Numerous protocols of cardiac differentiation have been established by essentially focusing on specific growth factors on human pluripotent stem cell (hPSC) differentiation efficiency. However, the optimal environmental factors to obtain cardiac myocytes in network are still unclear. The mesoderm germ layer differentiation is known to be enhanced by low oxygen exposure. Here, we hypothesized that low oxygen exposure enhances the molecular and functional maturity of the cardiomyocytes. We aimed at comparing the molecular and functional consequences of low (5% O2 or LOE) and high oxygen exposure (21% O2 or HOE) on cardiac differentiation of hPSCs in 2D- and 3D-based protocols. hPSC-CMs were differentiated through both the 2D (monolayer) and 3D (embryoid body) protocols using several lines. Cardiac marker expression and cell morphology were assessed. The mitochondrial localization and metabolic properties were evaluated. The intracellular Ca2+ handling and contractile properties were also monitored. The 2D cardiac monolayer can only be differentiated in HOE. The 3D cardiac spheroids containing hPSC-CMs in LOE further exhibited cardiac markers, hypertrophy, steadier SR Ca2+ release properties revealing a better SR Ca2+ handling, and enhanced contractile force. Preserved distribution of mitochondria and similar oxygen consumption by the mitochondrial respiratory chain complexes were also observed. Our results brought evidences that LOE is moderately beneficial for the 3D cardiac spheroids with hPSC-CMs exhibiting further maturity. In contrast, the 2D cardiac monolayers strictly require HOE.

• MeSH
• Biomarkers MeSH
• Cell Differentiation * MeSH
• Cell Culture Techniques MeSH
• Spheroids, Cellular MeSH
• Gene Expression MeSH
• Myocytes, Cardiac cytology   metabolism   MeSH
• Oxygen metabolism   MeSH
• Humans MeSH
• Pluripotent Stem Cells cytology   metabolism   MeSH
• Sarcoplasmic Reticulum metabolism   MeSH
• Mitochondria, Heart metabolism   MeSH
• Calcium metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

Carbon and its analogous nanomaterials are beneficial for toxic gas sensors since they are used to increase the electrochemically active surface region and improve the transmission of electrons. The present article addresses a detailed investigation on the potential of the monolayer PC3 compound as a possible sensor material for environmentally toxic nitrogen-containing gases (NCGs), namely NH3, NO, and NO2. The entire work is carried out under the frameworks of density functional theory, ab-initio molecular dynamics simulations, and non-equilibrium Green's function approaches. The monolayer-gas interactions are studied with the van der Waals dispersion correction. The stability of pristine monolayer PC3 is confirmed through dynamical, mechanical, and thermal analyses. The mobility and relaxation time of 2D PC3 sensor material with NCGs are obtained in the range of 101-104 cm2 V-1 s-1 and 101-103 fs for armchair and zigzag directions, respectively. Out of six possible adsorption sites for toxic gases on the PC3 surface, the most prominent site is identified with the highest adsorption energy for all the NCGs. Considering the most stable configuration site of the NCGs, we have obtained relevant electronic properties by utilizing the band unfolding technique. The considerable adsorption energies are obtained for NO and NO2 compared to NH3. Although physisorption is observed for all the NCGs on the PC3 surface, NO2 is found to convert into NO and O at 5.05 ps (at 300 K) under molecular dynamics simulation. The maximum charge transfer (0.31e) and work function (5.17 eV) are observed for the NO2 gas molecule in the series. Along with the considerable adsorption energies for NO and NO2 gas molecules, their shorter recovery time (0.071 s and 0.037 s, respectively) from the PC3 surface also identifies 2D PC3 as a promising sensor material for those environmentally toxic gases. The experimental viability and actual implications for PC3 monolayer as NCGs sensor material are also confirmed by examining the humidity effect and transport properties with modeled sensor devices. The transport properties (I-V characteristics) reflect the significant sensitivity of PC3 monolayer toward NO and NO2 molecules. These results certainly confirm PC3 monolayer as a promising sensor material for NO and NO2 NCG molecules.

• MeSH
• Adsorption MeSH
• Nitrogen MeSH
• Electrons MeSH
• Nanostructures * MeSH
• Gases * MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

The testis is a priority organ for developing alternative models to assess male reproductive health hazards of chemicals. This study characterized a 3D in vitro model of murine prepubertal Leydig TM3 cells with improved expression of steroidogenesis markers suitable for image-based screening of testicular toxicity. This 3D scaffold-free spheroid model was applied to explore the impact of prototypical endocrine-disrupting chemicals (EDCs) and environmental reprotoxicants (benzo[a]pyrene, 2- and 9-methylanthracenes, fluoranthene, triclosan, triclocarban, methoxychlor) on male reproductive health. The results were compared to the male reprotoxicity potential of EDCs assessed in a traditional monolayer (2D) culture. The testicular toxicity was dependent not only on the type of culture (2D vs. 3D models) but also on the duration of exposure. Benzo[a]pyrene and triclocarban were the most active compounds, eliciting cytotoxic effects in prepubertal Leydig cells at low micromolar concentrations, which might be a mechanism contributing to their male reprotoxicity.

• MeSH
• Benzo(a)pyrene toxicity   MeSH
• Endocrine Disruptors * chemistry   MeSH
• Leydig Cells * MeSH
• Mice MeSH
• Reproduction MeSH
• Testis MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

Three-dimensional cell culture systems are increasingly used for biological and anticancer drug screening as they mimic the structure and microenvironment of tumors more closely than conventional two-dimensional cell models. In this study, the growth kinetics of colon adenocarcinoma-derived spheroids (HT-29 cell line) cultivated in liquid marble micro-bioreactors and nonadherent PDMS-coated well plates was investigated in detail and enabled precise control of the spheroid size by the seed cell density and cultivation time. The therapeutic effect of 5-fluorouracil and irinotecan hydrochloride in 2D monolayer cell culture and 3D tumor spheroids revealed an unexpected twist in their efficacy due to different ability to penetrate through 3D microtissue. For 5-fluorouracil, the inhibitory concentration IC50 after 48 h exposure increased from 11.3 μM for a 2D cell culture to 707.7 μM for a 3D spheroid. In the case of irinotecan, IC50 increased from 24.9 μM to 77.8 μM. Despite its higher molar weight, irinotecan appeared to penetrate the 3D spheroid structure more efficiently than 5-fluorouracil. While 5-fluorouracil mainly caused a suppression of spheroid growth from the outside, irinotecan affected the entire spheroid and caused its originally compact structure to disintegrate. The acquired results highlight the need to screen cancer chemotherapeutics on 3D tumor models, as contrasting results can be obtained compared to standard 2D cell cultures.

• MeSH
• Adenocarcinoma * MeSH
• Spheroids, Cellular MeSH
• Cytostatic Agents * pharmacology   MeSH
• Fluorouracil pharmacology   MeSH
• Irinotecan pharmacology   MeSH
• Humans MeSH
• Cell Line, Tumor MeSH
• Tumor Microenvironment MeSH
• Colonic Neoplasms * drug therapy   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

Cardiovascular diseases remain the leading cause of death worldwide; hence there is an increasing focus on developing physiologically relevant in vitro cardiovascular tissue models suitable for studying personalized medicine and pre-clinical tests. Despite recent advances, models that reproduce both tissue complexity and maturation are still limited. We have established a scaffold-free protocol to generate multicellular, beating human cardiac microtissues in vitro from hiPSCs-namely human organotypic cardiac microtissues (hOCMTs)-that show some degree of self-organization and can be cultured for long term. This is achieved by the differentiation of hiPSC in 2D monolayer culture towards cardiovascular lineage, followed by further aggregation on low-attachment culture dishes in 3D. The generated hOCMTs contain multiple cell types that physiologically compose the heart and beat without external stimuli for more than 100 days. We have shown that 3D hOCMTs display improved cardiac specification, survival and metabolic maturation as compared to standard monolayer cardiac differentiation. We also confirmed the functionality of hOCMTs by their response to cardioactive drugs in long-term culture. Furthermore, we demonstrated that they could be used to study chemotherapy-induced cardiotoxicity. Due to showing a tendency for self-organization, cellular heterogeneity, and functionality in our 3D microtissues over extended culture time, we could also confirm these constructs as human cardiac organoids (hCOs). This study could help to develop more physiologically-relevant cardiac tissue models, and represent a powerful platform for future translational research in cardiovascular biology.

• MeSH
• Antineoplastic Agents * metabolism   MeSH
• Cell Differentiation physiology   MeSH
• Induced Pluripotent Stem Cells * MeSH
• Myocytes, Cardiac metabolism   MeSH
• Cardiovascular Agents * metabolism   MeSH
• Humans MeSH
• Heart physiology   MeSH
• Tissue Engineering methods   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Magnetic polymer nanocomposites are inherently multifunctional and harbor assorted physiochemical actions for applications thereof as novel drug nanocarriers. Herein, Fe3O4-nanoparticles were supported on rice straw cellulose for 5-fluorouracil carrier abbreviated as MC/5-FU for potential colorectal cancer treatments. Several analyses indicated the multifunctional properties of MC/5-FU bionanocomposites. Transmission and scanning electron microscopy study demonstrated that Fe3O4 nanofillers covered the cellulose matrix. The drug release from MC/5-FU was evaluated under various pH and temperature conditions, showing the maximum release at pH 7.4 and 44.2 °C. In in vitro anticancer assay, MC/5-FU exhibited enhanced selectivity and anticancer actions against 2D monolayer and 3D tumour spheroid models colorectal cancer cells. The anticancer effects of MC/5-FU with magnetic targeting and heat induction were also examined. This easily synthesized MC/5-FU indicated the potential in application as a low-cost drug formulation for colorectal cancer treatments.

• MeSH
• Antineoplastic Agents pharmacology   MeSH
• HT29 Cells MeSH
• Cellulose chemistry   MeSH
• Fluorouracil chemistry   pharmacology   MeSH
• HCT116 Cells MeSH
• Colorectal Neoplasms drug therapy   metabolism   MeSH
• Humans MeSH
• Magnetic Phenomena MeSH
• Magnetic Iron Oxide Nanoparticles chemistry   MeSH
• Microscopy, Electron, Scanning methods   MeSH
• Nanocomposites chemistry   MeSH
• Drug Carriers chemistry   MeSH
• Polymers chemistry   MeSH
• Drug Delivery Systems methods   MeSH
• Drug Liberation MeSH
• Cell Survival drug effects   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article 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
• Antineoplastic Agents * pharmacology   MeSH
• Apoptosis MeSH
• Phosphatidylinositol 3-Kinases MeSH
• Phosphorylcholine * analogs & derivatives   pharmacology   MeSH
• Protein Kinase Inhibitors pharmacology   MeSH
• Humans MeSH
• Cell Line, Tumor drug effects   MeSH
• Tumor Microenvironment MeSH
• Colonic Neoplasms * drug therapy   MeSH
• Proto-Oncogene Proteins c-akt metabolism   MeSH
• Drug Synergism MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

We present the synthesis and characterization of six new heteroleptic osmium(II) complexes of the type [Os(C^N)(N^N)2]OTf (N^N = 2,2'-bipyridine and dipyrido[3,2-d:2',3'-f]quinoxaline; C^N = deprotonated methyl 1-butyl-2aryl-benzimidazolecarboxylate) with varying substituents in the R3 position of the phenyl ring of the cyclometalating C^N ligand. The new compounds are highly kinetically inert and absorb a full-wavelength range of visible light. An investigation of the antiproliferative activity of the new compounds has been performed using a panel of human cancer and noncancerous 2D cell monolayer cultures under dark conditions and green light irradiation. The results demonstrate that the new Os(II) complexes are markedly more potent than conventional cisplatin. The promising antiproliferative activity of selected Os(II) complexes was also confirmed using 3D multicellular tumor spheroids, which have the characteristics of solid tumors and can mimic the tumor tissue microenvironment. The mechanism of antiproliferative action of complexes has also been investigated and revealed that the investigated Os(II) complexes activate the endoplasmic reticulum stress pathway in cancer cells and disrupt calcium homeostasis.

• MeSH
• Antineoplastic Agents * pharmacology   MeSH
• Benzimidazoles pharmacology   MeSH
• Homeostasis MeSH
• Coordination Complexes * pharmacology   MeSH
• Humans MeSH
• Cell Line, Tumor MeSH
• Neoplasms * MeSH
• Osmium pharmacology   MeSH
• Calcium MeSH
• Structure-Activity Relationship MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

The ability to simulate the 3D structure of a human body is essential to increase the efficiency of drug development. In vivo conditions are significantly different in comparison to in vitro conditions. A standardly used cell monolayer on tissue culture plastic (2D cell culture) is not sufficient to simulate the transfer phenomena occurring in living organisms, therefore, cell growth in a 3D space is desired. Drug absorption, distribution, metabolism, excretion and toxicity could be tested on 3D cell aggregates called spheroids, decrease the use of animal models and accelerate the drug development. In this work, the formation of spheroids from HT-29 human colorectal adenocarcinoma cells was successfully achieved by means of the so-called liquid marbles, which are liquid droplets encapsulated by a hydrophobic powder. During the cultivation in the medium inside the liquid marbles, cells spontaneously formed spherical agglomerates (spheroids) without the need of any supporting scaffold. The study focused on the influence of different parameters—namely liquid marble volume, seeding cell density and time of cultivation—on the final yield and quality of spheroids. This work has shown that using liquid marbles as microbioreactors is a suitable method for the cultivation of HT-29 cells in the form of spheroids.

The cell culture became an invaluable tool for studying cell behaviour, development, function, gene expression, toxicity of compounds and efficacy of novel drugs. Although most results were obtained from cell cultivation in two-dimensional (2D) systems, in which cells are grown in a monolayer, three-dimensional (3D) cultures are more promising as they correspond closely to the native arrangement of cells in living tissues. In our study, we focused on three types of 3D in vitro systems used for cultivation of one cell type. Cell morphology, their spatial distribution inside of resulting multicellular structures and changes in time were analysed with histological examination of samples harvested at different time periods. In multilayered cultures of WRL 68 hepatocytes grown on semipermeable membranes and non-passaged neurospheres generated by proliferation of neural progenitor cells, the cells were tightly apposed, showed features of cell differentiation but also cell death that was observable in short-term cultures. Biogenic scaffolds composed of extracellular matrix of the murine tibial anterior muscle were colonized with C2C12 myoblasts in vitro. The recellularized scaffolds did not reach high cell densities comparable with the former systems but supported well cell anchorage and migration without any signs of cell regression.

• MeSH
• Cell Culture Techniques MeSH
• Cell Line MeSH
• Spheroids, Cellular physiology   MeSH
• Hepatocytes physiology   MeSH
• Humans MeSH
• Myoblasts MeSH
• Mice MeSH
• Cell Proliferation physiology   MeSH
• Tissue Engineering methods   MeSH
• Tissue Scaffolds * MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH