Development and Validation of a Short Version of the Metric for the Observation of Decision-Making in Multidisciplinary Tumor Boards: MODe-Lite.

BACKGROUND: Evidence-based tools are necessary for scientifically improving the way MTBs work. Such tools are available but can be difficult to use. This study aimed to develop a robust observational assessment tool for use on cancer multidisciplinary tumor boards (MTBs) by health care professionals in everyday practice. METHODS: A retrospective cross-sectional observational study was conducted in the United Kingdom from September 2015 to July 2016. Three tumor boards from three teaching hospitals were recruited, with 44 members overall. Six weekly meetings involving 146 consecutive cases were video-recorded and scored using the validated MODe tool. Data were subjected to reliability and validity analysis in the current study to develop a shorter version of the MODe. RESULTS: Phase 1, a reduction of the original items in the MODe, was achieved through two focus group meetings with expert assessors based on previous research. The 12 original items were reduced to 6 domains, receiving full agreement by the assessors. In phase 2, the six domains were subjected to item reliability, convergent validation, and internal consistency testing against the MODe-Lite global score, the MODe global score, and the items of the MODe. Significant positive correlations were evident across all domains (p < 0.01), indicating good reliability and validity. In phase 3, feasibility and high inter-assessor reliability were achieved by two clinical assessors. Six domains measuring clinical input, holistic input, clinical collaboration, pathology, radiology, and management plan were integrated into MODe-Lite. CONCLUSIONS: As an evidence-based tool for health care professionals in everyday practice, MODe-Lite gives cancer MTBs insight into the way they work and facilitates improvements in practice.

Analysing Breast Cancer Multidisciplinary Patient Management: A Prospective Observational Evaluation of Team Clinical Decision-Making.

BACKGROUND: Multidisciplinary team (MDT)-driven cancer care is a mandatory UK national policy, widely used globally. However, few studies have examined how MDT members make decisions as a team. We report a single-centre prospective study on team working within breast cancer MDT. METHODS: This was a prospective observational study of 10 breast MDT meetings (MDM). Trained clinical observer scored quality of presented information and disciplinary contribution to case reviews in real time, using a validated tool, namely Metric for the Observation of Decision-Making. Data were analysed to evaluate quality of team working. RESULTS: Ten MDMs were observed (N = 346 patients). An average of 42 patients were discussed per MDM (range: 29-51) with an average 3 min 20 s (range: 31 s-9 min) dedicated to each patient. Management decision was made in 99% of cases. In terms of contribution to case reviews, breast care nurses scored significantly (p < 0.05) lower (M = 1.79, SD = 0.12) compared to other team members (e.g. surgeons, M = 4.65; oncologists, M = 3.07; pathologists, M = 4.51; radiologists, M = 3.21). Information on patient psychosocial aspects (M = 1.69, SD = 0.68), comorbidities (M = 1.36, SD = 0.39) and views on treatment options (M = 1.47, SD = 0.34) was also significantly (p < 0.05) less well represented compared to radiology (M = 3.62, SD = 0.77), pathology (M = 4.42, SD = 0.49) and patient history (M = 3.91, SD = 0.48). CONCLUSION: MDT evaluation via direct observation in a meeting is feasible and reliable. We found consistent levels of quality of information coverage and contribution within the team, but certain aspects could be improved. Contribution to patient review resides predominantly with surgeons, while presented patient information is largely of biomedical nature. These findings can be fed to cancer MDTs to identify potential interventions for improvement.

Enzymatic Isolation, Amplification and Characterization of Dental Pulp Stem Cells.

The dental pulp represents an easily accessible source of adult dental pulp stem cells (DPSCs). The preferred approach to DPSC isolation is enzymatic digestion. However, the duration of the enzymatic activity is crucial. The purpose of this study was to isolate the DPSC populations using this method, characterize their biological properties and proliferation capacity, and to determine their ability to differentiate into mature cells. Before enzymatic digestion using 0.05% trypsin, we used the homogenization method in order to obtain a fine homogenate from the solid pulp tissue. The stem cells were cultivated in modified cultivation medium for mesenchymal adult progenitor cells containing 2% foetal bovine serum, growth factors and insulin-transferrin-selenium supplement. We were successfully able to isolate 10 populations of DPSCs. The vitality of DPSCs did not drop below 90 %. However, the DPSCs showed a significant decrease in the relative telomere length number with increasing passaging (P < 0.05). Isolated DPSCs highly expressed the CD markers: CD29, CD44, CD90, CD13, CD73 and CD166. In contrast, CD markers CD31, CD106, CD34 and CD45 were negative or low positive. We confirmed the high osteogenic and chondrogenic potential of the isolated stem cells. Isolated DPSCs did not show signs of cell degeneration or spontaneous differentiation during the entire cultivation. In addition, we were able to shorten the enzyme activity duration, and we were the first to demonstrate trypsin as the enzyme used for the enzymatic digestion method with the viability over 90 % of isolated DPSCs using this method.

The Differentiation Potential of Human Natal Dental Pulp Stem Cells into Insulin-Producing Cells.

Mesenchymal stem cells have the ability to differentiate into insulin-producing cells, raising the hope for diabetes mellitus treatment. The aim of this research was to study the ability of stem cells from discarded natal teeth to differentiate into insulinproducing cells. Two vital human natal teeth were obtained from a healthy 2-day-old female. Stem cells from the dental pulp were isolated, cultured under xenogenic-free conditions, propagated and characterized. Proliferative activity, population doubling time and viability were measured, and the multipotent differentiation ability was investigated. A twostep protocol was used to induce the human natal dental pulp stem cells to differentiate into insulinproducing cells. Phenotypic analysis was done using flow cytometry. Immunohistochemistry was performed to detect insulin and C-peptide. PDX1, HES1 and Glut2 gene expression analysis was performed by quantitative reverse transcription-polymerase chain reaction. Human natal dental pulp stem cells were able to undergo osteogenic, chondrogenic and adipogenic differentiation upon exposure to the specific differentiation media for each lineage. Their differentiation into insulin-producing cells was confirmed by expression of C-peptide and insulin, as well as by 975.4 % higher expression of PDX-1 and 469.5 % higher expression of HES1 in comparison to the cells cultivated in standard cultivation media. Glut2 transporter mRNA was absent in the non-differentiated cells, and differentiation of the stem cells into insulin-producing cells induced appearance of the mRNA of this transporter. We were the first to demonstrate that stem cells obtained from the pulp of natal teeth could be differentiated into insulinproducing cells, which might prove useful in the stem cell therapy for type 1 diabetes.

Single-agent cytarabine is insufficient for the treatment of human mantle cell lymphoma in mouse xenograft model.

Mantle cell lymphoma is an aggressive type of B-cell non-Hodgkin lymphoma with adverse prognosis. It was demonstrated that alternation of CHOP and DHAP chemotherapy improved outcome of mantle cell lymphoma patients. However, which components of DHAP, cisplatin, cytarabine, or both, were responsible for the improved outcome remained unclear. To answer this question, antitumor efficacies of equally toxic doses of cytarabine, cisplatin, and three different combinations were compared in vivo using mouse xenograft models of mantle cell lymphoma. We demonstrated that cisplatin, alone or with cytarabine, is significantly superior to single-agent cytarabine in both eliminating lymphoma cells and suppressing their proliferation rate.

Proliferative Capacity and Phenotypical Alteration of Multipotent Ecto-Mesenchymal Stem Cells from Human Exfoliated Deciduous Teeth Cultured in Xenogeneic and Allogeneic Media.

Foetal calf serum (FCS) is a standard supplement used in media for in vitro stem cell cultivation. This xenogeneic supplement remains widely used for its favourable growth-promoting properties and ease of accessibility; however, it is inherently not fit for human medicine due to its capacity to temper with the cultured cell quality. For this reason, the international community encourages research and development of allogeneic sera, which would expunge this issue. This study aims to investigate the differences in proliferative capacity, phenotype, and differentiation capacity of ecto-mesenchymal stem cells from human exfoliated deciduous teeth (SHED) cultured in vitro in media supplemented with allogeneic and xenogeneic sera. To address these aims, we cultured three lineages of stem cells in media supplemented with FCS in a concentration of 2% + growth factors; human blood plasma and platelet-rich plasma in concentrations of 2% + growth factors, and 10%. Here, the xenogeneic cultivation was considered as a basis for comparison because this serum is commonly used in studies concerning ecto-mesenchymal stem cells. The study shows that multipotent ecto-mesenchymal SHED can be feasibly cultivated in media where the xenogeneic FCS is substituted by allogeneic platelet-rich plasma, considering the cultured cell proliferative and differentiation capacities. We have also proved that different sera impact the cultured cells' phenotype differently, which has major implications for previous and future stem cell research and regenerative therapy.

N-3 polyunsaturated fatty acids supplementation does not affect changes of lipid metabolism induced in rats by altered thyroid status.

Epidemiological studies have demonstrated that n-3 polyunsaturated fatty acid (PUFA) consumption is associated with a reduced risk of atherosclerosis and hyperlipidemia. It is well known that lipid metabolism is also influenced by thyroid hormones. The aim of our study was to test whether n-3 PUFA supplementation (200 mg/kg of body weight/day for 6 weeks given intragastrically) would affect lipid metabolism in Lewis male rats with altered thyroid status. Euthyroid, hypothyroid, and hyperthyroid status of experimental groups was well defined by plasma levels of triiodothyronine, the activity of liver mitochondrial glycerol-3-phosphate dehydrogenase, and by relative heart weight. Fasting blood glucose levels were significantly higher in the hyperthyroid compared to the euthyroid and hypothyroid rats (5.0±0.2 vs. 3.7±0.4 and 4.4±0.2 mmol/l, respectively). In hyperthyroid animals, the concentration of plasma postprandial triglycerides was also increased compared to euthyroid and hypothyroid rats (0.9±0.1 vs. 0.5±0.1 and 0.4±0.1 mmol/l, respectively). On the other hand, hypothyroidism compared to euthyroid and hyperthyroid status was associated with elevated plasma levels of total cholesterol (2.6±0.2 vs. 1.5±0.1 and 1.6±0.1 mmol/l, respectively), LDL cholesterol (0.9±0.1 vs. 0.4±0.1 and 0.2±0.1 mmol/l, respectively) as well as HDL cholesterol (1.6±0.1 vs. 1.0±0.1 and 1.3±0.1 mmol/l, respectively). Supplementation of n-3 PUFA in the present study did not significantly modify either relative heart weight or glucose and lipid levels in any thyroid status.

In vivo growth of mantle cell lymphoma xenografts in immunodeficient mice is positively regulated by VEGF and associated with significant up-regulation of CD31/PECAM1.

Mantle cell lymphoma (MCL) is an aggressive lymphoma subtype with dismal prognosis. New treatments are needed to improve outcome of relapsed/ refractory disease. Recently, several drugs targeting at least partially the process of angiogenesis have been successfully tested in the therapy of MCL. Molecular mechanisms that regulate MCL-induced angiogenesis and that might represent potential new druggable targets remain, however, incompletely understood. We established two mouse models of human MCL by subcutaneous xenotransplantation of JEKO-1 and HBL-2 cell lines into immunodeficient mice. Histological analyses of xenografts confirmed their neovascularization. The growth of xenografts was significantly suppressed by single-agent therapy with bevacizumab, monoclonal antibody targeting vascular endothelial growth factor (VEGF). Subsequently, we analysed expression of 94 angiogenesis related genes in ex vivo isolated JEKO-1 and HBL-2 cells compared to in vitro growing cells using TaqMan low-density arrays. The most up-regulated genes in both JEKO-1 and HBL-2 xenografts were genes encoding platelet/endothelial cell-adhesion molecule (CD31/PECAM1), VEGF receptor 1 (FLT1), hepatocyte growth factor (HGF), angiogenin (ANG) and transcription factor PROX1. The most downregulated genes in both JEKO-1 and HBL-2 xenografts were midkine (MDK) and ephrine B2 (EPHB2). In summary, our results demonstrate an important role of angiogenesis in the biology of MCL and provide preclinical evidence of potent anti-MCL activity of bevacizumab. In addition, gene expression profiling of 94 angiogenesis-related targets revealed several in vivo up-regulated and down-regulated transcripts. The most differentially expressed target in both MCL tumours was CD31/PECAM1. Whether any of these molecules might represent a potential druggable target in MCL patients remains to be elucidated.

Ionizing radiation induces senescence and differentiation of human dental pulp stem cells.

Head and neck cancer is one of the most common cancers in Europe. Many current anti-cancer treatments, including ionizing radiation, induce apoptosis via DNA damage. Unfortunately, such treatments are non-selective to cancer cells and produce similar toxicity in normal cells, including adult stem cells. One of the fundamental properties of an adult stem cell is that it does not have any tissue-specific structures that allow it to perform specialized functions. However, under certain stimuli, unspecialized adult stem cells can give rise to specialized cells to generate replacements for cells that are lost during one's life or due to injury or disease. Nevertheless, specialization of stem cells must be controlled by specific milieu and also initiated at the proper time, making the entire process beneficial for tissue recovery and maintaining it for a long time. In this paper we assess whether irradiated dental pulp stem cells have maintained open their options to mature into specialized cells, or whether they have lost their unspecialized (immature) state following irradiation. Our findings showed radiation-induced premature differentiation of dental pulp stem cells towards odonto-/osteoblast lineages in vitro. Matrix calcification was visualized from Day 6 or Day 9 following irradiation of cells expressing low or high levels of CD146, respectively.

The response of human ectomesenchymal dental pulp stem cells to cisplatin treatment.

AIM: To determine the response of dental pulp stem cells (DPSCs) to DNA-damaging cytostatic cisplatin and compare it with the response of normal human dermal fibroblasts (HDFs). METHODOLOGY: Dental pulp stem cells were exposed to 5, 10, 20 or 40 µmol L(-1) of cisplatin. The proliferation of affected cells was assessed by a Z2 Counter and viability was assessed by means of a Vi-Cell XR using Trypan blue exclusion staining. Cell cycle analysis and induction of apoptosis were performed by flow cytometry. Induction of apoptosis was determined by monitoring the activities of caspases. The expression of proteins was detected by electrophoresis and Western blotting. The descriptive statistics of the results was analyzed by Student's t-test. RESULTS: Dental pulp stem cells had a greater genotoxic stress response to cisplatin compared to HDFs. All three main Mitogen-activated protein kinases (MAPK) families - extracellular signal-regulated kinases (ERK), c-Jun-N-terminal kinase (JNK) and p38 were activated after treatment of DPSCs with cisplatin. The activation of MAPK pathways was not observed in HDFs exposed to cisplatin. The exposure of DPSCs and HDFs to cisplatin provoked an increase in p53 and p21 expression and p53 phosphorylation of serine 15. Higher concentrations of cisplatin reduced the viability of DPSCs and HDFs and induced the activation of caspases 3/7 and 9. CONCLUSION: Dental pulp stem cells had a greater genotoxic stress response to cisplatin compared to HDFs. Cisplatin in higher concentrations triggered activation of MAPK and apoptosis in DPSCs but not in HDFs.

[Bone healing capacity in patients undergoing total hip arthroplasty]. / Schopnost kostní reparace u pacientu podstupujících implantaci TEP kycelního kloubu.

PURPOSE OF THE STUDY: To identify some characteristics of bone repair capacity in elderly patients who undergo total hip arthroplasty, which requires good healing ability of bone for implant osteointegration and bone defect repair, particularly if revision arthroplasty is necessary. MATERIAL AND METHODS: In a group of 27 patients (mean age, 70 ± 7 years; range, 60 to 81 years) a coincidence of osteoarthritis and osteopenia/ osteoporosis was assessed, and mesenchymal stem cells (MSC) were isolated and their numbers, viability and proliferative capacity were evaluated. The MSC populations were examined for their behaviour on bone tissue scaffolds used in orthopaedic surgery for treatment of bone lesions. Each patient underwent bone densitometry examination before total hip arthroplasty. Bone marrow was harvested intra-operatively from the trochanteric region of the femur. From a portion of bone marrow, MSCs were isolated and cultured, and a mononuclear cell concentrate was obtained. Either whole bone marrow or a mononuclear cell concentrate was applied to selected matrices (allograft, demineralised bone matrix, porous beta-tricalcium phosphate (-TCP), pressed hydroxyapatite or calcium sulphate). The production of new collagen and extracellular mineralized matrix were first assessed in expansion medium and, when the production was low, differentiation medium was used. RESULTS: A coincidence of osteoarthritis and osteopenia/osteoporosis was found in 50% of the patients. All were women with a low body mass index and had been post-menopausal for an average of 23 years. The isolated MSCs contained a high percentage of viable cells (mean, 95%). The mesenchymal cells of patients with osteopenia, as compared with those having normal bone density, showed markedly lower numbers of fibroblastic colony forming units (CFU-F) per ml and had a lower proliferative capacity because the population doubling time during the first four passages was much longer. Of the scaffolds tested, allografts showed the most marked collagen and extracellular mineralized matrix production in expansion medium with either whole bone marrow or a monocyte concentrate; porous -TCP was the best of bone graft substitutes in collagen and extracellular mineralized matrix production by both whole bone marrow and a monocyte concentrate, but this was only in differential medium. DISCUSSION: The coincidence of ostearthritis with osteopenia/osteoporosis was found in a higher number of our patients than is reported in the literature. Also, a lower MSC proliferative capacity and a low number of CFU-F/ml in the patients with low bone density were interesting findings. Better bone regeneration would generally be achieved with higher MSC numbers and the use of growth factors for stimulation of osteoinduction and angiogenesis. Bone marrow harvesting for MSC isolation, cultivation and subsequent transplantation is currently feasible only in an experiment. A bone marrow aspirate can be applied, but it may not provide a sufficient number of MSCs. In addition to autologous bone grafts, the best collagen production was on allografts. In bone graft substitutes, the porous structure played an important role because on a non-porous material (calcium sulphate) the formation of collagen was very low. There was no difference in collagen and extracellular mineralized matrix production between whole bone marrow and monocyte concentrates. CONCLUSIONS: Elderly patients have reduced bone healing capacity also because of osteopenia/osteoporosis that occurs more often than it is generally diagnosed, including its coincidence with osteoarthritis. The mesenchymal stem cells isolated from osteopenic bone give a lower number of CFU-F/ml and have a lower proliferative capacity. Of the matrices for new bone formation, allografts showed the best results because collagen was produced already in expansion medium. Of the graft substitutes, porous -TCP was the best, but with collagen production in differential medium. The use of bone marrow aspirate is currently a method of choice in order to increase MSC numbers at the site of bone healing. The use of growth factors is an expensive treatment. To achieve the goal of reliable promotion of osteogenesis with cultured MSC transplantation and use of composite materials with pro-osteogenic and pro-angiogenic factors will still require many experimental and clinical studies.

[Drug compliance - a pitfall of therapy of rheumatoid arthritis?]. / Léková compliance - úskalí terapie revmatoidní artritidy?

Drug compliance is one of the conditions for effective treatment of various chronic diseases, including rheumatoid arthritis as well. This disease is characterized by a variable course, remissions and relapses, and also by specific treatment. The character of this illness and used medications may represent a risk in terms of non-compliance. As in other chronic diseases, scientists engage in identifying of the compliance rate for many years and the aim of this work is to summarize the state of knowledge in the field of drug compliance in rheumatoid arthritis.

Glycerol-3-phosphate dehydrogenase expression and oxygen consumption in liver mitochondria of female and male rats with chronic alteration of thyroid status.

In our chronic experiments (over several months), the activity and protein amount of glycerol-3-phosphate dehydrogenase (GPDH) in mitochondria isolated from the liver of adult male and female inbred Lewis strain euthyroid (EU), hyperthyroid (TH), and hypothyroid (HY) rats were analyzed by biochemical and Western blot methods. The TH status was induced by intraperitoneal injections of 3,3',5-triiodo- L-thyronine and the HY status with 0.05% solution of methimazole in drinking water. The TH status led to a significant increase and the HY status to a significant decrease of enzyme activity and protein amount in both male and female animals. These changes were, however, more pronounced in females. The EU and TH female rats also showed a significantly higher activity and the TH female rats showed also a significantly higher enzyme amount in comparison with males, while the HY rats showed low levels in both sexes. The glycerol-3-phosphate-dependent oxygen consumption of freshly isolated rat liver mitochondria from the TH animals was higher in comparison with the EU animals and it was activated by idebenone, a synthetic analogue of coenzyme Q, in both the EU and TH rats. Measurements of serum thyroid hormone levels and analysis of anatomical parameters (relative heart and thyroid gland weights) confirmed that our procedures inducing the TH and HY states are efficient and reliable and that determination of GPDH can serve as an additional criterion for the evaluation of the thyroid hormone status.

Biochemical and pharmacological effects of mitoxantrone and acetyl-L-carnitine in mice with a solid form of Ehrlich tumour.

BACKGROUND: Dysfunction of the carnitine system in non-tumour tissue following anticancer therapy has been reported. In this setting, supplementation with carnitine derivatives might increase the general metabolic activity of normal cells so that they might better withstand the adverse effects of chemotherapy aimed at tumour cells. Here we investigated the effect of acetyl-L-carnitine (ALC) alone and in combination with the antineoplastic agent mitoxantrone (MX) in an animal cancer model. METHODS: The effects of MX and MX-ALC were assessed based on gain or loss of body weight and on local growth of a solid form of Ehrlich tumour inoculated into mice. We also performed biochemical analyses like serum activities of some enzymes signalling the functioning of the liver, aspartate aminotransferase (AST), and alanine aminotransferase (ALT). Total protein, albumin and bilirubin were also determined in serum. Under favourable conditions, the Ehrlich tumour readily forms metastases, and this is the reason why we performed histological studies of samples of both the liver and heart in order to identify changes that may have mediated the observed effect of the treatment. In addition to those studies, the survival time of treated animals against controls was also noted. RESULTS: MX monotherapy was associated with lower body weight gain, fewer metastases, smaller tumour size, and lower dissemination. ALC alone promoted survival, but had no potentiating effect on MX therapy in terms of survival. Serum biochemistry changes associated with MX-ALC treatment consisted of a significant (p < 0.05) increase in AST with MX at 6 or 9 mg·kg(-1) plus ALC 200 mg·kg(-1) and a significant (p < 0.05) reduction in total protein compared to the corresponding MX group; serum albumin and bilirubin remained unchanged. CONCLUSION: ALC in combination with MX, regardless of the dose of MX, led to higher occurrences of metastases with dissemination to the kidneys, lungs, heart, and mediastinum compared to MX treatment alone. These histological findings indicate that ALC is inappropriate to combine with MX in the treatment of a solid cancer. The protective effect of ALC in combination therapy with the cytostatic drug MX was not supported in this study by our findings that the agent did not improve the therapeutic outcomes of MX therapy.

Individual, age and sex differences in fiber type composition of slow and fast muscles of adult Lewis rats: comparison with other rat strains.

We analyzed fiber type composition of soleus and extensor digitorum longus (EDL) muscles of 3- to 19-month-old male and female inbred Lewis rats using histochemical demonstration of mATPase activity. The rats were divided into four groups of the mean age of 3, 6, 9 and 14 months. We found that the soleus muscle of 3-month-old rats contained significantly more of fast 2A fibers and less of slow type 1 fibers compared to older rats, while no significant difference was found between female and male rats at any age group. In contrast, we found no significant difference in the EDL fiber type composition among the age groups, but we found that the EDL muscle of female rats contained significantly less 2A fibers and more 2B fibers than that of male animals. Our results thus revealed an age difference in the soleus muscle and a sex difference in the EDL muscle among postnatal Lewis rats. The number of slow type 1 fibers in the soleus muscle varied between 87 and 100% and that of 2A fibers between 13 and 0%. In the EDL the percentage of type 1 fibers varied between 2.6 and 8.7%, that of 2A fibers between 12.6 and 25.8% and that of 2B fibers between 70.4 and 81.6%. Both muscles thus exhibited a considerable degree of variability among individual animals even in the same age group. Furthermore, a comparison of the Lewis rats with literature data of other rat strains showed that the number of fast 2A fibers in the soleus muscle of 4-month-old and older animals decreased in this order: SHR>Lister Hooded>Fisher 344>Sprague-Dawley>Wistar>WBN/Kob>Lewis strain, being almost 20% in the SHR and less than 2% in the Lewis rats. In contrast, the "fastest" composition (judged according to the percentage of the fastest 2B fibers) of the EDL muscle was demonstrated by Lewis, Wistar and Fisher 344 rats (about 75%), while Sprague-Dawley and WBN/Kob rats contained only about 50% of 2B fibers. The percentage of slow type 1 fibers in the EDL was low in all strains (about 5%). Our results thus show that the individual, age and sex as well as inter-strain differences in muscle fiber type composition should not be ignored when comparing results of different studies. We also demonstrated that the inbred Lewis strain appears to have more "specialized" muscle composition, as its soleus is the "slowest" and its EDL is the "fastest" among the routinely used rat strains.

[Central hemiparesis as manifestation of systemic vasculitis]. / Centrální pravostranná hemiparéza jako dominující projev systémové vaskulitidy.

We present a case report of patient with systemic vasculitis with affection of heart, skin, gastrointestinal tract and rare involvement of central nervous system. Diagnosis of systemic vasculitis was based on clinical manifestations, blood hypereosinophilia and brain magnetic resonance imaging. Immunosuppressive therapy led to regression of symptoms including initially present neurologic manifestation.

Fiber type composition of unoperated rat soleus and extensor digitorum longus muscles after unilateral isotransplantation of a foreign muscle in long-term experiments.

We examined the effects of the unilateral heterochronous isotransplantation on the fiber type composition and myosin heavy chain (MyHC) isoform content of unoperated slow soleus and fast extensor digitorum longus muscles of female inbred Lewis strain rats. Comparison was made between "control" unoperated muscles of experimental rats (after intramuscular transplantation surgery) with the corresponding muscles of completely naive (unoperated) rats of three age groups (5-, 8- and 14-month-old). This was done in order to ascertain whether these muscles can be used as reliable controls to the transplanted and host muscles for our ongoing grafting experiments. The fiber type composition was determined by assessing the histochemical reaction for myofibrillar adenosine triphosphatase, the MyHC isoform content was determined immunocytochemically using monoclonal antibodies specific to different MyHC isoforms and by sodium dodecyl sulphate polyacrylamide gel electrophoresis. Our experiments show that the heterochronous intramuscular isotransplantation procedure had no significant effect on the fiber type composition and MyHC isoform content of the "control" unoperated muscles of the experimental rats when compared to the corresponding muscles of the naive animals. Furthermore, the duration and type of isotransplantation did not also lead to differences among corresponding "control" muscles of experimental animals. We conclude that the unoperated muscles of the experimental rats can be used as controls in our current transplantation project dealing with long-term grafting experiments.

Diabetes and thyroid hormones affect connexin-43 and PKC-epsilon expression in rat heart atria.

We have examined the changes of intercellular electrical coupling protein connexin-43 (Cx43) and of PKC-epsilon in heart atria of diabetic rats and/or after the treatment with triiodothyronine (T(3)). Diabetes was induced in Wistar-Kyoto rats by streptozotocin (50 mg/kg, i.v.) and atria were examined after 5 (acute stage) and 10 (chronic stage) weeks. T(3) (10 microg/100 g/day) was applied via a gastric tube for the last 10 days prior to the end of the experiments to non-diabetic and to the half of diabetic rats. Expression and phosphorylated status of Cx43, as well as expression of PKC-epsilon, were analyzed by Western blots using mouse monoclonal anti-Cx43 and rabbit polyclonal anti-PKC-epsilon antibodies. We found that the Cx43 expression was significantly increased after the treatment with T(3) and in the acute diabetes. Both in diabetes and after T(3) treatment the phosphorylation of Cx43 isoforms was markedly suppressed compared to the non-diabetic and T(3)-untreated controls. Such a down-regulation was less pronounced in diabetic rats after the T(3)-treatment. The expression of atrial PKC-epsilon was increased in diabetic rats. This increase was suppressed after T(3) administration and the expression was decreased in T(3)-treated non-diabetic rats. We suggest that the reduced Cx43 phosphorylation in diabetic and hyperthyroid rats can deteriorate a cell-to-cell coupling and consequently facilitate a development of atrial tachyarrhythmia in diabetic or hyperthyroid animals.

Real time RT-PCR with a newly designed set of primers confirmed the presence of 2b and 2x/d myosin heavy chain mRNAs in the rat slow soleus muscle.

In order to re-evaluate the presence and relative quantity of 2b and 2x/d myosin heavy chain (MyHC) transcripts in rat slow soleus muscle by using real time RT-PCR we have compared the available relevant cDNA sequences and designed a new set of primers having similar melting temperatures, matching separate MyHC exons in the regions of maximal differences in MyHC coding sequences, and containing G or C at the 3 -end. These also yielded PCR products of corresponding length, which is an important requirement for real time RT-PCR quantification. The experiments were performed on 8-month-old inbred female Lewis strain rats used in our current study of regenerating transplanted muscles. The real time RT-PCR measurement confirmed the expression of all four MyHC mRNAs (type 1, 2a, 2x/d and 2b) in both fast extensor digitorum longus and slow soleus muscles, although in the soleus muscle of adult rats, only type 1 and 2a protein isoforms can be usually detected.