• Klíčová slova
• RWE, BIG DIP, real time insights, business decisions, visual analytics,
• MeSH
• big data MeSH
• data mining MeSH
• elektronické zdravotní záznamy * MeSH
• farmaceutický průmysl MeSH
• informační management * MeSH
• lékařská informatika * MeSH
• lidé MeSH
• manažerské informační systémy MeSH
• metody pro podporu rozhodování MeSH
• sektor zdravotní péče trendy   MeSH
• šíření informací MeSH
• Check Tag
• lidé MeSH

AIMS: To explore the feasibility of subcutaneous continuous glucose monitoring (CGM) in perioperative settings and to evaluate the perioperative development of glycaemia in persons with diabetes mellitus or impaired glucose tolerance by means of CGM. METHODS: Monitoring by means of Guardian REAL-Time CGMS (Medtronic, Nortridge, USA) in 20 perioperative periods. Sensor was inserted on the day before surgery and continued for 3 days with some exceptions. RESULTS: Full implementation of the method was successful in the intensive care unit setting only. No electromagnetic interference and no side effects were found. The Wilcoxon signed-rank test revealed no significant difference between sensor and laboratory analyser values. Pearson's correlation coefficients of the values obtained by sensor and the Wellion Linus glucometer were 0.875 for the whole perioperative period, 0.866 for the intraoperative period and 0.903 for the first perioperative day. A decline in sensor accuracy on the 6(th) day was registered in one case. 16 monitored cases (80%) did not meet the criteria for safe plasma glucose range. Hypoglycaemia was found in 4 (20%) cases. There was an association between grade of the perioperative dysglycaemia and need for reoperation within the next 3 months. The most frequent perioperative glycaemic patterns are demonstrated. CONCLUSION: Subcutaneous CGM is safe offering detailed insight into glucose homeostasis in the dynamic perioperative period. Laboratory confirmation of sensor plasma glucose concentration by approved laboratory analyser is still necessary. The potentional benefits of maintaining patients within a safe glucose range should be comfirmed by future studies.

• MeSH
• biochemická analýza krve přístrojové vybavení   MeSH
• krevní glukóza analýza   MeSH
• lidé MeSH
• perioperační péče * MeSH
• pilotní projekty MeSH
• počítačové systémy MeSH
• protézy a implantáty * MeSH
• senioři MeSH
• studie proveditelnosti MeSH
• subkutánní tkáň MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• pozorovací studie MeSH
• práce podpořená grantem MeSH

Wilms' tumor gene 1 (WT1) functions including some contradictory effects may be explained by the presence and interactions of its isoforms, however, their evaluation has been so far complicated by several technical problems. We designed unique quantitative PCR systems for direct quantification of the major WT1 isoforms A[EX5-/KTS-], B[+/-], C[-/+] and D[+/+] and verified their sensitivity, specificity and reproducibility in extensive testing. With this method we evaluated WT1 total and isoform expression in 23 normal bone marrow (BM) samples, 73 childhood acute myeloid leukemia (AML), 20 childhood myelodysplastic syndrome (MDS), 9 childhood severe aplastic anemia (SAA), 30 adult AML and 29 adult MDS patients. WT1 isoform patterns showed differences among these samples and clustered them into groups representing the specific diagnoses (P<0.0001). Isoform profiles were independent of total WT1 expression and possess certain common features-overexpression of isoform D and EX5[+] variants. The KTS[+]/KTS[-] ratio was less variable than the EX5[+]/EX5[-] ratio and differed between children and adults (P<0.001); the EX5[+]/EX5[-] ratio varied between diagnoses (AML vs MDS, P<0.001). These findings bring new insights into WT1 isoform function and suggest that the ratio of WT1 isoforms, particularly EX5 variants, is probably crucial for the process of malignant transformation.

• MeSH
• akutní myeloidní leukemie genetika   MeSH
• dítě MeSH
• dospělí MeSH
• kojenec MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• lidé středního věku MeSH
• lidé MeSH
• messenger RNA genetika   MeSH
• mladiství MeSH
• mladý dospělý MeSH
• nádorové buňky kultivované MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• protein - isoformy MeSH
• proteiny WT1 genetika   MeSH
• senioři MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• kojenec MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• senioři MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Lactobacillus gasseri K7 is a probiotic strain that produces bacteriocins gassericin K7 A and K7 B. In order to develop a real-time quantitative PCR assay for the detection of L. gasseri K7, 18 reference strains of the Lactobacillus acidophilus group and 45 faecal samples of adults who have never consumed strain K7 were tested with PCR using 14 pairs of primers specific for gassericin K7 A and K7 B gene determinants. Incomplete gassericin K7 A or K7 B gene clusters were found to be dispersed in different lactobacilli strains as well as in faecal microbiota. One pair of primers was found to be specific for the total gene cluster of gassericin K7A and one for gassericin K7B. The real-time PCR analysis of faecal samples spiked with K7 strain revealed that primers specific for the gene cluster of the gassericin K7 A were more suitable for quantitative determination than those for gassericin K7 B, due to the lower detection level. Targeting of the gassericin K7 A or K7 B gene cluster with specific primers could be used for detection and quantification of L. gasseri K7 in human faecal samples without prior cultivation. The results of this study also present new insights into the prevalence of bacteriocin-encoding genes in gastrointestinal tract.

• MeSH
• bakteriociny genetika   MeSH
• bakteriologické techniky metody   MeSH
• DNA primery genetika   MeSH
• dospělí MeSH
• feces mikrobiologie   MeSH
• kvantitativní polymerázová řetězová reakce metody   MeSH
• Lactobacillus acidophilus genetika   izolace a purifikace   MeSH
• lidé MeSH
• senzitivita a specificita MeSH
• zdraví dobrovolníci pro lékařské studie MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• práce podpořená grantem MeSH

Reverse transcription quantitative PCR (RT-qPCR) has delivered significant insights in understanding the gene expression landscape. Thanks to its precision, sensitivity, flexibility, and cost effectiveness, RT-qPCR has also found utility in advanced single-cell analysis. Single-cell RT-qPCR now represents a well-established method, suitable for an efficient screening prior to single-cell RNA sequencing (scRNA-Seq) experiments, or, oppositely, for validation of hypotheses formulated from high-throughput approaches. Here, we aim to provide a comprehensive summary of the scRT-qPCR method by discussing the limitations of single-cell collection methods, describing the importance of reverse transcription, providing recommendations for the preamplification and primer design, and summarizing essential data processing steps. With the detailed protocol attached in the appendix, this tutorial provides a set of guidelines that allow any researcher to perform scRT-qPCR measurements of the highest standard.

• MeSH
• analýza jednotlivých buněk metody   normy   MeSH
• kvantitativní polymerázová řetězová reakce metody   normy   MeSH
• lidé MeSH
• reverzní transkripce genetika   MeSH
• senzitivita a specificita MeSH
• stanovení celkové genové exprese metody   normy   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

Background and Aims: The non-specific phospholipase C (NPC) is a new member of the plant phospholipase family that reacts to abiotic environmental stresses, such as phosphate deficiency, high salinity, heat and aluminium toxicity, and is involved in root development, silicon distribution and brassinolide signalling. Six NPC genes (NPC1-NPC6) are found in the Arabidopsis genome. The NPC2 isoform has not been experimentally characterized so far. Methods: The Arabidopsis NPC2 isoform was cloned and heterologously expressed in Escherichia coli. NPC2 enzyme activity was determined using fluorescent phosphatidylcholine as a substrate. Tissue expression and subcellular localization were analysed using GUS- and GFP-tagged NPC2. The expression patterns of NPC2 were analysed via quantitative real-time PCR. Independent homozygous transgenic plant lines overexpressing NPC2 under the control of a 35S promoter were generated, and reactive oxygen species were measured using a luminol-based assay. Key Results: The heterologously expressed protein possessed phospholipase C activity, being able to hydrolyse phosphatidylcholine to diacylglycerol. NPC2 tagged with GFP was predominantly localized to the Golgi apparatus in Arabidopsis roots. The level of NPC2 transcript is rapidly altered during plant immune responses and correlates with the activation of multiple layers of the plant defence system. Transcription of NPC2 decreased substantially after plant infiltration with Pseudomonas syringae, flagellin peptide flg22 and salicylic acid treatments and expression of the effector molecule AvrRpm1. The decrease in NPC2 transcript levels correlated with a decrease in NPC2 enzyme activity. NPC2-overexpressing mutants showed higher reactive oxygen species production triggered by flg22. Conclusions: This first experimental characterization of NPC2 provides new insights into the role of the non-specific phospholipase C protein family. The results suggest that NPC2 is involved in the response of Arabidopsis to P. syringae attack.

• MeSH
• Arabidopsis enzymologie   imunologie   mikrobiologie   MeSH
• fosfatidylcholiny metabolismus   MeSH
• fosfolipasy typu C fyziologie   MeSH
• Golgiho aparát enzymologie   MeSH
• imunita rostlin fyziologie   MeSH
• klonování DNA MeSH
• konfokální mikroskopie MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• nemoci rostlin imunologie   mikrobiologie   MeSH
• proteiny huseníčku fyziologie   MeSH
• protoplasty enzymologie   MeSH
• Pseudomonas syringae * MeSH
• reaktivní formy kyslíku MeSH
• regulace genové exprese u rostlin MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The similarity between humans and pigs, when it comes to tissue morphology, makes Sus scrofa not only a good research model, but also a potential source of cells for tissue engineering. Cell samples obtained from the pig donor, could be influenced in vitro, in order to become a source of tissue material for xenotransplantation, reconstructive and regenerative medicine. Significant amounts of data point to especially major similarities in pig and human reproductive systems. Because of that, particular scientific focus is centered on research concerning porcine COCs, theca and granulosa cells in primary cultures. One of the aspects of the reproductive process, that is still largely undiscovered, is the interaction between preimplantation blastocyst and maternal uterine tissues. In this study, we used molecular analysis techniques, such as RT-qPCR and immunocytochemistry, to analyze the expression and distribution of cytokeratin 18 and panCytokeratins 8, 18 and 19 and vimentin in porcine luminal endometrial epithelial cells, coupled with analysis of their behavior in RTCA. The results have confirmed the presence of epithelial, as well as stromal cell markers in the cells, varying in levels at different stages of culture. They have also given insight into the modes of proliferation and differentiation of studied cells in in vitro culture, as well as providing additional proof for the possible mesenchymal transdifferentiation of epithelial cells.

• MeSH
• biologické markery metabolismus   MeSH
• biologické modely MeSH
• buněčná diferenciace MeSH
• buňky stromatu cytologie   metabolismus   MeSH
• časové faktory MeSH
• endometrium cytologie   MeSH
• epitelové buňky cytologie   metabolismus   MeSH
• kultivované buňky MeSH
• lidé MeSH
• modely u zvířat MeSH
• prasata MeSH
• proliferace buněk * MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Conditions facilitating resistance to quaternary ammonium compounds (QACs) were investigated in Staphylococcus aureus SK982 exposed to benzalkonium chloride (BAC; a member of QACs) under various circumstances. S. aureus SK982 carrying the qacA gene encoding for resistance to QACs was grown in the presence of stable or gradually increasing concentrations of BAC, or it was exposed to this antiseptic in the exponential phase of growth. Bacteria cultivated in the highest BAC concentrations that did not retard their growth comparing to the untreated control were subjected to real-time quantitative polymerase chain reaction analysis for relative expression of the efflux genes qacA and norA. Under such conditions, S. aureus SK982 tolerated a relatively low stable concentration of BAC (1.22 mg/L) when compared with a gradually increasing antiseptic concentration (tolerance of 4.88 mg/L). However, in both cases, qacA expression was not significant. The culture exposed in the exponential phase of growth tolerated the highest concentration of BAC (9.76 mg/L) as also accompanied by significant overexpression of qacA. Expression of norA was relatively low regardless of the conditions tested. It seems that under the short-term conditions, the phase of bacterial growth is more important for the expression of BAC resistance than the capability to adapt to this antiseptic. This study provides a deeper insight into the relevance of the qac genes in conferring resistance to QACs.

• MeSH
• antibakteriální látky aplikace a dávkování   farmakologie   MeSH
• bakteriální léková rezistence genetika   MeSH
• bakteriální proteiny genetika   MeSH
• benzalkoniové sloučeniny aplikace a dávkování   farmakologie   MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• membránové transportní proteiny genetika   MeSH
• mikrobiální testy citlivosti MeSH
• proteiny spojené s mnohočetnou rezistencí k lékům genetika   MeSH
• regulace genové exprese u bakterií MeSH
• Staphylococcus aureus účinky léků   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Although the mechanism of action of leukemic oncogene Wilms' tumor gene 1 (WT1) remains unclear, WT1 has already been used in monitoring of patients with acute myeloid leukemia (AML) and it is being tested for immunotherapy. More detailed understanding of the role of WT1 in leukemia may improve its utilization. At least 36 isoforms may be produced. Four major variants denoted as -5/-KTS, -5/+KTS, +5/-KTS and +5/+KTS are produced by combining splicing of exon 5 and KTS sequence. In this study, we report applicability of newly developed real-time RT PCRs enabling for the first time full quantification of the four major WT1 splicing variants. Following careful optimization and testing of quantification reliability of four assays, we analyzed 34 samples of patients with AML and 12 samples of patients with chronic myeloid leukemia (CML) at the time of diagnosis. Analyses of five more CML patients provided insight into WT1 variants expression kinetics. We found predominance of +5/+KTS in both diagnoses. Comparison of WT1 variant expression in AML and CML patients' groups differing in response to therapy suggested possible importance of particular WT1 variant levels as markers of further disease course.

• MeSH
• akutní myeloidní leukemie diagnóza   metabolismus   terapie   MeSH
• alternativní sestřih MeSH
• buňky K562 MeSH
• chronická myeloidní leukemie diagnóza   metabolismus   terapie   MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• proteiny WT1 biosyntéza   MeSH
• regulace genové exprese u leukemie MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• klinické zkoušky MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH

BACKGROUND: Stage III non-small cell lung cancer (NSCLC) is a highly heterogeneous stage due to its subgroups (IIIA-IIIC) comprising both resectable and unresectable tumors. Accurate determination of the extent of the disease is essential for excluding stage IV and choosing the optimal treatment regimen. Whole body positron emission tomography and computed tomography scan (PET/CT) is recommended as an initial staging imaging in locally advanced NSCLC. Despite international guidelines for NSCLC diagnosis and treatment, they are not always adhered to due to various reasons. Even in such a groundbreaking study, the phase 3 trial PACIFIC investigating the efficacy of durvalumab as consolidation therapy in patients with stage III NSCLC PET/CT was not mandatory. With the premise that whole body PET/CT of the trunk is essential for diagnosing stage III NSCLC, we performed a retrospective study evaluating the relationship of the use of PET/CT versus conventional staging with CT of the chest and abdomen, in terms of survival. METHODS: This retrospective study of stage III NSCLC patients used the Czech lung cancer registry LUCAS, which was established in June 2018. As of the data export (up to February 9, 2022), a total of 703 patients were eligible for the analysis. Overall survival (OS) was compared using Kaplan-Meier analysis and a Cox regression model. Continuous variables were tested using the Mann-Whitney test, and categorical variables using the Pearson's Chi-square or Fisher's exact test. RESULTS: A total of 703 patients were included in the cohort with an average age of 69 years. PET/CT was performed on 354 patients, and conventional staging using chest and abdominal CT on 349 patients. The median OS among patients with PET/CT was 20.9 months [95% confidence interval (CI): 18.1-23.7], and it was statistically significantly higher (P<0.001) than among patients without PET/CT, where the median OS was 9.0 months (95% CI: 7.3-10.6). The observed effect of PET/CT was also statistically significant when comparing individual stages (IIIA, IIIB, IIIC). The multivariate Cox model confirmed the use of PET/CT as an independent prognostic factor. The most common reason for omission of PET/CT was the local or time unavailability of the examination. CONCLUSIONS: Omission of PET/CT can mean a significant decrement in survival for the patients in stage III NSCLC, likely due to poor staging and suboptimal treatment. Routine use of PET/CT is strictly recommended for the optimal management of stage III NSCLC patients even outside the high-income countries.

• Publikační typ
• časopisecké články MeSH