Thyroid Bethesda Category AUS/FLUS in Our Microscopes : Three-Year-Experience and Cyto-Histological Correlation

The Bethesda System for Reporting Thyroid Cytopathology (TBSRTC) introduced a new category: Atypia of Undetermined Significance/Follicular Lesion of Undetermined Significance (AUS/FLUS) comprising of heterogenous lesions with a lesser degree of atypia. Its routine use is a bit controversial. The study cohort included AUS/FLUS thyroid cytopathological diagnoses signed out at Fimlab Laboratories from the period of 1 October 2013 to 31 December 2016. We analyzed all the AUS/FLUS cases, their cytology subclassification, and their cyto-histological correlation, when available. In total, there were 331 AUS/FLUS cases from 252 patients. The mean age was 59.8 years and there were 196 females and 56 males. Repeated AUS/FLUS was diagnosed in 75 (29.8%) cases. Out of 252 patients, 118 (46.8%) were operated on. Sixty-eight were operated on after the first AUS/FLUS diagnosis, 46 after 2 AUS/FLUS diagnoses, and 4 after 3 AUS/FLUS diagnoses. In total, there were 37 (14.7%) malignancies and 40 benign tumors. The risk of malignancy for AUS/FLUS (14.7%) is in agreement with the original TBSRTC risk of malignancy. The risk of neoplasia was 30.6% in our series

Modern extraction techniques in connection with HPLC for analysis of contaminants

Univerzita Karlova, Farmaceutická fakulta v Hradci Králové Školicí pracoviště Katedra analytické chemie Doktorský studijní program Bioanalytické metody Kandidátka / kandidát Mgr. Kholová Aneta Školitelka / školitel prof. RNDr. Dalibor Šatínský, Ph.D. Konzultantka / konzultant PharmDr. Ivona Lhotská, Ph.D. Název disertační práce Moderní extrakční techniky ve spojení s HPLC pro analýzu kontaminantů V předložené disertační práci je prezentován komentovaný soubor šesti publikací zaměřujících se na využití a vzájemné porovnání extrakčních sorbentů k analýze kontaminujících látek v potravinách a v potravinových doplňcích. Hlavním cílem bylo představit nanovlákenné materiály jako vhodný extrakční sorbent pro využití v analytické chemii. Pro tento účel byla vlákna porovnávána s dalšími, většinou komerčními extrakčními sorbenty vhodnými pro testovaný typ matrice. Všechny experimenty byly prováděny v on-line zapojení extrakčního sorbentu přes selekční ventil s analytickou kolonou v chromatografickém přístroji. Nedílnou součástí bylo i využití zjištěných extrakčních vlastností nanovláken při analýze reálných vzorků. Nejdříve byla studována extrakční účinnost na jednoduchých kapalných matricích, kde není potřeba složité přečištění matrice a hlavním cílem je dostatečné zadržení a zakoncentrování analytů. Pro...Charles University, Faculty of Pharmacy in Hradec Králové Training Workplace Department of Analytical Chemistry Doctoral Degree Program Bioanalytical Methods Candidate Mgr. Kholová Aneta Supervisor prof. RNDr. Dalibor Šatínský, Ph.D. Advisor PharmDr. Ivona Lhotská, Ph.D. Title of Doctoral Thesis Modern extraction techniques in connection with HPLC for analysis of contaminants A compilation of six publications is presented in this dissertation thesis. The work focuses on applying and comparing extraction sorbents to analyze contaminating substances in food and food supplements. The main aim of this thesis was to present micro and nanofibers as suitable extraction sorbents for use in analytical chemistry. For this purpose, the fibers were compared with other, mostly commercial, extraction sorbents suitable for the type of matrix tested. All experiments were carried out in the on-line connection of the extraction sorbent through the selection valve with the analytical column in the chromatographic system. A crucial part was also the use of the extraction properties of nanofibers for the analysis of real-life samples. In the first part, the extraction efficiency was studied on liquid matrices, where no need for complex matrix purification was required. The main aim was sufficient retention and...Katedra analytické chemieDepartment of Analytical ChemistryFarmaceutická fakulta v Hradci KrálovéFaculty of Pharmacy in Hradec Králov

Understanding of terminal drought tolerance mechanisms in pearl millet (Pennisetum glaucum (L.) R. Br.)

The main objective of the thesis work was the detailed characterization of pearl millet (Pennisetum glaucum (L.) R. Br.) genotypes contrasting for terminal drought tolerance. For that work, we used a set of near isogenic lines (NIL-QTLs; carrying terminal drought tolerance quantitative trait locus (QTL) from a drougth tolerant donor parent on the genetic background of a sensitive parent) and a recombinant inbred lines population (RIL; developed from a cross between the tolerant and sensitive genotype). In these contrasting genotypes we investigated following physiological traits. Transpiration rate (Tr), transpiration efficiency (TE), transpiration response to increased vapor pressure deficit, threshold in volumetric soil moisture where transpiration begins to decline (FTSW threshold), stomatal density (SD), sensitivity of plants’ growth to VPD below and above 2kPa. Regarding biochemical traits, we followed content of chlorophyll (Chl), carotenoids (Car), abscisic acid (ABA), proline (Pro), we conducted isozyme analysis of antioxidative enzymes [superoxid dismutase (SOD), ascorbic peroxidase (APX), catalase (CAT)]. The main leading thread for understanding the drought tolerance mechanisms of pearl millet came from the analysis of traits related to the control of water losses under fully irrigated conditions. We could clearly distinguish drought tolerant genotypes from the sensitive ones based on: i) lower Tr in well-watered conditions measured on full plant basis and on detached leaves ii) higher leaf ABA content in well-watered conditons iii) sensitivity of transpiration to high VPD condition under well-watered conditions. Furthemore, the leaf expansion of tolerant genotypes was sensitive to VPD conditions in which plant development took place and these conditions determined the dynamics of water utilisation during plants development. Based on the biochemical parameters we could rarely distinquish between tolerant and sensitive genotype. Though we documented differences in the activity of APX5 isoenzyme and proline accumulation dynamics under water limiting conditions between tolerant/sensitive genotypes, this variation was probably not directly linked to the yield variation of these genotypes under terminal drought conditions. It is concluded that the major terminal drought tolerance mechanism of investigated tolerant pearl millet genotypes is linked to their lower Tr. Low Tr of these genotypes probably contribute to saving the water in the soil profile and so leaving a critical amount of water available for grain filling stage (in fact drought avoidance mechanism). It is further discussed that Tr could be influenced by the level of leaf ABA and the hydraulic properties of plant tissues. However, these “water saving” drought tolerance mechanisms seems to be specific to the environmental conditions in which plants` development took place. The importance of these water saving mechanisms is also being validated in RIL population. The biochemical parameters tested under drought conditions appeared to have no major significance for terminal drought tolerance

Cyto-Histopathological Correlations in Pathology Diagnostics

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Determination of mycotoxin citrin using on-line SPE HPLC on molecularly imprinted polymers

Univerzita Karlova Farmaceutická fakulta v Hradci Králové Katedra analytické chemie Kandidát: Bc. Aneta Kholová Školitel: Doc. RNDr. Dalibor Šatínský, Ph.D. Název diplomové práce: Stanovení mykotoxinu citrininu pomocí on-line SPE HPLC na molekulárně vtištěných polymerech Cílem této práce bylo vytvořit rychlou metodu pro stanovení mykotoxinu citrininu (CIT) za použití vysokoúčinné kapalinové chromatografie (HPLC) a techniky přepínání kolon v kombinaci s on-line extrakcí na pevné fázi (SPE) na molekulárně vtištěných polymerech (MIP). Nejvhodnější sorbent pro selektivní extrakci byl vybrán ze šesti nově nasyntetizovaných MIP po vyhodnocení jejich vazebné kapacity a selektivity. Selektivita byla testována porovnáním molekulárně vtištěných a nevtištěných polymerů a schopností polymeru selektivně oddělit citrinin od interferencí z testovaného vzorku. Nejlepším sorbentem MIP (funkční monomer metakrylamidu s porogenem acetonitrilem) byla naplněna předkolona (20 × 3 mm) a zapojena do systému přepínání kolon HPLC. Na tento chromatografický systém byly dávkovány tři různé typy matric (extrakt z potravních doplňků, červené fermentované rýže a obilovin). Dávkovaný objem vzorku 50 μl byl na MIP extrakční předkoloně promýván od interferencí methanolem / 0,5% vodným roztokem kyseliny octové v poměru 25:75 (v/v),...Charles University Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Bc. Aneta Kholová Supervisor: Doc. RNDr. Dalibor Šatínský, Ph.D. Title of Diploma Thesis: Determination of mycotoxin citrinin using on-line SPE HPLC on molecularly imprinted polymers The aim of this work was to develop a fast method for mycotoxin citrinin (CIT) determination using high performance liquid chromatography (HPLC) and column switching technique in combination with on-line solid phase extraction (SPE) based on citrinin-selective molecularly imprinted polymer (MIP). The most suitable sorbent for selective extraction was chosen from six newly synthetized MIP after evaluation of their binding capacity and selectivity. Selectivity was tested by comparing molecularly imprinted and non-imprinted polymers and the ability of the MIP polymer to selectively separate citrinin from interferences of the test sample. The best MIP (functional monomer of metacrylamide with porogen acetonitrile) was filled into pre-column (20 × 3 mm) and connected to HPLC column-switching system. Three different matrices were injected to chromatographic system. 50 μl of the sample was injected onto MIP extraction column and washed out of interferences with methanol / 0.5% water solution of acetic acid in a ratio of...Katedra analytické chemieDepartment of Analytical ChemistryFaculty of Pharmacy in Hradec KrálovéFarmaceutická fakulta v Hradci Králov

Salivary Gland FNA Diagnostics in a Real-Life Setting : One-Year-Experiences of the Implementation of the Milan System in a Tertiary Care Center

The Milan System for Reporting Salivary Gland Cytopathology (MSRSGC) was introduced in 2018 following other organ specific cytopathological reporting systems and it aimed at bringing a practical, evidence-based, user-friendly classification system with characterization and management algorithms. At the Department of Pathology, Fimlab Laboratories, Tampere, Finland all salivary fine needle aspirations (FNAs) have been given cytopathological diagnoses according to the MSRSGC since January 2018. Analyses of a one-year-period (January 2018–December 2018) consisted of 183 salivary FNA samples from 138 patients with correlation to histopathology in 90 cases with surgical follow-up. The MSRSGC performance in patient based analysis was as follows: accuracy was 90.9%, sensitivity was 61.5%, specificity was 100%, positive predictive value was 100%, and negative predictive value was 89.4%, respectively. Risks of malignancy (ROMs) in MSRSGC categories were: 0.0% (0/15) in non-diagnostic category, 100.0% (1/1) in non-neoplastic category biased by only one falsely-negative lymphoma case, 14.3% (1/7) in atypia of undetermined significance category, 0.0% (0/28) in benign neoplasm category, 27.3% (3/11) in neoplasm of uncertain malignant potential category, and 100% for both suspicious for malignancy (4/4) and malignancy (4/4) categories, respectively. The MSRSGC has been proven as a reliable classification system in salivary gland FNA routine diagnostics in a tertiary care center

Potassium in plant osmoregulation

Draslík je v rostlině velmi mobilní a je čile transportován přes membrány, za pomoci transportérů a kanálů. Vyskytuje se v rostlině výhradně ve formě iontu K+. Rovněž ve formě tohoto iontu je přijímán z půdního roztoku a to pomocí transportéru HAK5 a kanálu AKT1. Kanál SKOR a transportér KUP7 zprostředkovává přenos K+ do xylému. Klíčová je role draslíku jako osmoticky aktivního prvku, který reguluje množství vody v buňkách a tím i udržování turgoru, nezbytného pro udržení tvaru rostlinné buňku a její růst. Jako modelový systém pro popis procesů souvisejících s osmotickou funkcí K+ se rozšířilo využití průduchů. V průduchách, totiž pohyb K+ umožňuje regulaci otevírání a zavírání apertury na principu změn turgoru. Do svěracích buněk zajišťují transport K+ kanály KAT1, KAT2, AKT1 a AKT2, odvod K+ naopak obstarává kanál GORK na plazmatické membráně a TPK1 na tonoplastu. Draslík také podporuje transport asimilátů ve floému, v čemž hraje roli kanál AKT2. Právě osmotické funkce K+ jsou hlavním tématem této bakalářské práce, která shrnuje aktuální poznatky o transportních mechanismech K+ nezbytných pro funkci K+ jako osmoticky aktivní látky.Potassium is very mobile in plants and is transported across membranes using transporters and channels. It is present in the ionic form K+ in plants. Also, in the form of this ion, K+ is taken up by plants from the soil solution. The uptake is mediated by the transporter HAK5 and by the channel AKT1. The transfer of K+ to the xylem is provided by the channel SKOR and the transporter KUP7. The role of potassium as an osmotically active element is crucial. K+ helps to regulate the amount of water in the cells and thus to maintain the turgor. Turgor is important for maintaining the shape of the plant cells and for its growth. Stomatal guard cells are used as a model system for describing processes related to the K+ osmotic function. Movement of K+ is involved in the regulation of opening and closing of stomata on the principle of turgor changes. Transport of K+ into guard cells is provided by channels KAT1, KAT2, AKT1 and AKT2. On the contrary, K+ release is mediated by the channel GORK on the plasma membrane and TPK1 on the tonoplast. K+ supports also the transport of assimilates in the phloem, where the channel AKT2 plays a role. The osmotic functions of K+ are the main topic of this bachelor thesis, which summarizes current knowledge about transport mechanisms necessary for the function of K+ as osmotically...Department of Experimental Plant BiologyKatedra experimentální biologie rostlinFaculty of SciencePřírodovědecká fakult

Constitutive water-conserving mechanisms are correlated with the terminal drought tolerance of pearl millet [Pennisetum glaucum (L.) R. Br.]

Pearl millet, a key staple crop of the semi-arid tropics, is mostly grown in water-limited conditions, and improving its performance depends on how genotypes manage limited water resources. This study investigates whether the control of water loss under non-limiting water conditions is involved in the terminal drought tolerance of pearl millet. Two pairs of tolerant×sensitive pearl millet genotypes, PRLT 2/89-33–H77/833-2 and 863B-P2–ICMB 841-P3, and near-isogenic lines (NILs), introgressed with a terminal drought tolerance quantitative trait locus (QTL) from the donor parent PRLT 2/89-33 into H77/833-2 (NILs-QTL), were tested. Upon exposure to water deficit, transpiration began to decline at lower fractions of transpirable soil water (FTSW) in tolerant than in sensitive genotypes, and NILs-QTL followed the pattern of the tolerant parents. The transpiration rate (Tr, in g water loss cm−2 d−1) under well-watered conditions was lower in tolerant than in sensitive parental genotypes, and the Tr of NILs-QTL followed the pattern of the tolerant parents. In addition, Tr measured in detached leaves (g water loss cm−2 h−1) from field-grown plants of the parental lines showed lower Tr values in tolerant parents. Defoliation led to an increase in Tr that was higher in sensitive than in tolerant genotypes. The differences in Tr between genotypes was not related to the stomatal density. These results demonstrate that constitutive traits controlling leaf water loss under well-watered conditions correlate with the terminal drought tolerance of pearl millet. Such traits may lead to more water being available for grain filling under terminal drought

The diagnostic accuracy of pleomorphic adenoma in fine needle aspiration : Features leading to false positive and false negative diagnoses and the utility of cell blocks

Objective: Fine needle aspiration (FNA) is a well-established tool in preoperative diagnosis of salivary gland lesions with diagnostic accuracy of 90%. Pleomorphic adenoma (PA) is the most common salivary gland tumor comprising 45%–74% of all salivary gland tumors with FNA diagnostic accuracy of 89.5%–96.2%. The aim of the present study was to determine and analyze potential cytomorphological pitfalls and evaluate the diagnostic accuracy in FNA diagnosis of PA. Methods: Salivary gland specimens with both cytological and histological diagnoses were searched over a 10-year-period (2009–2018) from a laboratory information system of Pathology Department, Fimlab Laboratories, Tampere and matched to determine concordant and discordant PA cases. Sufficient material in histological and cytological sample was found in 401 cases. In 218 cases (54.4%) diagnosis was true-negative PA, in 169 cases (42.1%) diagnosis was true-positive PA and there were 14 discordant cases: 4 false-positive cases and 10 false-negative cases. False-negative cases were reclassified and subgrouped according to The Milan System for Reporting Salivary Gland Cytopathology (MSRSGC). Results: Cytomorphologically, cell type predominance was more often myoepithelial in true-positive cases (65%) and epithelial both in false-negative (70%, p =.007) and false-positive cases (75%, p =.027). Well-formed ducts were present in cytology in all true-positive cases (p <.001). Only 10% of true-positive cases did not show any matrix in cytology (p <.001). Nuclear changes were common in false-negative cases (80%, p =.002) and false-positive cases (75%, p =.003). Beneficial cell block (CB) was more common in true-positive cases (85%) than in false-negative cases (50%, p =.041) or in false-positive cases (50%, p =.116) and a lack of beneficial CB led more often to a false diagnosis (70% false diagnosis without beneficial CB versus 29% false diagnosis with beneficial CB). Conclusion: The present study showed diagnostic accuracy of 96.5% for FNA in PA diagnosis. Sensitivity, specificity, positive predictive value and negative predictive value were 94.4%, 98.2%, 97.7%, and 95.6%, respectively. The benefit of CBs was more evident in true-positive cases (85%).Peer reviewe

Transpiration difference under high evaporative demand in chickpea ( Cicer arietinum L.) may be explained by differences in the water transport pathway in the root cylinder

Terminal drought substantially reduces chickpea yield. Reducing water use at vegetative stage by reducing transpiration under high vapor pressure deficit (VPD), i.e. under dry/ hot conditions, contributes to drought adaptation. We hypothesized that this trait could relate to differences in a genotype’s dependence on root water transport pathways and hydraulics. • Transpiration rate responses in conservative and profligate chickpea genotypes were evaluated under increasing VPD in the presence/absence of apoplastic and cell-to-cell transport inhibitors. • Conservative genotypes ICC 4958 and ICC 8058 restricted transpiration under high VPD compared to the profligate genotypes ICC 14799 and ICC 867. Profligate genotypes were more affected by aquaporin inhibition of the cell-to-cell pathway than conservative genotypes, as measured by the root hydraulic conductance and transpiration under high VPD. Aquaporin inhibitor treatment also led to a larger reduction in root hydraulic conductivity in profligate than in conservative genotypes. In contrast, blockage of the apoplastic pathway in roots decreased transpiration more in conservative than in profligate genotypes. Interestingly, conservative genotypes had high early vigour, whereas profligate genotypes had low early vigour. • In conclusion, profligate genotypes depend more on the cell-to-cell pathway, which might explain their higher root hydraulic conductivity, whereas water-saving by restricting transpiration led to higher dependence on the apoplastic pathway. This opens the possibility to screen for conservative or profligate chickpea phenotypes using inhibitors, itself opening to the search of the genetic basis of these differences