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Transitioning from a fossil-based to a bio-based economy is crucial to climate action and achieving neutrality in greenhouse gas (GHG) emissions. Biofuel production is an essential land-based GHG mitigation alternative. However, it raises concerns about biodiversity conservation, competition with food production, and net GHG emissions associated with direct land-use change (dLUC). This study aims to assess how the location and conversion routes influence GHG emissions for sugarcane expansion in Brazil to supply ethanol demand projections for 2030. A consistent and significant reduction in GHG emissions is achievable by implementing a strategy that prioritizes the spatial distribution for ethanol biorefinery expansions based on georeferenced life cycle emissions, including dLUC emissions associated with sugarcane production. Because of conservative zoning for sugarcane expansion, dLUC emissions are not an overriding factor, representing less than 9.1 % of the total GHG mitigation potential. Despite that, accounting for georeferenced dLUC emissions when prioritizing expansion facilities leads to spatial differences. Regarding conversion routes and land requirements, using cellulosic biorefineries could meet future projected demand based on sugarcane production from 3.1 million hectares, mostly in currently degraded pastureland. Conventional refineries would require 5.5 million hectares to meet the same demand of 71 billion liters. Despite the 77 % higher land demand to produce the same volume of ethanol, conventional refineries with straw recovery could be considered if electricity generation is a priority. This study illustrates how Brazil can achieve GHG mitigation targets while attending to future energy demand and protecting areas with high biodiversity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Alongside the concerns of waste management, plastic production represents a future problem for managing greenhouse gas emissions. Advanced recycling and bio-based production are paramount to face this challenge. The sustainability of bio-based polyethylene (bioPE) depends on the feedstock, avoiding stress on natural resources. This work discusses Brazil's potential to meet future global bioPE demand by 2050, using sugarcane as feedstock and considering environmental sustainability for production expansion. From the assessed 35.6 Mha, 3.55 Mha would be exempt from trade-offs related to land use change (dLUC), biodiversity, and water availability. The scenario with the highest circularity efficiency would require 22.2 Mha to meet the global demand, which can be accommodated in areas with positive impacts in carbon stocks, neutral impacts in water availability, and medium impacts on biodiversity. Here, we show that dropping demand is essential to avoid trade-offs and help consolidate bioPE as a sustainable alternative for future net-zero strategies., (© 2024. The Author(s).)

Primordial germ cells (PGCs) are embryonic pluripotent cells that can differentiate into spermatogonia and oogonia, and therefore, PGCs are a genetic source for germplasm conservation through cryobanking and the generation of germline chimeras. The knowledge of PGC migration routes is essential for transplantation studies. In this work, the mRNA synthesized from the ddx4 3'UTR sequence of Pseudopimelodus mangurus, in fusion with gfp or dsred, was microinjected into zygotes of three neotropical species (P. mangurus, Astyanax altiparanae, and Prochilodus lineatus) for PGC labeling. Visualization of labeled PGCs was achieved by fluorescence microscopy during embryonic development. In addition, ddx4 and dnd1 expressions were evaluated during embryonic development, larvae, and adult tissues of P. mangurus, to validate their use as a PGC marker. As a result, the effective identification of presumptive PGCs was obtained. DsRed-positive PGC of P. mangurus was observed in the hatching stage, GFP-positive PGC of A. altiparanae in the gastrula stage, and GFP-positive PGCs from P. lineatus were identified at the segmentation stage, with representative labeling percentages of 29% and 16% in A. altiparanae and P. lineatus, respectively. The expression of ddx4 and dnd1 of P. mangurus confirmed the specificity of these genes in germ cells. These results point to the functionality of the P. mangurus ddx4 3'UTR sequence as a PGC marker, demonstrating that PGC labeling was more efficient in A. altiparanae and P. lineatus. The procedures used to identify PGCs in P. mangurus consolidate the first step for generating germinal chimeras as a conservation action of P. mangurus., Competing Interests: Declarations Ethics approval Pseudopimelodus mangurus were collected from the Moji Iguaçu River (Pirassununga–SP, Brazil) according to Brazilian law (Sisbio #31836-10), and the experiments were conducted following the Animal Ethics Committee from the National Center for Research and Conservation of Continental Aquatic Biodiversity (CEUA/CEPTA #010/02031.000114/2022-31). Competing interests The authors declare no competing interests., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

Primordial germ cells (PGCs) constitute an important cell lineage that directly impacts genetic dissemination and species conservation through the creation of cryobanks. In order to advance the field of animal genetic cryopreservation, this work aimed to recover intact PGCs cryopreserved in embryonic tissues during the segmentation phase for subsequent in vitro maintenance, using the yellow-tailed tetra (Astyanax altiparanae) as a model organism. For this, a total of 202 embryos were distributed in two experiments. In the first experiment, embryos in the segmentation phase were dissociated, and isolated PGCs were maintained in vitro. They were visualized using gfp-Pm-ddx4 3'UTR labeling. The second experiment aimed to vitrify PGCs using 3 cryoprotective agents or CPAs (dimethyl sulfoxide, ethylene glycol, and 1,2 propanediol) at 3 molarities (2, 3, and 4 M). The toxicity, somatic cell viability, and recovery of intact PGCs were evaluated. After cryopreservation and thawing, 2 M ethylene glycol produced intact PGCs and somatic cells (44 ± 11.52 % and 42.35 ± 0.33 %, respectively) post-thaw. The recovery of PGCs from frozen embryonic tissues was not possible without the use of CPAs. Thus, the vitrification of PGCs from an important developmental model and Neotropical species such as A. altiparanae was achieved, and the process of isolating and maintaining PGCs in a culture medium was successful. Therefore, to ensure the maintenance of genetic diversity, PGCs obtained during embryonic development in the segmentation phase between 25 and 28 somites were stored through vitrification for future applications in the reconstitution of species through germinal chimerism., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 Society for Cryobiology. Published by Elsevier Inc. All rights reserved.)

Spermatogonia cryopreservation can be a strategy for future conservation actions. The neotropical Siluriformes Pseudopimelodus mangurus was already classified as vulnerable on the Red List of Threatened Species. P. mangurus spermatogonial cells were isolated, assessed, and cryopreserved. Fragments of the testis were enzymatically dissociated, purified using Percoll density gradient, and submitted to differential plating. Fractionated cells were evaluated by microscopy, ddx4 (vasa) relative expression, and alkaline phosphatase activity. Cryopreservation was conducted using ethylene glycol, glycerol, dimethyl sulfoxide (DMSO), dimethylacetamide (DMA), and propanediol at 1 M, 1.5 M, and 2 M. Cell viability was evaluated and cell concentration was determined. Cell fractions from 20 % and 30 % Percoll gradient bands showed the highest concentrations of spermatogonia. The fraction mix showed 54 % purity and 93 % viability. After differential plating, 60 % purity and 92 % viability were obtained. Spermatogonial cells showed high alkaline phosphatase activity compared to spermatocytes and spermatids. The relative spermatogonial ddx4 expression from the Percoll density gradient was about twice as high as in samples from the testis and the differential plating. The increased ddx4 expression indicated the enrichment of spermatogonial cells by density gradient step and dead cells expressing ddx4 in differential plating, or ddx4 decreasing expression during cell culture. For this reason, cells from the Percoll gradient were chosen for cryopreservation. Propanediol at 1 M demonstrated the best condition for spermatogonial cell cryopreservation, presenting 98 % viability, while dimethylacetamide at 2 M represented the least favorable condition, with approximately 47 % viability. These findings are essential for P. mangurus spermatogonial cell cryopreservation, aiming to generate a spermatogonia cryobank for future conservation efforts., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Society for Cryobiology. Published by Elsevier Inc. All rights reserved.)

l-asparaginases play a crucial role in the treatment of acute lymphoblastic leukemia (ALL), a type of cancer that mostly affects children and teenagers. However, it is common for these molecules to cause adverse reactions during treatment. These downsides ignite the search for novel asparaginases to mitigate these problems. Thus, this work aimed to produce and characterize a recombinant asparaginase from Phaseolus vulgaris (Asp-P). In this study, Asp-P was expressed in Escherichia coli with high yields and optimum activity at 40 °C, pH 9.0. The enzyme K m and V max values were 7.05 mM and 1027 U/mg, respectively. Asp-P is specific for l-asparagine, showing no activity against l-glutamine and other amino acids. The enzyme showed a higher cytotoxic effect against Raji than K562 cell lines, but only at high concentrations. In silico analysis indicated that Asp-P has lower immunogenicity than a commercial enzyme. Asp-P induced biofilm formation by Candida sp. due to sublethal dose, showing an underexplored potential of asparaginases. The absence of glutaminase activity, lower immunogenicity and optimal activity similar to physiological temperature conditions are characteristics that indicate Asp-P as a potential new commercial enzyme in the treatment of ALL and its underexplored application in the treatment of other diseases., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Bruno Anderson Matias da Rocha reports financial support was provided by National Council for Scientific and Technological Development. Bruno Anderson Matias da Rocha reports financial support was provided by Coordination of Higher Education Personnel Improvement. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: To evaluate the influence of previous physical activity (PA) during childhood, adolescence, and current PA practice on the production of antibodies and inflammatory response between the first and second doses of the COVID-19 vaccine., Methods: Fifty-nine men and 56 women were evaluated before the first vaccine, and 12 weeks later, blood samples were taken to quantify production of anti-severe acute respiratory syndrome coronavirus-2 immunoglobulin G antibodies and cytokines. Previous PA during childhood and adolescence was self-referred, and current PA was assessed using the International Physical Activity Questionnaire., Results: A positive and significant association was observed only between PA practice during adolescence and an increase in antibody production in adulthood (β = 2012.077, 95% confidence interval, 257.7953-3766.358, P = .025). Individuals who practiced PA during adolescence showed higher production of antibodies between the first and second vaccine dose compared to nonpractitioners (P = .025) and those that accumulated ≥150 minutes per week of current moderate-vigorous PA (MVPA), and presented higher antibody production in relation to who did <150 minutes per week of MVPA (P = .046). Individuals that were practitioners during childhood produced higher G-CSF (P = .047), and those that accumulated ≥150 minutes per week of current MVPA demonstrated lower IP-10 levels (P = .033). However, PA practitioners during adolescence presented higher G-CSF (P = .025), IL-17 (P = .038), IL-1RA (P = .005), IL-1β (P = .020), and IL-2 (P = .026) levels., Conclusion: Our results suggest that adults that accumulated at least 150 minutes of MVPA per week or practiced PA during adolescence developed an improved immune and inflammatory response against COVID-19 vaccination.

The search for innovative anti-cancer drugs remains a challenge. Over the past three decades, antibodies have emerged as an essential asset in successful cancer therapy. The major obstacle in developing anti-cancer antibodies is the need for non-immunogenic antibodies against human antigens. This unique requirement highlights a disadvantage to using traditional hybridoma technology and thus demands alternative approaches, such as humanizing murine monoclonal antibodies. To overcome these hurdles, human monoclonal antibodies can be obtained directly from Phage Display libraries, a groundbreaking tool for antibody selection. These libraries consist of genetically engineered viruses, or phages, which can exhibit antibody fragments, such as scFv or Fab on their capsid. This innovation allows the in vitro selection of novel molecules directed towards cancer antigens. As foreseen when Phage Display was first described, nowadays, several Phage Display-derived antibodies have entered clinical settings or are undergoing clinical evaluation. This comprehensive review unveils the remarkable progress in this field and the possibilities of using clever strategies for phage selection and tailoring the refinement of antibodies aimed at increasingly specific targets. Moreover, the use of selected antibodies in cutting-edge formats is discussed, such as CAR (chimeric antigen receptor) in CAR T-cell therapy or ADC (antibody drug conjugate), amplifying the spectrum of potential therapeutic avenues.

Background: To evaluate oral conditions, demographic and socioeconomic characteristics of oral health-related quality of life (OHRQoL) in adolescents., Methods: Umbrella review, conducted according to the Preferred Reporting Items for Systematic Reviews and Meta Analyzes (PRISMA) checklist. The search strategy used a combination of words, applied in the electronic databases PubMed, WebScience, Embase, Lilacs, Scopus and Cochrane. Included publications until January 2022, without restrictions. Data collection took place with systematized practices and the eligibility criteria were studies focusing on OHRQoL; teenagers; adolescentes; present the term "systematic review" and/or "meta-analysis" in the title or abstract. The quality assessment followed the Assessment of Multiple Systematic Reviews (AMSTAR 2) and the adherence of the article to the PRISMA was verified., Results: Three hundred sixty-two articles were identified, and 22 were included, published between 2009 and 2022. 21 Systematic reviews focused on the English language. Most studies showed heterogeneity in the methodological structuring process: 10 articles were considered of low and 10 critically low quality. Clinical conditions associated with worsening in quality of life were dental caries, malocclusion, dental trauma, toothache, edentulism, need for orthodontic treatment, irregular brushing, and periodontal disease. Socioeconomic factors related to housing, parental education, access to health care, absence of siblings and nuclear family influence OHRQoL. Completion of orthodontic treatment, health promotion programs, dental care and safe housing all have a positive impact., Conclusion: Worse oral health status, older age, female sex and worse socioeconomic status were significantly associated with worse OHRQoL., Trial Registration: PROSPERO CRD4202129352., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Disruption of the epigenetic program of gene expression is a hallmark of cancer that initiates and propagates tumorigenesis. Altered DNA methylation, histone modifications and ncRNAs expression are a feature of cancer cells. The dynamic epigenetic changes during oncogenic transformation are related to tumor heterogeneity, unlimited self-renewal and multi-lineage differentiation. This stem cell-like state or the aberrant reprogramming of cancer stem cells is the major challenge in treatment and drug resistance. Given the reversible nature of epigenetic modifications, the ability to restore the cancer epigenome through the inhibition of the epigenetic modifiers is a promising therapy for cancer treatment, either as a monotherapy or in combination with other anticancer therapies, including immunotherapies. Herein, we highlighted the main epigenetic alterations, their potential as a biomarker for early diagnosis and the epigenetic therapies approved for cancer treatment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Costa, Sales, Pinheiro, Pontes, Maranhão, Pessoa, Furtado and Furtado.)

Objective.: Motivation for the study: it is important to assess the level of knowledge about healthy eating in university students as this allows raising awareness about the practice and maintenance of healthy eating habits. Main findings: most university students from the nine health careers had insufficient knowledge about healthy eating. The highest proportion of students with sufficient knowledge were found in the career of nutrition. Implications: there is a need for projects at the university level that encompass the triad of psychology, food, and body; which would improve healthy eating habits in university students. To assess the healthy eating (HE) knowledge of health students and the factors associated with the university environment., Materials and Methods.: This was a cross-sectional study of 512 university students (≥ 18 years) enrolled in nine undergraduate health careers. It was conducted from April to November 2017. The Instrument for Assessment of Health Promotion in Universities and the International Physical Activity Questionnaire were used. In addition, we measured weight, height and waist circumference. Bivariate and multivariate analyses were carried out with SPSS version 23.0., Results.: We found that most university students from the nine health careers had insufficient knowledge about healthy eating (71.9%; n=368). However, the highest proportion of students with sufficient knowledge was found in the career of nutrition (15.3%; n=22), followed by those in the physical education career (12.5%; n=18). The lowest percentage of students with sufficient knowledge was found in the career of medicine (8.3%; n=12). Multivariate analysis showed that sufficient knowledge about healthy eating was related to participation in healthy eating activities (p=0.012; PR=1.94), participation in activities addressing self-esteem and self-knowledge (p=0.046; PR=0.59) and being overweight (p=0.036; PR=1.53)., Conclusion.: A low percentage of health students had sufficient knowledge about healthy eating. However, participation in healthy eating, self-esteem and self-knowledge activities at the university managed to improve the level of knowledge. We recommend the development of university projects that include the psychological, food and body triad, thus involving all health careers, with the aim of improving the health and quality of life of university students.

Primordial germ cells (PGCs) are responsible for generating all germ cells. Therefore, they are essential targets to be used as a tool for the production of germline chimeras. The labeling and route of PGCs were evaluated during the initial embryonic development of Pseudopimelodus mangurus, using whole-mount in situ hybridization (WISH) and mRNA microinjection in zygotes. A specific antisense RNA probe constituted by a partial coding region from P. mangurus nanos3 mRNA was synthesized for the WISH method. RNA microinjection was performed using the GFP gene reporter regulated by translation regulatory P. mangurus buc and nanos3 3'UTR sequences, germline-specific markers used to describe in vivo migration of PGCs. Nanos3 and buc gene expression was evaluated in tissues for male and female adults and initial development phases and larvae from the first to seventh days post-hatching. The results from the WISH technique indicated the origin of PGCs in P. mangurus from the aggregations of nanos3 mRNA in the cleavage grooves and the signals obtained from nanos3 probes corresponded topographically to the migratory patterns of the PGCs reported for other fish species. Diffuse signals were observed in all blastomeres until the 16-cell stage, which could be related to the two sequences of the nanos3 3'UTR observed in the P. mangurus unfertilized egg transcriptome. Microinjection was not successful using GFP-Dr-nanos1 3'UTR mRNA and GFP-Pm-buc 3'UTR mRNA and allowed the identification of potential PGCs with less than 2% efficiency only and after hatching using GFP-Pm-nanos3 3'UTR. Nanos3 and buc gene expression was reported in the female gonads and from fertilized eggs until the blastula phase. These results provide information about the PGC migration of P. mangurus and the possible use of PGCs for the future generation of germline chimeras to be applied in the conservation efforts of Neotropical Siluriformes species. This study can contribute to establishing genetic banks, manipulating organisms, and assisting in biotechnologies such as transplanting germ cells in fish., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Here we show that selected nutritional feed additives reduce the adverse effects of transport stress on the immune system and hematology of tambaqui. We formulated a control diet to contain normal dietary levels of vitamin E (21.6 mg/ kg diet) and C (143 mg/kg), then we added supra levels of these vitamins (vit E - 264 mg/kg and vit C - 1000 mg/kg) to a second diet. Finally, a third diet was produced to contain similar levels of vitamins from diet 2 with 0.1% beta-glucan supplementation. Four hundred thirty-two tambaquis (20.91  g  ± 0.27 g) were randomly assigned to 12 aquaria and fed the diets for 15 days; then, all fish were transported for five h and then returned to the aquaria. Blood samples were collected before and after the transport and at the end of the trial (60 days). Transportation significantly increased blood glucose that returned to baseline levels at the end of the trial. However, cortisol seemed to be unresponsive to the stress. Surprisingly, the stress significantly increased the immunoglobulin level after transport. Additionally, the transport markedly reduced the red blood cell count and leukocyte and lymphocytes counts while increasing the control group's neutrophil number. These effects lasted until the end of the trial in the control group. Supra levels of the vitamins and glucan supplementation prevented the decrease in red blood cell and leukocyte count after the stress. Additionally, beta-glucan supplementation induced lower cortisol levels in all the sampling points. However, the effect on the immune parameters was limited, increasing only the lysozyme activity and serum protein levels in the beta-glucan supplemented group and the group fed only the supra levels of vitamins, respectively. In sum, our results indicated that transport for five h induced a limited effect on stress biomarkers. The use of supra levels of antioxidant vitamins alone or in combination with beta-glucan could restore or prevent the adverse effects of stress on hematology and the immune system., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Authors.)

The β-d-glucans are abundant cell wall polysaccharides in many cereals and contain both (1,3)- and (1,4)-bonds. The β-1,3-1,4-glucanases (EC 3.2.1.73) hydrolyze β-(1,4)-d-glucosidic linkages in glucans, and have applications in both animal and human food industries. A chimera between the family 11 carbohydrate-binding module from Ruminoclostridium (Clostridium)thermocellumcelH (RtCBM11), with the β-1,3-1,4-glucanase from Bacillus subtilis (BglS) was constructed by end-to-end fusion (RtCBM11-BglS) to evaluate the effects on the catalytic function and its application in barley β-glucan degradation for the brewing industry. The parental and chimeric BglS presented the same optimum pH (6.0) and temperature (50 °C) for maximum activity. The RtCBM11-BglS showed increased thermal stability and 30% higher hydrolytic efficiency against purified barley β-glucan, and the rate of hydrolysis of β-1,3-1,4-glucan in crude barley extracts was significantly increased. The enhanced catalytic performance of the RtCBM11-BglS may be useful for the treatment of crude barley extracts in the brewing industry., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The imprinted H19 long non-coding RNA, a knowing oncofetal gene, presents a controversial role during the carcinogenesis process since its tumor suppressor or oncogenic activity is not completely elucidated. Since H19 lncRNA is involved in many biological pathways related to tumorigenesis, we sought to develop a non-cancer lineage with CRISPR-Cas9-mediated H19 knockdown (H19-) and observe the changes in a cellular context. To edit the promoter region of H19, two RNA guides were designed, and the murine C2C12 myoblast cells were transfected. H19 deletion was determined by DNA sequencing and gene expression by qPCR. We observed a small deletion (~ 60 bp) in the promoter region that presented four predicted transcription binding sites. The deletion reduced H19 expression (30%) and resulted in increased proliferative activity, altered morphological patterns including cell size and intracellular granularity, without changes in viability. The increased proliferation rate in the H19- cell seems to facilitate chromosomal abnormalities. The H19- myoblast presented characteristics similar to cancer cells, therefore the H19 lncRNA may be an important gene during the initiation of the tumorigenic process. Due to CRISPR/Cas9 permanent edition, the C2C12 H19- knockdown cells allows functional studies of H19 roles in tumorigenesis, prognosis, metastases, as well as drug resistance and targeted therapy., (© 2021. The Author(s).)

Given the magnitude of the global snakebite crisis, strategies to ensure the quality of antivenom, as well as the availability and sustainability of its supply are under development by several research groups. Recombinant DNA technology has allowed the engineering of monoclonal antibodies and recombinant fragments as alternatives to conventional antivenoms. Besides having higher therapeutic efficacy, with broad neutralization capacity against local and systemic toxicity, novel antivenoms need to be safe and cost-effective. Due to the biological and physical chemical properties of camelid single-domain antibodies, with high volume of distribution to distal tissue, their modular format, and their versatility, their biotechnological application has grown considerably in recent decades. This article presents the most up-to-date developments concerning camelid single-domain-based antibodies against major toxins from snake venoms, the main venomous animals responsible for reported envenoming cases and related human deaths. A brief discussion on the composition, challenges, and perspectives of antivenoms is presented, as well as the road ahead for next-generation antivenoms based on single-domain antibodies., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Objective: L-Asparaginase (ASNase) is an enzyme used in the treatment of acute lymphoblastic leukemia (ALL). As the therapeutic ASNases has bacterial origin, severe side effects are associated with its use, among them hypersensitivity and inactivation of the enzyme. In this context, the objective of this work was to produce a recombinant ASNase of bacterial origin in human cells in order to determine the presence and consequences of potential post-translational modifications on the enzyme., Results: Recombinant ASNase was expressed in human cells with a molecular weight of 60 kDa, larger than in Escherichia coli, which is 35 kDa. N-glycosylation analysis demonstrated that the increased molecular weight resulted from the addition of glycans to the protein by mammalian cells. The glycosylated ASNase presented in vitro activity at physiological pH and temperature. Given that glycosylation can act to reduce antigenicity by masking protein epitopes, our data may contribute to the development of an alternative ASNase in the treatment of ALL in patients who demonstrate side effects to currently marketed enzymes.

Growing evidence suggests that aberrant epigenetic regulation of gene function is strongly related to the genesis of cancer. Unlike genetic mutations, the ability to reprogram the epigenetic landscape in the cancer epigenome is one of the most promising target therapies in both treatment and reversibility of drug resistance. Epigenetic alterations in cancer development and progression may be the basis for the individual variation in drug response. Thus, this review focuses on the emerging area of pharmaco(epi)genomics, specifically highlighting epigenetic reprogramming during tumorigenesis and how epigenetic markers are targeted as a therapy (epidrugs) and the clinical implications of this for cancer treatment.

Changes involving temporomandibular joint, masticatory musculature, and associated structures characterize temporomandibular dysfunction (TMD). The analgesic and anti-inflammatory effect produced by photobiomodulation has contributed to pain relief and functional improvement. However, the parameters to be used have not yet been well established. The aim of this study is to compare the efficacy of three different photobiomodulation dosimetries in the treatment of patients with TMD. A randomized, double-blind, placebo-controlled clinical trial with 44 subjects divided into the groups 8 J/cm 2 (n = 11), 60 J/cm 2 (n = 11), 105 J/cm 2 (n = 11), and control (n = 11). Pain, symptom severity, and joint mobility were evaluated before and after a ten-session protocol of photobiomodulation with AlGaAs laser (830 nm), at a power density of 30 mW/cm 2 . The mouth opening increased in the 8-J/cm 2 group from 10.49 ± 4.68 to 15.40 ± 6.43 degrees, and in the right protrusion from 9.80 ± 4.2 to 12.56 ± 5.40 degrees after the intervention protocol (p < 0.05). All groups significantly decreased pain (p < 0.05). 830-nm laser photobiomodulation was effective in reducing TMD pain and symptoms at all doses tested. Only the doses of 8 J/cm 2 were effective regarding maximal opening and protrusion of the mandible.

Carbohydrate binding modules (CBMs) are non-catalytic domains within larger multidomain polypeptides. The CelH from Ruminoclostridium (Clostridium) thermocellum contains a family 11 CBM (RtCBM11) with high binding affinity for the linear polysaccharide β-glucan, and low affinity for the branched xyloglucan. Screening a random RtCBM11 mutant phage library created by error prone PCR for xyloglucan binding identified RtCBM11 mutants with enhanced xyloglucan affinity. Subsequent recombination of the selected variants by site-directed mutagenesis generated the H102L/Y152F and Y46N/G52D/H102L/Y152F mutants. Fusion of the quadruple RtCBM11 mutant with the xyloglucanase from Aspergillus niveus increased the catalytic efficiency of the enzyme by 38%. Isothermal titration calorimetry demonstrated increased xyloglucan affinity for both mutants and reduced affinity for β-glucan in the H102L/Y152F mutant. Molecular dynamics simulations indicated that the increased xyloglucan specificity results both from formation of a xylosyl binding pocket in the carbohydrate binding cleft, and via modulation of a hydrogen bond network between the oligosaccharide ligand and the protein. These results explain the improved xyloglucan binding in the RtCBM11 H102L/Y152F mutant and advance the understanding of the structural determinants of CBMs binding that discriminate between branched and unbranched polysaccharides., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Due mainly to their high level of affinity and specificity, therapeutic monoclonal antibodies (mAbs) have been frequently selected as treatment for cancer, autoimmune or chronic inflammatory diseases. Despite the increasing number of mAbs and related products in the biopharmaceutical market, they are still expensive, can cause undesired side effects, and eventually cause resistance. Antibody engineering, which emerged to overcome limitations faced by mAb therapy, has supported the development of modified mAbs for immunotherapy. As part of this approach, researchers have invested in obtaining antibody fragments, as well as in Fc region modifications, since interactions with Fc receptors influence an antibody's half-life and mechanism of action. Thus, Fc engineering results in antibodies with more desirable characteristics and functions for which they are intended, creating "fit-for-purpose" antibodies with reduced side effects. Furthermore, aglycosylated antibodies, produced in bacterial cultivation, have been an alternative to create new effector functional human immunotherapeutics, while reducing mAb therapy costs. This review highlights some features that enhance mAb performance, related to the improvement of antibody half-life and effector responses by both Fc-engineering and glycoengineering., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Artocarpus incisa (breadfruit) seeds contain three different lectins (Frutalin, Frutapin (FTP) and Frutackin) with distinct carbohydrate specificities. The most abundant lectin is Frutalin, an α-D-galactose-specific carbohydrate-binding glycoprotein with antitumour properties and potential for tumour biomarker discovery as already reported. FTP is the second most abundant, but proved difficult to purify with very low yields and contamination with Frutalin frustrating its characterization. Here, we report for the first time high-level production and isolation of biologically active recombinant FTP in Escherichia coli BL21, optimizing conditions with the best set yielding >40 mg/l culture of soluble active FTP. The minimal concentration for agglutination of red blood cells was 62.5 µg/ml of FTP, a process effectively inhibited by mannose. Apo-FTP, FTP-mannose and FTP-glucose crystals were obtained, and they diffracted X-rays to a resolution of 1.58 (P2 1 2 1 2 1 ), 1.70 (P3 1 21) and 1.60 (P3 1 21) Å respectively. The best solution showed four monomers per asymmetric unit. Molecular dynamics (MD) simulation suggested that FTP displays higher affinity for mannose than glucose. Cell studies revealed that FTP was non-cytotoxic to cultured mouse fibroblast 3T3 cells below 0.5 mg/ml and was also capable of stimulating cell migration at 50 µg/ml. In conclusion, our optimized expression system allowed high amounts of correctly folded soluble FTP to be isolated. This recombinant bioactive lectin will now be tested in future studies for therapeutic potential; for example in wound healing and tissue regeneration., (© 2017 The Author(s).)

Background: Product inhibition can reduce catalytic performance of enzymes used for biofuel production. Different mechanisms can cause this inhibition and, in most cases, the use of classical enzymology approach is not sufficient to overcome this problem. Here we have used a semi-rational protein fusion strategy to create a product-stimulated enzyme., Results: A semi-rational protein fusion strategy was used to create a protein fusion library where the Bacillus subtilis GH11 xylanase A (XynA) was inserted at 144 surface positions of the Escherichia coli xylose binding protein (XBP). Two XynA insertions at XBP positions 209 ([209]XBP-Xyn-XBP) and 262 ([262]XBP-Xyn-XBP) showed a 20% increased xylanolytic activity in the presence of xylose, conditions where native XynA is inhibited. Random linkers of 1-4 Gly/Ala residues were inserted at the XynA N- and C-termini in the [209]XBP and [262]XBP, and the chimeras 2091A and 2621B were isolated, showing a twofold increased xylanolytic activity in the presence of xylose and k cat values of 200 and 240 s(-1) in the 2091A and 2621B, respectively, as compared to 70 s(-1) in the native XynA. The xylose affinity of the XBP was unchanged in the chimeras, showing that the ~3- to 3.5-fold stimulation of catalytic efficiency by xylose was the result of allosteric coupling between the XBP and XynA domains. Molecular dynamics simulations of the chimeras suggested conformation alterations in the XynA on xylose binding to the XBP resulted in exposure of the catalytic cavity and increased mobility of catalytic site residues as compared to the native XynA., Conclusions: These results are the first report of engineered glycosyl hydrolase showing allosteric product stimulation and suggest that the strategy may be more widely employed to overcome enzyme product inhibition and to improve catalytic performance. Graphical abstractProtein fusion of a GH11 xylanase (in red) and a xylose binding protein (XBP, in blue) results in a xylanase-XBP chimera that presents allosteric activation of the xylanase activity by xylose (shown as a space-filled molecule bound to the xylanase-XBP chimera).

Our ability to genetically manipulate living organisms is usually constrained by the efficiency of the genetic tools available for the system of interest. In this report, we present the design, construction and characterization of a set of four new modular vectors, the pHsal series, for engineering Halobacterium salinarum, a model halophilic archaeon widely used in systems biology studies. The pHsal shuttle vectors are organized in four modules: (i) the E. coli's specific part, containing a ColE1 origin of replication and an ampicillin resistance marker, (ii) the resistance marker and (iii) the replication origin, which are specific to H. salinarum and (iv) the cargo, which will carry a sequence of interest cloned in a multiple cloning site, flanked by universal M13 primers. Each module was constructed using only minimal functional elements that were sequence edited to eliminate redundant restriction sites useful for cloning. This optimization process allowed the construction of vectors with reduced sizes compared to currently available platforms and expanded multiple cloning sites. Additionally, the strong constitutive promoter of the fer2 gene was sequence optimized and incorporated into the platform to allow high-level expression of heterologous genes in H. salinarum. The system also includes a new minimal suicide vector for the generation of knockouts and/or the incorporation of chromosomal tags, as well as a vector for promoter probing using a GFP gene as reporter. This new set of optimized vectors should strongly facilitate the engineering of H. salinarum and similar strategies could be implemented for other archaea.

In this work, we present recent advances in the use of phage display technology for the preparation of antivenoms for animal toxin neutralization. Even though classical antivenoms have been used since the early 20th century, envenomation remains a global public health problem. Recently, the phage display technique has been used in an attempt to circumvent some of the difficulties associated with traditional preparations of antivenom. Here, we review studies that developed antibody fragments with potential inhibitory effects against animal toxins and discuss the most current technical issues and perspectives regarding phage display technology in this field., (Copyright © 2014. Published by Elsevier Ltd.)

This paper evaluated the occurrence and removal efficiency of four estrogenic hormones in five biological wastewater treatment plants (WWTPs), located in the State of Ceará, Brazil. The five WWTPs comprised: two systems consisted of one facultative pond followed by two maturation ponds, one facultative pond, one activated sludge (AS) system followed by a chlorination step, and one upflow anaerobic sludge blanket (UASB) reactor followed by a chlorination step. Estrogen occurrence showed a wide variation among the analyzed influent and effluent samples. Estrone (E1) showed the highest occurrence in the influent (76%), whereas both 17β-estradiol (E2) and 17α-ethynylestradiol (EE2) presented a 52% occurrence, and the compound 17β-estradiol 17-acetate (E2-17A), a 32% one. The occurrence in the effluent samples was 48% for E1, 28% for E2, 12% for E2-17A, and 40% for EE2. The highest concentrations of E1 and EE2 hormones in the influent were 3050 and 3180 ng L(-1), respectively, whereas E2 and E2-17A had maximum concentrations of 776 and 2300 ng L(-1), respectively. The lowest efficiencies for the removal of estrogenic hormones were found in WWTP consisted of waste stabilization ponds, ranging from 54 to 79.9%. The high-rate systems (AS and UASB), which have chlorination as post-treatment, presented removal efficiencies of approximately 95%., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The prime objective in the surgical treatment of basilar impression (BI), Chiari malformation (CM) and/or syringomyelia (SM) is based on the restoration of the normal cerebrospinal fluid (CSF) dynamics at the craniovertebral junction through the creation of a large artificial cisterna magna. A small suboccipital craniectomy has been emphasized to avoid caudal migration of the hindbrain structures into the vertebral canal. Nevertheless, the results showed downward migration of the hindbrain related to that type of craniectomy. The authors present, otherwise, the results of 104 cases of BI, CM and/or SM, whose surgical treatment was characterized by a large craniectomy with the patient in the sitting position, tonsillectomy, large opening of the fourth ventricle and duraplasty with creation of a large artificial cisterna magna. A significant upward migration of the posterior fossa structures was detected by postoperative magnetic resonance imaging.