Negative Charge-Carrying Glycans Attached to Exosomes as Novel Liquid Biopsy Marker.

Prostate cancer (PCa) is the second most common cancer. In this paper, the isolation and properties of exosomes as potential novel liquid biopsy markers for early PCa liquid biopsy diagnosis are investigated using two prostate human cell lines, i.e., benign (control) cell line RWPE1 and carcinoma cell line 22Rv1. Exosomes produced by both cell lines are characterised by various methods including nanoparticle-tracking analysis, dynamic light scattering, scanning electron microscopy and atomic force microscopy. In addition, surface plasmon resonance (SPR) is used to study three different receptors on the exosomal surface (CD63, CD81 and prostate-specific membrane antigen-PMSA), implementing monoclonal antibodies and identifying the type of glycans present on the surface of exosomes using lectins (glycan-recognising proteins). Electrochemical analysis is used to understand the interfacial properties of exosomes. The results indicate that cancerous exosomes are smaller, are produced at higher concentrations, and exhibit more nega tive zeta potential than the control exosomes. The SPR experiments confirm that negatively charged α-2,3- and α-2,6-sialic acid-containing glycans are found in greater abundance on carcinoma exosomes, whereas bisecting and branched glycans are more abundant in the control exosomes. The SPR results also show that a sandwich antibody/exosomes/lectins configuration could be constructed for effective glycoprofiling of exosomes as a novel liquid biopsy marker.

Genome-wide DNA methylome and transcriptome changes induced by inorganic nanoparticles in human kidney cells after chronic exposure.

The unique physicochemical properties make inorganic nanoparticles (INPs) an exciting tool in diagnosis and disease management. However, as INPs are relatively difficult to fully degrade and excrete, their unintended accumulation in the tissue might result in adverse health effects. Herein, we provide a methylome-transcriptome framework for chronic effects of INPs, commonly used in biomedical applications, in human kidney TH-1 cells. Renal clearance is one of the most important routes of nanoparticle excretion; therefore, a detailed evaluation of nanoparticle-mediated nephrotoxicity is an important task. Integrated analysis of methylome and transcriptome changes induced by INPs (PEG-AuNPs, Fe3O4NPs, SiO2NPs, and TiO2NPs) revealed significantly deregulated genes with functional classification in immune response, DNA damage, and cancer-related pathways. Although most deregulated genes were unique to individual INPs, a relatively high proportion of them encoded the transcription factors. Interestingly, FOS hypermethylation inversely correlating with gene expression was associated with all INPs exposures. Our study emphasizes the need for a more comprehensive investigation of INPs' biological safety, especially after chronic exposure.

Microfluidic In Vitro Platform for (Nano)Safety and (Nano)Drug Efficiency Screening.

Microfluidic technology is a valuable tool for realizing more in vitro models capturing cellular and organ level responses for rapid and animal-free risk assessment of new chemicals and drugs. Microfluidic cell-based devices allow high-throughput screening and flexible automation while lowering costs and reagent consumption due to their miniaturization. There is a growing need for faster and animal-free approaches for drug development and safety assessment of chemicals (Registration, Evaluation, Authorisation and Restriction of Chemical Substances, REACH). The work presented describes a microfluidic platform for in vivo-like in vitro cell cultivation. It is equipped with a wafer-based silicon chip including integrated electrodes and a microcavity. A proof-of-concept using different relevant cell models shows its suitability for label-free assessment of cytotoxic effects. A miniaturized microscope within each module monitors cell morphology and proliferation. Electrodes integrated in the microfluidic channels allow the noninvasive monitoring of barrier integrity followed by a label-free assessment of cytotoxic effects. Each microfluidic cell cultivation module can be operated individually or be interconnected in a flexible way. The interconnection of the different modules aims at simulation of the whole-body exposure and response and can contribute to the replacement of animal testing in risk assessment studies in compliance with the 3Rs to replace, reduce, and refine animal experiments.

Fibrous shape underlies the mutagenic and carcinogenic potential of nanosilver while surface chemistry affects the biosafety of iron oxide nanoparticles.

Nowadays engineered nanomaterials (ENMs) are increasingly used in a wide range of commercial products and biomedical applications. Despite this, the knowledge of human potential health risk as well as comprehensive biological and toxicological information is still limited. We have investigated the capacity of two frequently used metallic ENMs, nanosilver and magnetite nanoparticles (MNPs), to induce thymidine kinase (Tk +/-) mutations in L5178Y mouse lymphoma cells and transformed foci in Bhas 42 cells. Two types of nanosilver, spherical nanoparticles (AgNM300) and fibrous (AgNM302) nanorods/wires, and MNPs differing in surface modifications [MNPs coated with sodium oleate (SO-MNPs), MNPs coated with SO + polyethylene glycol (SO-PEG-MNPs) and MNPs coated with SO + PEG + poly(lactide-co-glycolic acid) SO-PEG-PLGA-MNPs] were included in this study. Spherical AgNM300 showed neither mutagenic nor carcinogenic potential. In contrast, silver nanorods/wires (AgNM302) increased significantly the number of both gene mutations and transformed foci compared with the control (untreated) cells. Under the same treatment conditions, neither SO-MNPs nor SO-PEG-PLGA-MNPs increased the mutant frequency compared with control cells though an equivocal mutagenic effect was estimated for SO-PEG-MNPs. Although SO-MNPs and SO-PEG-MNPs did not show any carcinogenic potential, SO-PEG-PLGA-MNPs increased concentration dependently the number of transformed foci in Bhas 42 cells compared with the control cells. Our results revealed that fibrous shape underlies the mutagenic and carcinogenic potential of nanosilver while surface chemistry affects the biosafety of MNPs. Considering that both nanosilver and MNPs are prospective ENMs for biomedical applications, further toxicological evaluations are warranted to assess comprehensively the biosafety of these nanomaterials.

Surface-modified magnetite nanoparticles act as aneugen-like spindle poison.

Iron oxide nanoparticles are one of the most promising types of nanoparticles for biomedical applications, primarily in the context of nanomedicine-based diagnostics and therapy; hence, great attention should be paid to their bio-safety. Here, we investigate the ability of surface-modified magnetite nanoparticles (MNPs) to produce chromosome damage in human alveolar A549 cells. Compared to control cells, all the applied MNPs increased the level of micronuclei moderately but did not cause structural chromosomal aberrations in exposed cells. A rise in endoreplication, polyploid and multinuclear cells along with disruption of tubulin filaments, downregulation of Aurora protein kinases and p53 protein activation indicated the capacity of these MNPs to impair the chromosomal passenger complex and/or centrosome maturation. We suppose that surface-modified MNPs may act as aneugen-like spindle poisons via interference with tubulin polymerization. Further studies on experimental animals revealing mechanisms of therapeutic-aimed MNPs are required to confirm their suitability as potential anti-cancer drugs.

Sustained induction of cytochrome P4501A1 in human hepatoma cells by co-exposure to benzo[a]pyrene and 7H-dibenzo[c,g]carbazole underlies the synergistic effects on DNA adduct formation.

To gain a deeper insight into the potential interactions between individual aromatic hydrocarbons in a mixture, several benzo[a]pyrene (B[a]P) and 7H-dibenzo[c,g]carbazole (DBC) binary mixtures were studied. The biological activity of the binary mixtures was investigated in the HepG2 and WB-F344 liver cell lines and the Chinese hamster V79 cell line that stably expresses the human cytochrome P4501A1 (hCYP1A1). In the V79 cells, binary mixtures, in contrast to individual carcinogens, caused a significant decrease in the levels of micronuclei, DNA adducts and gene mutations, but not in cell survival. Similarly, a lower frequency of micronuclei and levels of DNA adducts were found in rat liver WB-F344 cells treated with a binary mixture, regardless of the exposure time. The observed antagonism between B[a]P and DBC may be due to an inhibition of Cyp1a1 expression because cells exposed to B[a]P:DBC showed a decrease in Cyp1a1 mRNA levels. In human liver HepG2 cells exposed to binary mixtures for 2h, a reduction in micronuclei frequency was also found. However, after a 24h treatment, synergism between B[a]P and DBC was determined based on DNA adduct formation. Accordingly, the up-regulation of CYP1A1 expression was detected in HepG2 cells exposed to B[a]P:DBC. Our results show significant differences in the response of human and rat cells to B[a]P:DBC mixtures and stress the need to use multiple experimental systems when evaluating the potential risk of environmental pollutants. Our data also indicate that an increased expression of CYP1A1 results in a synergistic effect of B[a]P and DBC in human cells. As humans are exposed to a plethora of noxious chemicals, our results have important implications for human carcinogenesis.

Nanomedicine and epigenetics: New alliances to increase the odds in pancreatic cancer survival.

Pancreatic ductal adenocarcinoma (PDAC) is among the deadliest cancers worldwide, primarily due to its robust desmoplastic stroma and immunosuppressive tumor microenvironment (TME), which facilitate tumor progression and metastasis. In addition, fibrous tissue leads to sparse vasculature, high interstitial fluid pressure, and hypoxia, thereby hindering effective systemic drug delivery and immune cell infiltration. Thus, remodeling the TME to enhance tumor perfusion, increase drug retention, and reverse immunosuppression has become a key therapeutic strategy. In recent years, targeting epigenetic pathways has emerged as a promising approach to overcome tumor immunosuppression and cancer progression. Moreover, the progress in nanotechnology has provided new opportunities for enhancing the efficacy of conventional and epigenetic drugs. Nano-based drug delivery systems (NDDSs) offer several advantages, including improved drug pharmacokinetics, enhanced tumor penetration, and reduced systemic toxicity. Smart NDDSs enable precise targeting of stromal components and augment the effectiveness of immunotherapy through multiple drug delivery options. This review offers an overview of the latest nano-based approaches developed to achieve superior therapeutic efficacy and overcome drug resistance. We specifically focus on the TME and epigenetic-targeted therapies in the context of PDAC, discussing the advantages and limitations of current strategies while highlighting promising new developments. By emphasizing the immense potential of NDDSs in improving therapeutic outcomes in PDAC, our review paves the way for future research in this rapidly evolving field.

Rapid identification of in vitro cell toxicity using an electrochemical membrane screening platform.

This study compares the performance and output of an electrochemical phospholipid membrane platform against respective in vitro cell-based toxicity testing methods using three toxicants of different biological action (chlorpromazine (CPZ), colchicine (COL) and methyl methanesulphonate (MMS)). Human cell lines from seven different tissues (lung, liver, kidney, placenta, intestine, immune system) were used to validate this physicochemical testing system. For the cell-based systems, the effective concentration at 50 % cell death (EC50) values are calculated. For the membrane sensor, a limit of detection (LoD) value was extracted as a quantitative parameter describing the minimum concentration of toxicant which significantly affects the structure of the phospholipid sensor membrane layer. LoD values were found to align well with the EC50 values when acute cell viability was used as an end-point and showed a similar toxicity ranking of the tested toxicants. Using the colony forming efficiency (CFE) or DNA damage as end-point, a different order of toxicity ranking was observed. The results of this study showed that the electrochemical membrane sensor generates a parameter relating to biomembrane damage, which is the predominant factor in decreasing cell viability when in vitro models are acutely exposed to toxicants. These results lead the way to using electrochemical membrane-based sensors for rapid relevant preliminary toxicity screens.

Genotoxicity of 7H-dibenzo[c,g]carbazole and its methyl derivatives in human keratinocytes.

Differences between tissues in the expression of drug-metabolizing enzymes may substantially contribute to tissue-specificity of chemical carcinogens. To verify this hypothesis, the spontaneously immortalized human keratinocytes HaCaT were used, in order to evaluate the genotoxic potential of 7H-dibenzo[c,g]carbazole (DBC), a known hepatocarcinogen and sarcomagen, and its synthetic tissue-specific derivatives, 5,9-dimethyl-DBC (DiMeDBC) and N-methyl-DBC (N-MeDBC), which manifest specific tropism to the liver and skin, respectively. HaCaT cells mainly express cytochrome P4501A1 (CYP1A1), which is involved in metabolism of DBC and N-MeDBC, but not DiMeDBC [10]. Both DBC and the sarcomagen N-MeDBC induced significant levels of DNA strand-breaks, micronuclei, and DNA adducts followed by the phosphorylation of the p53 protein and histone H2AX in HaCaT cells. In contrast, the specific hepatocarcinogen DiMeDBC was devoid of any significant genotoxic activity in this cell line. Our study demonstrates that the absence of drug-metabolizing enzyme(s) involved in DiMeDBC metabolism may contribute substantially to the tissue-specific genotoxicity of this hepatocarcinogen.

The evaluation of the efficacy and potential genotoxic hazard of combined SAHA and 5-FU treatment in the chemoresistant colorectal cancer cell lines.

5-Fluorouracil (5-FU) is an essential chemotherapeutic drug for colorectal cancer (CRC) treatment. However, the frequent development of drug resistance has dramatically affected its clinical use. Therefore, novel treatment strategies are critical to improving patient outcomes. Herein, we investigated the ability of the epigenetic drug SAHA to increase the sensitivity of chemoresistant CRC cells to 5-FU. In addition, we evaluated the potential genotoxic risk of SAHA+5-FU combination treatment. As a model system, we used three CRC cell lines, HT-29, SW480, and HT-29/EGFP/FUR, differing in their resistance to 5-FU. CRC cell lines were exposed to sub-toxic SAHA concentrations for 24 h, followed by a 48 h treatment with 5-FU. The cytotoxicity of SAHA, 5-FU, and SAHA+5-FU was measured by the MTT test, the genotoxicity by the comet assay, and the micronucleus test. The apoptotic/necrotic activity was assessed using morphological criteria. We found a synergic decrease in the viability of HT-29 and SW480 cells, but not the most resistant HT-29/EGFP/FUR cells after combined SAHA+5-FU exposure compared to 5-FU. Remarkably, SAHA most efficiently induced apoptosis in HT-29/EGFP/FUR cells compared to HT-29 and SW480 cells. Combined SAHA+5-FU treatment resulted in a synergistic increase in apoptotic/necrotic cells in HT-29 cell line, while rather additive/sub-additive effect was determined in the SW480 and HT-29/EGFP/FUR cells. At the same time, however, a synergistic rise in micronuclei was found in CRC cell lines (at least at some concentrations). We have shown that SAHA can sensitize CRC cells to 5-FU; therefore, epigenetic and convential drug combinations could be beneficial for the patients. However, the increase in micronucleus formation after combined SAHA+5-FU treatment indicates a potential health hazard. The clastogenic activity could contribute to cancer heterogeneity, favoring progeny of such aberrant cells to clonal expansion. Therefore, developing new specific epigenetic drugs or nanocarriers for targeted drug delivery might reduce the potential genotoxic risk.

Plate reader spectroscopy as an alternative to atomic absorption spectroscopy for the assessment of nanoparticle cellular uptake.

Fundamental studies investigating the biological effects induced by nanoparticles (NPs) explicitly require the correct assessment of their intracellular concentration. Ultrasensitive atomic absorption spectroscopy (AAS) is perceived as one of the gold standard methods for quantifying internalized NPs. Besides its limitation to metal-based NPs though, AAS also requires specific infrastructure and tedious sample preparation and handling, making it time-consuming and cost-intensive. Herein we report on a reliable, rapid, and affordable alternative to AAS - plate reader spectroscopy (PRS), which offers an accessible option for everyday laboratory use without sophisticated instrumentation. Our results demonstrate, that following a proper methodological approach, data on intracellular concentration of NPs obtained by PRS are fully comparable to AAS results. Specifically, the intracellular concentration of magnetite NPs coated with sodium oleate and bovine serum albumin in human alveolar A549 cells was assessed by PRS and AAS in parallel, with a remarkable correlation coefficient of R = 0.9914.

Exosomes from prostate cancer cell lines: Isolation optimisation and characterisation.

Exosomes are considered to be a rich source of biomarkers, hence in this article we examine the best procedure for their isolation. We examine several isolation procedures, exosome storage conditions and other conditions affecting exosome production by prostate cell lines. We selected four different commercially available kits based on different principles to achieve exosome isolation, the best being magnetic-based. In addition, we found storage at - 20 °C to be good for storing isolated exosomes and that exosomes were produced from the cancerous prostate cell line 22Rv1 in much greater amounts than the non-cancerous prostate cell line RWPE1. We also found differences in the response of both cell lines in the production of exosomes as a result of stress, i.e. exposure to hydrogen peroxide and starvation. The effect of Triton X-100 on exosome lysis was examined using two different surfactant concentrations by analysis of the exosome count and change in the exosome size. The final part of the article details the advantages of the use of a 2D biochip prepared in-house over a commercially available 3D biochip for monitoring the interaction of exosomes via its surface receptors (CD63) with an immobilised ligand (anti-CD63 antibodies) using surface plasmon resonance. The final experiment shows the potential of lectin fluorescent microarrays for the analysis of glycans present in lysed exosomes.

Pharmacokinetics of PEGylated Gold Nanoparticles: In Vitro-In Vivo Correlation.

Data suitable for assembling a physiologically-based pharmacokinetic (PBPK) model for nanoparticles (NPs) remain relatively scarce. Therefore, there is a trend in extrapolating the results of in vitro and in silico studies to in vivo nanoparticle hazard and risk assessment. To evaluate the reliability of such approach, a pharmacokinetic study was performed using the same polyethylene glycol-coated gold nanoparticles (PEG-AuNPs) in vitro and in vivo. As in vitro models, human cell lines TH1, A549, Hep G2, and 16HBE were employed. The in vivo PEG-AuNP biodistribution was assessed in rats. The internalization and exclusion of PEG-AuNPs in vitro were modeled as first-order rate processes with the partition coefficient describing the equilibrium distribution. The pharmacokinetic parameters were obtained by fitting the model to the in vitro data and subsequently used for PBPK simulation in vivo. Notable differences were observed in the internalized amount of Au in individual cell lines compared to the corresponding tissues in vivo, with the highest found for renal TH1 cells and kidneys. The main reason for these discrepancies is the absence of natural barriers in the in vitro conditions. Therefore, caution should be exercised when extrapolating in vitro data to predict the in vivo NP burden and response to exposure.

Decitabine-induced DNA methylation-mediated transcriptomic reprogramming in human breast cancer cell lines; the impact of DCK overexpression.

Decitabine (DAC), a DNA methyltransferase (DNMT) inhibitor, is tested in combination with conventional anticancer drugs as a treatment option for various solid tumors. Although epigenome modulation provides a promising avenue in treating resistant cancer types, more studies are required to evaluate its safety and ability to normalize the aberrant transcriptional profiles. As deoxycytidine kinase (DCK)-mediated phosphorylation is a rate-limiting step in DAC metabolic activation, we hypothesized that its intracellular overexpression could potentiate DAC's effect on cell methylome and thus increase its therapeutic efficacy. Therefore, two breast cancer cell lines, JIMT-1 and T-47D, differing in their molecular characteristics, were transfected with a DCK expression vector and exposed to low-dose DAC (approximately IC20). Although transfection resulted in a significant DCK expression increase, further enhanced by DAC exposure, no transfection-induced changes were found at the global DNA methylation level or in cell viability. In parallel, an integrative approach was applied to decipher DAC-induced, methylation-mediated, transcriptomic reprogramming. Besides large-scale hypomethylation, accompanied by up-regulation of gene expression across the entire genome, DAC also induced hypermethylation and down-regulation of numerous genes in both cell lines. Interestingly, TET1 and TET2 expression halved in JIMT-1 cells after DAC exposure, while DNMTs' changes were not significant. The protein digestion and absorption pathway, containing numerous collagen and solute carrier genes, ranking second among membrane transport proteins, was the top enriched pathway in both cell lines when hypomethylated and up-regulated genes were considered. Moreover, the calcium signaling pathway, playing a significant role in drug resistance, was among the top enriched in JIMT-1 cells. Although low-dose DAC demonstrated its ability to normalize the expression of tumor suppressors, several oncogenes were also up-regulated, a finding, that supports previously raised concerns regarding its broad reprogramming potential. Importantly, our research provides evidence about the involvement of active demethylation in DAC-mediated transcriptional reprogramming.

Decitabine potentiates efficacy of doxorubicin in a preclinical trastuzumab-resistant HER2-positive breast cancer models.

Acquired drug resistance and metastasis in breast cancer (BC) are coupled with epigenetic deregulation of gene expression. Epigenetic drugs, aiming to reverse these aberrant transcriptional patterns and sensitize cancer cells to other therapies, provide a new treatment strategy for drug-resistant tumors. Here we investigated the ability of DNA methyltransferase (DNMT) inhibitor decitabine (DAC) to increase the sensitivity of BC cells to anthracycline antibiotic doxorubicin (DOX). Three cell lines representing different molecular BC subtypes, JIMT-1, MDA-MB-231 and T-47D, were used to evaluate the synergy of sequential DAC + DOX treatment in vitro. The cytotoxicity, genotoxicity, apoptosis, and migration capacity were tested in 2D and 3D cultures. Moreover, genome-wide DNA methylation and transcriptomic analyses were employed to understand the differences underlying DAC responsiveness. The ability of DAC to sensitize trastuzumab-resistant HER2-positive JIMT-1 cells to DOX was examined in vivo in an orthotopic xenograft mouse model. DAC and DOX synergistic effect was identified in all tested cell lines, with JIMT-1 cells being most sensitive to DAC. Based on the whole-genome data, we assume that the aggressive behavior of JIMT-1 cells can be related to the enrichment of epithelial-to-mesenchymal transition and stemness-associated pathways in this cell line. The four-week DAC + DOX sequential administration significantly reduced the tumor growth, DNMT1 expression, and global DNA methylation in xenograft tissues. The efficacy of combination therapy was comparable to effect of pegylated liposomal DOX, used exclusively for the treatment of metastatic BC. This work demonstrates the potential of epigenetic drugs to modulate cancer cells' sensitivity to other forms of anticancer therapy.

The role of human cytochrome P4503A4 in biotransformation of tissue-specific derivatives of 7H-dibenzo[c,g]carbazole.

The environmental pollutant 7H-dibenzo[c,g]carbazole (DBC) and its derivative, 5,9-dimethylDBC (DiMeDBC), produced significant and dose-dependent levels of micronuclei followed by a substantial increase in the frequency of apoptotic cells in the V79MZh3A4 cell line stably expressing the human cytochrome P450 (hCYP) 3A4. In contrast, neither micronuclei nor apoptosis were found in cells exposed to the sarcomagenic carcinogen, N-methylDBC (N-MeDBC). A slight but significant level of gene mutations and DNA adducts detected in V79MZh3A4 cells treated with N-MeDBC, only at the highest concentration (30µM), revealed that this sarcomagenic carcinogen was also metabolized by hCYP3A4. Surprisingly, DBC increased the frequency of 6-thioguanine resistant (6-TG(r)) mutations only at the highest concentration (30µM), while DiMeDBC failed to increase the frequency of these mutations. The resistance to 6-thioguanine is caused by the mutations in the hypoxanthine-guanine phosphoribosyltransferase (Hprt) gene. The molecular analysis of the coding region of Hprt gene showed a deletion of the entire exon 8 in DiMeDBC-induced 6-TG(r) mutants, while no changes in the nucleotide sequences were identified in 6-TG(r) mutants produced by DBC and N-MeDBC. Based on our results, we suggest that hCYP3A4 is involved in the metabolism of DBC and its tissue-specific derivatives. While hCYP3A4 probably plays an important role in biotransformation of the liver carcinogens, DBC and DiMeDBC, it might only have a marginal function in N-MeDBC metabolism.

Differences in surface chemistry of iron oxide nanoparticles result in different routes of internalization.

The efficient entry of nanotechnology-based pharmaceuticals into target cells is highly desired to reach high therapeutic efficiency while minimizing the side effects. Despite intensive research, the impact of the surface coating on the mechanism of nanoparticle uptake is not sufficiently understood yet. Herein, we present a mechanistic study of cellular internalization pathways of two magnetic iron oxide nanoparticles (MNPs) differing in surface chemistry into A549 cells. The MNP uptake was investigated in the presence of different inhibitors of endocytosis and monitored by spectroscopic and imaging techniques. The results revealed that the route of MNP entry into cells strongly depends on the surface chemistry of the MNPs. While serum bovine albumin-coated MNPs entered the cells via clathrin-mediated endocytosis (CME), caveolin-mediated endocytosis (CavME) or lipid rafts were preferentially involved in the internalization of polyethylene glycol-coated MNPs. Our data indicate that surface engineering can contribute to an enhanced delivery efficiency of nanoparticles.

Pharmacokinetics, Biodistribution, and Biosafety of PEGylated Gold Nanoparticles In Vivo.

Despite the obvious advantages of gold nanoparticles for biomedical applications, controversial and incomplete toxicological data hamper their widespread use. Here, we present the results from an in vivo toxicity study using gold nanoparticles coated with polyethylene glycol (PEG-AuNPs). The pharmacokinetics and biodistribution of PEG-AuNPs were examined in the rat's liver, lung, spleen, and kidney after a single i.v. injection (0.7 mg/kg) at different time intervals. PEG-AuNPs had a relatively long blood circulation time and accumulated primarily in the liver and spleen, where they remained for up to 28 days after administration. Increased cytoplasmic vacuolation in hepatocytes 24 h and 7 days after PEG-AuNPs exposure and apoptotic-like cells in white splenic pulp 24 h after administration has been detected, however, 28 days post-exposure were no longer observed. In contrast, at this time point, we identified significant changes in lipid metabolism, altered levels of liver injury markers, and elevated monocyte count, but without marked biological relevance. In blood cells, no DNA damage was present in any of the studied time intervals, with the exception of DNA breakage transiently detected in primary kidney cells 4 h post-injection. Our results indicate that the tissue accumulation of PEG-AuNPs might result in late toxic effects.

7H-Dibenzo[c,g]carbazole: Metabolic pathways and toxicity.

7H-Dibenzo[c,g]carbazole (DBC), a local and systemic carcinogen in animal studies, is a common environmental pollutant. It generally co-occurs in a variety of organic complex mixtures derived from incomplete combustion of organic matter. Despite high lipophilicity, DBC is more water-soluble and faster metabolized than the homocyclic aromatics. Moreover, greater polarity, high bioaccumulation potential, and persistence in the environment may imply DBC's higher biological significance and impact on human health, even at lower concentrations. The biotransformation pathways of DBC are incompletely known and the ultimate carcinogenic metabolite(s) are not clearly identified as yet. Structure-biological studies suggest two ways of activation: at the ring carbon atoms and at the pyrrole nitrogen. It is supposed that the particular pathway of biotransformation might be connected with the tissue/organ specificity of DBC. Cytochrome P450 (CYP) family of enzymes plays a pivotal role in the metabolism of DBC; though, the one-electron activation and the aldo-keto reductase-catalyzed oxidation are also involved in metabolic activation. Additionally, DBC can be photoactivated even at physiologically relevant doses of UVA light due to the extended aromatic ring system resulting in strong genotoxicity and oxidative stress. The goal of this review is to summarize current knowledge on mechanisms of DBC activation and possible implications for toxicity, genotoxicity, and carcinogenicity.

Surface coating determines the inflammatory potential of magnetite nanoparticles in murine renal podocytes and mesangial cells.

Drug-induced nephrotoxicity is a frequent adverse event and a dose-limiting factor in patient treatment and is a leading cause of prospective drug attrition during pharmaceutical development. Despite the obvious benefits of nanotherapeutics in healthcare strategies, the clearance of imaging agents and nanocarriers from the body following their therapeutic or diagnostic application generates concerns about their safety for human health. Considering the potency of nanoparticles and their massive utilization in biomedicine the impact of magnetic nanoparticles (MNPs) on cells forming the filtration apparatus of the kidney was studied. Using primary mouse renal glomerular podocytes and mesangial cells, we investigated their response to exposure to magnetic nanoparticles coated with polyethylene glycol and bovine serum albumin. Cultured podocytes were more sensitive to MNPs than mesangial cells displaying signs of cell damage and stronger inflammatory response. Both types of MNPs induced the remodeling of actin fibers, affected the cell shape and triggered expression of inflammatory cytokines TNFα and IL-6 in podocytes. On the other hand, iNOS was induced in both renal cell types but only by MNPs with a polyethylene glycol coating. Our results have revealed that the type of cell and the type of nanoparticle coating might be the strongest determinants of cellular response toward nanoparticle exposure. Differences in susceptibility of cells to MNPs might be evident also between neighboring renal cell subpopulations integrally forming functional sub-units of this organ.