VDR, SOD-2, and CYP24A1 Gene Expression in Different Genotypes of BsmI SNP of the Vitamin D Receptor Gene in Individuals with Hypovitaminosis.

BACKGROUND: Hypovitaminosis D is a public health problem due to its implications for various diseases. Vitamin D has numerous functions, such as modulating the metabolism of cellular tissues, and it is expressed through the vitamin D receptor (VDR) gene that may influence gene expression modulation, which plays an important role in vitamin D metabolism. OBJECTIVE: To evaluate the effect of the genotypes of BsmI single nucleotide polymorphism (SNP) of the VDR gene on VDR, SOD2, and CYP24A1 gene expression in individuals with low serum vitamin D levels. METHODS: This was a cross-sectional analytical study. After signing the informed consent form, individuals were invited to participate and answered a structured questionnaire with identification data. Blood was collected for biochemical analysis, and vitamin D was measured by chemiluminescence; BsmI polymorphism was determined using real-time polymerase chain reaction (PCR) assays with TaqMan allelic discrimination, and gene expression was conducted by qRT-PCR using QuantiFast SYBR® Green PCR Master Mix. Data were analyzed using the SPSS 20.0 software, and differences were considered significant at p < 0.05. RESULTS: 98 individuals with vitamin D ≤ 20 ng/dL were evaluated, and the BsmI SNP of the VDR gene showed CYP24A1 overexpression and low SOD2 expression. CONCLUSION: BsmI SNP of the VDR gene can modulate the expression of the genes evaluated without interfering with serum levels.

Obesity and its chronic inflammation as pain potentiation factor in rats with osteoarthritis.

BACKGROUND: Obesity produces the accumulation of adipose tissue and a chronic inflammatory process, while osteoarthritis (OA) is also an inflammatory disorder. OBJECTIVES: to evaluate whether obesity associated to OA may be a factor that increases inflammation and pain. METHODS: Male animals (M) were divided into groups: control (CM), OA-induced pain (MP), obese (OM) and obese with OA-induced pain (OMP). Similarly, females (F) were divided into groups: control (CF), OA-induced pain (FP), obese (OF) and obese with OA-induced pain (OFP). All the groups except for control and obese groups were submitted to OA induction by sodium monoiodoacetate injection and monitored until day 65. Their adiposity index, thermal, mechanical and spontaneous pain nociceptive profile were investigated. At the end of the experiment (t = 65 days) hematological parameters, biochemical parameters, andcytokines were assessed. RESULTS: Rats with obesity induction showed alterations in mechanical and thermal nociceptive profile, and increase in systemic inflammatory cytokines (TNF-α, IL-1ß, IL-6, IL-8 and leptin) with reduction in anti-inflammatory cytokines (adiponectin and IL-10). These profile changes were investigated by principal component analysis (PCA), in which the first two principal components explained near 90% of the data variability. Obesity, when present together with OA in OMP and OFP groups, yielded the highest levels of inflammatory cytokines and pain scores and the lowest levels on anti-inflamatory cytokines. CONCLUSION: Obesity modified the nociceptive profile when inflammatory process is produced. When obesity occurs concomitantly with OA, inflammatory progression is intensified, yelding increase in pain scores.

Aqueous extract of pummelo pulp (Citrus maxima) improves the biochemical profile and reduces the inflammation process in Wistar rats with non-alcoholic fatty liver disease.

This study investigated the effect of pummelo extract (Citrus maxima) on biochemical, inflammatory, antioxidant and histological changes in NAFLD rats. Forty male Wistar rats divided into four groups were used: (1) control group; (2) fructose associated with high-fat diet - DHF; (3) normal diet + pummelo extract (50 mg/kg); and (4) FHD + pummelo extract. This was administered at dose of 50 mg/kg of the animal's weight, by gavage, for 45 days. Significant improvement in lipid profile, liver and kidney function, inflammation, oxidative stress markers was identified in group 4 compared to group 2. Regarding TNF-α and IL-1ß, group 2 showed higher values (respectively 142, 5 ± 0.7 and 560.5 ± 2.7 pg/mg protein) compared to group 4 (respectively 91.4 ± 0.9 and 402.1.4 ± 0.9 pg/mg protein), p < 0.05. Significant increases were found in SOD and CAT activities, respectively 0.10 ± 0.06 and 8.62 ± 1.67 U/mg protein for group 2 and respectively 0.28 ± 0.08 and 21.52 ± 2.28 U/mg of protein for group 4. Decreases in triglycerides, hepatic cholesterol and fat droplets in hepatic tissue were observed in group 4 compared to group 2. Results highlight that pummelo extract may be useful for prevent the development of NAFLD.

Comparative study of oxidative stress and antioxidative markers in patients infected with Toxoplasma gondii.

Recently, oxidative stress and antioxidative compounds have been described as potential biomarkers. However, there is no consensus on the most appropriate oxidative and antioxidative biomarkers for patients with Toxoplasma gondii. In the present study, we evaluated the levels of lipid, protein, DNA oxidative damage and antioxidants in samples from patients infected with T. gondii with and without ocular toxoplasmosis. The levels of MDA, TBARS, micronuclei, carbonyl, GSH, vitamin C and vitamin E were measured on samples from 8 patients positive for T. gondii antibodies with ocular toxoplasmosis (OT), 20 patients positive for T. gondii antibodies without ocular toxoplasmosis (non OT), and 12 healthy individuals negative for T. gondii antibodies. The levels of MDA, TBARS, carbonyl and micronuclei were significantly higher in non OT patients, while MDA and TBARS levels were lower in OT patients. In contrast, the antioxidative factors, GSH and vitamin E levels were significantly lower in non OT patients, while vitamin C was lower in non OT and OT patients. Additionally, non OT patients were indicated to be high producers of oxidative markers (TBARS, MDA, micronuclei and carbonyl), while control group was indicated to be high producer of antioxidative markers (GSH, vitamins C and E). However, OT patients were not found as high producers of oxidative nor antioxidative markers. Our results provide a starting point of possible markers to better understand the disease pathogenesis in patients infected with T. gondii. Additional studies are needed to clarify the potential contribution of oxidative and antioxidative markers in these patients population.

The Val16Ala MnSOD gene polymorphism is associated with hypertension in self-declared black individuals.

Hypertension is the leading contributor to cardiovascular disease worldwide; the prevalence of hypertension is higher among black adults than other racial/ethnic groups. One of the cellular defense mechanisms against reactive oxygen species are the antioxidants, such as the enzyme superoxide dismutase (SOD). Therefore, this study aimed to analyze the influence of the SNP Val16Ala of the SOD2 gene on oxidative stress and hypertension in a community population of self-declared black individuals in southern Brazil. The 158 participants declared themselves black (black/brown) regarding their skin color, being 89 (56.3%) self-declared black and 69 (43.7%) brown. A real-time polymerase chain reaction determined the MnSOD Ala16Val polymorphism, and oxidative stress marker levels were significant, in addition to differences in the hypertensive group regarding the levels of carbonyl (p = .016), thiobarbituric acid reactive substances (p = .040), ischemia-modified albumin (p = .046), total antioxidant capacity (p = .011), and Nitric oxide metabolites (p = .029). The SOD Val/Val genotype was considered a risk factor regardless of the other variables for hypertension (p = .034). The Val16Ala polymorphism of the MnSOD gene presented an association with hypertension.

Association between carriers of the G allele of the + 45T> G variant of the ADIPOQ gene (rs 2241766) and the cardiometabolic profile in sickle cell trait.

AIMS: investigate the association between the +45T > G variant of the ADIPOQ gene and the metabolic syndrome (MS) in patients with sickle cell trait (SCT). 33 patients with SCT and 35 control group participated in the study. Lower levels of HDL and adiponectin were observed in patients with G allele and sickle cell trait. There were no differences between the prevalence of MS between the groups and there was no association between the +45T > G variant of the ADIPOQ gene and MS risk allele. MATERIALS AND METHODS: Participants with and without sickle cell anemia answered a questionnaire, performed anthropometric and laboratory analyzes. They were genotyped for the +45T > G variant of the ADIPOQ gene and evaluated for the presence or absence of metabolic syndrome. The study was approved by the Research Ethics Committee of UNIPAMPA (RS/Brazil). KEY FINDINGS: The GG + TG genetic model, it was associated with lower levels of adiponectin and HDL cholesterol in the SCT group. There was no association between the other studied markers and MS. SIGNIFICANCE: For the first time, an association was demonstrated between the G allele of the +45T > G variant of the ADIPOQ gene and a worse cardiometabolic profile (lower serum concentrations of adiponectin and HDL cholesterol) in patients with sickle cell trait.

Chaptalia nutans Polak: Root Extract Has High In Vitro Antioxidant Activity and Low Cytotoxicity In Vivo.

The Asteraceae family is widely known for its therapeutic, aromatic, and nutritional properties. Chaptalia nutans (C. nutans), a member of the family, is widely used in folk medicine in southern Brazil. In this study, we aim to assess compounds present in root extracts of C. nutans, and evaluate their antioxidant capacity and toxicity. To determine the chemical composition of the extract, was performed through Liquid Chromatography coupled with Mass Spectroscopy. Antioxidant capacity, toxicity (Artemia salina biosassay), cytotoxicity, genotoxicity (Allium cepa test), and neurotoxicity (Drosophila melanogaster model) were evaluated. A large number of bioactive phytoconstituents were determined to be present, such as alkaloids, coumarins, flavonoids, terpenes, and especially phenolic compounds, which may explain the antioxidant capacity of the extract. Extracts had the capacity to protect cells from protein and lipid damage, and inhibit the formation of oxygen radicals. The A. salina bioassay revealed that extracts were only slightly toxic. In A. cepa, cells exposed to 1.5 mg/mL extract were protected against chromosomal damage caused by glyphosate, and had mitotic index values that were reduced by 49%. A concentration of 10 mg/mL extract did not kill flies, and when coadministered with paraquat (PQ) (52.5%) produced a mortality rate of only 18.75%. These findings indicated that the extract had the potential to protect against PQ-induced neurotoxicity. Taken together, these data reveal for the first time that the root extract of C. nutans is a rich source of natural antioxidants. The extract may be useful in the food and pharmaceutical industries.

Mood disorders are associated with the reduction of brain derived neurotrophic factor in the hypocampus in rats submitted to the hipercaloric diet.

Adipose tissue accumulation, resulting from the consumption of hypercaloric foods, can cause a dysfunction of the endocrine system. Such endocrine changes can influence the expression of various neurochemicals including brain-derived neurotrophic factor (BDNF) - associated with cognitive and emotional problems. Here, we investigated the effects of a hypercaloric diet on depression- and anxiety-like behaviors in young rats along with concomitant changes in BDNF expression levels in the hippocampus. Eight week-old Wistar rats (n = 20) were divided in: control diet (CD) group which received industrial food (n = 8) and hypercaloric diet (HD) group which received animal fat and soybean oil (n = 12). After 45 days on the diet, the animals were evaluated: body weight and blood biochemical analisys. Changes in mood disposition were evaluated using forced swim test and the elevated plus-maze, whereas hippocampal BDNF expression levels were quantified by ELISA. After 45 weeks, the CD group showed a significant increase in body weight relative to the HD group. However, the HD rats had a body fat percentage and exhibited increased level of the biochemical markers. Furthermore, the animals in the HD group presented increased immobility time in the forced swimming test, as well as reduced response to plus-maze test suggesting a depression- and anxiety-like emotional state. In addition, the HD group also showed lower BDNF expression levels in the hippocampus. This study demonstrates that a hypercaloric diet induced increase in adipose tissue concentration in young rats was associated with reduced hippocampal BDNF expression and resulted in an increase in depression- and anxiety-like behaviors. Graphical abstract.

Evaluation In Vitro of Toxicity of Hydroalcoholic Extract of Leaves and Roots from Yacon (Smallanthus sonchifolius).

Yacon is an Andean plant that has been used in folk medicine for its medicinal properties. The beneficial effects of this plant are possibly due to the high content of phenolic compounds present in its leaves and roots. This study evaluated the in vitro toxicity of the hydroalcoholic extract of leaves and roots from yacon (1, 10, 50, and 100 µg/mL) through cell viability tests, genotoxic and mutagenic activity in leukocytes culture cells; and cytotoxicity and apoptosis cell death (1, 10, 50, 100, and 500 µg/mL) in cell line originally established from the primary mouse embryonic fibroblast cells that were cultured by the designated protocol, so-called 3T3 protocol "3-day transfer, inoculum 3 × 105 cells" (3T3 cell line). No mutagenic and cytotoxic activities were observed in leukocyte cultures. Cytotoxic activity was evidenced in the highest concentrations of yacon leaf extract (50 and 100 µg/mL), whereas all concentrations tested with yacon leaf extract there was induction for apoptosis in the 3T3 cells. Genotoxic potential was observed only at higher doses of leaf (50 and 100 µg/mL) and root (100 µg/mL) extract. These results suggest that yacon leaf at high concentrations may present toxic potential showing concentration-dependent behavior; however, in vivo studies should be performed to validate these results.

Phytochemical Analysis, Antioxidant Activity, Antimicrobial Activity, and Cytotoxicity of Chaptalia nutans Leaves.

OBJECTIVE: This work was to evaluate the chemical constitution of the hydromethanolic (30/70 methanol-water) macerating extract obtained from the leaves of C. nutans, as well as to study the antioxidant, antimicrobial, cytotoxic, and genotoxic activity of the species. Materials and methods. Phytochemical screening, antioxidant activity (total phenolic, total flavonoid, condensed tannins content, DPPH radical, and FRAP), antibacterial activity (P. aeruginosa, B. cereus, E. epidermidis, E. coli, S. aureus, E. faecalis, P. mirabilis, Candida glabrata (clinical isolate), Candida tropicalis (clinical isolate), C. krusei (clinical isolate), and C. albicans (clinical isolate)), and oxidative stress parameters (TBARS, carbonyl protein, and DCFH) were analyzed according to the literature. Toxicity of C. nutans was evaluated using an alternative method, D. melanogaster, as well as a locomotor assay. RESULTS: The phytochemical screening test of methanolic leaves extract revealed the presence of alkaloids, coumarins, quaternary bases, phenolics, flavonoids, tannins, and free steroids. A quantitative phytochemical study indicated the total phenol (30.17 ± 1.44 mg/g), flavonoid (21.64 ± 0.66 mg/g), and condensed tannins (9.58 ± 0.99 mg/g). DPPH (345.41 ± 5.35 µg/mL) and FRAP (379.98 ± 39.25 µM FeSO4/mg sample) show to extract of C. nutans leaves an intermediate value, indicating moderate antioxidant activity of the extract. Antibacterial results revealed only a positive result (antimicrobial activity) for the hexane fraction which significantly inhibited the microorganisms E. epidermidis, C. tropicalis, C. glabrata, and C. krusei at a concentration of 1000 µg/mL. TBARS, carbonyl protein, and DCFH demonstrate that the extract has the ability to protect the cell from protein and lipid damage, as well as the inhibition of oxygen-derived radicals at the three concentrations tested: 0.1, 1, and 10 mg/mL. Regarding the toxicity of C. nutans extract against D. melanogaster, it was found that until the concentration of 15 mg/mL, the extract showed no toxicity and that the LC50 obtained was 24 mg/mL. Results show that the C. nutans extract leaves used to prevent PQ damage were effective in reducing flies' mortality and improving locomotor capacity. CONCLUSION: Our studies demonstrated for the first time that C. nutans crude leaf extract has high antioxidant capacity both in vitro and in vivo through different analysis techniques. These results make it possible to infer future applications in the pharmacological area, evidenced by the low toxicity observed in D. melanogatser, as well as the ability to neutralize different sources of RONS.

Molecular detection of Leishmania spp. in Brazilian cross-border south region mammalian hosts.

Visceral leishmaniasis is an endemic zoonotic disease identified especially in developing territories. Brazil's northeast, southeast and midwest have been endemic for several years; currently, the infection is spreading to the south. Dogs are the main reservoirs; however, other mammal species have also been infected. Herein, we have identified the infecting Leishmania species in dogs and horses from the south of Brazil, a new outbreak of the infection. Blood samples were collected in the urban area of Uruguaiana city. DNA was extracted from peripheral blood, kinetoplast DNA (kDNA) and ribosomal DNA (rDNA) fragments were obtained by polymerase chain reaction (PCR) and sequenced. Out of 123 samples, 25 of them (14 dogs and 11 horses) were positive for Leishmania spp. Sequence alignment and phylogenetic analysis revealed that the kDNA in positive samples was similar to four species previously reported: L. infantum/L. chagasi, L. donovani, L. major. Despite kDNA minicircles regions are very useful due to high sensitivity to Leishmania spp. DNA detection, the sequence polymorphism among minicircles can be an obstacle to interspecific differentiation. Our results suggest that these strains are circulating in Brazil south region cross-border and indicate the susceptibility of new outbreak for visceral leishmaniasis infection in horses domiciled in endemic region for canine and human visceral leishmaniasis.

Polymorphism eNOS Glu298Asp modulates the inflammatory response of human peripheral blood mononuclear cells.

INTRODUCTION: Nitric oxide is a gaseous radical produced by the nitric oxide endothelial synthase (eNOS) whose most studied physiological action is the vasodilation. However, it also acts in the defense of the organism through the formation of cytotoxic radicals, which can potentiate the inflammatory lesion of the cells. The Glu298Asp is a single nucleotide polymorphism (SNP) in the eNOS gene related to the risk of cardiovascular disease. Blacks present a higher prevalence of hypertension and cardiovascular mortality. Then, we aimed to evaluate the influence of Glu298Asp polymorphism on inflammatory response in vitro and gene expression in blacks. MATERIAL AND METHODS: Peripheral blood mononuclear cells (PBMC) from blacks with different Glu298Asp genotypes were treated with phytohemagglutinin (PHA), a mitogen and activator of T cells. Oxidative, inflammatory markers, and expression of inflammation genes were evaluated. RESULTS: The genotype frequencies were TT 6.7%; TG 29.3% and GG 64.0%. Activation of PBMCs with 125 µg of PHA modulated the expression of inflammatory genes and increased levels of inflammatory cytokines. The T allele showed increased susceptibility to inflammation (higher levels of interleukin 1, interleukin 6 and tumor necrosis factor alpha; p < 0.001). The G allele exhibited protection through higher levels of nitric oxide (p < 0.001) and fewer inflammatory cytokines. CONCLUSION: Despite methodological limitations related to in vitro assays, the whole of results suggested that Glu298Asp modulates inflammatory genes, the T allele is more susceptible to inflammation and the G allele is protective.

Oxidative damage in mitochondrial fatty acids oxidation disorders patients and the in vitro effect of l-carnitine on DNA damage induced by the accumulated metabolites.

BACKGROUND: The mitochondrial fatty acids oxidation disorders (FAOD) are inherited metabolic disorders (IMD) characterized by the accumulation of fatty acids of different sizes of chain according to the affected enzyme. METHODS: This study evaluated the lipid peroxidation by the measurement of 8-isoprostanes, nitrosative stress parameters by the measurement of nitrite and nitrate content and DNA and RNA oxidative damage by the measurement of oxidized guanine species in urine samples from long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency (LCHADD), medium-chain acyl-CoA dehydrogenase deficiency (MCADD) and multiple acyl-CoA dehydrogenase deficiency (MADD) patients. Also, we analyzed the in vitro DNA damage by comet assay induced by adipic acid, suberic acid, hexanoylglycine and suberylglycine, separated and in combination, as well as the effect of l-carnitine in human leukocytes. RESULTS: An increase on 8-isoprostanes levels in all groups of patients was observed. The nitrite and nitrate levels were increased in LCHADD patients. DNA and RNA damage evaluation revealed increase on oxidized guanine species levels in LCHADD and MADD patients. The in vitro evaluation revealed an increase on the DNA damage induced by all metabolites, besides a potencialyzed effect. l-carnitine decreased the DNA damage induced by the metabolites. CONCLUSION: These results demonstrate that toxic metabolites accumulated could be related to the increased oxidative and nitrosative stress of FAOD patients and that the metabolites, separated and in combination, cause DNA damage, which was reduced by l-carnitine, demonstrating antioxidant protection. GENERAL SIGNIFICANCE: This work demonstrated oxidative stress in FAOD patients and the genotoxic potential of MCADD metabolites and the protective effect of l-carnitine.

Virgin Coconut Oil Associated with High-Fat Diet Induces Metabolic Dysfunctions, Adipose Inflammation, and Hepatic Lipid Accumulation.

Obesity reaches an epidemic level worldwide, and this condition is associated with chronic low-grade inflammation and secondary comorbidities, largely driven by global changes in lifestyle and diet. Various dietary approaches are proposed for the obesity treatment and its associated metabolic disorders. Good taste, antioxidant functions, and vitamins have been attributed to virgin coconut oil (VCO). However, VCO contains a large amount of saturated fatty acids, and the consumption of this fat is associated with a number of secondary diseases. We evaluate the effects of VCO supplementation on biochemical, inflammatory, and oxidative stress parameters in rats fed with high-fat diet (HFD). After feeding with HFD for 12 weeks, the animals were supplemented with VCO for 30 days. HFD+VCO group increased in diet intake, weight gain, low-density lipoprotein cholesterol level, and aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels. These findings were accompanied by increased in hepatic lipid profile and fat deposition in the liver. Adipocyte hypertrophy was observed in the HFD+VCO group, which was associated with elevated expression of tumor necrosis factor alpha (TNF-α) in adipose tissue. These results revealed that VCO associated with HFD induced important metabolic alterations, adipose inflammation, and hepatic lipid accumulation in rats.

Gamma-hexalactone flavoring causes DNA lesion and modulates cytokines secretion at non-cytotoxic concentrations.

BACKGROUND: The γ-hexalactone is a flavoring agent for alcoholic beverages, teas, breads, dairy products, coffees, buttery products among others. It presents low molecular weight and exhibits sweet fruity aroma with nuances of nuts. As far as we know, both literature and government regulations have gaps regarding the safe use of the γ-hexalactone. In this context, the main objective of this work was to evaluate the effects of γ-hexalactone through in silico and in vitro approaches. METHODS: The in silico analysis was performed through four free online platforms (admetSAR, Osiris Property Explorer®, pkCSM platform and PreADMET) and consisted of comparative structural analysis with substances present in databases. The computational prediction was performed in the sense of complement and guide the in vitro tests. Regarding in vitro investigations, screening of cytotoxicity (assessed by cell proliferation and viability parameters) in lymphocytes exposed to γ-hexalactone for 72 h were carried out previously to determine non-cytotoxic concentrations. Following this screening, concentrations of 5.15, 0.515, and 0.0515 µM were selected for the study of the respective potentials: genotoxic (assessed by DNA comet assay), chromosomal mutation (analysis of micronucleus frequency) and immunomodulatory (cytokine quantification using ELISA immunoassay). The results of in vitro assays were compared by one-way analysis of variance (ANOVA), followed by Bonferroni's post hoc test, conducted by statistic software. RESULTS: The platform PreADMET pointed out that γ-hexalactone is potentially mutagenic and carcinogenic. The comet assay data corroborate with these results demonstrating that γ-hexalactone at 5.15 µM caused lymphocytes DNA damage. In relation to cytokine secretion, the results indicate that lymphocytes were activated by γ-hexalactone at non-cytotoxic concentrations, involving an increase in the IL-1 levels in all tested concentrations, ranging from approximately 56 to 93%. The γ-hexalactone only at 5.15 µM induced increase in the levels of IL-6 (~ 60%), TNF-α (~ 68%) and IFN-γ (~ 29%), but decreased IL-10 (~ 46%) in comparison with the negative control (p < 0.05). No change was observed in total lymphocytes or in cell viability at the concentrations tested. CONCLUSIONS: In summary, the γ-hexalactone demonstrated immunomodulatory and genotoxic effects at non-cytotoxic concentrations in healthy lymphocytes.

Assessment of Leishmania infantum infection in equine populations in a canine visceral leishmaniosis transmission area.

BACKGROUND: Leishmaniosis, zoonosis that produces significant public health impacts, is caused by Leishmania infantum. Canines are the main domestic reservoir and, besides humans, other species of mammals could be infected when living in endemic areas. In this study, we detected equine Leishmania infantum infections in a canine visceral leishmaniosis transmission area and evaluated the clinical, haematological, biochemical and oxidative stress disorders. This study was conducted in Uruguaiana, Rio Grande do Sul, south of Brazil. Peripheral blood samples were collected from 124 animals (98 horses and 26 dogs) of both genders and several breeds after they underwent general and dermatologic examinations. RESULTS: Twenty five Leishmania infantum infected animals (20.16%), 14 horses and 11 dogs were detected by PCR (Polymerase Chain Reaction) amplification of kinetoplast DNA regions with 96% homology to Leishmania infantum (GenBank Accession No. L 19877.1). The clinical and haematological alterations of infected equines were skin lesions, nodules, lymphadenopathy, decreased levels in red blood cells and haematocrit (p < 0.05) and increase in urea serum concentration (p < 0.05), while CVL presented a decrease in red blood cells counts (p < 0.05), increase in lymphocytes (p < 0.05), and decrease in neutrophil-lymphocyte ratio (p < 0.05). Oxidative stress markers of plasma protein carbonyl and plasma lipid peroxidation were not statistically significant (p > 0.05) in both species. CONCLUSIONS: To our knowledge, this has been the first leishmaniosis equine survey performed in south of Brazil, caused by Leishmania infantum that were able to initially identify haematological and biochemical changes in the species, even in asymptomatic animals. We present evidence supporting those findings of haematological and biochemical changes could be related to infection. Surprisingly, the clinical manifestations of equine infection were similar to those found in canine visceral leishmaniosis. The equine population could be play an important role in the cycle of leishmaniosis in south Brazil and consequently indicates a great risk of public health. This evaluation of infected animals is important to establish the clinical and laboratory parameters involved in the disease progression.

The neuroprotective role of melatonin in a gestational hypermethioninemia model.

Elevated levels of methionine in blood characterize the hypermethioninemia, which may have genetic or non-genetic origin, as for example from high protein diet. Born rats from hypermethioninemic mothers presented cerebral oxidative stress, inhibition of Na+,K+-ATPase, memory deficit and ultrastructure cerebral changes. Melatonin is a hormone involved in circadian rhythm and has antioxidant effects. The aim of this study was to verify the possible neuroprotective effects of melatonin administration in hypermethioninemic pregnant rats on damage to biomolecules (Na+,K+-ATPase, sulfhydryl content and DNA damage index) and behavior (open field, novel object recognition and water maze tasks), as well as its effect on cells morphology by electron microscopy in offspring. Wistar female rats received methionine (2.68 µmol/g body weight) and/or melatonin (10 mg/kg body weight) by subcutaneous injections during entire pregnancy. Control rats received saline. Biochemical analyzes were performed at 21 and 30 days of life of offspring and behavioral analyzes were performed only at 30 days of age in male pups. Results showed that gestational hypermethioninemia diminished Na+,K+-ATPase activity and sulfhydryl content and increased DNA damage at 21 and 30 days of life. Melatonin was able to totally prevent Na+,K+-ATPase activity alteration at 21 days and partially prevent its alteration at 30 days of rats life. Melatonin was unable in to prevent sulfhydryl and DNA damage at two ages. It also improved DNA damage, but not at level of saline animals (controls). Regarding to behavioral tests, data showed that pups exposed to gestational hypermethioninemia decreased reference memory in water maze, spent more time to the center of the open field and did not differentiate the objects in the recognition test. Melatonin was able to prevent the deficit in novel object recognition task. Electron microscopy revealed ultrastructure alterations in neurons of hypermethioninemic at both ages of offspring, whose were prevented by melatonin. These findings suggest that melatonin may be a good neuroprotective to minimize the harmful effects of gestational hypermethioninemia on offspring.

Prevention by L-carnitine of DNA damage induced by 3-hydroxy-3-methylglutaric and 3-methylglutaric acids and experimental evidence of lipid and DNA damage in patients with 3-hydroxy-3-methylglutaric aciduria.

3-hydroxy-3-methylglutaric aciduria (HMGA) is an inherited disorder of the leucine catabolic pathway in which occurs a deficiency of the 3-hydroxy-3-methylglutaryl-CoA lyase enzyme. Therefore, the organic acids 3-hydroxy-3-methylglutaric (HMG) and 3-methylglutaric (MGA), mainly, accumulate in tissues of affected patients. Lately, much attention has been focused on free radicals as mediators of tissue damage in human diseases, causing lipid peroxidation, protein oxidation and DNA damage. The treatment of this disease is based in a restricted protein ingest and supplementation with l-carnitine (LC), an antioxidant and detoxifying agent. In the present work, we investigated the in vitro oxidative damage to DNA induced by the accumulation of organic acids and oxidative stress parameters in vivo of patients with 3-HMG, as well as the effect of the recommended therapy. The in vitro DNA damage was analyzed by the alkaline comet assay in leukocytes incubated with HMG and MGA (1 mM, 2.5 mM and 5 mM) and co-incubated with LC (90 µM and 150 µM). The in vivo urinary 15-F2t-isoprostane levels and urinary oxidized guanine species were measured by ELISA kits in patient's urine before and after the treatment with LC. HMG and MGA induced a DNA damage index (DI) significantly higher than that of the control group. The DI was significantly reduced in the presence of LC. It was also verified a significant increase of oxidized guanine species and urinary isoprostane levels, biomarker of oxidative DNA damage and lipid peroxidation respectively, in patients before treatment. After the treatment and supplementation with LC, patients presented significantly lower levels of those biomarkers. Analyzing the data together, we can conclude that HMGA patients present oxidative lipid and DNA damage, which is induced by HMG and MGA, and the antioxidant therapy with LC can prevent that kind of injuries.

Chronic mild Hyperhomocysteinemia impairs energy metabolism, promotes DNA damage and induces a Nrf2 response to oxidative stress in rats brain.

Homocysteine (HCY) has been linked to oxidative stress and varied metabolic changes that are dependent on its concentration and affected tissues. In the present study we evaluate parameters of energy metabolism [succinate dehydrogenase (SDH), complex II and IV (cytochrome c oxidase), and ATP levels] and oxidative stress [DCFH oxidation, nitrite levels, antioxidant enzymes and lipid, protein and DNA damages, as well as nuclear factor erythroid 2-related (Nrf2) protein abundance] in amygdala and prefrontal cortex of HCY-treated rats. Wistar male rats were treated with a subcutaneous injection of HCY (0.03 µmol/g of body weight) from the 30th to 60th post-natal day, twice a day, to induce mild hyperhomocysteinemia (HHCY). The rats were euthanatized without anesthesia at 12 h after the last injection, and amygdala and prefrontal cortex were dissected for biochemical analyses. In the amygdala, mild HHCY increased activities of SDH and complex II and decreased complex IV and ATP level, as well as increased antioxidant enzymes activities (glutathione peroxidase and superoxide dismutase), nitrite levels, DNA damage, and Nrf 2 protein abundance. In the prefrontal cortex, mild HHCY did not alter energy metabolism, but increased glutathione peroxidase, catalase and DNA damage. Other analyzed parameters were not altered by HCY-treatment. Our findings suggested that chronic mild HHCY changes each brain structure, particularly and specifically. These changes may be associated with the mechanisms by which chronic mild HHCY has been linked to the risk factor of fear, mood disorders and depression, as well as in neurodegenerative diseases.

Resistance exercise protocol does not cause acute genotoxic effects in trained individuals / Protocolo de exercício resistido não provoca efeito genotóxico agudo em sujeitos treinados / Protocolo de ejercicio resistido no provoca efecto genotóxico agudo en sujetos entrenados

ABSTRACT Introduction: Resistance exercise, particularly strength training, has been progressively gaining more and more followers worldwide. Despite a considerable increase in the amount of research and literature available on this topic, resistance training is undergoing important developments. Anaerobic metabolism, which characterizes resistance training, enhances the ischemic process and blood reperfusion, thereby generating reactive oxygen species (ROS). The imbalance between the production of free radicals and antioxidant defenses may induce oxidative stress with subsequent protein oxidation, lipid peroxidation, DNA damage in several cells, and other effects. This process may be intensified at rest because the O2 deficit is counteracted by a process known as excess post-exercise oxygen consumption. Objective: To analyze the effects of ROS in strength training on the DNA of human lymphocyte, biomarkers of lipid damage (TBARS) and metabolism (triglycerides, protein, glycose, albumin and urea). Methods: Comet assay involving a count of 100 cells, which were divided into five classes of damage (no damage = 0, maximum damage = 4), thereby constituting an indication of DNA damage, and the micronucleus test, where the cell samples were centrifuged at 1000-1500 RPM for ten minutes at room temperature for the micronuclei analysis. Results: An elevation in triglyceride concentrations was observed 5h post-exercise (p=0.018), probably due to nutrition. There were no significant differences in the other biochemical parameters. In terms of the DNA damage measured by the Comet assay and micronucleus test, no statistical differences were observed until 5h post-exercise. Conclusion: The proposed training session did not cause oxidative or genotoxic damage in trained individuals under the proposed conditions. Level of Evidence II; Prognostic studies-Investigation of the effect of patient characteristics on the disease outcome.

RESUMO Introdução: O treinamento resistido, principalmente a musculação, vem progressivamente ganhando novos adeptos em todo o mundo. Apesar de haver um significativo aumento no número de pesquisas e na literatura disponível sobre o assunto, o treinamento resistido vem passando por um importante processo de evolução. O metabolismo anaeróbico, que caracteriza o treinamento resistido, acentua o processo de isquemia e a reperfusão sanguínea, gerando espécies reativas do oxigênio (ERO). O desequilíbrio entre a produção de radicais livres e as defesas antioxidantes pode desencadear estresse oxidativo e levar, entre outros, à oxidação de proteínas e à peroxidação de lipídios, além de danos no DNA de diversas células. Isso pode ser amplificado durante o repouso, pois o déficit de O2 é reposto por um processo denominado excesso de oxigênio consumido após exercícios. Objetivo: Analisar os efeitos de ERO em um treinamento de musculação sobre o DNA de linfócitos humanos, de biomarcadores de dano lipídico (TBARS) e de metabolismo (triglicerídeos, proteínas, glicose, albumina e ureia). Métodos: Teste Cometa mediante contagem de 100 células, as quais foram classificadas em cinco classes de dano (sem dano = 0, dano máximo = 4), constituindo, dessa maneira, um índice de dano no DNA e o teste de micronúcleo, no qual as amostras das células foram centrifugadas a 1000-1500 RPM por dez minutos em temperatura ambiente para a análise de micronúcleos. Resultados: Constatou-se elevação das concentrações de triglicerídeos após 5 horas do treinamento (p = 0,018), relacionada, provavelmente, à alimentação. Os demais parâmetros bioquímicos não mostraram diferenças significantes. Com relação ao dano provocado no DNA medido pelos testes Cometa e de micronúcleo, não se constatou diferença estatística até 5 horas após o treinamento. Conclusão: A sessão de treinamento proposto não provocou danos oxidativos nem genotóxicos em indivíduos treinados, nas condições propostas. Nível de Evidência II; Estudos prognósticos - Investigação do efeito de característica de um paciente sobre o desfecho da doença.

RESUMEN Introducción: El entrenamiento de resistencia, especialmente la musculación viene ganando progresivamente nuevos adeptos en todo el mundo. A pesar de haber un aumento significativo en el número de investigaciones y en la literatura sobre el tema, el entrenamiento de resistencia viene pasando por un importante proceso de evolución. El metabolismo anaeróbico, que caracteriza el entrenamiento de resistencia, acentúa el proceso de isquemia y la reperfusión sanguínea, generando especies reactivas de oxígeno (ERO). El desequilibrio entre la producción de radicales libres y las defensas antioxidantes puede desencadenar el estrés oxidativo y llevar, entre otros, a la oxidación de proteínas y peroxidación de lípidos, además de daño en el ADN de diferentes células. Eso puede ser amplificado durante el reposo, para el déficit de O2 es restablecido por un proceso denominado exceso de oxígeno consumido después de ejercicios. Objetivo: Analizar los efectos de ERO en un entrenamiento de musculación sobre el ADN de linfocitos humanos, de biomarcadores de daño lipídico (TBARS) y de metabolismo (triglicéridos, proteínas, glucosa, albúmina y urea). Métodos: Prueba Cometa a través de un recuento de 100 células, las cuales se clasificaron en cinco clases de daño (sin daño = 0, daño máximo = 4), constituyendo así un índice de daño en el ADN y la prueba de micronúcleo, en el cual las muestras de las células fueron centrifugadas a 1000-1500 RPM por diez minutos a temperatura ambiente para el análisis de micronúcleos. Resultados: Se constató elevación de las concentraciones de triglicéridos después de 5 horas del entrenamiento (p = 0,018), probablemente relacionada con la alimentación. Los demás parámetros bioquímicos no mostraron diferencias significativas. Con respecto al daño provocado en el ADN medido por las pruebas Cometa y de micronúcleos, no se constató diferencia estadística hasta 5 horas después del entrenamiento. Conclusión: La sesión de entrenamiento propuesto no provocó daño oxidativo ni genotóxico en individuos entrenados, en las condiciones propuestas. Nivel de evidencia II; Estudios pronósticos - Investigación del efecto de característica de un paciente sobre el desenlace de la enfermedad.