OBJECTIVE: Approximately one third of patients diagnosed with muscle-invasive urinary bladder cancer (UBC) have undetected metastases at the time of treatment of the primary tumor. Currently there are no reliable specific serum markers for monitoring and evaluating risk profiles of urothelial cancers. Several studies suggest that detection of circulating tumor cells (CTCs) may correlate with the disease status and prognosis at baseline and early in the treatment of cancers. In this study a new way of isolation and in vitro cultivation of CTCs of urinary bladder cancer was introduced. MATERIALS AND METHODS: Peripheral blood (PB) samples from 53 patients who had undergone urological procedure were evaluated using the MetaCell device (MetaCell s.r.o., Ostrava, Czech Republic). The patients enrolled in the study were both oncological patients with UBC and non-oncological patients with inflammation (14 patients). The sensitivity and quantification of CTCs were evaluated. The separated CTCs were cultured in vitro. RESULTS: 39 patients with confirmed UBC were enrolled in the study. CTCs were detected in 25 (64%) patients, and most of these patients had between 6 and 10 cells. The separated CTCs were successfully cultured in vitro. CONCLUSION: CTCs were detected in a higher percentage of patients than in other studies. This paper describes the first successful culturing of human UBC cells. The MetaCell approach used in this study enabled the capture of viable intact virgin CTCs (virgin CTC) suitable for next in vitro culturing, single cell analysis or drug testing.

• Klíčová slova
• CTC, MetaCell, Urinary bladder cancer, circulating tumor cells, urothelial tumors,
• MeSH
• lidé středního věku MeSH
• lidé MeSH
• nádorové buňky kultivované MeSH
• nádorové cirkulující buňky patologie   MeSH
• nádory močového měchýře krev   patologie   MeSH
• papilární karcinom krev   patologie   MeSH
• proliferace buněk MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• separace buněk metody   MeSH
• staging nádorů MeSH
• studie případů a kontrol MeSH
• stupeň nádoru MeSH
• viabilita buněk MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Endophytic fungi live inside vegetal tissues without causing damage to the host plant and may provide lead compounds for drug discovery. The co-culture of two or more endophytic fungi can trigger silent gene clusters, which could lead to the isolation of bioactive compounds. In this study, two endophytic strains isolated from Handroanthus impetiginosus leaves, identified as Talaromyces purpurogenus H4 and Phanerochaete sp. H2, were grown in mixed and axenic cultures. The meroterpenoid austin was detected only in the extracts from the mixed culture. Once isolated, austin displayed very interesting trypanocidal activity, with an IC50 value of 36.6 ± 1.2 μg/mL against Trypanosoma cruzi in the epimastigote form. The results obtained highlight the importance of the co-culturing of endophytic fungi to obtain natural bioactive products. The findings also enhance our understanding of the ecological relationships between endophytic fungi.

• MeSH
• endofyty chemie   genetika   růst a vývoj   MeSH
• kokultivační techniky MeSH
• listy rostlin mikrobiologie   MeSH
• Phanerochaete chemie   genetika   růst a vývoj   metabolismus   MeSH
• Tabebuia mikrobiologie   MeSH
• Talaromyces chemie   genetika   růst a vývoj   metabolismus   MeSH
• terpeny analýza   metabolismus   farmakologie   MeSH
• trypanocidální látky analýza   metabolismus   farmakologie   MeSH
• Trypanosoma cruzi účinky léků   růst a vývoj   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• austin MeSH Prohlížeč
• terpeny MeSH
• trypanocidální látky MeSH

PURPOSE: Dysfunction of the retinal pigment epithelium (RPE) causes numerous forms of retinal degeneration. RPE replacement is a modern option to save vision. We aimed to test the results of transplanting cultured RPEs on biocompatible membranes. METHODS: We cultivated porcine primary RPE cells isolated from cadaver eyes from the slaughterhouse on two types of membranes: commercial polyester scaffolds Transwell (Corning Inc., Kenneburg, ME, USA) with 0.4 µm pore size and prepared Poly (L-lactide-co-DL-lactide) (PDLLA) nanofibrous membranes with an average pore size of 0.4 µm. RESULTS: Five types of assays were used for the analysis: immunocytochemistry (ICC), phagocytosis assay, Western blotting, real-time qPCR (RT-qPCR) and electron microscopy. RT-qPCR demonstrated that RPEs cultured on nanofibrous membranes have higher expressions of BEST1 (bestrophin 1), RLBP1 (retinaldehyde-binding protein 1), RPE65 (retinal pigment epithelium-specific 65 kDa protein), PAX6 (transcription factor PAX6), SOX9 (transcription factor SOX9), DCT (dopachrome tautomerase) and MITF (microphthalmia-associated transcription factor). ICC of the RPEs cultured on nanofibrous membranes showed more intensive staining of markers such as BEST1, MCT1 (monocarboxylate transporter 1), Na+ /K+ ATPase, RPE65 and acetylated tubulin in comparison with commercial ones. Additionally, the absence of α-SMA proved the stability of the RPE polarization state and the absence of epithelial-to-mesenchymal transition. RPE possessed high phagocytic activity. Electron microscopy of both membranes confirmed a confluent layer of RPE cells and their genuine morphological structure, which was comparable to native RPEs. CONCLUSIONS: Retinal pigment epitheliums cultured on polylactide nanofibrous membranes improved the final quality of the cell product by having better maturation and long-term survival of the RPE monolayer compared to those cultured on commercial polyester scaffolds. PDLLA-cultured RPEs are a plausible source for the replacement of non-functioning RPEs during cell therapy.

• Klíčová slova
• AMD, RPE, eye, nanofibrous membrane, retina, retinal pigment epithelium,
• MeSH
• bestrofiny metabolismus   MeSH
• degenerace retiny * metabolismus   MeSH
• kultivované buňky MeSH
• nanovlákna * chemie   MeSH
• polyestery metabolismus   MeSH
• prasata MeSH
• retinální pigmentový epitel metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bestrofiny MeSH
• polyestery MeSH

Protists are key players in the biosphere. Here, we provide a perspective on integrating protist culturing with omics approaches, imaging, and high-throughput single-cell manipulation strategies, concluding with actions required for a successful return of the golden age of protist culturing.

• Klíčová slova
• culturing, culturomics, protists,
• MeSH
• Eukaryota * genetika   MeSH
• multiomika MeSH
• Publikační typ
• časopisecké články MeSH

BACKGROUND: Human induced pluripotent stem cells (hiPSCs) play roles in both disease modelling and regenerative medicine. It is critical that the genomic integrity of the cells remains intact and that the DNA repair systems are fully functional. In this article, we focused on the detection of DNA double-strand breaks (DSBs) by phosphorylated histone H2AX (known as γH2AX) and p53-binding protein 1 (53BP1) in three distinct lines of hiPSCs, their source cells, and one line of human embryonic stem cells (hESCs). METHODS: We measured spontaneously occurring DSBs throughout the process of fibroblast reprogramming and during long-term in vitro culturing. To assess the variations in the functionality of the DNA repair system among the samples, the number of DSBs induced by γ-irradiation and the decrease over time was analysed. The foci number was detected by fluorescence microscopy separately for the G1 and S/G2 cell cycle phases. RESULTS: We demonstrated that fibroblasts contained a low number of non-replication-related DSBs, while this number increased after reprogramming into hiPSCs and then decreased again after long-term in vitro passaging. The artificial induction of DSBs revealed that the repair mechanisms function well in the source cells and hiPSCs at low passages, but fail to recognize a substantial proportion of DSBs at high passages. CONCLUSIONS: Our observations suggest that cellular reprogramming increases the DSB number but that the repair mechanism functions well. However, after prolonged in vitro culturing of hiPSCs, the repair capacity decreases.

• Klíčová slova
• 53BP1, DNA double-strand breaks, DNA repair, Human induced pluripotent stem cells, Long-term in vitro culture, γH2AX,
• MeSH
• 53BP1 genetika   metabolismus   MeSH
• buněčné linie MeSH
• DNA genetika   metabolismus   MeSH
• dvouřetězcové zlomy DNA * účinky záření   MeSH
• exprese genu MeSH
• fibroblasty cytologie   metabolismus   účinky záření   MeSH
• fosforylace účinky záření   MeSH
• histony genetika   metabolismus   MeSH
• indukované pluripotentní kmenové buňky cytologie   metabolismus   účinky záření   MeSH
• kontrolní body fáze G1 buněčného cyklu genetika   MeSH
• kontrolní body fáze G2 buněčného cyklu genetika   MeSH
• lidé MeSH
• lidské embryonální kmenové buňky cytologie   metabolismus   účinky záření   MeSH
• oprava DNA genetika   MeSH
• přeprogramování buněk MeSH
• stárnutí buněk genetika   účinky záření   MeSH
• záření gama MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• 53BP1 MeSH
• DNA MeSH
• H2AX protein, human MeSH Prohlížeč
• histony MeSH
• TP53BP1 protein, human MeSH Prohlížeč

Most of the small-molecule drugs approved for the treatment of cancer over the past 40 years are based on natural compounds. Bacteria provide an extensive reservoir for the development of further anti-cancer therapeutics to meet the challenges posed by the diversity of these malignant diseases. While identifying cytotoxic compounds is often easy, achieving selective targeting of cancer cells is challenging. Here we describe a novel experimental approach (the Pioneer platform) for the identification and development of 'pioneering' bacterial variants that either show or are conduced to exhibit selective contact-independent anti-cancer cytotoxic activities. We engineered human cancer cells to secrete Colicin M that repress the growth of the bacterium Escherichia coli, while immortalised non-transformed cells were engineered to express Chloramphenicol Acetyltransferase capable of relieving the bacteriostatic effect of Chloramphenicol. Through co-culturing of E. coli with these two engineered human cell lines, we show bacterial outgrowth of DH5α E. coli is constrained by the combination of negative and positive selection pressures. This result supports the potential for this approach to screen or adaptively evolve 'pioneering' bacterial variants that can selectively eliminate the cancer cell population. Overall, the Pioneer platform demonstrates potential utility for drug discovery through multi-partner experimental evolution.

• MeSH
• antitumorózní látky * farmakologie   MeSH
• buněčné linie MeSH
• Escherichia coli genetika   MeSH
• kokultivační techniky MeSH
• lidé MeSH
• nádory * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antitumorózní látky * MeSH

Biomaterials containing citric acid as a building unit show potential for use as blood vessel and skin tissue substitutes. The success in commercializing implants containing a polymer matrix of poly(1,8-octanediol citrate) provides a rationale for exploring polycitrates based on other diols. Changing the aliphatic chain length of the diol allows functional design strategies to control the implant's mechanical properties, degradation profile and surface energy. In the present work, poly(1,2-ethanediol citrate) was synthesized and used as an additive to polylactide in the electrospinning process. It was established that the content of polycitrate greatly influences the nonwovens' properties: an equal mass ratio of polymers resulted in the best morphology. The obtained nonwovens were characterized by surface hydrophilicity, tensile strength, and thermal properties. L929 cell cultures were carried out on their surface. The materials were found to be non-cytotoxic and the degree of porosity was suitable for cell colonization. On the basis of the most important parameters for assessing the condition of cultured cells (cell density and viability, cell metabolic activity and lactate dehydrogenase activity), the potential of PLLA + PECit nonwovens for application in tissue engineering was established.

• Klíčová slova
• biomaterials, citrate-based polyesters, citric acid, electrospinning, nonwovens,
• Publikační typ
• časopisecké články MeSH

Natural cytokinins as well as the majority of their synthetic derivatives show negative effects on root growth and development. Changes in morphology, primarily linked to the inhibition of the cell division in the meristematic zone, are manifested as thickening and shortening of the primary root and impaired lateral root branching. Rational design of cytokinin derivatives can partially overcome these drawbacks and reduce the negative effects. Using our database of cytokinin derivatives, we selected several aromatic cytokinin analogs with modifications at the N9 atom of the adenine moiety. We found that tetrahydropyranyl and tetrahydrofuranyl substitutions at the N9 atom led to enhanced acropetal transport of the modified cytokinin, and both derivatives also showed weak anticytokinin activity. Consequently, changes in the distribution of the active cytokinin pool together with gradual metabolic conversion of the modified cytokinin to its free form prevent root growth inhibition that limits cytokinin utilization in micropropagation techniques.

• Klíčová slova
• N9-substituted cytokinins, biotechnology, cytokinin, inhibition, micropropagation, root, tissue culture,
• MeSH
• cytokininy chemie   farmakologie   MeSH
• kořeny rostlin účinky léků   MeSH
• kukuřice setá MeSH
• kultivační techniky * MeSH
• vývoj rostlin účinky léků   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• cytokininy MeSH

Clostridium perfringens is a Validated Biological Agent and a pathogen of medical, veterinary, and military significance. Gas gangrene is the most destructive of all the clostridial diseases and is caused by C. perfringens type A strains wherein the infection spreads quickly (several inches per hour) with production of gas. Influence of repeated in vitro cultivation on the infectivity of C. perfringens was investigated by comparing the surface proteins of laboratory strain and repository strains of the bacterium using 2DE-MS approach. In order to optimize host-pathogen interaction during experimental gas gangrene infection, we also explored the role of particulate matrix on ability of C. perfringens to cause gas gangrene.

• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• Clostridium perfringens genetika   růst a vývoj   fyziologie   MeSH
• gangréna plynatá mikrobiologie   MeSH
• interakce hostitele a patogenu MeSH
• lidé MeSH
• modely nemocí na zvířatech MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální proteiny MeSH

Optimising diagnostic methods in shelters so that they are as economical as possible for the shelter is especially important because shelters often have a significant lack of funds and so usually do not carry out preventive screening of cats. Dermatophyte fungi spread quickly and can infect shelter staff. The aim of our work was to identify the occurrence of Microsporum canis in shelter cats. It aimed to determine the prevalence of M. canis in cats at the selected shelter and compare the efficiency of detection using a Wood's lamp and culturing on Sabouraud's agar. All cats present in the shelter at the time of the study (n = 70) were examined with Wood's lamp and hair sampling followed by subsequent culturing on Sabouraud's agar. Identification of fungi was based on microscopic proof of macroconidia and microconidia. The prevalence of M. canis by diagnosis on Sabouraud's agar was 64.29% of cats, with the help of Wood's lamp 48.57% of cats showed positive fluorescence. The sensitivity of the Wood lamp examination was 71% and the specificity was 92%. Our study suggests that Wood's lamp could be used by trained shelter personnel for the first examination of cats at reception and could significantly reduce the risk of spreading M. canis in shelters.

• Klíčová slova
• Sabouraud’s agar, culturing, dermatophyte, skin disease,
• Publikační typ
• časopisecké články MeSH