Silver and Graphenic Carbon Nanostructures Differentially Influence the Morphology and Viability of Cardiac Progenitor Cells.

The characteristic features of nanomaterials provide rich opportunities for a broad range of applications due to their different physicochemical properties. Nanocolloidal silver and graphenic carbon materials differ in most physicochemical characteristics, except for their nanodimensions. Since there is a growing demand for stem cell therapies for coronary disorders, examining cardiac progenitor cells (CPC) in terms of their response to nanostructure treatment seems to be a reasonable approach. Morphological studies and viability assessments were performed with CPC in vitro, treated with small concentrations of silver nanoparticles (AgNP), hierarchical nanoporous graphenic carbon (HNC) and their mixtures. A viability test confirmed the morphological assessment of CPC treated with AgNP and HNC; moreover, the action of both nanomaterials was time-dependent and dose-dependent. For AgNP, between the two of the applied concentrations lies a border between their potential beneficial effect and toxicity. For HNC, at a lower concentration, strong stimulation of cell viability was noted, whereas a higher dosage activated their differentiation. It is necessary to perform further research examining the mechanisms of the action of AgNP and especially of unexplored HNC, and their mixtures, on CPC and other cells.

Graphene Oxide-Based Nanocomposites Decorated with Silver Nanoparticles as an Antibacterial Agent.

One of the most promising methods against drug-resistant bacteria can be surface-modified materials with biocidal nanoparticles and nanocomposites. Herein, we present a nanocomposite with silver nanoparticles (Ag-NPs) on the surface of graphene oxide (GO) as a novel multifunctional antibacterial and antifungal material. Ultrasonic technologies have been used as an effective method of coating polyurethane foils. Toxicity on gram-negative bacteria (Escherichia coli), gram-positive bacteria (Staphylococcus aureus and Staphylococcus epidermidis), and pathogenic yeast (Candida albicans) was evaluated by analysis of cell morphology, assessment of cell viability using the PrestoBlue assay, analysis of cell membrane integrity using the lactate dehydrogenase assay, and reactive oxygen species production. Compared to Ag-NPs and GO, which have been widely used as antibacterial agents, our nanocomposite shows much higher antimicrobial efficiency toward bacteria and yeast cells.

Diamond, graphite, and graphene oxide nanoparticles decrease migration and invasiveness in glioblastoma cell lines by impairing extracellular adhesion.

The highly invasive nature of glioblastoma is one of the most significant problems regarding the treatment of this tumor. Diamond nanoparticles (ND), graphite nanoparticles (NG), and graphene oxide nanoplatelets (nGO) have been explored for their biomedical applications, especially for drug delivery. The objective of this research was to assess changes in the adhesion, migration, and invasiveness of two glioblastoma cell lines, U87 and U118, after ND, NG, and nGO treatment. All treatments affected the cell surface structure, adhesion-dependent EGFR/AKT/mTOR, and ß-catenin signaling pathways, decreasing the migration and invasiveness of both glioblastoma cell lines. The examined nanoparticles did not show strong toxicity but effectively deregulated cell migration. ND was effectively taken up by cells, whereas nGO and NG strongly interacted with the cell surface. These results indicate that nanoparticles could be used in biomedical applications as a low toxicity active compound for glioblastoma treatment.

Toxicity studies of six types of carbon nanoparticles in a chicken-embryo model.

In the present study, the toxicity of six different types of carbon nanoparticles (CNPs) was investigated using a chicken-embryo model. Fertilized chicken eggs were divided into the following treatment groups: placebo, diamond NPs, graphite NPs, pristine graphene, small graphene oxide, large graphene oxide, and reduced graphene oxide. Experimental solutions at a concentration of 500 µg/mL were administrated into the egg albumin. Gross pathology and the rate of survival were examined after 5, 10, 15, and 20 days of incubation. After 20 days of incubation, blood samples were collected and the weight of the body and organs measured. The relative ratio of embryo survival decreased after treatment all treatments except diamond NPs. There was no correlation between the rate of survival and the ζ-potential or the surface charge of the CNPs in solution. Body and organ weight, red blood-cell morphology, blood serum biochemical parameters, and oxidative damage in the liver did not differ among the groups. These results indicate that CNPs can remain in blood circulation without any major side effects, suggesting their potential applicability as vehicles for drug delivery or active compounds per se. However, there is a need for further investigation of their properties, which vary depending on production methods and surface functionalization.

Diamond Nanoparticles Modify Curcumin Activity: In Vitro Studies on Cancer and Normal Cells and In Ovo Studies on Chicken Embryo Model.

Curcumin has been studied broadly for its wide range of biological activities, including anticancer properties. The major problem with curcumin is its poor bioavailability, which can be improved by the addition of carriers, such as diamond nanoparticles (DN). They are carbon allotropes, and are therefore biocompatible and easily taken up by cells. DN are non-toxic and have antiangiogenic properties with potential applications in cancer therapy. Their large surface makes them promising compounds in a drug delivery system for bioactive agents, as DN create bio-complexes in a fast and simple process of self-organisation. We investigated the cytotoxicity of such bio-complexes against liver cancer cells and normal fibroblasts, revealing that conjugation of curcumin with DN significantly improves its activity. The experiment performed in a chicken embryo model demonstrated that neither curcumin nor DN nor bio-complexes affect embryo development, even though DN can form deposits in tissues. Preliminary results confirmed the applicability of DN as an efficient carrier of curcumin, which improves its performance against cancer cells in vitro, yet is not toxic to an organism, which makes the bio-complex a promising anticancer agent.

Long Term Influence of Carbon Nanoparticles on Health and Liver Status in Rats.

Due to their excellent biocompatibility, carbon nanoparticles have been widely investigated for prospective biomedical applications. However, their impact on an organism with prolonged exposure is still not well understood. Here, we performed an experiment investigating diamond, graphene oxide and graphite nanoparticles, which were repeatedly administrated intraperitoneally into Wistar rats for four weeks. Some of the animals was sacrificed after the last injection, whereas the rest were sacrificed twelve weeks after the last exposure. We evaluated blood morphology and biochemistry, as well as the redox and inflammatory state of the liver. The results show the retention of nanoparticles within the peritoneal cavity in the form of prominent aggregates in proximity to the injection site, as well as the presence of some nanoparticles in the mesentery. Small aggregates were also visible in the liver serosa, suggesting possible transportation to the liver. However, none of the tested nanoparticles affected the health of animals. This lack of toxic effect may suggest the potential applicability of nanoparticles as drug carriers for local therapies, ensuring accumulation and slow release of drugs into a targeted tissue without harmful systemic side effects.

Graphene Functionalized with Arginine Decreases the Development of Glioblastoma Multiforme Tumor in a Gene-Dependent Manner.

Our previous studies revealed that graphene had anticancer properties in experiments in vitro with glioblastoma multiforme (GBM) cells and in tumors cultured in vivo. We hypothesized that the addition of arginine or proline to graphene solutions might counteract graphene agglomeration and increase the activity of graphene. Experiments were performed in vitro with GBM U87 cells and in vivo with GBM tumors cultured on chicken embryo chorioallantoic membranes. The measurements included cell morphology, mortality, viability, tumor morphology, histology, and gene expression. The cells and tumors were treated with reduced graphene oxide (rGO) and rGO functionalized with arginine (rGO + Arg) or proline (rGO + Pro). The results confirmed the anticancer effect of graphene on GBM cells and tumor tissue. After functionalization with amino acids, nanoparticles were distributed more specifically, and the flakes of graphene were less agglomerated. The molecule of rGO + Arg did not increase the expression of TP53 in comparison to rGO, but did not increase the expression of MDM2 or the MDM2/TP53 ratio in the tumor, suggesting that arginine may block MDM2 expression. The expression of NQO1, known to be a strong protector of p53 protein in tumor tissue, was greatly increased. The results indicate that the complex of rGO + Arg has potential in GBM therapy.

Biodistribution of a High Dose of Diamond, Graphite, and Graphene Oxide Nanoparticles After Multiple Intraperitoneal Injections in Rats.

Carbon nanoparticles have recently drawn intense attention in biomedical applications. Hence, there is a need for further in vivo investigations of their biocompatibility and biodistribution via various exposure routes. We hypothesized that intraperitoneally injected diamond, graphite, and graphene oxide nanoparticles may have different biodistribution and exert different effects on the intact organism. Forty Wistar rats were divided into four groups: the control and treated with nanoparticles by intraperitoneal injection (4 mg of nanoparticles/kg body weight) eight times during the 4-week period. Blood was collected for evaluation of blood morphology and biochemistry parameters. Photographs of the general appearance of each rat's interior were taken immediately after sacrifice. The organs were excised and their macroscopic structure was visualized using a stereomicroscope. The nanoparticles were retained in the body, mostly as agglomerates. The largest agglomerates (up to 10 mm in diameter) were seen in the proximity of the injection place in the stomach serous membrane, between the connective tissues of the abdominal skin, muscles, and peritoneum. Numerous smaller, spherical-shaped aggregates (diameter around 2 mm) were lodged among the mesentery. Moreover, in the connective and lipid tissue in the proximity of the liver and spleen serosa, small aggregates of graphite and graphene oxide nanoparticles were observed. However, all tested nanoparticles did not affect health and growth of rats. The nanoparticles had no toxic effects on blood parameters and growth of rats, suggesting their potential applicability as remedies or in drug delivery systems.

In Ovo Administration of Silver Nanoparticles and/or Amino Acids Influence Metabolism and Immune Gene Expression in Chicken Embryos.

Due to their physicochemical and biological properties, silver nanoparticles (NanoAg) have a wide range of applications. In the present study, their roles as a carrier of nutrients and an immunomodulator were tested in chicken embryos. Cysteine (Cys)+NanoAg injected embryos had smaller livers but heavier breasts on the 19th day of embryogenesis. Cys injected embryos had lower oxygen consumption compared to threonine (Thr) or NanoAg injected embryos. The energy expenditure in Thr+NanoAg, or NanoAg injected embryos was higher than Cys or Cys+NanoAg but was not different from uninjected control embryos. Relative expression of the hepatic insulin-like growth factor-I (IGF-I) gene was higher in Cys or NanoAg injected embryos after lipopolysaccharide (LPS) induction. The gene expression of hepatic tumour necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) did not differ among amino acids, NanoAg and uninjected controls in the non-LPS groups, but increased by many folds in the LPS treated NanoAg, Cys and Cys+NanoAg groups. In LPS treated spleens, TNF-α expression was also up-regulated by NanoAg, amino acids and their combinations, but interleukin-10 (IL-10) expression was down-regulated in Thr, Cys or Thr+NanoAg injected embryos. Toll like receptor-2 (TLR2) expression did not differ in NanoAg or amino acids injected embryos; however, toll like receptor-4 (TLR4) expression was higher in all treated embryos, except for Cys+NanoAg, than in uninjected control embryos. We concluded that NanoAg either alone or in combination with amino acids did not affect embryonic growth but improved immunocompetence, indicating that NanoAg and amino acid complexes can act as potential agents for the enhancement of innate and adaptive immunity in chicken.

Interaction of graphene family materials with Listeria monocytogenes and Salmonella enterica.

Graphene family materials have unique properties, which make them valuable for a range of applications. The antibacterial properties of graphene have been reported; however, findings have been contradictory. This study reports on the antimicrobial proprieties of three different graphene materials (pristine graphene (pG), graphene oxide (GO), and reduced graphene oxide (rGO)) against the food-borne bacterial pathogens Listeria monocytogenes and Salmonella enterica. A high concentration (250 µg/mL) of all the analyzed graphenes completely inhibited the growth of both pathogens, despite their difference in bacterial cell wall structure. At a lower concentration (25 µg/mL), similar effects were only observed with GO, as growth inhibition decreased with pG and rGO at the lower concentration. Interaction of the nanoparticles with the pathogenic bacteria was found to differ depending on the form of graphene. Microscopic imaging demonstrated that bacteria were arranged at the edges of pG and rGO, while with GO, they adhered to the nanoparticle surface. GO was found to have the highest antibacterial activity.

Structural damage of chicken red blood cells exposed to platinum nanoparticles and cisplatin.

Side effects and resistance of cancer cells to cisplatin are major drawbacks to its application, and recently, the possibility of replacing cisplatin with nanocompounds has been considered. Most chemotherapeutic agents are administered intravenously, and comparisons between the interactions of platinum nanoparticles (NP-Pt) and cisplatin with blood compartments are important for future applications. This study investigated structural damage, cell membrane deformation and haemolysis of chicken embryo red blood cells (RBC) after treatment with cisplatin and NP-Pt. Cisplatin (4 µg/ml) and NP-Pt (2,6 µg/ml), when incubated with chicken embryo RBC, were detrimental to cell structure and induced haemolysis. The level of haemolytic injury was increased after cisplatin and NP-Pt treatments compared to the control group. Treatment with cisplatin caused structural damage to cell membranes and the appearance of keratocytes, while NP-Pt caused cell membrane deformations (discoid shape of cells was lost) and the formation of knizocytes and echinocytes. This work demonstrated that NP-Pt have potential applications in anticancer therapy, but potential toxic side effects must be explored in future preclinical research.

Toxicity of pristine graphene in experiments in a chicken embryo model.

Evaluation of the potential cytotoxicity of graphene is a key factor for medical applications, where flakes or a surface of graphene may be used as bioactive molecules, drug carriers, or biosensors. In the present work, effects of pristine graphene (pG) on the development of a living organism, with an emphasis on morphological and molecular states of the brain, were investigated using a chicken embryo model. Fertilized chicken eggs were divided into the control group and groups administered with pG suspended in milli-Q water at concentrations of 50 µg/L, 100 µg/L, 500 µg/L, 1,000 µg/L, 5,000 µg/L, and 10,000 µg/L (n=30 per group). The experimental solutions were injected in ovo into the albumin and then the eggs were incubated. After 19 days of incubation, the survival, weight of the body and organs, and blood serum biochemical indices were measured. The brain samples were collected for microscopic examination of brain ultrastructure and measurements of gene and protein expression. Survival of embryos was significantly decreased after treatment with pG, but the body and organ weights as well as biochemical indices were not affected. In all treatment groups, some atypical ultrastructures of the brain were observed, but they were not enhanced by the increasing concentrations of pG. Expression of proliferating cell nuclear antigen at the messenger ribonucleic acid level was downregulated, and the number of proliferating cell nuclear antigen-positive nuclei was significantly reduced in the 500-10,000 µg/L groups compared with the control group, indicating a decreased rate of deoxyribonucleic acid synthesis in the brain. The present results demonstrate some harmful effects of the applied pG flakes on the developing organism, including brain tissue, which ought to be considered prior to any medical applications.