Ilona Grabowska-Jadach

Multi-function microsystem for cells migration analysis and evaluation of photodynamic therapy procedure in coculture

Cell migration is an important physiological process, which is involved in cancer metastasis. Therefore, the investigation of cell migration may lead to the development of novel therapeutic approaches. In this study, we have successfully developed a microsystem for culture of two cell types (non-malignant and carcinoma) and for analysis of cell migration dependence on distance between them. Finally, we studied quantitatively the influence of photodynamic therapy (PDT) procedures on the viability of pairs of non-malignant (MRC5 or Balb/3T3) and carcinoma (A549) cells coculture. The proposed geometry of the microsystem allowed for separate introduction of two cell lines and analysis of cells migration dependence on distance between the cells. We found that a length of connecting microchannel has an influence on cell migration and viability of non-malignant cells after PDT procedure. Summarizing, the developed microsystem can constitute a new tool for carrying out experiments, which offers a few functions: cell migration analysis, carcinoma and non-malignant cells coculture, and evaluation of PDT procedure in the various steps of cell migration.

Studies on influence of polymer modifiers for fluorescent nanocrystals’ cytotoxicity

The presented studies aimed at investigation of the effect of CdSeS/ZnS quantum dots (QDs) stabilized with hyperbranched polyglycidol and its carboxylated derivative on adenocarcinomic human alveolar basal epithelial cells (A549). The first stage of studies concerned the modification of quantum dots with both types of the tested polymers with the use of pyridine as an intermediate agent. Subsequently, cytotoxic effect of the prepared nanoparticles was examined after various incubation time using MTT test (cell metabolic activity assay). Our studies revealed that CdSeS/ZnS with a diameter of 6nm, which were stabilized with hyperbranched polymers do not penetrate into cells, even after prolonged incubation time. Moreover, the cytotoxic effect of the tested QDs was observed over a range of tested concentrations (5-90μM of Cd(2+)). It was confirmed that tested nanoparticles had significant influence on cell culture viability. The examined cytotoxic effect of the tested quantum dots was dependent on the type of polymer applied and the experiments indicated, that the one bearing carboxylic moieties is more toxic to A549 cells.

Cytotoxicity studies of CdSeS/ZnS quantum dots on cell culture in microfluidic system

Quantum dots (QDs) semi-conducting nanocrystals have found numerous applications in many fields of science. Nowadays one can observe a growing perspective to use them in biomedicine. Thanks to QDs unique fluorescence properties (narrow emission spectra, high extinction coefficients, high quantum yields, photostability) and possibility to form conjugates with bioactive molecules, they can become a chance for better cancer cells imaging in cancer therapy. Therefore there is a need for better understanding of biological interactions between QDs and cancer cells in vitro. For this purpose we performed cytotoxicity tests of CdSeS/ZnS quantum dots stabilized with mercaptopropionic acid (MPA) ligand, on human lung cancer cell line (A549) in vitro in macro- (96-well plate) and micro-scale (a specially designed and fabricated microfluidic device). The results obtained demonstrated a little extent of cytotoxic effect of selected solutions of QDs to A549 cells.

Possible role of hydrolytic enzymes (Sap, Kex2) in Candida albicans response to aromatic compounds bearing a sulfone moiety

Hydrolytic enzymes e.g., Saps and Kex2 are, due to their role in Candida virulence, considered important targets for new synthetic inhibitors. MICTI and MICPI values indicate that disruption of SAP1-3 significantly increases the resistance of Candida mutants to β-ketosulfone (1). Contrariwise, sap123Δ showed sensitive phenotype to halogenated methylphenyl sulfone (2). Anticandidal potency of 2 differed in the Candida cells of kex2Δ. Sulfone is the most effective agent against the Candida albicans kex2Δ double mutant (MICTI of 0.5 µg mL−1). Up-regulation of KEX2 mediated the resistance of sap4-6Δ towards 2. Both sulfones tested reduced the adhesion of the wild type cells significantly (P ≤ 0.05). Contrariwise, sap123Δ showed significantly enhanced adhesion capability when 1 was used (P ≤ 0.05). Both sulfones had weak fungicidal effect on mature C. albicans biofilms. It was shown that the uptake of IP correlates with the membrane perturbations caused by 1 in the blastoconidial cells. Sulfones were found to disturb the basic developmental phases of biofilm growth: adhesion and morphogenesis. Altered KEX2 levels for 1 can be caused by the compensatory mechanism for the maintenance of cell wall integrity and morphogenesis. KEX2 decreases the antifungal activity of sulfones. Sulfones affecting the crucial virulence factors of Candida can even eliminate these fungal infections.

Lab-on-a-chip Systems for Cellomics—Materials and Technology

The main advantage of the microsystems is their ability to imitate in vivo conditions which are missing in macroscale cell cultures. The materials which find applications in Lab-on-a-chip devices for cellomics, their properties, and microfabrication techniques are presented in this chapter. Such microfluidic devices are useful tools in many fields involving cell culture studies, e.g., cell trapping, counting or sorting, cell lysis and fusion, cultivation, and drug screening. Construction materials, not only the most commonly used poly(dimethyl siloxane) (PDMS) and glass, but also such polymers as polystyrene (PS), poly(methyl methacrylate) (PMMA), polycarbonate (PC), and cyclic olefin copolymer (COC), are presented. There are many materials which are utilized to create spatial arrangement of the cells in the developed microsystems. For this purpose, natural (e.g., collagen), synthetic (e.g., poly(ethylene glycol)—PEG), and hybrid (e.g., gelatin methacryloyl—GelMA) hydrogels as well as nanofibrous scaffolds are applied. We present short description and some examples of the usage of above materials. This chapter also describes the most common fabrication methods of Lab-on-a-chip (LOC) devices for cellomics. These considerations are extended to potential mass production of cell-based microsystems, using a range of materials. Adopting more time and cost-effective fabrication methods is critical for the integration of LOCs into mainstream applications, and therefore, factors such as quality control or device repeatability were detailed.

Cytotoxicity studies of selected cadmium-based quantum dots on 2D vs. 3D cell cultures

In our work, the cytotoxicity of selected, cadmium-based quantum dots with various surface architectures was studied on 3D spheroids. A specially designed microsystem as a tool for three-dimensional cell culture and nanoparticle toxicity evaluation was used for this aim. Two types of hydrophilic quantum dots with different surface charges at physiological pH were examined: CdTe-capped with ω-mercaptocarboxylic acid (COO−-terminated) and CdSeS/ZnS-glucosamine (–OH-terminated). We studied the influence of five different concentrations of nanoparticles within the range of 5 to 100 μM in order to assess dose-dependent toxic effects of the selected quantum dots on cellular spheroids as a more realistic (in vivo-like) model of human tissue. The obtained results of cytotoxicity were compared with the results for a standard, two-dimensional model – a cell monolayer. In the case of three-dimensional structures of various cell lines (normal MRC-5 and tumor A549) no significant differences in cytotoxicity caused by the tested nanoparticles have been noticed. The comparative studies revealed the enhanced biocompatibility of CdSeS/ZnS-OH quantum dots resulting from the presence of an uncharged ligand of biomimetic character on their surface. It was also found that the cytotoxicity of the same quantum dots depends on the cell culture model on which the tests were conducted. Significantly higher cytotoxicity of the tested nanomaterial was observed when experiments were carried out with the use of cell monolayers. Based on the obtained results, we claim that cytotoxicity studies of nanomaterials conducted on standard 2D cell monolayers are overestimated. In our opinion, reliable in vitro studies on the biological activity of nanoparticles require application of 3D cell cultures.

Comparative studies of biological activity of cadmium-based quantum dots with different surface modifications

This paper presents a modification of the surface of CdS/ZnS and CdSexS1−x/ZnS quantum dots (QDs) with 3-mercaptopropionic and 6-mercaptohexanoic acid. The obtained QDs were characterized using TEM, DLS, UV–Vis, and fluorescence spectroscopy. Flow cytometry was applied to evaluate the cytotoxicity of QDs and examine the type of death caused by the tested nanoparticles. In addition, the generation of reactive oxygen species after incubation of the tested cells with CdSexS1−x/ZnS–MPA and CdSexS1−x/ZnS–MHA QDs was evaluated. The study was conducted on three cell lines: adherent (A549 and MRC-5) and suspension ones (K562). The conducted research demonstrated that the tested nanoparticles exhibit concentration-dependent toxicity. It was observed that the surface modification influences the toxicity level of the examined QDs, and modification of their surface with the use of the ligand of longer carbon chain (MHA) reduces the toxicity in comparison with QDs–MPA. It was also found that all tested QDs caused the death of cells in the course of necrosis. Based on obtained results, it was concluded that the cytotoxicity of QDs is to a large extent related to reactive oxygen species (ROS) generation.