair abnormalities. Without therapy, the disease is fatal in early infancy. Rapid diagnosis of Menkes disease and early start of copper therapy is critical for the effectiveness of treatment. We report a molecular biology-based strategy that allows early diagnosis of copper transport defects and implementation of individual therapies before the full development of pathological symptoms. Low serum copper and decreased activity of copperdependent mitochondrial cytochrome c oxidase in isolated platelets found in three patients indicated a possibility of functional defects in copper-transporting proteins, especially in the ATPA7 protein, a copper- transporting P-type ATPase. Rapid mutational screening of the ATP7A gene using high-resolution melting analysis of DNA indicated presence of mutations in the patients. Molecular investigation for mutations in the ATP7A gene revealed three nonsense mutations: c.2170C>T (p.Gln724Ter); c.3745G>T (p.Glu1249Ter); and c.3862C>T (p.Gln1288Ter). The mutation c.3745G>T (p.Glu1249Ter) has not been identified previously. Molecular analysis of the ATOX1 gene as a possible modulating factor of Menkes disease did not reveal presence of pathogenic mutations. Molecular diagnostics allowed early onset of individual therapies, adequate genetic counselling and prenatal diagnosis in the affected families.

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.

Genotype-phenotype correlation in 44 Czech, Slovak, Croatian and Serbian patients with mucopolysaccharidosis type II.

Mucopolysaccharidosis type II (Hunter syndrome, MPS II, OMIM 309900) is an X-linked lysosomal storage disorder caused by deficiency of iduronate-2-sulfatase (IDS). We analyzed clinical and laboratory data from 44 Slavic patients with this disease. In total, 21 Czech, 7 Slovak, 9 Croatian and 7 Serbian patients (43 M/1 F) were included in the study (median age 11.0 years, range 1.2-43 years). Birth prevalence ranged from 1:69,223 (Serbia) to 1:192,626 (Czech Rep.). In the majority of patients (71%), the disease manifested in infancy. Cognitive functions were normal in 10 patients. Four, six and 24 patients had mild, moderate, and severe developmental delay, respectively, typically subsequent to developmental regression (59%). Residual enzyme activity showed no predictive value, and estimation of glycosaminoglycans (GAGs) had only limited importance for prognosis. Mutation analysis performed in 36 families led to the identification of 12 novel mutations, eight of which were small deletions/insertions. Large deletions/rearrangements and all but one small deletion/insertion led to a severe phenotype. This genotype-phenotype correlation was also identified in six cases with recurrent missense mutations. Based on patient genotype, the severity of the disease may be predicted with high probability in approximately half of MPS II patients.

[Traumatic Thrombosis of the Renal Artery following Seat-Belt Injury]. / Traumatická trombóza renální tepny pri seat-belt injury.

Renal artery thrombosis is a rare complication of blunt abdominal injury. It occurs most frequently in car accidents in which sudden deceleration results in multiple internal injuries. Renal artery occlusion occurs rarely as an isolated blunt trauma. This report presents a case of traumatic occlusion of the left renal artery with complete ischaemia of the left kidney in a haemodynamically stable girl who suffered multiple seat-belt injuries in a car crash. Ambiguous findings on a CT scan of the abdominal cavity indicated surgical inspection of the abdomen and, since the injury-to-surgery interval was short, an attempt at revascularisation of the left kidney was made. This was performed using laparotomy through a lateral left-side incision, as an uncommon approach to the injured organs. A seat-belt fracture of the L2/3 spine was stabilised at secondstage surgery. Key words: renal artery thrombosis, seat-belt fracture of the lumbar spine.

The phenotypic spectrum of fifty Czech m.3243A>G carriers.

BACKGROUND: Mitochondrial myopathy, Encephalopathy, Lactic Acidosis and Stroke-like episodes syndrome (MELAS) is a common mitochondrial disorder with varying multisystemic clinical manifestation. We present a comprehensive clinical picture of 50 Czech m.3243A>G carriers with emphasis on the sequence of symptoms in symptomatic patients. RESULTS: Symptoms developed in 33 patients (66%) and 17 carriers remained unaffected (34%). The age of onset varied from 1month to 47years of age, with juvenile presentation occurring in 53% of patients. Myopathy was the most common presenting symptom (18%), followed by CPEO/ptosis and hearing loss, with the latter also being the most common second symptom. Stroke-like episodes (SLE) occurred in fourteen patients, although never as a first symptom, and were frequently preceded by migraines (58%). Rhabdomyolysis developed in two patients. The second symptom appeared 5.0±8.3years (range 0-28years) after the first, and the interval between the second and third symptom was 2.0±6.0years (range 0-21years). Four of our patients remained monosymptomatic up to 12years of follow-up. The sequence of symptoms according to their time of manifestation was migraines, myopathy, seizures, CPEO/ptosis, SLE, hearing loss, and diabetes mellitus. The average age at death was 32.4±17.7years (range 9-60years) in the juvenile form and 44.0±12.7years (range 35-53years) in the adult form. Some patients with SLE harboured very low heteroplasmy levels in various tissues. No threshold for any organ dysfunction could be determined based on these levels. CONCLUSIONS: Sufficient knowledge of the timeline of the natural course of MELAS syndrome may improve the prediction and management of symptoms in patients with this mitochondrial disease.

X-Chromosome Inactivation Analysis in Different Cell Types and Induced Pluripotent Stem Cells Elucidates the Disease Mechanism in a Rare Case of Mucopolysaccharidosis Type II in a Female.

Mucopolysaccharidosis type II (MPS II) is an X-linked lysosomal storage disorder resulting from deficiency of iduronate-2-sulphatase activity. The disease manifests almost exclusively in males; only 16 symptomatic heterozygote girls have been reported so far. We describe the results of X-chromosome inactivation analysis in a 5-year-old girl with clinically severe disease and heterozygous mutation p.Arg468Gln in the IDS gene. X inactivation analysed at three X-chromosome loci showed extreme skewing (96/4 to 99/1) in two patient's cell types. This finding correlated with exclusive expression of the mutated allele. Induced pluripotent stem cells (iPSC) generated from the patient's peripheral blood demonstrated characteristic pluripotency markers, deficiency of enzyme activity, and mutation in the IDS gene. These cells were capable of differentiation into other cell types (cardiomyocytes, neurons). In MPS II iPSC clones, the X inactivation ratio remained highly skewed in culture conditions that led to partial X inactivation reset in Fabry disease iPSC clones. Our data, in accordance with the literature, suggest that extremely skewed X inactivation favouring the mutated allele is a crucial condition for manifestation of MPS II in females. This suggests that the X inactivation status and enzyme activity have a prognostic value and should be used to evaluate MPS II in females. For the first time, we show generation of iPSC from a symptomatic MPS II female patient that can serve as a cellular model for further research of the pathogenesis and treatment of this disease.

Analysis of expression profiles of genes involved in F(o)F(1)-ATP synthase biogenesis during perinatal development in rat liver and skeletal muscle.

During the process of intra-uterine mammalian fetal development, the oxygen supply in growing fetus is low. A rapid switch from glycolysis-based metabolism to oxidative phosphorylation (OXPHOS) must proceed during early postnatal adaptation to extra-uterine conditions. Mitochondrial biogenesis and mammalian mitochondrial F(o)F(1)-ATP synthase assembly (complex V, EC 3.6.3.14, ATPase) are complex processes regulated by multiple transcription regulators and assembly factors. Using RNA expression analysis of rat liver and skeletal tissue (Rattus norvegicus, Berkenhout, 1769), we describe the expression profiles of 20 genes involved in mitochondrial maturation and ATP synthase biogenesis in detail between the 16th and 22nd day of gestation and the first 4 days of life. We observed that the most important expression shift occurred in the liver between the 20th and 22nd day of gestation, indicating that the fetus prepares for birth about two days before parturition. The detailed mechanism regulating the perinatal adaptation process is not yet known. Deeper insights in perinatal physiological development will help to assess mitochondrial dysfunction in the broader context of cell metabolism in preterm newborns or neonates with poor adaptation to extra-uterine life.

[Augmentation of the Anterior Cruciate Ligament in Patients with Symptomatic Isolated Tear of Anteromedial or Posterolateral Bundle: Evaluation of Two-Year Clinical Results]. / Augmentace predního zkrízeného vazu u pacientu se symptomatickou izolovanou rupturou anteromediální nebo posterolaterální porce: zhodnocení dvouletých klinických výsledku.

PURPOSE OF THE STUDY: We present the results of a prospective study of patients with symptomatic partial ACL tears comparing the pre-operative findings with the clinical results at two years after anterior cruciate ligament (ACL) augmentation. MATERIAL AND METHODS: A total of 29 patients (7 women, 22 men; average age, 27.8 years) who were diagnosed with an isolated tear of the posterolateral (PL) bundle (18 patients) or the anteromedial (AM) bundle (11 patients) at 9 to 24 weeks after injury, and underwent ACL augmentation by isolated PL or AM bundle replacement, were evaluated. The characteristics assessed before surgery and at two years after it included the Lysholm and subjective IKDC scores and knee laxity measurement with a GNRB arthrometer (at 134 N and 250 N) and its assessment by the Lachman, anterior drawer and pivot shift tests. In addition, the presence of cyclop syndrome, graft failure and post-operative complications were evaluated. RESULTS: The patients with isolated reconstruction of the PL bundle showed post-operatively statistically significant improvement in the degree of rotational knee laxity (p < 0.05) and the ventral knee laxity assessed by the Lachman test (p < 0.05). Postoperative improvement in the anterior drawer test results was not statistically significant (p = 0.07). The median value of side-to-side difference in knee laxity measured with the GNRB arthrometer decreased at 134 N from 1.7 mm pre-operatively to 0.8 mm at two years post-operatively (p < 0.05) and, at 250 N, from 2.8 mm to 1.5 mm (p < 0.05). The median Lysholm score increased from 74 to 91 points at two post-operative years and the median IKDC score improved from 76 to 92 points (p < 0.05). Graft failure was reported in one patient (5.6%) and 14 subjects (77.8%) reported return to pre-injury sports activities. The patients undergoing isolated reconstruction of the AM bundle achieved, at two years after surgery, a statistically significant decrease in positivity of the Lachman and anterior drawer tests (p < 0.05), while the results of the pivot shift test did not improve significantly (p = 0.09). The decrease in median values of side-to-side difference in knee laxity measured with the GNRB arthrometer was from pre-operative 3.1 mm to 1.2 mm at 134 N (p < 0.05) and from 6.2 mm to 1.9 mm at 250N (p < 0.05). The median Lysholm and IKDC scores increased from 68 to 92 points and from 70 to 94 points, respectively (p < 0.05). Nine patients (81.9%) reported return to pre-injury participation in sports. Apart from early wound bleeding in one patient, no complications were recorded. DISCUSSION: By permitting maintenance of a healthy bundle and replacement of only a torn one, ACL augmentation provides several benefits. It allows for accelerated revascularization and re-innervation of the graft through mechanoreceptors of the healthy portion; it enables the surgeon to get a good anatomical orientation and achieve precise tunnel reaming; in addition the healthy bundle provides protection for the graft in the early post-operative period. Thus rehabilitation can be faster and also return to sports activities. CONCLUSIONS: Our results show that ACL augmentation using isolated replacement of either the AM or the PL bundle brings about statistically significant improvement of all subjective and most of the objective criteria by two years after surgery.