Iêda Maria Martinez Paino

Cyto and genotoxicity of gold nanoparticles in human hepatocellular carcinoma and peripheral blood mononuclear cells.

Engineered nanomaterials have been extensively applied as active materials for technological applications. Since the impact of these nanomaterials on health and environment remains undefined, research on their possible toxic effects has attracted considerable attention. It is known that in humans, for example, the primary site of gold nanoparticles (AuNps) accumulation is the liver. The latter has motivated research regarding the use of AuNps for cancer therapy, since specific organs can be target upon appropriate functionalization of specific nanoparticles. In this study, we investigate the geno and cytotoxicity of two types of AuNps against human hepatocellular carcinoma cells (HepG2) and peripheral blood mononuclear cells (PBMC) from healthy human volunteers. The cells were incubated in the presence of different concentrations of AuNps capped with either sodium citrate or polyamidoamine dendrimers (PAMAM). Our results suggest that both types of AuNps interact with HepG2 cells and PBMC and may exhibit in vitro geno and cytotoxicity even at very low concentrations. In addition, the PBMC were less sensitive to DNA damage toxicity effects than cancer HepG2 cells upon exposure to AuNps.

Synthesis and characterization of jacalin-gold nanoparticles conjugates as specific markers for cancer cells.

New nanobiocomposites that combine nanoparticles and biomolecules have been shown very relevant for medical applications. Recently, cancer diagnostics and treatment have benefited from the development of nanobiocomposites, in which metallic or magnetic nanoparticles are conjugated with specific biomolecules for selective cell uptake. Despite recent advances in this area, the biomedical applications of these materials are still limited by the low efficiency of functionalization, low stability, among other factors. In this study, we report the synthesis of jacalin-conjugated gold nanoparticles, a nanoconjugate with potential application in medical areas, especially for cancer diagnosis. Jacalin is a lectin protein and it was employed due to its ability to recognize the Galβ1-3GalNAc disaccharide, which is highly expressed in tumor cells. Gold nanoparticles (AuNPs) were synthesized in the presence of generation 4 polyamidoamine dendrimer (PAMAM G4) and conjugated with fluorescein isothiocyanate (FITC)-labeled jacalin. The AuNPs/jacalin nanoconjugates were characterized by transmission electron microscopy (TEM), dynamic light scattering (DLS) and vibrational spectroscopy (FTIR). We also performed an investigation using isothermal titration calorimetry (ITC) and fluorescence quenching measurements to understand the interactions occurring between the AuNPs and jacalin, which revealed that the nanoconjugate formation is driven by an entropic process with good affinity. Furthermore, in vitro tests revealed that the AuNPs/jacalin-FITC nanoconjugates exhibited higher affinity for leukemic K562 cells than for healthy mononuclear blood cells, which could be useful for biomedical applications, including cancer cells imaging.

Poly(vinyl alcohol)-coated silver nanoparticles: Activation of neutrophils and nanotoxicology effects in human hepatocarcinoma and mononuclear cells

Silver nanoparticles (AgNps) have been described as important for their excellent biocompatibility, biomedical applications. Nevertheless, AgNps can interact with the immune system which is essential to analyze human exposure to assess their potential risk to health and environment. In general, the primary site for accumulation of nanoparticles has been demonstrated to be the liver. Furthermore, the direct activation of neutrophils or oxidative burst by a given nanoparticle is poorly documented. In this paper, we investigated the cell uptake, apoptosis, necrosis, DNA damage in human hepatocarcinoma cells (HepG2), primary normal human peripheral blood mononuclear cells (PBMC) and the direct activation of primary isolated neutrophils through the oxidative burst on exposure to AgNps coated with Polyvinyl-alcohol (PVA). All cell types were incubated in the presence of 1.0 and 50.0 μM of AgNps-PVA for 24h. A significant cyto- and genotoxic-response and the activation of human neutrophils were induced by AgNps-PVA (p<0.05). Our results revealed that AgNps can interact with the normal isolated neutrophils, PBMC and HepG2 cells in vitro, which opens the way for further studies on the toxicological effects of AgNps in the human immune system response and cancer cells.

In vitro nanotoxicity of single-walled carbon nanotube–dendrimer nanocomplexes against murine myoblast cells

Single-wall carbon nanotubes (SWCNTs) and polyamidoamine dendrimers (PAMAM) have been proposed for a variety of biomedical applications. The combination of both molecules makes this new composite nanomaterial highly functionalizable and versatile to theranostic and drug-delivery systems. However, recent toxicological studies have shown that nanomaterials such as SWCNTs and PAMAM may have high toxicity in biological environments. Aiming to elucidate such behavior, in vitro studies with different cultured cells have been conducted in the past few years. This study focuses on the effects of SWCNT-PAMAM nanomaterials and their individual components on the C2C12 murine cell line, which is a mixed population of stem and progenitor cells. The interactions between the cells and the nanomaterials were studied with different techniques usually employed in toxicological analyses. The results showed that SWCNT-PAMAM and PAMAM inhibited the proliferation and caused DNA damage of C2C12 cells. Data from flow cytometry revealed a less toxicity in C2C12 cells exposed to SWCNT compared to the other nanomaterials. The results indicated that the toxicity of SWCNT, SWCNT-PAMAM and PAMAM in C2C12 cells can be strongly correlated with the charge of the nanomaterials.

Phagocytosis, Oxidative Burst, and Produced Reactive Species are Affected by Iron Deficiency Anemia and Anemia of Chronic Diseases in Elderly

Iron and oxidative stress have a regulatory interplay. During the oxidative burst, phagocytic cells produce free radicals such as hypochlorous acid (HOCl). Nevertheless, scarce studies evaluated the effect of either iron deficiency anemia (IDA) or anemia of chronic disease (ACD) on phagocyte function in the elderly. The aim of the present study was to determine the oxidative burst, phagocytosis, and nitric oxide (•NO) and HOCl, reactive species produced by monocytes and neutrophils in elderly with ACD or IDA. Soluble transferrin receptor, serum ferritin, and soluble transferrin receptor/log ferritin (TfR-F) index determined the iron status. The study was constituted of 39 patients aged over 60 (28 women and 11 men) recruited from the Brazilian Public Health System. Oxidative burst fluorescence intensity per neutrophil in IDA group and HOCl generation in both ACD and IDA groups were found to be lower (pu2009<u20090.05). The percentages of neutrophils and monocytes expressing phagocytosis in ACD group were found to be higher (pu2009<u20090.05). There was an overproduction of •NO from monocytes, whereas the fundamental generation of HOCl appeared to be lower. Phagocytosis, oxidative burst, and •NO and HOCl production are involved in iron metabolism regulation in elderly patients with ACD and IDA.

Phagocytosis and nitric oxide levels in rheumatic inflammatory states in elderly women.

Background: Very few studies have investigated, in the elderly, the effect of rheumatic inflammatory states on phagocyte function and free radical production. The objective of this article is to evaluate phagocytosis by neutrophils and the production of nitric oxide (·NO) by monocytes in elderly women recruited among patients of the Brazilian Public Health System. Methods: Forty patients aged more than 60 years with rheumatic inflammatory diseases were studied. Phagocytosis was measured by flow cytometry. ·NO production was measured by the total nitrite assay and conventional inflammation markers were determined. Data were analyzed with the Mann–Whitney nonparametric test and P<0.05 was considered significant. Results: C‐reactive protein levels and white blood cell counts were significantly higher in inflammation than in the control group (P<0.05). The phagocytosis fluorescence intensity per neutrophil and the percentual of neutrophils expressing phagocytosis were significantly higher (P<0.05) in the test than in the control group. Furthermore, there was significant ·NO overproduction by monocytes, (P<0.05). Conclusion: Phagocytosis and ·NO production are affected by rheumatic states. This suggests that the increased ·NO levels may play a part in the increased oxidative stress in rheumatic diseases in elderly women. J. Clin. Lab. Anal. 25:47–51, 2011.

Toxicological effects of graphene oxide on adult zebrafish (Danio rerio)

Graphene exhibits unique physical and chemical properties that facilitate its application in many fields, including electronics and biomedical areas. However, the use of graphene and its derivatives could result in accumulation in aquatic environments, and the risks posed by these compounds for organisms are not completely understood. In this study, we investigated the effects of graphene oxide (GO) on adult zebrafish (Danio rerio). Experimental fish were exposed to 2, 10 or 20mgL-1 GO, and the cytotoxicity, genotoxicity and oxidative stress were assessed. The morphology of the gills and liver tissues was also analyzed. Graphene oxide exposure led to an increase in the number of gill cells that were in early apoptotic and necrotic stages, but genotoxicity was not observed in blood cells. We also observed the generation of Reactive Oxygen Species (ROS) in gill cells. Structural analysis revealed injuries to gill tissues, including a dilated marginal channel, lamellar fusion, clubbed tips, swollen mucocytes, epithelial lifting, aneurysms, and necrosis. Liver tissues also presented lesions such as peripherally located nuclei. Furthermore, hepatocytes exhibited a non-uniform shape, picnotic nuclei, vacuole formation, cell rupture, and necrosis. Our results showed that sub-lethal doses of graphene oxide could be harmful to fish species and thus represent risks for the aquatic food chain.

Biocompatibility and toxicology effects of graphene oxide in cancer, normal and primary immune cells

Graphene oxide (GO) has received enormous attention regarding its possible applications in medical areas including cancer treatment. Nevertheless, graphene biocompatibility and its interactions with cancer, normal and immune system cells still remain a major issue. In the current study, we focused on the immunological impact of GO in the oxidative burst by GO produced in fresh isolated primary human neutrophils, the most abundant leukocyte of immune system. We also studied the GO cytotoxicity, cell uptake, and genotoxicity in fresh isolated primary human monocyte, neutrophil, human carcinoma cervical (HeLa) and L929 cells. GO biocompatibility was also analyzed in human red blood cell (hemocompatibility) and in primary human T lymphocytes (T cell). We observed that GO can interact with HeLa in vitro and immune system cells, but in major extension with cancer cells. The latter opened the way for further studies on the effects of GO on immune system pathways and treatments for human cancer at lower concentrations.

Collagen-based silver nanoparticles: Study on cell viability, skin permeation, and swelling inhibition

Collagen is considered the most abundant protein in the animal kingdom, comprising 30% of the total amount of proteins and 6% of the human body by weight. Studies that examine the interaction between silver nanoparticles and proteins have been highlighted in the literature in order to understand the stability of the nanoparticle system, the effects observed in biological systems, and the appearance of new chemical pharmaceutical products. The objective of this study was to analyze the behavior of silver nanoparticles stabilized with collagen (AgNPcol) and to check the skin permeation capacity and action in paw edema induced by carrageenan. AgNPcol synthesis was carried out using solutions of reducing agent sodium borohydride (NaBH4), silver nitrate (AgNO3) and collagen. Characterization was done by using dynamic light scattering (DLS) and X-ray diffraction (XRD) and AFM. Cellular viability testing was performed by using flow cytometry in human melanoma cancer (MV3) and murine fibroblast (L929) cells. The skin permeation study was conducted using a Franz diffusion cell, and the efficiency of AgNPcol against the formation of paw edema in mice was evaluated. The hydrodynamic diameter and zeta potential of AgNPcol were 140.7±7.8nm and 20.1±0.7mV, respectively. AgNPcol failed to induce early apoptosis, late apoptosis, and necrosis in L929 cells; however, it exhibited enhanced toxicity in cancer cells (MV3) compared to normal cells (L929). AgNPcol demonstrated increased toxicological effects in cancer MV3 cells, promoting skin permeation, and preventing paw edema.

Synthesis and evaluation of the potential deleterious effects of ZnO nanomaterials (nanoneedles and nanoflowers) on blood components, including albumin, erythrocytes and human isolated primary neutrophils

The application of zinc oxide (ZnO) nanoparticles in biomaterials has increased significantly in the recent years. Here, we aimed to study the potential deleterious effects of ZnO on blood components, including human serum albumin (HSA), erythrocytes and human isolated primary neutrophils. To test the influence of the morphology of the nanomaterials, ZnO nanoneedles (ZnO-nn) and nanoflowers (ZnO-nf) were synthesized. The zeta potential and mean size of ZnO-nf and ZnO-nn suspensions in phosphate-buffered saline were −10.73xa0mV and 3.81xa0nm and −5.27xa0mV and 18.26xa0nm, respectively. The incubation of ZnO with HSA did not cause its denaturation as verified by the absence of significant alterations in the intrinsic and extrinsic fluorescence and in the circular dichroism spectrum of the protein. The capacity of HSA as a drug carrier was not affected as verified by employing site I and II fluorescent markers. Neither type of ZnO was able to provoke the activation of neutrophils, as verified by lucigenin- and luminol-dependent chemiluminescence and by the extracellular release of hydrogen peroxide. ZnO-nf, but not ZnO-nn, induced the haemolysis of erythrocytes. In conclusion, our results reinforce the concept that ZnO nanomaterials are relatively safe for usage in biomaterials. A potential exception is the capacity of ZnO-nf to promote the lysis of erythrocytes, a discovery that shows the importance of the morphology in the toxicity of nanoparticles.