BACKGROUND: Natural killer (NK) cell-based therapies represent a promising approach for acute myeloid leukemia (AML) relapse, yet their efficacy is hindered by immunosuppressive factors such as transforming growth factor beta (TGF-β) in the tumor microenvironment. This study investigated the effects of TGF-β on NK cell cytotoxicity and migration using 2D and 3D co-culture models that mimic the leukemic microenvironment. METHODS: TGF-β production was evaluated in AML-derived leukemic cell lines and mesenchymal stromal cells (hTERT-MSCs) using ELISA. Bulk RNA sequencing (RNA-seq) was performed to analyze global gene expression changes in TGF-β-treated primary human NK cells. NK cell cytotoxicity and migration were assessed in 2D monolayer and 3D spheroid co-cultures containing hTERT-MSCs and leukemic cells using flow cytometry and confocal microscopy. RESULTS: Both leukemic cells and MSCs produced TGF-β, with increased levels observed in MSCs after co-culture with primary AML blasts. RNA sequencing revealed that TGF-β altered key gene pathways associated with NK cell cytotoxicity, adhesion, and migration, supporting its immunosuppressive role. In functional assays, TGF-β exposure significantly reduced NK cell-mediated cytotoxicity in a time-dependent manner and impaired NK cell infiltration into 3D spheroids, particularly in models incorporating MSCs. Additionally, MSCs themselves provided a protective environment for leukemic cells, further reducing NK cell effectiveness in 2D co-cultures. CONCLUSION: TGF-β suppresses both NK cell cytotoxicity and migration, limiting their ability to eliminate leukemic cells and infiltrate the bone marrow niche (BMN). These findings provide novel insights into TGF-β-mediated immune evasion mechanisms and provide important insights for the future design of NK-based immunotherapies and clinical trials.

• Publikační typ
• časopisecké články MeSH

BACKGROUND AND OBJECTIVE: Microbial selenium (Se) supplementation is an essential area of biotechnological research due to differences in the bioavailability and toxicity of different forms of selenium. To date, research has focused mainly on the use of selenized yeast. However, in recent years, scientific interest has also increased in other microorganisms, such as lactic acid bacteria (LAB), which have several unique properties that can affect the quality and bioavailability of selenium. LAB, unlike yeast, can also act as probiotics, which may bring additional health benefits related to improving the intestinal microbiota and supporting the health of the gastrointestinal tract. METHODS: This study investigates the in vitro bioaccessibility and bioavailability of Se from two lactic acid bacterial strains, Streptococcus thermophilus CCDM 144 and Enterococcus faecium CCDM 922 A. We evaluated Se accumulation, speciation, and stability during simulated gastrointestinal digestion and Se permeation through a Caco-2 cell monolayer model. RESULTS: Both strains accumulated Se, metabolizing it predominantly into selenium nanoparticles (SeNPs, 64-77 % of total Se), with only a minor fraction (<5 % of total Se) of organic Se species. Experiments revealed that while organic Se species had high bioavailability (up to 90 %), their bioaccessibility during digestion was very low (<0.1 % of total Se). In contrast, SeNPs showed high bioaccessibility (∼90 %) and moderate transport efficiency through the intestinal model (16-19 % after 4 hours). CONCLUSION: These results highlight the potential of SeNPs produced by lactic acid bacteria as a bioaccessible form of Se for dietary supplementation. Further research is required to explore the behavior of SeNPs within the human body to fully understand how they can be used safely and effectively in nutrition or other applications.

• MeSH
• biologická dostupnost * MeSH
• biologické modely MeSH
• Caco-2 buňky MeSH
• funkce střevní bariéry MeSH
• Lactobacillales metabolismus   MeSH
• lidé MeSH
• permeabilita MeSH
• selen * metabolismus   MeSH
• Streptococcus thermophilus metabolismus   MeSH
• trávení MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

While two-dimensional (2D) cell cultures, such as Caco-2 and Madin-Darby canine kidney (MDCK) cells are widely used in a variety of biological models, these two-dimensional in vitro systems present inherent limitations in replicating the complexities of in vivo biology. Recent progress in three-dimensional organoid technology has the potential to address these limitations. In this study, the characteristics of conventional 2D cell culture systems were compared to those of canine intestinal organoids (enteroids, ENT, and colonoids, COL). Light microscopy and transmission electron microscopy were employed to evaluate the microanatomy of ENT, COL, Caco-2, and MDCK cell monolayers, while transepithelial electrical resistance (TEER) values were measured to assess monolayer integrity. The TEER values of canine ENT monolayers more closely approximated reported TEER values for human small intestines compared to Caco-2 and MDCK monolayers. Additionally, canine ENT demonstrated greater monolayer stability than Caco-2 and MDCK cells. Notably, while all systems displayed desmosomes, canine ENT and COL exclusively produced mucus. These findings highlight the potential of the canine organoid system as a more biologically relevant model for in vitro studies, addressing the limitations of conventional 2D cell culture systems.

• Publikační typ
• časopisecké články MeSH

Medulloblastoma, the most prevalent brain tumor among children, requires a comprehensive understanding of its cellular characteristics for effective research and treatment. In this study, we focused on DAOY, a permanent cell line of medulloblastoma, and investigated the unique properties of DAOY cells when cultured as floating multicellular aggregates called spheres, as opposed to adherent monolayers. Through our comprehensive analysis, we identified distinct characteristics associated with DAOY spheres. Our findings demonstrate that DAOY spheres express markers for both neural stem cells, such as CD133 (PROM1), and differentiated neurons, exemplified by MAP2. Additionally, our investigation revealed that spheres-derived cells exhibit heightened resistance to ionizing radiation compared to adherent cells. Consequently, our results indicate that caution is advised when interpreting experimental results obtained from adherent cell cultures and extrapolating them to in vivo situations.

• Publikační typ
• časopisecké články MeSH

Herein, we describe the general design, synthesis, characterization, and biological activity of new multitargeting Pt(IV) prodrugs that combine antitumor cisplatin and dasatinib, a potent inhibitor of Src kinase. These prodrugs exhibit impressive antiproliferative and anti-invasive activities in tumor cell lines in both two-dimensional (2D) monolayers of cell cultures and three-dimensional (3D) spheroids. We show that the cisplatin moiety and dasatinib in the investigated Pt(IV) complexes are both involved in the mechanism of action in MCF7 breast cancer cells and act synergistically. Thus, combining dasatinib and cisplatin into one molecule, compared to using individual components in a mix, may bring several advantages, such as significantly higher activity in cancer cell lines and higher selectivity for tumor cells. Most importantly, Pt(IV)-dasatinib complexes hold significant promise for potential anticancer therapies by targeting epithelial-mesenchymal transition, thus preventing the spread and metastasis of tumors, a value unachievable by a simple combination of both individual components.

• MeSH
• cisplatina * farmakologie   MeSH
• dasatinib * farmakologie   chemie   chemická syntéza   MeSH
• lidé MeSH
• MFC-7 buňky MeSH
• nádorové buněčné linie MeSH
• organoplatinové sloučeniny farmakologie   chemie   chemická syntéza   MeSH
• prekurzory léčiv * farmakologie   chemie   chemická syntéza   MeSH
• proliferace buněk účinky léků   MeSH
• protinádorové látky * farmakologie   chemie   chemická syntéza   MeSH
• screeningové testy protinádorových léčiv MeSH
• synergismus léků * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Statins, the drugs used for the treatment of hypercholesterolemia, have come into the spotlight not only as chemoadjuvants, but also as potential stem cell modulators in the context of regenerative therapy. In our study, we compared the in vitro effects of all clinically used statins on the viability of human pancreatic cancer (MiaPaCa-2) cells, non-cancerous human embryonic kidney (HEK 293) cells and adipose-derived mesenchymal stem cells (ADMSC). Additionally, the effect of statins on viability of MiaPaCa-2 and ADMSC cells spheroids was tested. Furthermore, we performed a microarray analysis on ADMSCs treated with individual statins (12 μM) and compared the importance of the effects of statins on gene expression between stem cells and pancreatic cancer cells. Concentrations of statins that significantly affected cancer cells viability (< 40 μM) did not affect stem cells viability after 24 h. Moreover, statins that didn ́t affect viability of cancer cells grown in a monolayer, induce the disintegration of cancer cell spheroids. The effect of statins on gene expression was significantly less pronounced in stem cells compared to pancreatic cancer cells. In conclusion, the low efficacy of statins on non-tumor and stem cells at concentrations sufficient for cancer cells growth inhibition, support their applicability in chemoadjuvant tumor therapy.

• MeSH
• buněčné sféroidy účinky léků   MeSH
• HEK293 buňky MeSH
• lidé MeSH
• mezenchymální kmenové buňky * účinky léků   metabolismus   MeSH
• nádorové buněčné linie MeSH
• nádory slinivky břišní * farmakoterapie   patologie   metabolismus   MeSH
• statiny * farmakologie   MeSH
• viabilita buněk * účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Portable household air purifiers are widely used devices designed to maintain a high-quality level of indoor air. Portable air purifiers equipped with the high-efficiency air (HEPA) filter served 100 h in a household space occupied by two adults without any symptoms of respiratory tract infection. The main objective of the study was to determine microbial contamination on the HEPA filter and to investigate if the selected nanotextile monolayer made of polyamide 6 (PA6) nanofibers can capture potential microorganisms when installed downstream of the HEPA filter as the final filtration medium. Samples were taken from the inlet and outlet surfaces. Samples from the nanotextile were collected in the same manner as from the HEPA filter. QIAStat DX® 1.0 Analyzer using the Respiratory SARS CoV-2 Panel multiplex PCR detection system was selected for microorganism detection. Adenovirus was detected on the inlet surface of the HEPA filter. The outlet surface of the filter contained no viruses included in the Respiratory SARS CoV-2 Panel portfolio. The nanotextile monolayer was replaced twice during the 100 h of operation, so three pieces were used and all contained coronavirus 229 E. Coronavirus 229 E was then detected in the nasopharynx of one of the members of the household as well. It may be assumed that the selected nanotextile is capable of capturing a virus of a small size.

• MeSH
• filtrace MeSH
• lidé MeSH
• pilotní projekty MeSH
• SARS-CoV-2 MeSH
• syndrom akutního respiračního selhání * MeSH
• vzduchové filtry * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

The placenta plays a critical role in maternal-fetal nutrient transport and fetal protection against drugs. Creating physiological in vitro models to study these processes is crucial, but technically challenging. This study introduces an efficient cell model that mimics the human placental barrier using co-cultures of primary trophoblasts and primary human umbilical vein endothelial cells (HUVEC) on a Transwell®-based system. Monolayer formation was examined over 7 days by determining transepithelial electrical resistance (TEER), permeability of Lucifer yellow (LY) and inulin, localization of transport proteins at the trophoblast membrane (immunofluorescence), and syncytialization markers (RT-qPCR/ELISA). We analysed diffusion-based (caffeine/antipyrine) and transport-based (leucine/Rhodamine-123) processes to study the transfer of physiologically relevant compounds. The latter relies on the adequate localization and function of the amino-acid transporter LAT1 and the drug transporter P-glycoprotein (P-gp) which were studied by immunofluorescence microscopy and application of respective inhibitors (2-Amino-2-norbornanecarboxylic acid (BCH) for LAT1; cyclosporine-A for P-gp). The formation of functional monolayer(s) was confirmed by increasing TEER values, low LY transfer rates, minimal inulin leakage, and appropriate expression/release of syncytialization markers. These results were supported by microscopic monitoring of monolayer formation. LAT1 was identified on the apical and basal sides of the trophoblast monolayer, while P-gp was apically localized. Transport assays confirmed the inhibition of LAT1 by BCH, reducing both intracellular leucine levels and leucine transport to the basal compartment. Inhibiting P-gp with cyclosporine-A increased intracellular Rhodamine-123 concentrations. Our in vitro model mimics key aspects of the human placental barrier. It represents a powerful tool to study nutrient and drug transport mechanisms across the placenta, assisting in evaluating safer pregnancy therapies.

• MeSH
• biologické modely MeSH
• biologický transport MeSH
• endoteliální buňky pupečníkové žíly (lidské) * metabolismus   MeSH
• inulin metabolismus   MeSH
• isochinoliny MeSH
• kokultivační techniky MeSH
• leucin metabolismus   MeSH
• lidé MeSH
• maternofetální výměna látek * MeSH
• P-glykoprotein metabolismus   MeSH
• placenta * metabolismus   MeSH
• rhodamin 123 metabolismus   MeSH
• těhotenství MeSH
• trofoblasty * metabolismus   MeSH
• Check Tag
• lidé MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

An acidic environment and hypoxia within the tumour are hallmarks of cancer that contribute to cell resistance to therapy. Deregulation of the PI3K/Akt pathway is common in colon cancer. Numerous Akt-targeted therapies are being developed, the activity of Akt-inhibitors is, however, strongly pH-dependent. Combination therapy thus represents an opportunity to increase their efficacy. In this study, the cytotoxicity of the Akt inhibitor perifosine and the Bcl-2/Bcl-xL inhibitor ABT-737 was tested in colon cancer HT-29 and HCT-116 cells cultured in monolayer or in the form of spheroids. The efficacy of single drugs and their combination was analysed in different tumour-specific environments including acidosis and hypoxia using a series of viability assays. Changes in protein content and distribution were determined by immunoblotting and a "peeling analysis" of immunohistochemical signals. While the cytotoxicity of single agents was influenced by the tumour-specific microenvironment, perifosine and ABT-737 in combination synergistically induced apoptosis in cells cultured in both 2D and 3D independently on pH and oxygen level. Thus, the combined therapy of perifosine and ABT-737 could be considered as a potential treatment strategy for colon cancer.

• MeSH
• apoptóza MeSH
• fosfatidylinositol-3-kinasy MeSH
• fosforylcholin * analogy a deriváty   farmakologie   MeSH
• inhibitory proteinkinas farmakologie   MeSH
• lidé MeSH
• nádorové buněčné linie účinky léků   MeSH
• nádorové mikroprostředí MeSH
• nádory tračníku * farmakoterapie   MeSH
• protinádorové látky * farmakologie   MeSH
• protoonkogenní proteiny c-akt metabolismus   MeSH
• synergismus léků MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Helicobacter pylori infection is a major risk factor for the development of gastric cancer. The bacteria reside in close proximity to gastric surface mucous as well as stem and progenitor cells. Here, we take advantage of wild-type and genetically engineered murine gastric organoids and organoid-derived monolayers to study the cellular targets of H. pylori-induced DNA damage and replication stress and to explore possible interactions with preexisting gastric cancer driver mutations. We find using alkaline comet assay, single-molecule DNA fiber assays, and immunofluorescence microscopy of DNA repair foci that H. pylori induces transcription-dependent DNA damage in actively replicating, Leucine-rich-repeat containing G-Protein-Coupled Receptor 5 (Lgr5)-positive antral stem and progenitor cells and their Troy-positive corpus counterparts, but not in other gastric epithelial lineages. Infection-dependent DNA damage is aggravated by Apc inactivation, but not by Trp53 or Smad4 loss, or Erbb2 overexpression. Our data suggest that H. pylori induces DNA damage in stem and progenitor cells, especially in settings of hyperproliferation due to constitutively active Wnt signaling.

• MeSH
• Helicobacter pylori * genetika   MeSH
• infekce vyvolané Helicobacter pylori * genetika   mikrobiologie   MeSH
• kmenové buňky MeSH
• lidé MeSH
• myši MeSH
• nádory žaludku * patologie   MeSH
• poškození DNA MeSH
• receptory spřažené s G-proteiny genetika   MeSH
• tumor supresorové geny MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH