MiRNAs that target amyloid precursor protein processing machinery in extracellular vesicles and particles derived from oral squamous cells carcinoma.

BACKGROUND: Considering that microRNAs (miRNAs), extracellular vesicles and particles (EVPs) and the amyloid precursor protein (APP) processing have been shown to be altered in oral squamous cells carcinoma (OSCC), it is possible that miRNAs that target APP processing pathways in EVPs are impacted in tumor cells. Our aim was to evaluate miRNAs that target APP itself or disintegrin and metalloproteinase domain 10 (ADAM10), which generate a trophic compound, sAPPα, in EVPs derived from OSCC cell lines, an aggressive and non-invasive, compared to normal keratinocytes. METHODS: We used two OSCC cell lines, an aggressive human oral squamous cell carcinoma cell line (SCC09) and a less aggressive cell line (CAL27) compared with a keratinocyte lineage (HaCaT). Cells were maintained in cell media, from which we isolated EVPs. EVPs were evaluated regarding their size and concentration using Nanotracking Analysis. We measured the levels of miRNAs which had as potential downstream target APP or ADAM10, specifically miR-20a-5p, miR-103a-3p, miR-424-5p, miR-92b-3p, miR-31-5p, and miR-93-5. RESULTS: There were no differences on size distributions and concentration of isolated EVPs. OSCC cell lines-derived EVPs miR-20a-5p, miR-92b-3p, and miR-93-5p were upregulated in comparison to HaCaT-derived EVPs; while miR-31-5p was reduced in EVPs obtained from CAL27 cells. CONCLUSION: Our results indicate changes in miRNAs that target APP machinery processing in EVPs derived from OSCC cell lines of different aggressiveness, which may be involved with abnormal miRNA expression in OSCC tissue and/or releasing tumor suppressor miRNA.

The adverse effects of injected functionalized multi-walled carbon nanotube (f-MWCNT) on in vivo neurosecretory brain cells of Jamaican field cricket, Gryllus assimilis.

Carbon nanotubes (CNTs) have been increasingly more prevalent due to their use in product technology owing to their exceptional electrical and thermal conductivity and tensile strength because of their nanostructure and strength of the bonds among carbon atoms. The potential increase of CNTs in the environment is a concern, and studies to assess the toxic effects of these nanomaterials (NMs) are needed. However, so far, most of the studies are focused on aquatic species and much less is understood about the effects of NM in terrestrial organisms. This investigation used a functionalized multi-walled carbon nanotube (f-MWCNT) and the Jamaican cricket Gryllus assimilis to assess the effects of this NM. Cricket nymphs were injected with f-MWCNT suspension-at three different concentrations. The insecticide Fipronil was used as a positive control. Survival was monitored, and histological analysis was made in the brains. Pyknotic cells were quantified in two brain regions, a neurosecretory called Pars intercerebralis (PI), and an associative region called mushroom body (MB). No mortality was observed in any f-MWCNT concentration tested. A significant increase in pyknotic cells was observed as sub-lethal effect for the intermediate concentration of f-MWCNT, at PI, while any significant change was observed at the Kenyon cells of the MB. These results are discussed in the context of agglomeration and dispersion of the f-MWCNT at different concentrations, and availability of the f-MWCNT on the circulatory system, as well as the natural decay of pyknotic cells with time and different patterns of adult cricket neurogenesis. Our results showed that f-MWCNT had negative effects in the neurosecretory region of the brain.

Effects of exercise modalities on BDNF and IL-1ß content in circulating total extracellular vesicles and particles obtained from aged rats.

There are evidences about the involvement of systemic factors, such as brain-derived neurotrophic factor (BDNF), on functional exercise effects. Although aerobic exercise can impact circulating extracellular vesicles and particles (EVPs) cargo, other exercise modalities were not studied. Taken that BDNF and anti-inflammatory effects have been related to functional outcomes, and BDNF and IL-1ß have been detected in circulating EVPs, our aim was to evaluate circulating total EVPs profile from adult and aged Wistar rats submitted to exercise modalities, namely aerobic, acrobatic, resistance or combined for 20 min, 3 times a week, during 12 weeks. A modality- and age-dependent effect on total EVPs cargo was observed; aerobic exercise induced an augment in BDNF and IL-1ß in EVPs from aged rats, while acrobatic and combined exercise modalities reduced IL-1ß content in EVPs from adult ones. Besides, all exercise modalities attenuated aging-induced CD63 changes in circulating total EVPs; this finding can be involved with reduced mortality rate and improved memory performance previously observed. Changes on EVPs profile, such as increased CD63 levels can be related, at least in part, to an exercise-induced healthier global status. Additionally, aerobic exercise-induced effects on BDNF and IL-1ß levels might indicate additional benefits in aged individuals.

Multi-walled carbon nanotubes functionalized with pyrene-PEG via π-π interactions: toxicological assessment in zebrafish embryos.

Multi-walled carbon nanotubes (MWCNT) have many promising biological applications, even though functionalization is needed for better biocompatibility. Functionalization of MWCNT with polyethylene glycol (PEG) is a promising and widely studied approach, but the best PEGylation method is still under investigation. In this work, we have tested the biological implications of MWCNT functionalized via π-stacking with pyrene-PEG (MWCNT-Pyr-PEG) in zebrafish embryos. As Pyr toxicity is well documented and represents a major concern for the safety of the proposed approach, we have also tested the effects of the exposure to the isolated conjugate (Pyr-PEG). The resulting suspensions were stable in saline medium and well dispersed. Zebrafish embryos at 24 h post-fertilization (hpf) were dechorionated and randomly assigned to seven experimental groups (n = 50 per group): control, MWCNT-Pyr-PEG at 0.2, 2.0, and 20.0 mg l-1, and Pyr-PEG at the same concentrations, and exposures were performed in 96-well plates. Specimens were observed for heart rate, malformations, body length, mortality, traveled distance, and number of new movements. Heart rate was reduced in embryos exposed to any tested concentration of MWCNT-Pyr-PEG, while this effect was observed with Pyr-PEG from 2 mg l-1. The highest concentration of MWCNT-Pyr-PEG also led to increased occurrence of malformations, shortened body length and reduced traveled distance. The functionalization approach shows promise due to the stability in saline media, even though toxic effects were observed in the highest tested concentrations, being the MWCNT the main actors underlying these outcomes.

Toxicological assessment of PEGylated single-walled carbon nanotubes in early developing zebrafish.

Functionalization of single-walled carbon nanotubes (SWCNT) with polyethylene glycol (PEG) is among the most promising strategies to avoid SWCNT aggregation in aqueous media, improving its interactions with biological systems. However, the best molecular PEG weight and functionalization strategy remain under investigation. In this work we assessed the toxicological effects of SWCNT functionalized with PEG at 600 Da in zebrafish embryos. Embryos were exposed to SWCNT at 0.01, 0.1 and 1 mg/L from 3 to 96 h post-fertilization (hpf). At the highest concentration, SWCNT led to toxic effects at several endpoints, including mortality, delayed hatching, malformations, reduced body length, increased ROS production and DNA damage. Even with these effects, SWCNT could not be detected within the bodily tissues of the larvae. Our results give evidence that the tested PEGylation approach was unsuitable to avoid SWCNT aggregation in aqueous media, and that SWCNT can induce toxicity even without being absorbed by the organism by obstructing the chorion pores.

Fullerene (C60) particle size implications in neurotoxicity following infusion into the hippocampi of Wistar rats.

The buckminsterfullerene (C60) is considered as a relevant candidate for drug and gene delivery to the brain, once it has the ability to cross the blood-brain barrier. However, the biological implications of this nanomaterial are not fully understood, and its safety for intracerebral delivery is still debatable. In this study, we investigated if C60 particle size could alter its biological effects. For this, two aqueous C60 suspensions were used with maximum particle size up to 200nm and 450nm. The suspensions were injected in the hippocampus, the main brain structure involved in memory processing and spatial localization. In order to assess spatial learning, male Wistar rats were tested in Morris water maze, and the hippocampal BDNF protein levels and gene expression were analyzed. Animals treated with C60 up to 450nm demonstrated impaired spatial memory with a significant decrease in BDNF protein levels and gene expression. However, an enhanced antioxidant capacity was observed in both C60 treatments. A decrease in reactive oxygen species levels was observed in the treatments with suspensions containing particles measuring with up to 450nm. Thiobarbituric acid reactive substances, glutamate cysteine ligase, and glutathione levels showed no alterations among the different treatments. In conclusion, different particle sizes of the same nanomaterial can lead to different behavioral outcomes and biochemical parameters in brain tissue.

Implications of PEGylation of Carbon Nanotubes for Central Nervous System Bioavailability.

BACKGROUND AND OBJECTIVE: In this mini-review, we have compiled the most recent and comparable information to shed light on the action of PEGylation in the biodistribution of carbon nanotubes (CNT) in the central nervous system (CNS). It is well known that due to the complexity of the CNS and the severity of the outcome following changes in this system, this is one of the areas where there are more investments in research to develop new technologies and approaches for more effective and less invasive treatments. The CNS is highly protected against toxic and invasive microorganisms thanks to the blood brain barrier (BBB), but this protection also prevents the passage of potentially beneficial molecules for the treatment of neurological disorders. Nanotechnology attempts to develop nanocompounds that are biocompatible and non-immunogenic, and that are able to cross the BBB in therapeutic amounts without causing damage and to diffuse through nerve tissue. These compounds should also be cleared and biodistributed properly, being capable of performing drug delivery exclusively for CNS pathologies, such as neurodegenerative diseases (Parkinson's and Alzheimer's) and brain tumors. CONCLUSION: In this way, this review focuses on CNT PEGylation, aiming to help in the development of viable and effective nanomedicines for neuroscience applications.

Effects of learning on mTOR pathway gene expression in the brain of zebrafish (Danio rerio) of different ages.

Target of rapamycin (TOR) is a protein kinase involved in the modulation of mRNA translation and, therefore, in the regulation of protein synthesis. In neurons, the role of TOR is particularly important in the consolidation of long-term memory (LTM). One of the modulators of TOR is brain-derived neurotrophic factor (BDNF), which activates the TOR signaling pathway to promote protein synthesis, synapse strengthening, and the creation of new neural networks. We investigated the gene expression pattern of this pathway during memory consolidation in zebrafish of different ages. Our findings demonstrate that TOR activation in old animals occurs in the early phase of consolidation, and follows a pattern identical to that of BDNF expression. In younger animals, this increase in activation did not occur, and changes in BDNF expression were also not so remarkable. Furthermore, the expression of the main proteins regulated by the synthesis of TOR (i.e., 4EBP and p70S6K) remained identical to that of TOR in all age groups.