Blueberry Extract Modulates Brain Enzymes Activities and Reduces Neuroinflammation: Promising Effect on Lipopolysaccharide-Induced Depressive-Like Behavior.

Major depressive disorder (MDD) is one of the most common neuropsychiatric disorders with high rates of prevalence and mortality. MDD is pathophysiologically complex, and treatment options are limited. Blueberries are rich in polyphenols and have neuroprotective potential. The aim of this study was to investigate the effects of blueberry extract on neuroinflammatory and neuroplasticity parameters, as well as Na+/K+-ATPase, monoamine oxidase-A (MAO-A), and acetylcholinesterase (AChE) activities in the cerebral cortex and hippocampus of mice subject to lipopolysaccharide (LPS)-induced depressive-like behavior. We also analyzed the interaction between anthocyanins and indoleamine 2 3-dioxygenase (IDO). Male Swiss mice (60-day-old) received vehicle, fluoxetine (20 mg/kg), or blueberry extract (100 or 200 mg/kg) intragastrically for 7 days before intraperitoneal LPS (0.83 mg/kg) injection. Twenty-four hours after LPS administration, the mice were subjected to behavioral tests. Both fluoxetine and blueberry extract (200 mg/kg) decreased the immobility time in the forced swim test, without affecting locomotor activity. Fluoxetine attenuated the decrease of Na+/K+-ATPase in the cerebral cortex, while blueberry extract promoted this same effect in the hippocampus. Additionally, fluoxetine and blueberry extract attenuated the decrease in the activity of MAO-A in the hippocampus. Blueberry extract (200 mg/kg) also prevented LPS-induced increase in AChE activity in the hippocampus as well as LPS upregulation of relative mRNA expression of tumor necrosis factor alpha, interleukin (IL)-1ß, and IL-10 in the cerebral cortex. Molecular docking analysis revealed binding sites for malvidin 3-galactoside (- 7.8 kcal/mol) and malvidin 3-glucoside (- 7.9 kcal/mol) residues with IDO. Taken together, these results indicate that blueberry extract improved depression-like behavior and attenuated the neurochemical and molecular changes in the brains of mice challenged with LPS.

A Purine Derivative Containing an Organoselenium Group Protects Against Memory Impairment, Sensitivity to Nociception, Oxidative Damage, and Neuroinflammation in a Mouse Model of Alzheimer's Disease.

In the present study, the effect of 6-((4-fluorophenyl) selanyl)-9H-purine (FSP) was tested against memory impairment and sensitivity to nociception induced by intracerebroventricular injection of amyloid-beta peptide (Aß) (25-35 fragment), 3 nmol/3 µl/per site in mice. Memory impairment was determined by the object recognition task (ORT) and nociception by the Von-Frey test (VFT). Aß caused neuroinflammation with upregulation of glial fibrillary acidic protein (GFAP) (in hippocampus), nuclear factor-κB (NF-κB), and the proinflammatory cytokines interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) in cerebral cortex and hippocampus. Additionally, Aß increased oxidant levels and lipid peroxidation in cerebral cortex and hippocampus, but decreased heme oxygenase-1 (HO-1) and peroxiredoxin-1 (Prdx1) expression in the hippocampus. Anti-neuroinflammatory effects of FSP were demonstrated by a decrease in the expression of GFAP and NF-κB in the hippocampus, as well as a decrease in proinflammatory cytokines in both the hippocampus and cerebral cortex FSP protected against oxidative stress by decreasing oxidant levels and lipid peroxidation and by increasing HO-1 and Prdx1 expressions in the hippocampus of mice. Moreover, FSP prevented the activation of nuclear factor erythroid 2-related factor 2 (Nrf-2) in the hippocampus of mice induced by Aß. In conclusion, treatment with FSP attenuated memory impairment, nociception sensitivity by decreasing oxidative stress, and neuroinflammation in a mouse model of Alzheimer's disease.

Investigating the Effect of Inosine on Brain Purinergic Receptors and Neurotrophic and Neuroinflammatory Parameters in an Experimental Model of Alzheimer's Disease.

Alzheimer's disease (AD) is a neurodegenerative pathology characterized by progressive impairment of memory, associated with neurochemical alterations and limited therapy. The aim of this study was to evaluate the effects of inosine on memory, neuroinflammatory cytokines, neurotrophic factors, expression of purinergic receptors, and morphological changes in the hippocampus and cerebral cortex of the rats with AD induced by streptozotocin (STZ). Male rats were divided into four groups: I, control; II, STZ; III, STZ plus inosine (50 mg/kg); and IV, STZ plus inosine (100 mg/kg). The animals received intracerebroventricular injections of STZ or buffer. Three days after the surgical procedure, animals were treated with inosine (50 mg/kg or 100 mg/kg) for 25 days. Inosine was able to prevent memory deficits and decreased the immunoreactivity of the brain A2A adenosine receptor induced by STZ. Inosine also increased the levels of brain anti-inflammatory cytokines (IL-4 and IL-10) and the expression of brain-derived neurotrophic factor and its receptor. Changes induced by STZ in the molecular layer of the hippocampus were attenuated by treatment with inosine. Inosine also protected against the reduction of immunoreactivity for synaptophysin induced by STZ in CA3 hippocampus region. However, inosine did not prevent the increase in GFAP in animals exposed to STZ. In conclusion, our findings suggest that inosine has therapeutic potential for AD through the modulation of different brain mechanisms involved in neuroprotection.

Effect of a purine derivative containing selenium to improve memory decline and anxiety through modulation of the cholinergic system and Na+/K+-ATPase in an Alzheimer's disease model.

Alzheimer's disease (AD) is a worldwide problem, and there are currently no treatments that can stop this disease. To investigate the binding affinity of 6-((4-fluorophenyl) selanyl)-9H-purine (FSP) with acetylcholinesterase (AChE), to verify the effects of FSP in an AD model in mice and to evaluate the toxicological potential of this compound in mice. The binding affinity of FSP with AChE was investigated by molecular docking analyses. The AD model was induced by streptozotocin (STZ) in Swiss mice after FSP treatment (1 mg/kg, intragastrically (i.g.)), 1st-10th day of the experimental protocol. Anxiety was evaluated in an elevated plus maze test, and memory impairment was evaluated in the Y-maze, object recognition and step-down inhibitory avoidance tasks. The cholinergic system was investigated based on by looking at expression and activity of AChE and expression of choline acetyltransferase (ChAT). We evaluated expression and activity of Na+/K+-ATPase. For toxicological analysis, animals received FSP (300 mg/kg, i.g.) and aspartate aminotransferase, alanine aminotransferase activities were determined in plasma and δ-aminolevulinate dehydratase activity in brain and liver. FSP interacts with residues of the AChE active site. FSP mitigated the induction of anxiety and memory impairment caused by STZ. FSP protected cholinergic system dysfunction and reduction of activity and expression of Na+/K+-ATPase. FSP did not modify toxicological parameters evaluated and did not cause the death of mice. FSP protected against anxiety, learning and memory impairment with involvement of the cholinergic system and Na+/K+-ATPase in these actions.

Toxicological evaluation of 3-(4-Chlorophenylselanyl)-1-methyl-1H-indole through the bovine oocyte in vitro maturation model.

The development of new bioactive molecules based on the molecular hybridization has been widely explored. In line with this, reliable tests should be employed to give information about the toxicology of these new molecules. In this sense, the use of in vitro tests is a valuable tool, especially the in vitro maturation of oocytes (IVM), which is an efficient resource to discover the potential toxicity of synthetic molecules. Thus, the aim of the present study was to evaluate the toxicological effects of the selenium-containing indolyl compound 3-(4-Chlorophenylselanyl)-1-methyl-1H-indole (CMI), on different quality parameters of bovine oocytes through the IVM. Different concentrations of the CMI compound (0, 25, 50, 100, 200 µM) were supplemented during the in vitro maturation process. After, the oocyte maturation rate, glutathione (GSH) levels, reactive oxygen species (ROS) levels, membrane, and mitochondrial integrity were evaluated. The results showed that the lowest concentration of CMI induced the highest GSH production (P < 0.05), an important marker of cytoplasmic quality and maturation. All treatments increased ROS production in relation to non-supplementation (P < 0.05). In addition, oocyte maturation was reduced only with the highest concentration of CMI (P < 0.05). Supplementation with CMI did not impact mitochondrial activity, integrity and cell membrane. To our knowledge, this is the first study that evaluates CMI on the oocyte in vitro maturation process. Importantly, our results did not find any toxic effect of CMI on bovine oocytes. CMI was efficient for cytoplasmic maturation by promoting an increase in the intracellular levels of glutathione.

Heterologous expression and characterization of a new galactose-binding lectin from Bauhinia forficata with antiproliferative activity.

In this study, we describe the characterization of a new lectin, BfL-II, purified from the seeds of Bauhinia forficata, which is distinct, at sequence-level, from the previously reported lectin from the same specie (BfL). In addition, the gene for this lectin was cloned and expressed in Escherichia coli and its antiproliferative activity was evaluated against human breast and colorectal cancer cells (MCF-7 and HT-29, respectively). The treatment with 100 µg/µL of either native or recombinant BfL (nBfL or rBfL) significantly reduced the proliferation of both cancer cell lines (p < 0.01). The inhibition of HT-29 cell proliferation was as high as 82.5% and 93.6% with 100 µg/µL of rBfL-I and nBfL after 24 h, respectively. Therefore, BfL-II presents a promising antiproliferative activity, which may be applied in the development of new anticancer treatments.

Revitalizing the AZT Through of the Selenium: An Approach in Human Triple Negative Breast Cancer Cell Line.

Triple-negative breast cancer represents about 15% of all cases of breast cancer, and still represents a therapeutic challenge. 3'-Azido-3'-deoxythymidine (AZT) is a nucleoside reverse transcriptase inhibitor with antitumor activity. Chalcogenides compounds, such as selenium, are very important intermediates applied in organic synthesis. Our objective was to investigate the effect and the underlying cell death mechanisms of AZT and its derivatives, in human breast cancer cell lines. The inhibitory effect of AZT and derivatives (1072, 1073, and 1079) was determined by MTT assay (0.1, 1, 10, 50, and 100 µM for concentrations and times 4, 24, 48, and 72 h) and Live/Dead in tumor cell lines MCF-7, MDA-MB 231 and also in non-tumor cell line CHO. Gene expression profiles related to apoptosis were investigated by qRT-PCR and induction of apoptosis was investigated by flow cytometry. MTT and Live/Dead assays showed that AZT derivatives decreased the rate of cell proliferation at concentrations of 50 and 100 µM in tumor cell lines MCF-7 and MDA-MB 231 while the commercial AZT presented a low antitumoral potential in all strains tested. In flow cytometry analysis we demonstrated that derivatives of AZT induced apoptosis, with an increase in both initial and late stages in both tumor cell lines evaluated, especially in MDA-MB 231. Our data show that the AZT derivative 1072 increased the expression of transcripts of the genes caspase 3 and 8 in MDA-MB 231 cell line when compared to control, suggesting that the extrinsic pathway of apoptosis was activated. In conclusion, derivatives of AZT, especially 1072, induce cytotoxicity in vitro in the triple negative breast cancer cell line through activation of the extrinsic pathway of apoptosis. These compounds containing selenium in its formulation are potential therapeutic agents for breast cancer.

Whole-genome sequencing of Leptospira interrogans from southern Brazil: genetic features of a highly virulent strain

BACKGROUND Leptospirosis is the most widespread zoonotic disease. It is caused by infection with pathogenic Leptospira species, of which over 300 serovars have been described. The accurate identification of the causative Leptospira spp. is required to ascertain the pathogenic status of the local isolates. OBJECTIVES This study aimed to obtain the complete genome sequence of a virulent Leptospira interrogans strain isolated from southern Brazil and to describe its genetic features. METHODS The whole genome was sequenced by next-generation sequencing (Ion Torrent). The genome was assembled, scaffolded, annotated, and manually reviewed. Mutations were identified based on a variant calling analysis using the genome of L. interrogans strain Fiocruz L1-130 as a reference. FINDINGS The entire genome had an average GC content of 35%. The variant calling analysis identified 119 single nucleotide polymorphisms (SNPs), from which 30 led to a missense mutation. The structural analyses identified potential evidence of genomic inversions, translocations, and deletions in both the chromosomes. MAIN CONCLUSIONS The genome properties provide comprehensive information about the local isolates of Leptospira spp., and thereby, could facilitate the identification of new targets for the development of diagnostic kits and vaccines.

Melatonin delivery by nanocapsules during in vitro bovine oocyte maturation decreased the reactive oxygen species of oocytes and embryos.

In this work, a promising approach to increase the advantageous properties of melatonin through its encapsulation into lipid-core nanocapsules (LNC) was examined. Oocytes were treated during in vitro maturation with non-encapsulated melatonin (Mel), melatonin-loaded lipid-core nanocapsules (Mel-LNC), and unloaded LNC. Cytotoxicity, meiotic maturation rate, development to the blastocyst stage, reactive oxygen species (ROS) and glutathione levels, mean cell number and apoptotic cell/blastocyst, and mRNA quantification were evaluated. Both Mel and Mel-LNC enhanced in vitro embryo production, however, Mel-LNC proved to be more effective at decreasing ROS levels and the apoptotic cell number/blastocyst, increasing the cleavage and blastocyst rates, up-regulating the GPX1 and SOD2 genes, and down-regulating the CASP3 and BAX genes. Mel-LNC could penetrate into oocytes and remain inside the cells until they reach the blastocyst stage. In conclusion, when melatonin was encapsulated in LNC and applied during in vitro oocyte maturation, some quality aspects of the blastocysts were improved.

Brazilian red propolis induces apoptosis-like cell death and decreases migration potential in bladder cancer cells.

Natural products continue to be an invaluable resource of anticancer drug discovery in recent years. Propolis is known for its biological activities such as antimicrobial and antitumor effects. This study assessed the effects of Brazilian red propolis (BRP) on apoptosis and migration potential in human bladder cancer cells. The effect of BRP ethanolic extract (25, 50, and 100 µg/mL) on 5637 cells was determined by MTT, LIVE/DEAD, and migration (scratch assay) assays. Apoptosis induction was investigated through flow cytometry and gene expression profile was investigated by qRT-PCR. Results showed cytotoxicity on MTT and LIVE/DEAD assays, with IC50 values of 95 µg/mL in 24 h of treatment. Cellular migration of 5637 cells was significantly inhibited through lower doses of BRP ethanolic extract (25 and 50 µg/mL). Flow cytometry analyses showed that BRP induced cytotoxicity through apoptosis-like mechanisms in 5637 cells and qRT-PCR revealed increased levels of Bax/Bcl-2 ratio, p53, AIF, and antioxidant enzymes genes. Data suggest that BRP may be a potential source of drugs to bladder cancer treatment.

Encapsulation in lipid-core nanocapsules overcomes lung cancer cell resistance to tretinoin.

Tretinoin is a retinoid derivative that has an antiproliferative effect on several kinds of tumours. Human lung adenocarcinoma epithelial cell lines (A549) exhibit a profound resistance to the effects of tretinoin. Nanocarriers seem to be a good alternative to overcomecellular resistance to drugs. The aim of this study was to test whether tretinoin-loaded lipid-core nanocapsules exert anantitumor effect on A549 cells. A549 cells were incubated with free tretinoin (TTN), blank nanocapsules (LNC) and tretinoin-loaded lipid-core nanocapsules (TTN-LNC). Data from evaluation of DNA content and Annexin V binding assay by flow cytometry showed that TTN-LNC induced apoptosis and cell cycle arrest at the G1-phase while TTN did not. TTN-LNC showed higher cytotoxic effects than TTN on A549 cells evaluated by MTT and LIVE/DEAD cell viability assay. Gene expression profiling identified up-regulated expression of gene p21 by TTN-LNC, supporting the cell cycle arrest effect. These results showed for the first time that TTN-LNC are able to overcome the resistance of adenocarcinoma cell line A549 to treatment with TTN by inducing apoptosis and cell cycle arrest, providing support for their use in applications in lung cancer therapy.

Effect of porcine somatotropin on metabolism and testicular characteristics of prepubertal pigs / Efeito da somatotrofina suína sobre o metabolismo e características testiculares de suínos pré-púberes

The effect of pST on the testicular characteristics and metabolic parameters of prepubertal pigs was evaluated. Experiment 1 aimed to determine the interval between applications of pST based on the concentrations of circulating IGF-I. Experiment 2 aimed to evaluate the effect of pST on metabolic parameters, testicular characteristics, and expression of GHR, IGF-I and PCNA. In Experiment 1 twelve piglets with 30 days of age were used. The pST Group (n = 6) was submitted to one i.m. injection of pST and the Control Group (n = 6) to one placebo injection. Blood collections were performed until day 7 post pST application to determine IGF-I concentration and metabolic profile. In Experiment 2 twelve piglets with 22 days of age were used. The pST Group was submitted to pST injections every three days, and the Control Group received placebo doses during 30 days. Blood collections were performed every 3 days. Samples of liver and testicular tissue were collected to determine gene expression and testicular characteristics. In Experiment 1 IGF-I concentration was higher for the pST Group (P = 0.02). In Experiment 2 the pST Group had higher body and testicular weight (P=0.06) and increased gene expression of PCNA in testes (P < 0.05). However, a reduction in the number of seminiferous tubules, and Sertoli cells, and in GHR expression (P < 0.05) was observed. Thus, pST administration increased body and testis development in prepubertal pigs, however it reduced the density of seminiferous tubules and Sertoli cells.

Foi investigado o efeito da pST sobre características testiculares e metabolismo de suínos pré-púberes. O Experimento 1 determinou o intervalo entre aplicações de pST, baseado nas concentrações de IGF-I. O Experimento 2 avaliou o efeito da pST sobre o metabolismo, características testiculares e expressão gênica de GHR, IGF-I e PCNA. No Experimento 1, foram usados 12 leitões com 30 dias de idade. O grupo pST (n = 6) foi submetido a uma injeção IM de pST e o grupo Controle (n = 6) a uma injeção de placebo. Coletas de sangue foram realizadas até o dia sete após a aplicação de pST para determinação dos níveis de IGF-I e parâmetros metabólicos. No Experimento 2, foram usados 12 leitões com 22 dias de idade. O grupo pST foi submetido às aplicações de pST a cada 3 dias, e o grupo Controle, às doses de placebo, durante 30 dias. Coletas de sangue foram realizadas a cada três dias. Amostras de fígado e testículo foram coletadas para determinar a expressão gênica e características testiculares. No Experimento 1, a concentração de IGF-I foi maior no grupo pST (P = 0,02). No Experimento 2, o grupo pST teve maior peso corporal e testicular (P = 0,06) e aumento na expressão de PCNA no testículo (P < 0,05). Contudo, foi observada uma redução no número de túbulos seminíferos, células de Sertoli e GHR (P < 0,05). Assim, a administração de pST aumentou o desenvolvimento testicular e corporal de suínos pré-púberes, porém reduziu a densidade de túbulos seminíferos e células de Sertoli.

Auxotrophic recombinant Mycobacterium bovis BCG overexpressing Ag85B enhances cytotoxicity on superficial bladder cancer cells in vitro.

BCG therapy remains at the forefront of immunotherapy for treating patients with superficial bladder cancer. The high incidence of local side effects and the presence of non-responder diseases have led to efforts to improve the therapy. Hence, we proposed that an auxotrophic recombinant BCG strain overexpressing Ag85B (BCG ∆leuD/Ag85B), could enhance the cytotoxicity to the human bladder carcinoma cell line 5637. The rBCG was generated using an expression plasmid encoding the mycobacterial antigen Ag85B to transform a BCG ∆leuD strain. The inhibitory effect of BCG ∆leuD/Ag85B on 5637 cells was determined by the MTT method, morphology observation and a LIVE/DEAD assay. Gene expression profiles for apoptotic, cell cycle-related and oxidative stress-related genes were investigated by qRT-PCR. Bax, bcl-2 and p53 induction by BCG ∆leuD/Ag85B treatment was evaluated by Western blotting. BCG ∆leuD/Ag85B revealed a superior cytotoxicity effect compared to the control strains used in this study. The results showed that the expression level of pro-apoptotic and cell cycle-related genes increased after BCG ∆leuD/Ag85B treatment, whereas the mRNA levels of anti-apoptotic genes decreased. Interestingly, BCG ∆leuD/Ag85B also increased the mRNA level of antioxidant enzymes in the bladder cancer cell line. Bax and p53 proteins levels increased following treatment. In conclusion, these results suggest that treatment with BCG ∆leuD/Ag85B enhances cytotoxicity for superficial bladder cancer cells in vitro. Therefore, rBCG therapy may have potential benefits in the treatment of bladder cancer.

Cytotoxic and apoptotic effects of chalcone derivatives of 2-acetyl thiophene on human colon adenocarcinoma cells.

Recent studies report that chalcones exhibit cytotoxicity to human cancer cell lines. Typically, the form of cell death induced by these compounds is apoptosis. In the context of the discovery of new anticancer agents and in light of the antitumour potential of several chalcone derivatives, in the present study, we synthesized and tested the cytotoxicity of six chalcone derivatives on human colon adenocarcinoma cells. Six derivatives of 3-phenyl-1-(thiophen-2-yl) prop-2-en-1-one were prepared and characterized on the basis of their (1) H and (13) C NMR spectra. HT-29 cells were treated with synthesized chalcones on two concentrations by three different incubation times. Cells were evaluated by cell morphology, Tetrazolium dye (MTT) colorimetric assay, live/dead, flow cytometry (annexin V) and gene expression analyses to determine the cytotoxic way. Chalcones 3-(4-bromophenyl)-1-(thiophen-2-yl)prop-2-en-1-one (C06) and 3-(2-nitrophenyl)-1-(thiophen-2-yl)prop-2-en-1-one (C09) demonstrated higher cytotoxicity than other chalcones as shown by cell morphology, live/dead and MTT assays. In addition, C06 induced apoptosis on flow cytometry annexin V assay. These data were confirmed by a decreased expression of anti-apoptotic genes and increased pro-apoptotic genes. Our findings indicate in summary that the cytotoxic activity of chalcone C06 on colorectal carcinoma cells occurs by apoptosis.

NPY and sbGnRH gene expression in juvenile and adult male Brazilian flounder Paralichthys orbignyanus / Expressão gênica do NPY e do sbGnRH em machos juvenis e adultos de linguado Paralichthys orbignyanus

The objective of this study was to evaluate neuropeptide Y (NPY) and sea bream gonadotropin-release hormone (sbGnRH) gene expression in juvenile and adult males of Brazilian flounder. Hypothalamuses from fish were sampled for total RNA extraction. After cDNA synthesis, real-time PCR was used to measure gene expression. NPY showed approximately 2-fold increases in their mRNA levels while sbGnRH showed 3-fold increases in adult fish. These results suggest that these peptides could be involved on hypothalamic regulation of Brazilian flounder sexual maturation.

O objetivo deste estudo foi avaliar a expressão gênica do neuropeptídeo Y (NPY) e da variante sea bream do hormônio liberador de gonadotrofinas (sbGnRH) em linguados machos juvenis e adultos. O hipotálamo foi isolado para a extração de RNA total. Após a síntese de cDNA, a PCR em tempo real foi usada para avaliar a expressão gênica. Foi observado um aumento de aproximadamente duas vezes nos níveis de NPY e de aproximadamente três vezes nos níveis de sbGnRH nos peixes adultos. Esses resultados demonstram que estes peptídeos podem estar envolvidos na regulação, via hipotálamo, da maturação sexual no linguado.

MicroRNAs e seu papel no desenvolvimento embrionário / MicroRNAs and its role in embryonic development

MicroRNAs (miRNAs) são pequenas moléculas de RNA com aproximadamente 22 nucleotídeos incapazes de codificar proteínas e que apresentam função na regulação pós-transcricional da expressão gênica. Vários estudos vêm demonstrando o importante papel dos miRNAs na regulação do desenvolvimento embrionário de diferentes espécies, desde o controle da expressão de RNAs mensageiros durante o desenvolvimento inicial embrionário até a determinação de linhagens celulares durante a organogênese. Esta revisão irá abordar os principais miRNAs e seu papel na biologia reprodutiva, com ênfase no desenvolvimento embrionário de mamíferos.

MicroRNAs (miRNAs) are small RNA molecules with around 22 nucleotides that are unable to encode proteins and play a key role on post-transcription regulation process. Several studies have demonstrated the relevant role of miRNAs on the regulation of embryonic development on different species, from the control of gene expression during the early embryo development to determination of cellular lineages over the organogenesis. This review will present the miRNAs and its role on reproductive biology focusing on the mammalian embryo development.