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Aim: To evaluate the clinical effectiveness of ozonated sunflower oil (Oz) as an adjunctive of non-surgical periodontal therapy in patients with type 2 diabetes mellitus (DM2), on fibroblast cell viability and migration and the effectiveness of Oz on a Candida albicans (C. albicans) culture., Methodology: In total, 32 sites in 16 DM2 with moderate to advanced periodontal disease with periodontal pocket depths ≥5mm were selected. The treatments were divided into two groups: control, saline solution (SS) as an adjunctive of scaling and root planing (SRP+SS), and test, Oz as an adjunctive of SRP (SRP+Oz). Hematological [fasting glucose level (FGL) and hemoglobin A1c (HbA1c)] and microbiological samples were collected from the participants at baseline and three months after periodontal treatment and the microbiological samples were analyzed by PCR. C. albicans was previously tested by the agar diffusion test. The effect of Oz was tested on cell viability and fibroblast migration., Results: The groups showed no statistically significant differences (paired t-test-p>0.05) regarding hematological parameters, FGL (median - baseline 171.41, 3 months 164mg/dL), and HbA1c (baseline 8%, 3 months 7.5%) (Kruskal-Wallis One-Way Nonparametric-p>0.05) after periodontal therapy. The groups showed statistical differences for periodontal parameters between baseline and three months (paired t-test-p<0.05). PCR analysis showed a reduction in the percentage of C. albicans in the SRP+Oz group after three months (McNemar's test-p=0.002). Cell viability was lower in the high glucose Dulbecco's modified Eagle's medium (4500 mg/L) than in low glucose (1000 mg/L) (RM-ANOVA-p<0.0001). The wound healing test showed reduced fibroblast migration (one-way ANOVA with Dunnett's post-test-p<0.01). Oz showed high C. albicans antifungal inhibition (Kruskal-Wallis test-p=0.0001)., Conclusions: SRP+Oz effectively reduced C. albicans in-vitro and in-vivo but showed no clinical improvements compared to the control. Cell viability and wound healing of fibroblasts showed no improvements.

Purpose: To evaluate the effect of the topical application of the ethanol extract (EESL) and the hydroethanolic fraction (HFSL) of ripe Solanum lycocarpum fruit on the healing of experimentally-induced wounds in mice., Methods: The EESL and HFSL obtained from ripe fruit of the species S. lycocarpum were obtained by percolation with ethanol. They were tested in the healing of excisional wounds in mice, which were induced in anesthetized animals using a 7-mm dermatological punch. They were divided according to the treatment period and group, n = 6, and received topical application of the EESL or HFSL daily or saline solution for one, five, seven, or 21 days. At the end of this period, the animals were euthanized, and the lesions were subjected to histopathological processing and analysis to evaluate macroscopic area of the wound and microscopic analysis by morphometry of the number of leukocytes, mast cells, fibroblasts, re-epithelialization, and matrix deposition., Results: The application of the EESL and HFSL reduced the number of leukocytes after one, five, and seven days of treatment. EESL improved re-epithelialization and tissue proliferation in wound healing from day 7., Conclusion: The study demonstrated that daily administration of EESL and HFSL exhibit wound healing activity, with this effect being more pronounced by EESL. To our knowledge, this is the first report of the anti-inflammatory and healing activity of this species through topical application.

Objective: The objective was to identify validated instruments from the literature that assess the mental health of police officers., Methods: This is a systematic review of validated instruments used to assess the mental health of police officers. Searches were conducted in the MEDLINE, Web of Science, Scopus, Embase, CINAHL/EBSCO, and Virtual Health Library databases. This review follows the JBI Manual for Systematic Reviews and the PRISMA statement. The methodological quality of the articles and the risk of bias were assessed., Results: A total of 1530 studies were identified across the six databases, with 158 studies read in full by the authors after excluding duplicates and those that did not meet the inclusion criteria. The final 29 studies were analyzed for methodological quality and risk of bias using the AXIS and SFS-D tools., Conclusion: This review identified 27 self-administered validated instruments useful for assessing various mental health outcomes in police officers, with the most frequently used being the Police Stress Questionnaire. These findings may help guide security force administration, occupational health professionals, and mental health researchers in selecting and implementing psychometrically reliable instruments for screening the mental health of police officers.

Neurodegenerative disorders, including traumatic injuries to the central nervous system (CNS) and neurodegenerative diseases, are characterized by early axonal damage, which does not regenerate in the adult mammalian CNS, leading to permanent neurological deficits. One of the primary causes of the loss of regenerative ability is thought to be a developmental decline in neurons' intrinsic capability for axon growth. Different molecules are involved in the developmental loss of the ability for axon regeneration, including many transcription factors. However, the function of microRNAs (miRNAs), which are also modulators of gene expression, in axon re-growth is still unclear. Among the various miRNAs recently identified with roles in the CNS, miR-17, which is highly expressed during early development, emerges as a promising target to promote axon regeneration. Here, we used adeno-associated viral (AAV) vectors to overexpress miR-17 (AAV.miR-17) in primary cortical neurons and evaluate its effects on neurite and axon regeneration in vitro. Although AAV.miR-17 had no significant effect on neurite outgrowth and arborization, it significantly enhances neurite regeneration after scratch lesion and axon regeneration after axotomy of neurons cultured in microfluidic chambers. Target prediction and functional annotation analyses suggest that miR-17 regulates gene expression associated with autophagy and cell metabolism. Our findings suggest that miR-17 promotes regenerative response and thus could mitigate neurodegenerative effects.

Coumarins have great pharmacotherapeutic potential, presenting several biological and pharmaceutical applications, like antibiotic, fungicidal, anti-inflammatory, anticancer, anti-HIV, and healing activities, among others. These molecules are practically insoluble in water, and for biological applications, it became necessary to complex them with cyclodextrins (CDs), which influence their bioavailability in the target organism. In this work, we studied two coumarins, and it was possible to conclude that there were structural differences between 4,7-dimethyl-2 H -chromen-2-one (DMC) and 7-methoxy-4-methyl-2 H -chromen-2-one (MMC)/β-CD that were solubilized in ethanol, frozen, and lyophilized (FL) and the mechanical mixtures (MM). In addition, the inclusion complex formation improved the solubility of DMC and MMC in an aqueous medium. According to the data, the inclusion complexes were formed and are more stable at a molar ratio of 2:1 coumarin/β-CD, and hydrogen bonds along with π-π stacking interactions are responsible for the better stability, especially for (MMC) 2 @β-CD. In vivo wound healing studies in mice showed faster re-epithelialization and the best deposition of collagen with the (DMC) 2 @β-CD (FL) and (MMC) 2 @β-CD (FL) inclusion complexes, demonstrating clearly that they have potential in wound repair. Therefore, (DMC) 2 @β-CD (FL) deserves great attention because it presented excellent results, reducing the granulation tissue and mast cell density and improving collagen remodeling. Finally, the protein binding studies suggested that the anti-inflammatory activities might exert their biological function through the inhibition of MEK, providing the possibility of development of new MEK inhibitors.

Background: Patients with hematological malignancies (HM) frequently present thrombocytopenia and higher risk of bleeding. Although transfusion is associated with higher risk of adverse events and poor outcomes, prophylactic transfusion of platelets is a common practice to prevent hemorrhagic complications. Thromboelastometry has been considered a better predictor for bleeding than isolated platelet counts in different settings. In early stages of sepsis, hypercoagulability may occur due to higher fibrinogen levels., Objectives: To evaluate the behavior of coagulation in patients with HM who develop sepsis and to verify whether a higher concentration of fibrinogen is associated with a proportional increase in maximum clot firmness (MCF) even in the presence of severe thrombocytopenia., Methods: We performed a unicentric analytical cross-sectional study with 60 adult patients with HM and severe thrombocytopenia, of whom 30 had sepsis (sepsis group) and 30 had no infections (control group). Coagulation conventional tests and specific coagulation tests, including thromboelastometry, were performed. The main outcome evaluated was MCF., Results: Higher levels of fibrinogen and MCF were found in sepsis group. Both fibrinogen and platelets contributed to MCF. The relative contribution of fibrin was significantly higher (60.5 ± 12.8% vs 43.6 ± 9.7%; P  < .001) and that of platelets was significantly lower (39.5 ± 12.8% vs 56.4 ± 9.7%; P  < .001) in the sepsis group compared with the control group., Conclusion: Patients with sepsis and HM presented higher concentrations of fibrinogen than uninfected patients, resulting in greater MCF amplitudes even in the presence of thrombocytopenia., (© 2024 The Author(s).)

Introduction: Chronic low back pain may be associated with pathoanatomical, neurophysiological, physical, psychological and social factors; thus, treatments to reduce symptoms are important to improve the quality of life of this population. We aimed to evaluate the effects of transcranial direct current stimulation (tDCS) combined with Pilates-based exercises compared with sham stimulation on pain, quality of life and disability in patients with chronic non-specific low back pain., Methods and Analysis: This is a protocol for a double-blind randomised controlled trial with participants, outcome assessor and statistician blinded. We will include 36 individuals with a history of non-specific chronic low back pain for more than 12 weeks and minimum pain intensity of 3 points on the Numerical Pain Rating Scale. Individuals will be randomised into two groups: (1) active tDCS combined with Pilates-based exercises and (2) sham tDCS combined with Pilates-based exercises. Three weekly sessions of the protocol will be provided for 4 weeks, and individuals will be submitted to three assessments: the first (T0) will be performed before the intervention protocol, the second (T1) immediately after the intervention protocol and the third (T2) will be a follow-up 1 month after the end of the intervention. We will assess pain, disability, central sensitisation, quality of life, pressure pain threshold, global impression of change, adverse events and medication use. The Numerical Pain Rating Scale and the Roland-Morris Disability Questionnaire will be used at T1 to assess pain and disability, respectively, as primary outcome measures., Ethics and Dissemination: This trial was prospectively registered in ClinicalTrials.gov website and ethically approved by the Ethics and Research Committee of the Faculty of Health Sciences of Trairi (report number: 5.411.244) before data collection. We will publish the results in a peer-reviewed medical journal and on institution websites., Trial Registration Number: ClinicalTrials.gov (NCT05467566)., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Bone fractures cause acute inflammation that, despite being important for initial repair, may delay the healing of the damaged bone. Parenteral injection of dietary protein has been shown to decrease inflammation and accelerate the repair of skin wounds and other inflammatory pathologies. Thus, our aim was to evaluate whether the intraperitoneal (i.p.) immunization with zein, an abundant protein in rodent chow, would favor bone healing. Wistar rats received i.p. immunization: saline (SG), adjuvant (AG) and zein associated with adjuvant (ZG). Then, a 2 mm of defect bone was performed on the right tibia, and on days 7, 14, 28 and 45 thereafter, analyses were performed. The results showed that the injection of zein reduced inflammation without impairing bone mineralization. Moreover, biomechanical tests demonstrated higher levels of maximum force (N) in ZG, indicating better mechanical resistance in relation to the others. The computerized tomography also indicated lower levels of medullary content in the ZG than in the SG, suggesting the absence of trabeculae in the medullary region in the ZG. These findings suggest that the injection of zein in previously tolerated animals may improve bone repair, leading to mechanically functional bone formation., (© 2023. The Author(s).)

Introduction: Low back pain in military pilots is a frequent condition which constantly leads to absences from work, decreased concentration and performance during flight, as well as changes in work functions., Methods: This is a cross-sectional analytical study including 28 fighter pilots who underwent an evaluation comprising muscle strength and fatigue resistance, trunk mobility and application of questionnaires to identify associated clinical factors., Results: It was observed that 68% of the pilots reported low back pain with an average pain intensity of 3.7 at numerical pain scale in the last week. No significant differences were observed regarding the range of motion and trunk muscle strength when pilots with low back pain were compared with asymptomatic pilots. However, lateral right trunk muscle (mean difference=16, 95% CI 0.6 to 33.0]) and lateral left trunk muscle (mean difference=22, 95% CI 1 to 44) fatigue sooner in pilots with low back pain when the two groups were compared (p=0.04 for both)., Conclusion: There was a high rate of low back pain complaints among fighter pilots. There was also a significant reduction in fatigue resistance of the lateral trunk muscles in symptomatic pilots when compared with asymptomatic pilots. These factors must be considered in the physical training of this population., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. No commercial re-use. See rights and permissions. Published by BMJ.)

Objective: This study analyzed socioeconomic inequality in self-rated health for older adults (aged fifty or over) in Brazil., Methods: Data from the 2015-2016 Brazilian Longitudinal Study of Aging (ELSI-Brazil). Socioeconomic inequality in self-rated health was measured using the concentration index, which was decomposed to analyze the contribution of different factors., Results: This study revealed that 11.5% of the older adults interviewed reported their health as poor and very poor. For the complete sample, the estimated concentration index, -0.2434, indicated that there is a concentration of poor and very poor self-rated health among older and poorer adults. Income, education and having a private health insurance plan are the factors that contributed most to the observed inequality., Discussion: The decomposition showed that there are avoidable inequalities in relation to socioeconomic status for older adults in Brazil. These factors can guide the formulation of social and health policies aimed at reducing health inequalities., Competing Interests: None., (© 2022 The Authors.)

Toxin-antitoxin (TA) gene pairs are ubiquitous in microbial chromosomal genomes and plasmids as well as temperate bacteriophages. They act as regulatory switches, with the toxin limiting the growth of bacteria and archaea by compromising diverse essential cellular targets and the antitoxin counteracting the toxic effect. To uncover previously uncharted TA diversity across microbes and bacteriophages, we analyzed the conservation of genomic neighborhoods using our computational tool FlaGs (for flanking genes), which allows high-throughput detection of TA-like operons. Focusing on the widespread but poorly experimentally characterized antitoxin domain DUF4065, our in silico analyses indicated that DUF4065-containing proteins serve as broadly distributed antitoxin components in putative TA-like operons with dozens of different toxic domains with multiple different folds. Given the versatility of DUF4065, we have named the domain Panacea (and proteins containing the domain, PanA) after the Greek goddess of universal remedy. We have experimentally validated nine PanA-neutralized TA pairs. While the majority of validated PanA-neutralized toxins act as translation inhibitors or membrane disruptors, a putative nucleotide cyclase toxin from a Burkholderia prophage compromises transcription and translation as well as inducing RelA-dependent accumulation of the nucleotide alarmone (p)ppGpp. We find that Panacea-containing antitoxins form a complex with their diverse cognate toxins, characteristic of the direct neutralization mechanisms employed by Type II TA systems. Finally, through directed evolution, we have selected PanA variants that can neutralize noncognate TA toxins, thus experimentally demonstrating the evolutionary plasticity of this hyperpromiscuous antitoxin domain., Competing Interests: The authors declare no competing interest., (Copyright © 2022 the Author(s). Published by PNAS.)

The use of edible coatings in meat is currently being investigated in several studies. However, consumers' attitudes toward this technology are not clear. This study evaluated consumers' intention to purchase vacuum-packaged meat with a chitosan coating based on the label information. Three factors combined with different levels were considered: type of cut ("loin", "French rack", or "leg steak"), coating/information (no information, chitosan without information explained, and chitosan with information explained), and price (low or high). Consumers' purchase intentions were evaluated using a 7-point structured scale (1 = certainly would not buy; 7 = certainly would buy). The data were analyzed by conjoint analysis and cluster analysis. The average purchase intention value was 4.3 ± 0.5. The "French rack" cut showed the highest value for purchase intention and a high price was also a relevant factor. The packaging with the label stating the presence of a "chitosan" coating without giving any further information obtained higher purchase intentions than the one with the more detailed information label. In the cluster analysis, the factor "type of cut" showed the highest relative importance in two groups, while "price" had a higher impact in another. The type of cut was the main factor affecting consumers' purchase intentions. However, different attitudes were observed depending on the group.

Stilbenes are a class of natural compounds with a wide variety of biological effects, such as antitumor activity. The best-known stilbene is resveratrol, whose clinical application is limited due to its low bioavailability. Methoxylated derivatives of this stilbene, including cis-trimethoxystilbene (cis-TMS) and trans-trimethoxystilbene (trans-TMS) have demonstrated more pronounced cytotoxic and anti-proliferative effects than resveratrol. Thus, the objective of this study is to evaluate and compare the cytotoxicity and antiproliferative effects of cis- and trans-TMS in MCF-7 and its normal counterpart MCF-10A. Both compounds were cytotoxic, genotoxic, and induced G2-M accumulation and cell death in the two cell lines. These results suggested that the genotoxicity of cis- and trans-TMS is involved in the reduction of cellular proliferation of MCF-7 and MCF-10A cells, but notably, such antiproliferative effects are more pronounced for cis- than trans-TMS.

Introduction: Propolis is widely used in folk medicine, and many factors can affect its chemical composition, including abiotic factors that can influence plants and bees. Therefore, analytical methods are powerful techniques in the quality control of such products., Objective: Develop and validate an analytical method for quantifying volatile compounds in Brazilian brown propolis, and evaluate its biological activities., Methods: A gas chromatography flame ionisation detector (GC-FID) analytical method was validated, attending the parameters of international validation guidelines as ANVISA 2017 and ICH 2005, for quantification of compounds present in volatile oils from propolis. Evaluation of cytotoxic, antimicrobial, and leishmanicidal activities of the oil., Results: The compounds 1,8-cineole, terpinen-4-ol, α-copaene, β-caryophyllene, γ-muurolene, nerolidol, spathulenol, and γ-palmitolactone were isolated from the volatile fraction of a Brazilian brown propolis and used in the method validation. All the validation parameters of the method were satisfactory. The volatile fraction displayed a significant leishmanicidal activity, with half maximal inhibition concentration (IC 50 ) = 21.3 μg/mL against amastigote forms and IC 50 = 25.1 μg/mL against promastigote forms of Leishmania amazonensis. The oil also displayed an antibacterial effect by inhibiting the growth of Streptococcus mutans and Staphylococcus aureus at 25 μg/mL and 50 μg/mL, respectively, but it was not cytotoxic against AGP-01, He-La and CHO-K1cell lines, with IC 50 > 100 μg/mL., Conclusion: The GC-FID method can be a useful tool in the quality control of propolis material. The southeast brown propolis showed a high chemical complexity in its volatile fraction, which displayed leishmanicidal activity and bactericidal activity., (© 2020 John Wiley & Sons, Ltd.)

Axonal damage is an early step in traumatic and neurodegenerative disorders of the central nervous system (CNS). Damaged axons are not able to regenerate sufficiently in the adult mammalian CNS, leading to permanent neurological deficits. Recently, we showed that inhibition of the autophagic protein ULK1 promotes neuroprotection in different models of neurodegeneration. Moreover, we demonstrated previously that axonal protection improves regeneration of lesioned axons. However, whether axonal protection mediated by ULK1 inhibition could also improve axonal regeneration is unknown. Here, we used an adeno-associated viral (AAV) vector to express a dominant-negative form of ULK1 (AAV.ULK1.DN) and investigated its effects on axonal regeneration in the CNS. We show that AAV.ULK1.DN fosters axonal regeneration and enhances neurite outgrowth in vitro. In addition, AAV.ULK1.DN increases neuronal survival and enhances axonal regeneration after optic nerve lesion, and promotes long-term axonal protection after spinal cord injury (SCI) in vivo. Interestingly, AAV.ULK1.DN also increases serotonergic and dopaminergic axon sprouting after SCI. Mechanistically, AAV.ULK1.DN leads to increased ERK1 activation and reduced expression of RhoA and ROCK2. Our findings outline ULK1 as a key regulator of axonal degeneration and regeneration, and define ULK1 as a promising target to promote neuroprotection and regeneration in the CNS.

The Buccal Micronucleus Cytome Assay (BMCyt) has become an important biomonitoring tool for assessing cytogenetic damage in many studied populations. Each laboratory applies protocols that vary according to the method of collecting and preparing samples. Besides, Brazil is a country of great territorial extensions that received immigrants from various parts of the world with different genetic backgrounds. Therefore, the present study aimed to evaluate the inter-laboratory variation in scoring the same set of slides using the more comprehensive scoring criteria, to standardize the BMCyt protocol, to observe the basal alterations in populations of different Brazilian regions and to compare it with other places around the world. Our results showed that a valuable number of laboratories participated, ten laboratories from different regions of the country, for the validation of the BMCyt in human biomonitoring studies, resulting in the 804 healthy individuals. This was possible because we observed: a range of measures needs to be considered, such as the baseline frequency of DNA damage and cell death in non-exposed individuals; age when grouped showed an influence on DNA damage, although when evaluated by group we did not see an influence; association between smoking habit and all endpoints of the BMCyt (except karyolytic cells) was evident; the basal MN frequency, in the majority of groups, follows those around the world; and the BMCyt was confirmed as a good health status biomarker. We emphasize the need for constant discussions on the parameters of cell death due to greater difficulty among the analyzers., Competing Interests: Declaration of Competing Interest The authors declare no potential conflicts of interest concerning the research, authorship, and/or publication of this article., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Type 2 diabetes mellitus (T2DM), dyslipidemia and periodontitis are frequently associated pathologies; however, there are no studies showing the peripheral blood transcript profile of these combined diseases. Here we identified the differentially expressed genes (DEGs) of circulating lymphocytes and monocytes to reveal potential biomarkers that may be used as molecular targets for future diagnosis of each combination of these pathologies (compared to healthy patients) and give insights into the underlying molecular mechanisms of these diseases. Study participants (n = 150) were divided into groups: (H) systemically and periodontal healthy (control group); (P) with periodontitis, but systemically healthy; (DL-P) with dyslipidemia and periodontitis; (T2DMwell-DL-P) well-controlled type 2 diabetes mellitus with dyslipidemia and periodontitis; and (T2DMpoorly-DL-P) poorly-controlled type 2 diabetes mellitus with dyslipidemia and periodontitis. We preprocessed the microarray data using the Robust Multichip Average (RMA) strategy, followed by the RankProd method to identify candidates for DEGs. Furthermore, we performed functional enrichment analysis using Ingenuity Pathway Analysis and Gene Set Enrichment Analysis. DEGs were submitted to pairwise comparisons, and selected DEGs were validated by quantitative polymerase chain reaction. Validated DEGs verified from T2DMpoorly-DL-P versus H were: TGFB1I1, VNN1, HLADRB4 and CXCL8; T2DMwell-DL-P versus H: FN1, BPTF and PDE3B; DL-P versus H: DAB2, CD47 and HLADRB4; P versus H: IGHDL-P, ITGB2 and HLADRB4. In conclusion, we identified that circulating lymphocytes and monocytes of individuals simultaneously affected by T2DM, dyslipidemia and periodontitis, showed an altered molecular profile mainly associated to inflammatory response, immune cell trafficking, and infectious disease pathways. Altogether, these results shed light on novel potential targets for future diagnosis, monitoring or development of targeted therapies for patients sharing these conditions.

Under stressful conditions, bacterial RelA-SpoT Homolog (RSH) enzymes synthesize the alarmone (p)ppGpp, a nucleotide second messenger. (p)ppGpp rewires bacterial transcription and metabolism to cope with stress, and, at high concentrations, inhibits the process of protein synthesis and bacterial growth to save and redirect resources until conditions improve. Single-domain small alarmone synthetases (SASs) are RSH family members that contain the (p)ppGpp synthesis (SYNTH) domain, but lack the hydrolysis (HD) domain and regulatory C-terminal domains of the long RSHs such as Rel, RelA, and SpoT. We asked whether analysis of the genomic context of SASs can indicate possible functional roles. Indeed, multiple SAS subfamilies are encoded in widespread conserved bicistronic operon architectures that are reminiscent of those typically seen in toxin-antitoxin (TA) operons. We have validated five of these SASs as being toxic (toxSASs), with neutralization by the protein products of six neighboring antitoxin genes. The toxicity of Cellulomonas marina toxSAS FaRel is mediated by the accumulation of alarmones ppGpp and ppApp, and an associated depletion of cellular guanosine triphosphate and adenosine triphosphate pools, and is counteracted by its HD domain-containing antitoxin. Thus, the ToxSAS-antiToxSAS system with its multiple different antitoxins exemplifies how ancient nucleotide-based signaling mechanisms can be repurposed as TA modules during evolution, potentially multiple times independently., Competing Interests: The authors declare no competing interest., (Copyright © 2020 the Author(s). Published by PNAS.)

A new method of screening was developed to generate 770 organic and water-soluble fractions from extracts of nine species of marine sponges, from the growth media of 18 species of marine-derived fungi, and from the growth media of 13 species of endophytic fungi. The screening results indicated that water-soluble fractions displayed significant bioactivity in cytotoxic, antibiotic, anti- Leishmania , anti- Trypanosoma cruzi , and inhibition of proteasome assays. Purification of water-soluble fractions from the growth medium of Penicillium solitum IS1-A provided the new glutamic acid derivatives solitumine A ( 1 ), solitumine B ( 2 ), and solitumidines A-D ( 3 - 6 ). The structures of compounds 1 - 6 have been established by analysis of spectroscopic data, chemical derivatizations, and vibrational circular dichroism calculations. Although no biological activity could be observed for compounds 1 - 6 , the new structures reported for 1 - 6 indicate that the investigation of water-soluble natural products represents a relevant strategy in finding new secondary metabolites.

Melanin is produced by melanocytes and protects against DNA damage by ultraviolet light. Unfortunately, the melanin protein present in melanoma tumor cells is often co-purified during DNA extraction, and this contamination may inhibit subsequent PCR methods, which directly impacts research applications and the molecular diagnostic tests needed for targeted therapeutics. There are presently no described purification protocols that efficiently remove melanin from genomic DNA. In this study, we compare six different methods for melanin removal from genomic DNA: Agarose Gel Electrophoresis, 1mg Chelex®-100, Chelex®-100 5%, centrifugation, OneStep™ PCR Inhibitor Removal Kit and centrifugation plus OneStep™ PCR Inhibitor Removal Kit. Each comparison was made using 16 formalin-fixed paraffin-embedded (FFPE) and 11 fresh cell line samples. All samples were initially tested using the multiplex PCR reaction for GAPDH gene that generates different sized amplified products: 100, 200, 300 and 400 base pairs, which could be inhibited by the addition of exogenous melanin. Six purification protocols were then applied, and all samples that amplified at least one GAPDH fragment were sequenced to analyze the presence of the BRAF V600E mutation. The efficiencies of amplification decreased for larger sized fragments in all methods. Our comparisons showed that centrifugation combined with the OneStep™ PCR Inhibitor Removal Kit was superior to all other methods for successful BRAF sequencing with 100% (100bp), 75% (200bp), 50% (300bp), and 31.3% (400bp) amplification efficiencies for the different amplicon sizes. In conclusion, this genomic DNA extraction method is highly efficient for successful PCR when tumor samples are contaminated with melanin.

During amino acid starvation the Escherichia coli stringent response factor RelA recognizes deacylated tRNA in the ribosomal A-site. This interaction activates RelA-mediated synthesis of alarmone nucleotides pppGpp and ppGpp, collectively referred to as (p)ppGpp. These two alarmones are synthesized by addition of a pyrophosphate moiety to the 3' position of the abundant cellular nucleotide GTP and less abundant nucleotide GDP, respectively. Using untagged native RelA we show that allosteric activation of RelA by pppGpp increases the efficiency of GDP conversion to achieve the maximum rate of (p)ppGpp production. Using a panel of ribosomal RNA mutants, we show that the A-site finger structural element of 23S rRNA helix 38 is crucial for RelA binding to the ribosome and consequent activation, and deletion of the element severely compromises (p)ppGpp accumulation in E. coli upon amino acid starvation. Through binding assays and enzymology, we show that E. coli RelA does not form a stable complex with, and is not activated by, deacylated tRNA off the ribosome. This indicates that in the cell, RelA first binds the empty A-site and then recruits tRNA rather than first binding tRNA and then binding the ribosome.

This paper is the first report on the in vitro effects of licochalcone A, a chalcone isolated from Glycyrrhiza inflate Batalin (Leguminosae), on Schistosoma mansoni adult worms. In vitro, licochalcone A afforded lethal concentrations for 50% of parasites (LC 50 ) of 9.12±1.1 and 9.52±0.9μM against female and male adult worms, respectively, at 24h. Additionally, the compound reduced the total number of S. mansoni eggs and affected the development of eggs produced by S. mansoni adult worms. Together, the results achieved after 24h showed that licochalcone A was 55.7- and 53.3-fold more toxic to S. mansoni female and male adult worms than to Chinese hamster ovary fibroblasts cells, respectively. Treatment with licochalcone A elicited drastic changes in the tegument of S. mansoni adult worms, as well as mitochondrial alteration and chromatin condensation. Licochalcone A also increased the superoxide anion level and decreased the superoxide dismutase activity in S. mansoni adult worms. Overall, our results indicated that licochalcone A displays in vitro schistosomicidal activity. This effect may result from increased production of reactive oxygen species (ROS) induced by the action of licochalcone A. The resulting ROS could act on the S. mansoni tegument and membranes and help induce the death of S. mansoni adult worms., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

Despite advances in the field of biomaterials for bone repair/regeneration, some challenges for developing an ideal bone substitute need to be overcome. Herein, this study synthesized and evaluated in vitro a nanocomposite based on bacterial cellulose (BC), collagen (COL), apatite (Ap) and osteogenic growth peptide (OGP) or its C-terminal pentapeptide [OGP(10-14)] for bone regeneration purposes. The BC-COL nanocomposites were successfully obtained by carbodiimide-mediated coupling as demonstrated by spectroscopy analysis. SEM, FTIR and 31 P NMR analyses revealed that in situ synthesis to apatite was an effective route for obtaining of bone-like apatite. The OGP-containing (BC-COL)-Ap stimulated the early development of the osteoblastic phenotype. Additionally, the association among collagen, apatite, and OGP peptides enhanced cell growth compared with OGP-containing BC-Ap. Furthermore, none of the nanocomposites showed cytotoxic, genotoxic or mutagenic effects. These promising results suggest that the (BC-COL)-Ap associated with OGP peptides might be considered a potential candidate for bone tissue engineering applications., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Here we describe an HPLC-based method to quantify bacterial housekeeping nucleotides and the signaling messengers ppGpp and pppGpp. We have replicated and tested several previously reported HPLC-based approaches and assembled a method that can process 50 samples in three days, thus making kinetically resolved experiments feasible. The method combines cell harvesting by rapid filtration, followed by acid extraction, freeze-drying with chromatographic separation. We use a combination of C18 IPRP-HPLC (GMP unresolved and co-migrating with IMP; GDP and GTP; AMP, ADP and ATP; CTP; UTP) and SAX-HPLC in isocratic mode (ppGpp and pppGpp) with UV detection. The approach is applicable to bacteria without the requirement of metabolic labelling with 32P-labelled radioactive precursors. We applied our method to quantify nucleotide pools in Escherichia coli BW25113 K12-strain both throughout the growth curve and during acute stringent response induced by mupirocin. While ppGpp and pppGpp levels vary drastically (40- and ≥8-fold, respectively) these changes are decoupled from the quotients of the housekeeping pool and guanosine and adenosine housekeeping nucleotides: NTP/NDP/NMP ratio remains stable at 6/1/0.3 during both normal batch culture growth and upon acute amino acid starvation.

Oral tolerance refers to the specific inhibition of immune responsiveness to T-cell-dependent antigens contacted through the oral route before parenteral immunization. Oral tolerance to one protein does not inhibit immune responses to other unrelated proteins, but parenteral injection of tolerated antigens plus adjuvant into tolerant, but not normal, mice inhibits immune responses to antigens injected concomitantly or soon thereafter. The inhibitory effect triggered by parenteral injection of tolerated proteins is known as bystander suppression or indirect effects of oral tolerance. Intraperitoneal injection of ovalbumin (OVA) plus alum adjuvant in OVA-tolerant mice soon before skin injury inhibits inflammation and improves cutaneous wound healing. However, as OVA is not a regular component of mouse chow, we tested whether indirect effects could be triggered by zein, the main protein of corn that is regularly present in mouse chow. We show that intraperitoneal injection of a single dose (10 μg) of zein plus alum adjuvant soon before skin injury in mice reduces leucocyte infiltration but increase the number of T cells and the expression of resistin-like molecule-α (a marker of alternatively activated macrophages) in the wound bed, increases the expression of transforming growth factor-β 3 in the newly formed epidermis and reduces cutaneous scar formation. These results suggest that indirect effects of oral tolerance triggered by parenteral injection of regular dietary components may be further explored as one alternative way to promote scarless wound healing., (© 2017 John Wiley & Sons Ltd.)

The cytotoxicity and genotoxicity of the stilbenes (E)-methyl-4-(3-5-dimethoxystyryl)benzoate (ester), (E)-4-(3-5-dimethoxystyryl)aniline (amino), (Z)-1,3-dimethoxy-5-(4-methoxystyryl)benzene (cis-TMS) and (E)-1,3-dimethoxy-5-(4-methoxystyryl)benzene (trans-TMS) were investigated in this work. Structural modifications of resveratrol, a naturally occurring stilbene, have been previously performed, including the replacement of hydroxyl by different functional groups. Such modifications resulted in significant improvement of target-specific effects on cell death and antiproliferative responses. The parameters were evaluated using XTT assay, clonogenic survival assay and the cytokinesis-block micronucleus assay in CHO-K1 and HepG2 cell lines. The results showed that cis-TMS is approximately 250-fold more cytotoxic than the amino and ester, and 128-fold more cytotoxic than trans-TMS. When genotoxicity was evaluated, only the trans-TMS did not significantly increase the frequency of micronucleus (MN). While the cis-TMS induced a mean of 5.2 and 5.9 MN/100 cells at 0.5 μM in CHO-K1 and HepG2, respectively, the amino and ester induced 3.1 and 3.6 MN/100 cells at 10 μM in CHO-K1, respectively, and 3.5 and 3.8 in HepG2. Trans-TMS is genotoxic only in HepG2 cells. Based on these results, the cis-TMS was the most cytotoxic and genotoxic compound in both cell lines.

The goal of our study was to evaluate the effect of kaurenoic acid, obtained from copaiba oil resin, in gastric cancer (GC) and a normal mucosa of stomach (MNP01) cell lines. The compound was tested at concentrations of 2.5, 5, 10, 30 and 60μg/mL. Comet and micronucleus assays were used to access its potential genotoxicity in vitro. Moreover, we evaluated the effect of kaurenoic acid in cell cycle progression and in the transcription of genes involved in the control of the cell cycle: MYC, CCND1, BCL2, CASP3, ATM, CHK2 and TP53. Kaurenoic acid induced an increase on cell DNA damage or micronucleus frequencies on GC cell lines in a dose-dependent manner. The GC and MNP01 cell lines entering DNA synthesis and mitosis decreased significantly with kaurenoic acid treatment, and had an increased growth phase compared with non-treated cells. The treatment induced apoptosis (or necrosis) even at a concentration of 2.5μg/mL in relation to non-treated cells. GC cell lines presented reduced MYC, CCND1, BCL2 and CASP3 transcription while ATM, CHK2 and TP53 increased in transcription in relation to non-treated cells, especially at a concentration above 10μg/mL. The gene transcription in the MNP01 (non-treated non-cancer cell line) was designated as a calibrator for all the GC cell lines. In conclusion, our results showed that kaurenoic acid obtained from Copaifera induces DNA damage and increases the micronuclei frequency in a dose-dependent manner in GC cells, with a significant genotoxicity observed above the concentration of 5μg/mL. Moreover, this compound seems to be able to induce cell cycle arrest and apoptosis in GC cells., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

The diterpene kaurenoic acid (KA) has vasorelaxant, antimicrobial, anti-tumoural and anti-leishmanial effects. Semi-synthetic derivatives were obtained to achieve more satisfactory responses. The assessment of genotoxicity is part of the toxicological evaluation of therapeutic compound candidates. The present study investigated the cytotoxicity and genotoxicity of KA and its semi-synthetic derivatives methoxy kaurenoic acid (MKA) and kaurenol (KRN) using the CHO-K1 cell line. The cytotoxicity evaluation demonstrated that treatments with 200 and 400 μM KA reduced cellular proliferation to 36.5 and 4.43%, respectively, and that 100 and 200 μM KA reduced the survival fraction (SF) to 48.1 and 5.5%, respectively. MKA and KRN at concentrations of 400 μM reduced proliferation to 81 and 86.8%, respectively, while 100 and 200 μM KRN reduced the SF to 50%, and 200 μM MKA reduced the SF to 74%. No genotoxicity was observed for KA or MKA. However, 100 μM KRN increased the DNA damage index, as detected by comet assay, although a micronucleus assay did not confirm these data. The results demonstrated that KA and its semi-synthetic derivative MKA were not genotoxic when tested at noncytotoxic concentrations, but KRN was genotoxic at the highest concentration that was tested, as demonstrated by the comet assay., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The nucleotide (p)ppGpp is a key regulator of bacterial metabolism, growth, stress tolerance, and virulence. During amino acid starvation, the Escherichia coli (p)ppGpp synthetase RelA is activated by deacylated tRNA in the ribosomal A-site. An increase in (p)ppGpp is believed to drive the formation of antibiotic-tolerant persister cells, prompting the development of strategies to inhibit (p)ppGpp synthesis. We show that in a biochemical system from purified E. coli components, the antibiotic thiostrepton efficiently inhibits RelA activation by the A-site tRNA. In bacterial cultures, the ribosomal inhibitors thiostrepton, chloramphenicol, and tetracycline all efficiently abolish accumulation of (p)ppGpp induced by the Ile-tRNA synthetase inhibitor mupirocin. This abolishment, however, does not reduce the persister level. In contrast, the combination of dihydrofolate reductase inhibitor trimethoprim with mupirocin, tetracycline, or chloramphenicol leads to ampicillin tolerance. The effect is independent of RelA functionality, specific to β-lactams, and not observed with the fluoroquinolone norfloxacin. These results refine our understanding of (p)ppGpp's role in antibiotic tolerance and persistence and demonstrate unexpected drug interactions that lead to tolerance to bactericidal antibiotics., (Copyright © 2017 Kudrin et al.)

Introduction: The clinical manifestations of sickle cell disease are related to the polymerization of hemoglobin S. The chronic hemolysis caused by this condition often causes the formation of gallstones that can migrate and block the common bile duct leading to acute abdomen., Objective: This study aimed to evaluate the profile of patients with sickle cell disease and cholelithiasis., Methods: Patients with sickle cell disease were separated into groups according to the presence or absence of cholelithiasis. Socioepidemiological and clinical characteristics, such as gender, age, use of hydroxyurea and the presence of other hemoglobinopathies were researched in the medical records of patients., Results: A hundred and seven patients with sickle cell anemia were treated at the institution. Of these, 27 (25.2%) had cholelithiasis. The presence of cholelithiasis was higher in the 11-29 age group than in younger than 11 years and over 29 years. No association was found for the presence of cholelithiasis with gender, use of hydroxyurea or type of hemoglobinopathy (hemoglobin SS, hemoglobin SC or sickle beta-thalassemia). Sixteen of the patients had to be submitted to cholecystectomy with 14 of the surgeries being performed by laparoscopy. Complications were observed in three patients and one patient died for reasons unrelated to the surgery., Conclusion: A quarter of patients with sickle cell disease had gallstones, more commonly in the 11- to 29-year age range. Patients should be monitored from childhood to prevent cholelithiasis with preoperative, intra-operative and postoperative care being crucial to reduce the risk of complications in these patients., (Copyright © 2016 Associação Brasileira de Hematologia, Hemoterapia e Terapia Celular. Published by Elsevier Editora Ltda. All rights reserved.)

Despite increasing research in type 2 diabetes mellitus (T2D), there are few studies showing the impact of the poor glycemic control on biological processes occurring in T2D. In order to identify potential genes related to poorly/well-controlled patients with T2D, our strategy of investigation included a primary screen by microarray (Human Genome U133) in a small group of individuals followed by an independent validation in a greater group using RT-qPCR. Ninety patients were divided as follows: poorly controlled T2D (G1), well-controlled T2D (G2), and normoglycemic individuals (G3). After using affy package in R, differentially expressed genes (DEGs) were prospected as candidate genes potentially relevant for the glycemic control in T2D patients. After validation by RT-qPCR, the obtained DEGs were as follows-G1 + G2 versus G3: HLA-DQA1 , SOS1 , and BRCA2 ; G2 versus G1: ENO2 , VAMP2 , CCND3 , CEBPD , LGALS12 , AGBL5 , MAP2K5 , and PPAP2B ; G2 versus G3: HLA-DQB1 , MCM4 , and SEC13 ; and G1 versus G3: PPIC . This demonstrated a systemic exacerbation of the gene expression related to immune response in T2D patients. Moreover, genes related to lipid metabolisms and DNA replication/repair were influenced by the glycemic control. In conclusion, this study pointed out candidate genes potentially associated with adequate glycemic control in T2D patients, contributing to the knowledge of how the glycemic control could systemically influence gene expression.

A high percentage of type 2 diabetes mellitus (T2D) patients are also affected by dyslipidemia and chronic periodontitis (CP), but no studies have determined the gene expression in patients that are simultaneously affected by all three diseases. We investigated the systemic expression of immune-related genes in T2D, dyslipidemia, and CP patients. One hundred and fifty patients were separated into five groups containing 30 individuals each: (G1) poorly controlled T2D with dyslipidemia and CP; (G2) well-controlled T2D with dyslipidemia and CP; (G3) normoglycemic individuals with dyslipidemia and CP; (G4) healthy individuals with CP; (G5) systemic and periodontally healthy individuals. Blood analyses of lipid and glycemic profiles were carried out. The expression of genes, including IL10, JAK1, STAT3, SOCS3, IP10, ICAM1, IFNA, IFNG, STAT1, and IRF1, was investigated by RT-qPCR. Patients with dyslipidemia demonstrated statistically higher expression of the IL10 and IFNA genes, while IFNG, IP10, IRF1, JAK1, and STAT3 were lower in comparison with nondyslipidemic patients. Anti-inflammatory genes, such as IL10 , positively correlated with parameters of glucose, lipid, and periodontal profiles, while proinflammatory genes, such as IFNG , were negatively correlated with these parameters. We conclude that dyslipidemia appears to be the primary disease that is associated with gene expression of immune-related genes, while parameters of T2D and CP were correlated with the expression of these important immune genes., Competing Interests: The authors declare that they have no competing interests.

Breast cancer incidence rises worldwide and new chemotherapeutical strategies have been investigated to overcome chemoresistance. (-)-Hinokinin is a dibenzylbutyrolactone lignan derived from the partial synthesis of (-)-cubebin extracted from Piper cubeba seeds. Biological effects of dibenzylbutyrolactone lignans include antiviral, antitumor, anti-inflammatory, and trypanocidal activities. In the present study, we evaluated the ability of (-)-hinokinin to modulate the antiproliferative effects of doxorubicin intumoral (MCF-7 and SKBR-3) and normal (MCF-10 A) breast cell lines. Treatment with (-)-hinokinin did not affect the cellular proliferation or contribute to the antitproliferative effects of doxorubicin in MCF-10 A cells. After 24 and 48 hours of treatment with (-)-hinokinin, MCF-7 and SKBR-3 were accumulated in G2/M and, when combined with doxorubicin, (-)-hinokinin contributed to the antiproliferative effects of this chemotherapic by modulation of the cyclin-dependent kinase inhibitor 1. Apoptotic cell death was observed in response to (-)-hinokinin alone in MCF-7, but not in SKBR-3 even 72 hours after treatment. In MCF-7, doxorubicin-induced apoptosis was not increased by (-)-hinokinin. The findings of the present study suggest (-)-hinokinin as an antiproliferative agent that contributes to the effects of doxorubicin. (-)-Hinokinin modulates apoptotic cell death via the molecular regulation of the cell cycle and apoptotic control genes, but the cellular genetic background directly affects the cell fate decision in response to treatment., (Georg Thieme Verlag KG Stuttgart · New York.)

Background: Immunological tolerance refer to the inhibition of specific immune responsiveness and the ingestion of proteins previous to immunization is a reliable method to induce (oral) tolerance. Parenteral exposure to tolerated antigens, in adjuvant, trigger indirect and systemic effects that inhibits concomitant immune responses to other unrelated antigens and also decrease unrelated inflammatory responses. Interesting, intraperitoneal (i.p.) exposure to orally-tolerated proteins soon before an incisional linear skin wound improves the healing by primary intention in mice. An important clinical and surgical objective is to identify strategies to improve wound healing and reduce scarring., Objective: To evaluate whether i.p. injection of an orally-tolerated protein improves wound healing by secondary intention and reduce scarring of full-thickness excisional skin injury., Methods: C57Bl/6 mice were turned tolerant to ovalbumin (OVA) by drinking a solution containing OVA; seven days later, they received an i.p. injection of OVA plus Al(OH)3 adjuvant immediately before two full-thickness excisional skin wounds, under anesthesia. The wound healing process was evaluated macro and microscopically after H&E, toluidine blue and Gomori's Trichrome staining. The presence of granulocytes, macrophages, miofibroblasts, fibronectin, collagen I and collagen III was investigated by immunofluorescence and the levels of cytokines by flow cytometry or ELISA. Mice not tolerant to OVA were included as controls., Results: The i.p. injection of OVA+Al(OH)3 in mice orally tolerant to OVA reduced the subsequent inflammatory response in the wound bed and the cutaneous scarring. There was a change in the pattern of collagen deposition making it more similar to the pattern observed in intact skin. In tolerant mice, mast cells and granulocytes (Ly-6C/G+), were reduced, while lymphocytes (CD3+) were increased in the wound bed. Time course analysis of Th1/Th2/Th17 cytokines and growth factors showed slightly differences between tolerant and control groups., Conclusion: Parenteral injection of an orally-tolerated protein has systemic consequences that impair the inflammatory response triggered by skin injury and reduce the cutaneous scarring., (Copyright © 2015 Elsevier GmbH. All rights reserved.)

The transition of Escherichia coli from the exponential into the stationary phase of growth induces the stringent response, which is mediated by the rapid accumulation of the alarmone nucleotide (p)ppGpp produced by the enzyme RelA. The significance of RelA's functionality during the transition in the opposite direction, i.e. from the stationary phase into new exponential growth, is less well understood. Here we show that the relaxed strain, i.e. lacking the relA gene, displays a relative delay in regrowth during the new exponential growth phase in comparison with the isogenic wild type strain. The severity of the effect is a function of both the carbon source and amino acid composition of the outgrowth media. As a result, the loss of RelA functionality increases E. coli tolerance to the bactericidal antibiotic ampicillin during growth resumption in fresh media in a medium-specific way. Taken together, our data underscore the crucial role of medium composition and growth conditions for studies of the role of individual genes and regulatory networks in bacterial phenotypic tolerance to antibiotics.

Skin wound healing is a complex process involving many types of cells and molecules and often results in scar tissue formation in adult mammals. However, scarless healing occurs in foetal skin and minimal scars may occur after cutaneous healing in the adult with reduced inflammation. Alpha-melanocyte-stimulating hormone (α-MSH) is widely distributed within the central nervous system and in other body regions, such as the skin, and has strong anti-inflammatory activity. The aim in the present experiments was to learn whether intraperitoneal (i.p) injection of α-MSH just before skin wounds antagonize inflammation and improves skin wound healing in adult mice. C57BL/6 young adult mice received an i.p. injection of 1 mg/kg of α-MSH and, 30 min later, two circular through-and-through holes (6.5 mm diameter) were made in their dorsal skin under anaesthesia. Control mice were wounded after vehicle injection. The wound healing process was analysed macroscopically and microscopically at 3, 7, 40 and 60 days. Skin samples were fixed in formalin, embedded in paraffin, sectioned at 5 μm, stained with H&E or toluidine blue for cell analysis or Gomori's trichrome for extracellular matrix (ECM) analysis. Other samples were fixed in DMSO+methanol, embedded in paraplast and incubated with anti-CD45, antismooth muscle actin, anticollagen-I and anticollagen-III for immunofluorescence analysis. Alpha-MSH significantly reduced the number of leucocytes, mast cells and fibroblasts at 3 and 7 days after injury. On days 40 and 60, α-MSH reduced scar area and improved the organization of the collagen fibres indicating that it may direct the healing into a more-regenerative/less-scarring pathway., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The use of poly(L-lactic acid) (PLA) has been considered an important alternative for medical devices once this polyester presents biomechanical, optical and biodegradable properties. Moreover, the use of PLA results in less inflammatory reactions and more recently it has been proposed its application in drug delivery systems. Genotoxicological evaluations are considered part of the battery assays in toxicological analysis. Considering the wide applications of PLA, the present work evaluated the potential cytotoxic and genotoxic effects of PLA in CHO-K1 cells, as well as its physicochemical properties. No cytotoxic effects of PLA were detected by colorimetric tetrazolium assay (XTT) analysis, and the clonogenic survival assay showed that PLA did not disrupt the replicative cell homeostasis, neither exhibited genotoxic effects as evidenced by comet and micronucleus assays. Thermogravimetric properties of PLA were not altered after contact with cells and this film exhibited ability in absorb and release Europium(III) complex. All these data suggest genotoxicological safety of PLA for further applications in drug delivery systems., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Sesquiterpene lactones are known to be active, but are also known to present high cytotoxicity. In the present work an evaluation of how slight structural alterations affect the cytotoxicity and the schistosomicidal activity of sesquiterpene lactones was undertaken. More specifically, we assessed the activity of budlein-A, a furanoheliangolide sesquiterpene lactone, and four of its derivatives. The structural modifications of budlein-A, presented in this work, diminished the cytotoxicity and changed the antiparasitary behavior of the molecule. They also provided data for a better understanding of the sesquiterpene lactone cytotoxicity. The establishment of the structures of three synthesized sesquiterpene lactones on the basis of NMR and HRESIMS data is also presented here. Complete and detailed (1)H and (13)C 1D and 2D NMR data, with measurements of all J values and all multiplicities clarified, are presented for five sesquiterpene lactones for the first time.

In this study, we analysed the frequency of micronuclei (MN), nucleoplasmic bridges (NPBs) and nuclear buds (NBUDs) and evaluated mutagen-induced sensitivity in the lymphocytes of patients chronically infected with hepatitis B virus (HBV) or hepatitis C virus (HCV). In total, 49 patients with chronic viral hepatitis (28 HBV-infected and 21 HCV-infected patients) and 33 healthy, non-infected blood donor controls were investigated. The frequencies (‰) of MN, NPBs and NBUDs in the controls were 4.41 ± 2.15, 1.15 ± 0.97 and 2.98 ± 1.31, respectively. The frequencies of MN and NPBs were significantly increased (p < 0.0001) in the patient group (7.01 ± 3.23 and 2.76 ± 2.08, respectively) compared with the control group. When considered separately, the HBV-infected patients (7.18 ± 3.57) and HCV-infected patients (3.27 ± 2.40) each had greater numbers of MN than did the controls (p < 0.0001). The HCV-infected patients displayed high numbers of NPBs (2.09 ± 1.33) and NBUDs (4.38 ± 3.28), but only the HBV-infected patients exhibited a significant difference (NPBs = 3.27 ± 2.40, p < 0.0001 and NBUDs = 4.71 ± 2.79, p = 0.03) in comparison with the controls. Similar results were obtained for males, but not for females, when all patients or the HBV-infected group was compared with the controls. The lymphocytes of the infected patients did not exhibit sensitivity to mutagen in comparison with the lymphocytes of the controls (p = 0.06). These results showed that the lymphocytes of patients who were chronically infected with HBV or HCV presented greater chromosomal instability.

Recently, while studying erythrocytic apoptosis during Plasmodium yoelii infection, we observed an increase in the levels of non-parasitised red blood cell (nRBC) apoptosis, which could be related to malarial anaemia. Therefore, in the present study, we attempted to investigate whether nRBC apoptosis is associated with the peripheral RBC count, parasite load or immune response. To this end, BALB/c mice were infected with P. yoelii 17XL and nRBC apoptosis, number of peripheral RBCs, parasitaemia and plasmatic levels of cytokines, nitric oxide and anti-RBC antibodies were evaluated at the early and late stages of anaemia. The apoptosis of nRBCs increased at the late stage and was associated with parasitaemia, but not with the intensity of the immune response. The increased percentage of nRBC apoptosis that was observed when anaemia was accentuated was not related to a reduction in peripheral RBCs. We conclude that nRBC apoptosis in P. yoelii malaria appears to be induced in response to a high parasite load. Further studies on malaria models in which acute anaemia develops during low parasitaemia are needed to identify the potential pathogenic role of nRBC apoptosis.

Background: Hemovigilance is an organized system of surveillance throughout the transfusion chain intended to evaluate information in order to prevent the appearance or recurrence of adverse reactions related to the use of blood products., Objective: The aims of this study were to assess the late reporting of incidents related to possible seroconversion in respect to age, marital status and ethnical background, annual variations in late reporting, the number of reports opened and closed, seroconversion of donors and transfusions of blood products within the window period., Methods: This retrospective, descriptive study used data on blood donations in the blood bank in Uberaba during the period from 2004 to 2011. Some socio-epidemiological characteristics of the donors and serology test results of donors and recipients were analyzed in respect to the late reporting of incidents related to possible seroconversion. The Chi-square test, odds ratio and a regression model were used for statistical analysis., Results: From 2004 to 2011, the blood bank in Uberaba collected 117,857 blood bags, 284 (0.24%) of which were investigated for late reported incidents. The profile of the donors was less than 29 years old, unmarried and non-Whites. Differences in age (p-value < 0.0001), marital status (p-value = 0.0002) and ethnical background (p-value < 0.0001) were found to be statistically significant. There was no statistical difference between men and women (0.24% and 0.23% respectively; p-value = 0.951). The number of late reported incidents increased until 2008 followed by a downward trend until 2011. There were twelve cases of seroconversion in subsequent donations (seven human immunodeficiency virus, four hepatitis B and one hepatitis C) with proven human immunodeficiency virus infection after screening of only one recipient., Conclusion: The twelve cases of seroconversion in donors with subsequent infection proven in one recipient underscores the importance of this tool to increase transfusion safety.

The aim of this study was to characterize the physicochemical properties of bacterial cellulose (BC) membranes functionalized with osteogenic growth peptide (OGP) and its C-terminal pentapeptide OGP[10-14], and to evaluate in vitro osteoinductive potential in early osteogenesis, besides, to evaluate cytotoxic, genotoxic and/or mutagenic effects. Peptide incorporation into the BC membranes did not change the morphology of BC nanofibers and BC crystallinity pattern. The characterization was complemented by Raman scattering, swelling ratio and mechanical tests. In vitro assays demonstrated no cytotoxic, genotoxic or mutagenic effects for any of the studied BC membranes. Culture with osteogenic cells revealed no difference in cell morphology among all the membranes tested. Cell viability/proliferation, total protein content, alkaline phosphatase activity and mineralization assays indicated that BC-OGP membranes enabled the highest development of the osteoblastic phenotype in vitro. In conclusion, the negative results of cytotoxicity, genotoxicity and mutagenicity indicated that all the membranes can be employed for medical supplies, mainly in bone tissue engineering/regeneration, due to their osteoinductive properties.

The high consumption of fructose is linked to the increase in various characteristics of the metabolic syndrome. Fish oil is beneficial for the treatment of these comorbidities, such as insulin resistance, dyslipidemia, and hepatic steatosis. The objective of this study was to evaluate the consequences of the administration of fish oil concomitant to fructose ingestion during the experiment (45 days) and during the final 15 days in high-fructose-fed rats. Male Wistar rats were divided into 5 groups: control; those receiving 10% fish oil (FO); those receiving 60% fructose (Fr); those receiving 60% fructose and 10% fish oil for 45 days (FrFO); and those receiving fructose plus soybean oil for 30 days and fish oil for the final 15 days of the study (FrFO15). There was an increase in triacylglycerol, serum total cholesterol, and hepatic volume in the Fr group. The FO and FrFO groups experienced an increase in lipid peroxidation and a decrease in serum reduced glutathione. The FrFO group suffered greater hepatic injury, with increased alanine aminotransferase levels and DNA damage. Marked n-3 incorporation occurred in the groups receiving fish oil, favoring a better response to the oral glucose tolerance test. Fructose induced comorbidities of the metabolic syndrome, and the use of fish oil promoted a better glucose tolerance, although it was accompanied by more hepatocyte damage.

Parenteral injection of tolerated proteins into orally tolerant mice inhibits the initiation of immunological responses to unrelated proteins and blocks severe chronic inflammatory reactions of immunological origin, such as autoimmune reactions. This inhibitory effect which we have called "indirect effects of oral tolerance" is also known as "bystander suppression." Herein, we show that i.p. injection of OVA + Al(OH)(3) minutes before i.v. injection of Schistosoma mansoni eggs into OVA tolerant mice blocked the increase of pulmonary granulomas. In addition, the expression of ICAM-1 in lung parenchyma in areas outside the granulomas of OVA-orally tolerant mice was significantly reduced. However, at day 18 after granuloma induction there was no difference in immunofluorescency intensity to CD3, CD4, F4/80, andα-SMA per granuloma area of tolerant and control groups. Reduction of granulomas by reexposure to orally tolerated proteins was not correlated with a shift in Th-1/Th-2 cytokines in serum or lung tissue extract.

In the present study, we investigated the relationship between polymorphisms in the estrogen-metabolizing genes CYP17, CYP1B1, CYP1A1, and COMT and genomic instability in the peripheral blood lymphocytes of 62 BC patients and 62 controls considering that increased or prolonged exposure to estrogen can damage the DNA molecule and increase the genomic instability process in breast tissue. Our data demonstrated increased genomic instability in BC patients and that individuals with higher frequencies of MN exhibited higher risk to BC when belonging Val/Met genotype of the COMT gene. We also observed that CYP17 and CYP1A1 polymorphisms can modify the risk to BC depending on the menopause status. We can conclude that the genetic background in estrogen metabolism pathway can modulate chromosome damage in healthy controls and patients and thereby influence the risk to BC. These findings suggest the importance to ally biomarkers of susceptibility and effects to estimate risk groups.

ABH and Lewis antigen expression has been associated with cancer development and prognosis, tumor differentiation, and metastasis. Considering that invasive ductal breast carcinoma (IDC) presents multiple molecular alterations, the aim of the present study was to determine whether the polymorphism of ABO, Lewis, and Secretor genes, as well as ABO phenotyping, could be associated with tumor differentiation and lymph nodes metastasis. Seventy-six women with IDC and 78 healthy female blood donors were submitted to ABO phenotyping/genotyping and Lewis and Secretor genotyping. Phenotyping was performed by hemagglutination and genotyping by the polymerase chain reaction with sequence-specific primers. ABO, Lewis, and Secretor genes were classified by individual single nucleotide polymorphism at sites 59, 1067, 202, and 314 of the Lewis gene, 428 of the Secretor gene, and 261 (O1 allele), 526 (O2 and B allele), and 703 (B allele). No association was found between breast cancer and ABO antigen expression (P = 0.9323) or genotype (P = 0.9356). Lewis-negative genotype was associated with IDC (P = 0.0126) but not with anatomoclinical parameters. Nonsecretor genotype was associated with axillary lymph node metastasis (P = 0.0149). In conclusion, Lewis and Secretor genotyping could be useful to predict respectively breast cancer susceptibility and axillary lymph nodes metastasis.

A very appropriate method for antigenotoxicity evaluation of antioxidants is the comet assay, since this analytical method detects initial DNA lesions that are still subject to repair; in other words, lesions that are very associated to damages resulting from the generation and subsequent action of reactive species. However, a solid evaluation should be developed in order to avoid inexact interpretations. In our study, besides the association of curcumin with cisplatin, curcumin and cisplatin agents were also tested separately. Classical genotoxic compounds, when tested by the comet assay, present an increase in the nucleoid tail; however, the cisplatin treatment has resulted in a decrease of DNA migration. This was an expected effect, as the cross-links between cisplatin and DNA decrease the DNA electrophoretic mobility. A similar effect was observed with the curcumin treatment, which decreased the nucleoid tail. Such effect was not expected and reinforced the necessity of including in the study, separate treatment groups with potentially antigenotoxic substances. The comet assay results have been analyzed using specific software for image analysis, as well as the classical visual analysis, and we have observed that the effect of decrease in DNA electrophoretic mobility was more easily observed when the data were analyzed by the software.

Introduction: A retrospective study was conducted in order to assess the prevalence and factors associated with seropositivity for HTLV-1/2 between 1995 and 2008 in Uberaba Regional Blood Center, and to describe the seropositive blood donors in relation to gender, age, marital status, skin color and origin., Methods: Descriptive statistical analysis, chi-square tests and odds ratios were produced to compare proportions, along with scatter charts with linear correlation coefficients., Results: Among the donors tested, the prevalence of seropositivity for HTLV was found to be 0.02%, with indeterminate results in 0.09%. There was a significant reduction in seropositivity for HTLV between 2002 and 2008, compared with the period from 1995 to 2001. Among the seropositive individuals, females were significantly predominant., Conclusions: The gradual decrease in seropositivity over this period was attributed to the permanent exclusion of seropositive repeat donors and improvement in the clinical screening methods and serological tests over the years, with a positive impact on transfusion safety.

The objective of the present study was to evaluate the integrity of the peripheral and central auditory systems of sickle cell disease (SCD) patients, through electrophysiological evaluation utilizing auditory evoked potentials, and comparing the results obtained in SCD patients with individuals without SCD. A total of 80 individuals were evaluated: 40 SCD patients; and 40 healthy age- and sex-matched controls. Brainstem auditory evoked response (BAER) was used to check neural integrity and electrophysiological thresholds, and cognitive potential (P300) to analyse the auditory selective attention. Despite the exclusion of individuals with comorbidities typical of SCD, the predominance of hearing loss among the patients was detected in 16 ears (20%). The absolute latencies of the BAER were within the expected range but the SCD group showed a small but statistically significant reduction of the interpeaks I-V, indicative of cochlear alteration. P300 latency and amplitude were adequate for both groups suggesting the absence of central auditory system abnormalities. The present findings suggest that SCD causes variable degree of cochlear abnormalities without evidence of neural problems.

The present study evaluated the basal DNA damage and the cellular response to this damage induced by in vitro administration of Etoposide in lymphocytes donated by twenty untreated breast cancer (BC) patients and twenty age-matched healthy women. Micronucleus (MN) and alkaline Comet assays were performed in cultured peripheral blood lymphocytes (PBL) according to a standard protocol for in vitro treatment with various concentrations of Etoposide or a control. For the Comet Assay, three samples of cells were collected: T(0) (immediately preceding treatment of the cultures), T(1) (immediately after completion of the treatment) and T(2) (four hours after completion of the treatment). MN frequency in the BC group treated with 25 muM Etoposide (19.1 +/- 7.35) was significantly higher than the control (10.9 +/- 9.87) group. In the alkaline Comet Assay, both the BC group and the healthy women showed the ability to repair Etoposide-induced DNA damage within 4 hours of reincubation.