Pancreatic cancer, due to its asymptomatic development and drug-resistance, is difficult to cure. As many metallic and carbon-based nanomaterials have shown anticancer properties, we decided to ...investigate their potential use as anticancer agents against human pancreatic adenocarcinoma. The objective of the study was to evaluate the toxic properties of the following nanomaterials: silver (Ag), gold (Au), platinum (Pt), graphene oxide (GO), diamond (ND), and fullerenol (C
(OH)
) against the cell lines BxPC-3, AsPC-1, HFFF-2, and HS-5. The potential cytotoxic properties were evaluated by the assessment of the cell morphology, cell viability, and cell membrane damage. The cancer cell responses to GO and ND were analysed by determination of changes in the levels of 40 different pro-inflammatory proteins. Our studies revealed that the highest cytotoxicity was obtained after the ND treatment. Moreover, BxPC-3 cells were more sensitive to ND than AsPC-1 cells due to the ND-induced ROS production. Furthermore, in both of the cancer cell lines, ND caused an increased level of IL-8 and a decreased level of TIMP-2, whereas GO caused only decreased levels of TIMP-2 and ICAM-1 proteins. This work provides important data on the toxicity of various nanoparticles against pancreatic adenocarcinoma cell lines.
There are numerous applications of graphene in biomedicine and they can be classified into several main areas: delivery systems, sensors, tissue engineering and biological agents. The growing ...biomedical field of applications of graphene and its derivates raises questions regarding their toxicity. We will demonstrate an analysis of the toxicity of two forms of graphene using four various biological models: zebrafish (Danio rerio) embryo, duckweed (Lemna minor), human HS-5 cells and bacteria (Staphylococcus aureus). The toxicity of pristine graphene (PG) and graphene oxide (GO) was tested at concentrations of 5, 10, 20, 50 and 100 µg/mL. Higher toxicity was noted after administration of high doses of PG and GO in all tested biological models. Hydrophilic GO shows greater toxicity to biological models living in the entire volume of the culture medium (zebrafish, duckweed, S. aureus). PG showed the highest toxicity to adherent cells growing on the bottom of the culture plates—human HS-5 cells. The differences in toxicity between the tested graphene materials result from their physicochemical properties and the model used. Dose-dependent toxicity has been demonstrated with both forms of graphene.
Finding an effective muscle regeneration technique is a priority for regenerative medicine. It is known that the key factors determining tissue formation include cells, capable of proliferating ...and/or differentiating, a niche (surface) allowing their colonization and growth factors. The interaction between these factors, especially between the surface of the artificial niche and growth factors, is not entirely clear. Moreover, it seems that the use of a complex of complementary growth factors instead of a few strictly defined ones could increase the effectiveness of tissue maturation, including muscle tissue. In this study, we evaluated whether graphene oxide (GO) nanofilm, chicken embryo muscle extract (CEME), and GO combined with CEME would affect the differentiation and functional maturation of muscle precursor cells, as well as the ability to spontaneously contract a pseudo-tissue muscle. CEME was extracted on day 18 of embryogenesis. Muscle cells obtained from an 8-day-old chicken embryo limb bud were treated with GO and CEME. Cell morphology and differentiation were observed using different microscopy methods. Cytotoxicity and viability of cells were measured by lactate dehydrogenase and Vybrant Cell Proliferation assays. Gene expression of myogenic regulatory genes was measured by Real-Time PCR. Our results demonstrate that CEME, independent of the culture surface, was the main factor influencing the intense differentiation of muscle progenitor cells. The present results, for the first time, clearly demonstrated that the cultured tissue-like structure was capable of inducing contractions without externally applied impulses. It has been indicated that a small amount of CEME in media (about 1%) allows the culture of pseudo-tissue muscle capable of spontaneous contraction. The study showed that the graphene oxide may be used as a niche for differentiating muscle cells, but the decisive influence on the maturation of muscle tissue, especially muscle contractions, depends on the complexity of the applied growth factors.
Graphene (GN) and its derivatives (rGOs) show anticancer properties in glioblastoma multiforme (GBM) cells in vitro and in tumors in vivo. We compared the anti-tumor effects of rGOs with different ...oxygen contents with those of GN, and determined the characteristics of rGOs useful in anti-glioblastoma therapy using the U87 glioblastoma line. GN/ExF, rGO/Term, rGO/ATS, and rGO/TUD were structurally analysed via transmission electron microscopy, Raman spectroscopy, FTIR, and AFM. Zeta potential, oxygen content, and electrical resistance were determined. We analyzed the viability, metabolic activity, apoptosis, mitochondrial membrane potential, and cell cycle. Caspase- and mitochondrial-dependent apoptotic pathways were investigated by analyzing gene expression. rGO/TUD induced the greatest decrease in the metabolic activity of U87 cells. rGO/Term induced the highest level of apoptosis compared with that induced by GN/ExF. rGO/ATS induced a greater decrease in mitochondrial membrane potential than GN/ExF. No significant changes were observed in the cytometric study of the cell cycle. The effectiveness of these graphene derivatives was related to the presence of oxygen-containing functional groups and electron clouds. Their cytotoxicity mechanism may involve electron clouds, which are smaller in rGOs, decreasing their cytotoxic effect. Overall, cytotoxic activity involved depolarization of the mitochondrial membrane potential and the induction of apoptosis in U87 glioblastoma cells.
Background
The extracellular matrix (ECM) is a mosaic of various structural and functional proteins that cooperate with the cell, regulate adhesion, and consequently manage its further fate. Liver ...destruction is accompanied by a disruption of the physicochemical properties of the ECM which deregulates the cell–ECM interaction and can lead to uncontrolled proliferation and neoplastic transformation of cells. Therefore, it can be assumed that ECM modification and restoration of its characteristics for healthy tissue may counteract uncontrolled cell proliferation. The purpose of the presented research model was to optimise the physical characteristics of ECM by introducing a graphene oxide plane/nanofilm (nfGO) and enriching the cell environment with potentially missing proteins by adding a functional protein cocktail (chicken embryo liver extract) and determine the impact of these factors on cell–ECM cooperation and its consequences on adhesion, proliferation, and cell phase, which are factors of the invasiveness of cancer cells.
Results
Experiments were performed with non-cancer HS-5 cells and liver cancer cells HepG2 and C3A. The cells were divided into four groups: (1) control, (2) cultured on nfGO, (3) cultured with the addition of chicken embryo liver extract (CELE) and (4) cultured on the nfGO with the addition of CELE. CELE contained 1735 proteins; the top 57 of these proteins have been presented. The use of nfGO as well as CELE and nfGO + CELE reduced the proliferation of HepG2 cancer cells to the greatest extent; this is in contrast to non-cancer cells and also to C3A cancer cells. Furthermore, the combined use of the CELE protein cocktail and GO substrate effectively resulted in a decrease in the population of HepG2 cells in the G0/G1 phase and an increase of the population in G2/M. Molecular analysis of HepG2 cancer cells also showed an increase in the expression of genes responsible for adhesion such as
focal adhesion kinase
(
fak
),
e-cadherin,
and
n-cadherin
and a decrease in
β-catenin
, which is considered a proto-oncogene.
Conclusions
Studies have shown that both the GO surface structure on which the cells are grown as well as the presence of a multi-component natural cocktail of regulatory proteins, can modify the expression of integrins, increase adhesion and, as a consequence, proliferation and the cell cycle—entering the resting phase. For the first time, it has been documented that hepatic cancer cells of the HepG2 line under the influence of stimuli derived from mimic ECM (graphene oxide) in interaction with a unique protein complex derived from chicken liver embryo decreased of the invasiveness of cancer cells.
Aggressive invasiveness is a common feature of malignant gliomas, despite their high level of tumor heterogeneity and possible diverse cell origins. Therefore, it is important to explore new ...therapeutic methods. In this study, we evaluated and compared the effects of graphene (GN) and reduced graphene oxides (rGOs) on a highly invasive and neoplastic cell line, U87. The surface functional groups of the GN and rGO flakes were characterized by X-ray photoelectron spectroscopy. The antitumor activity of these flakes was obtained by using the neutral red assay and their anti-migratory activity was determined using the wound healing assay. Further, we investigated the mRNA and protein expression levels of important cell adhesion molecules involved in migration and invasiveness. The rGO flakes, particularly rGO/ATS and rGO/TUD, were found highly toxic. The migration potential of both U87 and Hs5 cells decreased, especially after rGO/TUD treatment. A post-treatment decrease in mobility and FAK expression was observed in U87 cells treated with rGO/ATS and rGO/TUD flakes. The rGO/TUD treatment also reduced β-catenin expression in U87 cells. Our results suggest that rGO flakes reduce the migration and invasiveness of U87 tumor cells and can, thus, be used as potential antitumor agents.
Abstract
The increasing emergence of antibiotic-resistant bacteria and the need to reduce the use of antibiotics call for the development of safe alternatives
,
such as silver nanoparticles. However, ...their potential cytotoxic effect needs to be addressed. Graphene oxide provides a large platform that can increase the effectiveness and safety of silver nanoparticles. Graphene oxide and silver nanoparticles complex applied as a part of an innovative material might have direct contact with human tissues, such as skin, or might be inhaled from aerosol or exfoliated pieces of the complex. Thereby, the safety of the prepared complex has to be evaluated carefully, employing a range of methods. We demonstrated the high cytocompatibility of graphene oxide and the graphene oxide–silver nanoparticles complex toward human cell lines, fetal foreskin fibroblasts (HFFF2), and lung epithelial cells (A549). The supporting platform of graphene oxide also neutralized the slight toxicity of bare silver nanoparticles. Finally, in studies on
Staphylococcus aureus
and
Pseudomonas aeruginosa
, the number of bacteria reduction was observed after incubation with silver nanoparticles and the graphene oxide–silver nanoparticles complex. Our findings confirm the possibility of employing a graphene oxide–silver nanoparticles complex as a safe agent with reduced silver nanoparticles’ cytotoxicity and antibacterial properties.
The development of nanotechnology based on graphene and its derivatives has aroused great scientific interest because of their unusual properties. Graphene (GN) and its derivatives, such as reduced ...graphene oxide (rGO), exhibit antitumor effects on glioblastoma multiforme (GBM) cells in vitro. The antitumor activity of rGO with different contents of oxygen-containing functional groups and GN was compared. Using FTIR (fourier transform infrared) analysis, the content of individual functional groups (GN/exfoliation (ExF), rGO/thermal (Term), rGO/ammonium thiosulphate (ATS), and rGO/ thiourea dioxide (TUD)) was determined. Cell membrane damage, as well as changes in the cell membrane potential, was analyzed. Additionally, the gene expression of voltage-dependent ion channels (
,
,
,
,
,
,
, and
) and extracellular receptors was determined. A reduction in the potential of the U87 glioma cell membrane was observed after treatment with rGO/ATS and rGO/TUD flakes. Moreover, it was also demonstrated that major changes in the expression of voltage-dependent ion channel genes were observed in
,
, and
after treatment with rGO/ATS and rGO/TUD flakes. Furthermore, the GN/ExF, rGO/ATS, and rGO/TUD flakes significantly reduced the expression of extracellular receptors (uPar, CD105) in U87 glioblastoma cells. In conclusion, the cytotoxic mechanism of rGO flakes may depend on the presence and types of oxygen-containing functional groups, which are more abundant in rGO compared to GN.
Degradation of the extracellular matrix (ECM) changes the physicochemical properties and dysregulates ECM-cell interactions, leading to several pathological conditions, such as invasive cancer. ...Carbon nanofilm, as a biocompatible and easy to functionalize material, could be used to mimic ECM structures, changing cancer cell behavior to perform like normal cells.
Experiments were performed in vitro with HS-5 cells (as a control) and HepG2 and C3A cancer cells. An aqueous solution of fullerene C
was used to form a nanofilm. The morphological properties of cells cultivated on C
nanofilms were evaluated with light, confocal, electron and atomic force microscopy. The cell viability and proliferation were measured by XTT and BrdU assays. Immunoblotting and flow cytometry were used to evaluate the expression level of proliferating cell nuclear antigen and determine the number of cells in the G2/M phase.
All cell lines were spread on C
nanofilms, showing a high affinity to the nanofilm surface. We found that C
nanofilm mimicked the niche/ECM of cells, was biocompatible and non-toxic, but the mechanical signal from C
nanofilm created an environment that affected the cell cycle and reduced cell proliferation.
The results indicate that C
nanofilms might be a suitable, substitute component for the niche of cancer cells. The incorporation of fullerene C
in the ECM/niche may be an alternative treatment for hepatocellular carcinoma.
Currently, carbon nanostructures are vastly explored materials with potential for future employment in biomedicine. The possibility of employment of diamond nanoparticles (DN), graphene oxide (GO) or ...graphite nanoparticles (GN) for in vivo applications raises a question of their safety. Even though they do not induce a direct toxic effect, due to their unique properties, they can still interact with molecular pathways. The objective of this study was to assess if DN, GO and GN affect three isoforms of cytochrome P450 (CYP) enzymes, namely, CYP1A2, CYP2D6 and CYP3A4, expressed in the liver.
Dose-dependent effect of the DN, GO and GN nanostructures on the catalytic activity of CYPs was examined using microsome-based model. Cytotoxicity of DN, GO and GN, as well as the influence of the nanostructures on mRNA expression of CYP genes and CYP-associated receptor genes were studied in vitro using HepG2 and HepaRG cell lines.
All three nanostructures interacted with the CYP enzymes and inhibited their catalytic activity in microsomal-based models. CYP gene expression at the mRNA level was also downregulated in HepG2 and HepaRG cell lines. Among the three nanostructures, GO showed the most significant influence on the enzymes, while DN was the most inert.
Our findings revealed that DN, GO and GN might interfere with xenobiotic and drug metabolism in the liver by interactions with CYP isoenzymes responsible for the process. Such results should be considered if DN, GO and GN are used in medical applications.
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