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Abstract: Objective: Protein–energy malnutrition (PEM) is an important public health problem affecting millions of people worldwide. Hematopoietic tissue requires a high nutrient supply, and a reduction in leukocytes, especially lymphocytes, suggests that some nutritional deficiencies might be altering bone marrow function and decreasing its ability to produce lymphocytes. In this study, we evaluated the effect that PEM has on lymphocyte subtypes and the cell cycle of CD5+ cells. Methods: Swiss mice were subjected to PEM using a low-protein diet containing 4% protein. When the experimental group had lost about 20% of their original body weight, we collected blood and bone marrow cells and evaluated the hemogram, the myelogram, bone marrow lymphoid markers using flow cytometry, and the cell cycle in CD5+ bone marrow. Results: Malnourished animals presented anemia, reticulocytopenia, and leukopenia with lymphopenia. The bone marrow was hypocellular, and flow cytometric analyses of bone marrow cells showed cells that were CD45+ (91.2%), CD2+ (84.9%), CD5+ (37.3%), CD3+ (23.5%), CD19+ (43.3%), CD22+ (34.7%), CD19+/CD2+ (51.2%), CD19+/CD3+ (24.0%), CD19+/CD5+ (13.2%), CD22+/CD2+ (40.1%), CD22+/CD3+ (30.3%), and CD22+/CD5+ (1.1%) in malnourished animals and CD45+ (97.5%), CD2+ (42.9%), CD5+ (91.5%), CD3+ (92.0%), CD19+ (52.0%), CD22+ (75.6%), CD19+/CD2+ (62.0%), CD19+/CD3+ (55.4%), CD19+/CD5+ (6.7%), CD22+/CD2+ (70.3%), CD22+/CD3+ (55.9%), and CD22+/CD5+ (8.4%) in control animals. Malnourished animals also presented more CD5+ cells in the G0 phase of cell cycle development. Conclusion: Malnourished animals presented bone marrow hypoplasia, maturation interruption, prominent lymphopenia with depletion in the lymphoid lineage, and changes in cellular development. We suggest that these changes are some of the primary causes of lymphopenia in cases of PEM and partly explain the increase in susceptibility to infections found in malnourished individuals. [ABSTRACT FROM AUTHOR]

Malaria continues to afflict hundreds of millions of lives annually, causing substantial fatalities despite available vaccines endorsed by the World Health Organization (WHO). However, these vaccines lack efficacy against Plasmodium vivax (Pv). Concomitantly, a considerable part of residents from several Pv-endemic areas face malnutrition, compromising their immunity to diseases, including malaria. Since our group developed an immunogenic yeast-expressing recombinant Pv circumsporozoite protein (yPvCSP-All CT epitopes) capable of protecting mice against lethal transgenic parasites, we investigated the influence of malnutrition on vaccine-derived responses in C57Bl/6 mice. Animals subjected to a protein-restricted diet presented protein-energetic malnutrition, diminished vaccine-specific IgG-secreting cells in the bone marrow, and reduced IgG and IgG1 serum titers compared to mice under a control diet. IgM titers remained consistent across groups, suggesting that the nutrition status may influence the antibody affinity maturation. These findings emphasize the pivotal role of proper nutrition in enhancing vaccination immunity against Pv malaria., Competing Interests: Competing interests ISS is co-inventor of the potential P. vivax vaccine evaluated in this study. The patent is under evaluation process, application number BR102022005915-2. The remaining authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

There has been a global increase in the older population in recent decades and, as age advances, complex metabolic and epigenetic changes occur in the organism, and these may trigger some health complications commonly found among this population. Additionally, several changes occur in older people that can reduce the dietary intake or the process of nutrient absorption. In this way, tissues with high nutrient requirements are more affected. Hematopoiesis is the process of formation, development, and maturation of blood cells and is a process with a high turnover. This high demand makes the integrity of the hematopoietic process susceptible to various factors that impair physiological function, such as aging and micronutrient bioavailability. Among these micronutrients, Zinc is considered an important micronutrient, playing diverse roles across various tissues and cell types. Some of the alterations in hematopoiesis that appear as a consequence of aging and due to insufficient micronutrient intake are well described in the literature; however, not much is known about how zinc deficiency contributes towards the development of diseases seen in aging. Considering the importance of zinc to act on several biological processes, this narrative review discusses several studies related to the physiological requirements, deficiency, or excess of zinc, including studies in experimental models and humans, and aimed to shed light on the relationship between zinc and the regulation of hematopoietic tissue, exploring possible links between this mineral with common disorders that appear during aging., (© The Author(s) 2023. Published by Oxford University Press on behalf of the International Life Sciences Institute. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Hypertriglyceridemia (HTG) is a common dyslipidemia associated with an increased risk of cardiovascular disease and pancreatitis. It is well stablished that the severe cases of disease often present with an underlying genetic cause. In this study, we determined the frequency and variation spectrum of genes involved in the triglyceride metabolism in a series of Brazilian patients with severe HTG. A total of 212 patients with very high HTG, defined with fasting triglycerides (TG) ≥ 880 mg/ dL, that underwent a multi-gene panel testing were included in this research. Germline deleterious variants (i.e. Pathogenic/Likely Pathogenic (P/LP) variants) were identified in 28 out of 212 patients, reflecting an overall diagnostic yield of 13% in our cohort. Variants of unknown significance (VUS) were identified in 87 patients, and represent 80% of detected variants in this dataset. We confirm the LPL as the most frequently mutated gene in patients with severe HTG, and we had only one suspected case of familial chylomicronemia syndrome, caused by a homozygous variant in LMF1, in our cohort. Notably, we report 16 distinct and novel variants (P/LP and VUS), each of them representing a single case, not previously reported in any public databases or other studies. Our data expand our knowledge of genetic variation spectrum in patients with severe HTG in the Brazilian population, often underrepresented in public genomic databases, being also a valuable clinical resource for genetic counseling and healthcare programs in the country., Competing Interests: M.H.C.G. reported that she received payment from PTC Therapeutics, Brasil for teaching activities. All other authors report no conflict of interest relevant to this article., (© 2024 The Authors. Published by Elsevier Inc.)

The bone marrow is a haven for hematopoietic and non-hematopoietic cells, creating complex micro-anatomical regions called niches. These distinct niches all participate in an intricate orchestra of cellular interactions that regulates the hematopoietic stem cell and its progenies. In this review, we provide a detailed description of the three most well-known bone marrow niches and their participation in hematopoiesis. We use pre-clinical data, including different in vitro and in vivo studies to discuss how a group of proteins called Semaphorins could potentially modulate both hematopoietic and non-hematopoietic cells, establishing links between the niches, semaphorins, and hematopoietic regulation. Thus, here we provide a deep dive into the inner functioning of the bone marrow and discuss the overarching implications that semaphorins might have on blood formation., 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 Ltd. All rights reserved.)

Overweight and obesity are typical conditions of chronic low-intensity systemic inflammatory responses, and both have become more common in recent decades, which emphasizes the necessity for healthier diet intake. Fruits such as grapes are rich in anthocyanins, one of which is delphinidin, a promising chemopreventive agent with anti-inflammatory properties. Considering that polymorphonuclear cells (PMNs) are rapidly mobilized to tissues when the inflammatory process is initiated, this study aimed to understand the impact of grape juice intake and delphinidin on the migration properties of PMNs. Overweight women ingested 500 mL of grape juice for 28 days, and then lipid and inflammatory profiles, as well as the white blood cell count (WBC), were evaluated. Additionally, the gene expression of inflammatory markers and quantified migration molecules such as CD11/CD18, ICAM-1 and VCAM-1 were evaluated in PMNs. The influence of delphinidin-3-O-glucoside in vitro on some migration properties was also evaluated. Grape juice intake did not influence the lipid profile or affect the WBC. However, NFκB gene expression was reduced in PMNs, also reducing the circulating values of IL-8, sICAM-1, and sVCAM-1. The in vitro results demonstrated that delphinidin significantly reduced the migration potential of cells and reduced CD11-/CD18-positive cells, the gene expression of ICAM-1, and the phosphorylation and gene expression of NFκB. Additionally, delphinidin also reduced the production of IL-6, IL-8, and CCL2. Grape juice, after 28 days of intervention, influenced some properties related to cell migration, and delphinidin in vitro can modify the cell migration properties., 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 © 2023 Elsevier Ltd. All rights reserved.)

Protein malnourishment (PM) is common among the elderly, but how aging and PM impact hematopoiesis is not fully understood. This study aimed to assess how aging and PM affect the hematopoietic regulatory function of bone marrow (BM) mesenchymal stem cells (MSCs). Young and aged male C57BL/6J mice were fed with normoproteic or hypoproteic diets and had their nutritional, biochemical, and hematological parameters evaluated. BM MSCs were characterized and had their secretome, gene expression, autophagy, reactive oxygen species production (ROS), and DNA double-stranded breaks evaluated. The modulation of hematopoiesis by MSCs was assayed using in vitro and in vivo models. Lastly, BM invasiveness and mice survival were evaluated after being challenged with leukemic cells of the C1498 cell line. Aging and PM alter biochemical parameters, changing the peripheral blood and BM immunophenotype. MSC autophagy was affected by aging and the frequencies for ROS and DNA double-stranded breaks. Regarding the MSCs' secretome, PM and aging affected CXCL12, IL-6, and IL-11 production. Aging and PM up-regulated Akt1 and PPAR-γ while down-regulating Cdh2 and Angpt-1 in MSCs. Aged MSCs increased C1498 cell proliferation while reducing their colony-forming potential. PM and aging lowered mice survival, and malnourishment accumulated C1498 cells at the BM. Finally, aged and/or PM MSCs up-regulated Sox2, Nanog, Pou5f1, and Akt1 expression while down-regulating Cdkn1a in C1498 cells. Together, aging and PM can induce cell-intrinsic shifts in BM MSCs, creating an environment that alters the regulation of hematopoietic populations and favoring the development of malignant cells., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Branched-chain amino acids (BCAA) are essential for maintaining intestinal mucosal integrity. However, only a few studies have explored the role of BCAA in the modulation of intestinal inflammation. In this study, we investigated in vitro effects of BCAA on the inflammatory response induced by lipopolysaccharide (LPS) (1 µg/mL) in Caco-2 cells. Caco-2 cells were assigned to six groups: control without BCAA (CTL0), normal BCAA (CTL; 0.8 mM leucine, 0.8 mM isoleucine, and 0.8 mM valine); leucine (LEU; 2 mM leucine), isoleucine (ISO; 2 mM isoleucine), valine (VAL; 2 mM valine), and high BCAA (LIV; 2 mM leucine, 2 mM isoleucine, and 2 mM valine). BCAA was added to the culture medium 24 h before LPS stimulation. Our results indicated that BCAA supplementation did not impair cell viability. The amino acids leucine and isoleucine attenuated the synthesis of IL-8 and JNK and NF-kB phosphorylation induced by LPS. Furthermore, neither BCAA supplementation nor LPS treatment modulated the activity of glutathione peroxidase or the intracellular reduced glutathione/oxidized glutathione ratio. Therefore, leucine and isoleucine exert anti-inflammatory effects in Caco-2 cells exposed to LPS by modulating JNK and NF-kB phosphorylation and IL-8 production. Further in vivo studies are required to validate these findings and gather valuable information for potential therapeutic or dietary interventions.

The bone marrow is responsible for producing an incredible number of cells daily in order to maintain blood homeostasis through a process called hematopoiesis. Hematopoiesis is a greatly demanding process and one entirely dependent on complex interactions between the hematopoietic stem cell (HSC) and its surrounding microenvironment. Zinc (Zn 2+ ) is considered an important trace element, playing diverse roles in different tissues and cell types, and zinc finger proteins (ZNF) are proteins that use Zn 2+ as a structural cofactor. In this way, the ZNF structure is supported by a Zn 2+ that coordinates many possible combinations of cysteine and histidine, with the most common ZNF being of the Cys 2 His 2 (C2H2) type, which forms a family of transcriptional activators that play an important role in different cellular processes such as development, differentiation, and suppression, all of these being essential processes for an adequate hematopoiesis. This review aims to shed light on the relationship between ZNF and the regulation of the hematopoietic tissue. We include works with different designs, including both in vitro and in vivo studies, detailing how ZNF might regulate hematopoiesis., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2023 Elsevier GmbH. All rights reserved.)

Protein restriction (PR) leads to bone marrow hypoplasia with changes in stromal cellularity components of the extracellular matrix in hematopoietic stem cells (HSCs). However, the underlying signaling mechanisms are poorly understood. We hypothesize that PR impairs the HSC mitogen-activated protein kinase (MAPK) signaling pathway response activation. Our aim is to evaluate the activation of MAPK and interleukin-3 (IL-3) proteins in HSC to explain PR-induced bone marrow hypoplasia, which causes altered proliferation and differentiation. C57BL/6 male mice were subjected to a low-protein diet (2% protein) or normoproteic (12% protein). PKC, PLCγ2, CaMKII, AKT, STAT3/5, ERK1/2, JNK, and p38d phosphorylation were evaluated by flow cytometry, and GATA1/2, PU.1, C/EBPα, NF-E2, and Ikz-3 genes (mRNAs) assessed by quantitative real-time-polymerase chain reaction. Pathway proteins, such as PLCγ2, JAK2, STAT3/5, PKC, and RAS do not respond to the IL-3 stimulus in PR, leading to lower activation of ERK1/2 and Ca2+ signaling pathways, consequently lowering the production of hematopoietic transcription factors. Colony forming units granulocyte-macrophage and colony forming units macrophage formation are impaired in PR even after being stimulated with IL-3. Long-term hematopoietic stem cells, short-term hematopoietic stem cells, granulocyte myeloid progenitor, and megakaryocyte-erythroid progenitor cells were significantly reduced in PR animals. This study shows for the first time that activation of MAPK pathway key proteins in HSCs is impaired in cases of PR. Several pathway proteins, such as PLCγ2, JAK2, STAT3, PKC, and RAS do not respond to IL-3 stimulation, leading to lower activation of extracellular signal-regulated protein kinase 1/2 and consequently lower production of hematopoietic transcription factors GATA1/2, PU.1, C/EBPa, NF-E2, and Ikz3. These changes result in a reduction in colony-forming units, proliferation, and differentiation, leading to hypocellularity., 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 © 2023 Elsevier Inc. All rights reserved.)

Blood orange consumption presents potential health benefits and may modulate epigenetic mechanisms such as microRNAs (miRNAs) expression. MiRNAs are non-coding RNAs responsible for post-transcriptional gene regulation, and these molecules can also be used as biomarkers in body fluids. This study was designed to investigate the effect of chronic blood orange juice (BOJ) intake on the inflammatory response and miRNA expression profile in plasma and blood cells in overweight women. The study cohort was comprised of twenty women aged 18-40 years old, diagnosed as overweight, who consumed 500 mL/d of BOJ for four weeks. Clinical data were collected at baseline and after 4 weeks of juice consumption, e.g., anthropometric and hemodynamic parameters, food intake, blood cell count, and metabolic and inflammatory biomarkers. BOJ samples were analyzed and characterized. Additionally, plasma and blood cells were also collected for miRNA expression profiling and evaluation of the expression of genes and proteins in the MAPK and NFκB signaling pathways. BOJ intake increased the expression of miR-144-3p in plasma and the expression of miR-424-5p, miR-144-3p, and miR-130b-3p in peripheral blood mononuclear cells (PBMC). Conversely, the beverage intake decreased the expression of let-7f-5p and miR-126-3p in PBMC. Computational analyses identified different targets of the dysregulated miRNA on inflammatory pathways. Furthermore, BOJ intake increased vitamin C consumption and the pJNK/JNK ratio and decreased the expression of IL6 mRNA and NFκB protein. These results demonstrate that BOJ regulates the expression of genes involved in the inflammatory process and decreases NFкB-protein expression in PBMC., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interest., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Objective: Anthocyanins are polyphenols that are promising chemopreventive agents. They stand out for their anti-inflammatory properties, with specific modulatory actions on the immune system. Additionally, regarding the immune system, a group of cells identified as mesenchymal stem cells (MSCs) have been attracting attention, mainly because of their capacity to migrate to sites of inflammation and produce potent immunomodulatory effects. Considering the ability of these cells to act on the immune system, as well as the properties of anthocyanins, especially delphinidin, in modulating the immune system, the aim of this study was to investigate the effects of delphinidin in influencing some immunoregulatory properties of MSCs., Methods: MSCs were cultivated in the presence of delphinidin 3-O-β-d-glycoside and cell viability, the cell cycle and the production of soluble factors (interleukin [IL]-1β, IL-6, IL-10, transforming growth factor [TGF]-β, prostaglandin E 2 [PGE 2 ] and nitric oxide [NO]) were evaluated, as was the expression of the transcription factors nuclear factor (NF)-κB and STAT3. Additionally, the effects of conditioned media from MSCs on macrophage activation were assessed., Results: Delphinidin at 50 µM does not affect cell viability. In association with lipopolysaccharide, delphinidin was able to induce MSC proliferation. Additionally, delphinidin modulated the MSC immune response, showing increased levels of anti-inflammatory cytokines such as IL-10 and TGF-β as well as lower expression of NF-κB. Furthermore, conditioned media from MSCs inhibited macrophage metabolism, reducing the production of IL-1β, IL-12, and TNF-α and increasing IL-10., Conclusions: Overall, this work showed that delphinidin can modify the immunomodulatory properties of MSCs, increasing the IL-10 production by macrophages., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Malnutrition is considered one of the most common problems in the elderly population worldwide and can significantly interfere in health evolution in these individuals, predisposing them to increased infection susceptibility. The immune response triggered by infections comprises several mechanisms, and macrophages play important roles in this response. This study aimed to evaluate mechanisms related to macrophage function in a model of protein malnutrition in the elderly. Two age groups (young: 3-5 months and elderly: 18-19 months) male C57BL/6NTac mice were subjected to protein malnutrition with a low-protein diet (2 %). The nutritional status, hemogram and number of peritoneal cells were affected by both age and nutritional status. Additionally, the spreading capacity as well as the phagocytic and fungicidal activity of peritoneal macrophages were affected by the nutritional status and age of the animal. Interestingly, the percentages of F4/80 + /CD11b + and CD86 + cells were reduced mostly in elderly animals, while the TLR-4 + population was more affected by nutritional status than by age. The production of pro-inflammatory cytokines such as TNF-α, IL-1α, and IL-6 was also influenced by nutritional status and/or by age, and malnourished animals of advanced age produced higher amounts of the anti-inflammatory cytokine IL-10. Furthermore, the phosphorylation ratio of the transcription factor NFκB (pNFκB/NFκB) was directly affected by the nutritional status, independently of age. Thus, these results allow us to conclude that aging and protein malnutrition compromise macrophage function, likely affecting their immune function, and in aged protein-malnourished animals, this impairment tends to be more pronounced., 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 © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: Dietary protein deficiency is common in the elderly, compromising hematopoiesis and the immune response, and may cause a greater susceptibility to infections. Mesenchymal stem cells (MSCs) have immunomodulatory properties and are essential to hematopoiesis. Therefore, this study aimed to investigate, in an aging model subjected to malnutrition due a reduced protein intake, aspects related to the immunomodulatory capacity of MSCs., Methods: Male C57BL/6 mice from young and elderly groups were fed with normoproteic or hypoproteic diets (12% and 2% of protein, respectively) and nutritional, biochemical and hematological parameters were evaluated. MSCs from bone marrow were isolated, characterized and their secretory parameters evaluated, along with gene expression. Additionally, the effects of aging and protein malnutrition on MSC immunomodulatory properties were assessed., Results: Malnourished mice lost weight and demonstrated anemia, leukopenia, and bone marrow hypoplasia. MSCs from elderly animals from both groups showed reduced CD73 expression and higher senescence rate; also, the malnourished state affected CD73 expression in young animals. The production of IL-1β and IL-6 by MSCs was affected by aging and malnutrition, but the IL-10 production not. Aging also increased the expression of NFκB, reducing the expression of STAT-3. However, MSCs from malnourished groups, regardless of age, showed decreased TGF-β and PGE 2 production. Evaluation of the immunomodulatory capacity of MSCs revealed that aging and malnutrition affected, mainly in lymphocytes, the production of IFN-γ and IL-10., Conclusion: Aging and reduced protein intake are factors that, alone or together, influence the immunomodulatory properties of MSCs and provide basic knowledge that can be further investigated to explore whether MSCs' therapeutic potential may be affected., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany.)

Pathogenic variants in A Disintegrin And Metalloproteinase (ADAM) 22, the postsynaptic cell membrane receptor for the glycoprotein leucine-rich repeat glioma-inactivated protein 1 (LGI1), have been recently associated with recessive developmental and epileptic encephalopathy. However, so far, only two affected individuals have been described and many features of this disorder are unknown. We refine the phenotype and report 19 additional individuals harbouring compound heterozygous or homozygous inactivating ADAM22 variants, of whom 18 had clinical data available. Additionally, we provide follow-up data from two previously reported cases. All affected individuals exhibited infantile-onset, treatment-resistant epilepsy. Additional clinical features included moderate to profound global developmental delay/intellectual disability (20/20), hypotonia (12/20) and delayed motor development (19/20). Brain MRI findings included cerebral atrophy (13/20), supported by post-mortem histological examination in patient-derived brain tissue, cerebellar vermis atrophy (5/20), and callosal hypoplasia (4/20). Functional studies in transfected cell lines confirmed the deleteriousness of all identified variants and indicated at least three distinct pathological mechanisms: (i) defective cell membrane expression; (ii) impaired LGI1-binding; and/or (iii) impaired interaction with the postsynaptic density protein PSD-95. We reveal novel clinical and molecular hallmarks of ADAM22 deficiency and provide knowledge that might inform clinical management and early diagnostics., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Guarantors of Brain.)

Reliable serological tests for the detection of SARS-CoV-2 antibodies among infected or vaccinated individuals are important for epidemiological and clinical studies. Low-cost approaches easily adaptable to high throughput screenings, such as Enzyme-Linked Immunosorbent Assays (ELISA) or electrochemiluminescence immunoassay (ECLIA), can be readily validated using different SARS-CoV-2 antigens. A total of 1,119 serum samples collected between March and July of 2020 from health employees and visitors to the University Hospital at the University of São Paulo were screened with the Elecsys ® Anti-SARS-CoV-2 immunoassay (Elecsys) (Roche Diagnostics) and three in-house ELISAs that are based on different antigens: the Nucleoprotein (N-ELISA), the Receptor Binding Domain (RBD-ELISA), and a portion of the S1 protein (ΔS1-ELISA). Virus neutralization test (CPE-VNT) was used as the gold standard to validate the serological assays. We observed high sensitivity and specificity values with the Elecsys (96.92% and 98.78%, respectively) and N-ELISA (93.94% and 94.40%, respectively), compared with RBD-ELISA (90.91% sensitivity and 88.80% specificity) and the ΔS1-ELISA (77.27% sensitivity and 76% specificity). The Elecsys ® proved to be a reliable SARS-CoV-2 serological test. Similarly, the recombinant SARS-CoV-2 N protein displayed good performance in the ELISA tests. The availability of reliable diagnostic tests is critical for the precise determination of infection rates, particularly in countries with high SARS-CoV-2 infection rates, such as Brazil. Collectively, our results indicate that the development and validation of new serological tests based on recombinant proteins may provide new alternatives for the SARS-CoV-2 diagnostic market., 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 © 2022 Andreata-Santos, Machado, Alves, Sales, Soares, Rodrigues, Silva, Favaro, Rodrigues-Jesus, Yamamoto, Andrade, Fock, Margarido, Carvalho, Boscardin, Durigon and Ferreira.)

Purpose: This study aimed to evaluate the radiation-induced direct and bystander (BYS) responses of mesenchymal stem cells (MSCs) and to characterize these cells radiobiologically. Methods and materials: MSCs were irradiated (IR) and parameters related to DNA damage and cellular signaling were verified in a dose range from 0.5 to 15 Gy; also a transwell insert co-culture system was used to study medium-mediated BYS effects. Results: The main effects on directly IR cells were seen at doses higher than 6 Gy: induction of cell death, cell cycle arrest, upregulation of p21, and alteration of redox status. Irrespective of a specific dose, induction of micronuclei formation, H2AX phosphorylation, and decreased Akt expression also occurred. Thus, mTOR expression, cell senescence, nitric oxide generation, and calcium levels, in general were not significantly modulated by radiation. Data from the linear-quadratic model showed a high alpha/beta ratio, which is consistent with a more exponential survival curve. BYS effects from the unirradiated MSCs placed into companion wells with the directly IR cells, were not observed. Conclusions: The results can be interpreted as a positive outcome, meaning that the radiation damage is restricted to the directed IR MSCs not leading to off-target cell responses.

The conditions of aquatic environments have a great influence on the microbiota of several animals, many of which are a potential source of microorganisms of biotechnological interest. In this study, bacterial strains isolated from aquatic environments were bioprospected to determine their probiotic profile and antimicrobial effect against fish and food pathogens. Two isolates, identified via 16S rRNA sequencing as Lactococcus lactis (L1 and L2) and one as Enterococcus faecium 135 (EF), produced a bacteriocin-like antimicrobial substance (BLIS), active against Listeria monocytogenes, Salmonella Choleraesuis and Salmonella Typhimurium. Antimicrobial activity of BLIS was reduced when exposed to high temperatures and proteolytic enzymes (trypsin, pepsin, papain and pancreatin). All strains were sensitive to 7 types of antibiotics (vancomycin, clindamycin, streptomycin, gentamicin, chloramphenicol, rifampicin and ampicillin), exhibited a high rate of adherence to Caco-2 cells and expressed no hemolysin and gelatinase virulence factors. EF showed some resistance at pH 2.5 and 3.0, and L2/EF showed higher resistance to the action of bile salts. Finally, the presence of bacteriocin genes encoding for proteins, including Nisin (L1 and L2), Enterocin A, B, P, and Mundticin KS (EF) was detected. The molecular and physiological evidence suggests that the bacterial isolates in this study could be used as natural antimicrobial agents and may be considered safe for probiotic application., (© 2022. The Author(s).)

In the course of infection and intense endotoxemia processes, induction of a catabolic state leading to weight loss is observed in mice and humans. However, the late effects of acute inflammation on energy homeostasis, regulation of body weight and glucose metabolism are yet to be elucidated. Here, we addressed whether serial intense endotoxemia, characterized by an acute phase response and weight loss, could be an aggravating or predisposing factor to weight gain and associated metabolic complications. Male Swiss Webster mice were submitted to 8 consecutive doses of lipopolysaccharide (10 mg/kg LPS), followed by 10 weeks on a high-fat diet (HFD). LPS-treated mice did not show changes in weight when fed standard chow. However, when challenged by a high-fat diet, LPS-treated mice showed greater weight gain, with larger fat depot areas, increased serum leptin and insulin levels and impaired insulin sensitivity when compared to mice on HFD only. Acute endotoxemia caused a long-lasting increase in mRNA expression of inflammatory markers such as TLR-4, CD14 and serum amyloid A (SAA) in the adipose tissue, which may represent the key factors connecting inflammation to increased susceptibility to weight gain and impaired glucose homeostasis. In an independent experimental model, and using publicly available microarray data from adipose tissue from mice infected with Gram-negative bacteria, we performed gene set enrichment analysis and confirmed upregulation of a set of genes responsible for cell proliferation and inflammation, including TLR-4 and SAA. Together, we showed that conditions leading to intense and recurring endotoxemia, such as common childhood bacterial infections, may resound for a long time and aggravate the effects of a western diet. If confirmed in humans, infections should be considered an additional factor contributing to obesity and type 2 diabetes epidemics.

Endothelial cells play an essential role in inflammation through synthesis and secretion of chemoattractant cytokines and expression of adhesion molecules required for inflammatory cell attachment and infiltration. The mechanisms by which endothelial cells control the pro-inflammatory response depend on the type of inflammatory stimuli, endothelial cell origin, and tissue involved. In the present study, we investigated the role of the transcription factor c-Myc in inflammation using a conditional knockout mouse model in which Myc is specifically deleted in the endothelium. At a systemic level, circulating monocytes, the chemokine CCL7, and the extracellular-matrix protein osteopontin were significantly increased in endothelial c-Myc knockout (EC-Myc KO) mice, whereas the cytokine TNFSF11 was downregulated. Using an experimental model of steatohepatitis, we investigated the involvement of endothelial c-Myc in diet-induced inflammation. EC-Myc KO animals displayed enhanced pro-inflammatory response, characterized by increased expression of pro-inflammatory cytokines and leukocyte infiltration, and worsened liver fibrosis. Transcriptome analysis identified enhanced expression of genes associated with inflammation, fibrosis, and hepatocellular carcinoma in EC-Myc KO mice relative to control (CT) animals after short-exposure to high-fat diet. Analysis of a single-cell RNA-sequencing dataset of human cirrhotic livers indicated downregulation of MYC in endothelial cells relative to healthy controls. In summary, our results suggest a protective role of endothelial c-Myc in diet-induced liver inflammation and fibrosis. Targeting c-Myc and its downstream pathways in the endothelium may constitute a potential strategy for the treatment of inflammatory disease., (© 2021 Federation of American Societies for Experimental Biology.)

Anemia is a common feature of chronic kidney disease (CKD). It is a process related to erythropoietin deficiency, shortened erythrocyte survival, uremic erythropoiesis inhibitors, and disordered iron homeostasis. Animal models of CKD-induced anemia are missing and would be desirable in order to study anemia mechanisms and facilitate the development of novel therapeutic tools. We induced three different models of CKD in mice and evaluated the development of anemia characteristics. Mice were subjected to unilateral ischemia-reperfusion or received repeated low doses of cisplatin or folic acid to induce nephropathy. Renal function, kidney injury and fibrotic markers were measured to confirm CKD. Moreover, serum hemoglobin, ferritin and erythropoietin were analyzed. Renal mRNA levels of HIF-2α , erythropoietin, hepcidin, GATA-2 , and GATA-2 target genes were also determined. All three CKD models presented increased levels of creatinine, urea, and proteinuria. Renal up-regulation of NGAL , KIM-1 , and TNF-α mRNA levels was observed. Moreover, the three CKD models developed fibrosis and presented increased fibrotic markers and α-SMA protein levels. CKD induced decreased hemoglobin and ferritin levels and increased erythropoietin levels in the serum. Renal tissue showed decreased erythropoietin and HIF-2α mRNA levels, while an increase in the iron metabolism regulator hepcidin was observed. GATA-2 transcription factor (erythropoietin repressor) mRNA levels were increased in all CKD models, as well as its target genes. We established three models of CKD-induced anemia, regardless of the mechanism and severity of kidney injury.

Protein malnutrition causes anemia and leukopenia as it reduces hematopoietic precursors and impairs the production of mediators that regulate hematopoiesis. Hematopoiesis occurs in distinct bone marrow niches that modulate the processes of differentiation, proliferation and self-renewal of hematopoietic stem cells (HSCs). Mesenchymal stem cells (MSCs) contribute to the biochemical composition of bone marrow niches by the secretion of several growth factors and cytokines, and they play an important role in the regulation of HSCs and hematopoietic progenitors. In this study, we investigated the effect of protein malnutrition on the hematopoietic regulatory function of MSCs. C57BL/6NTaq mice were divided into control and protein malnutrition groups, which received, respectively, a normal protein diet (12% casein) and a low protein diet (2% casein). The results showed that protein malnutrition altered the synthesis of SCF, TFG-β, Angpt-1, CXCL-12, and G-CSF by MSCs. Additionally, MSCs from the protein malnutrition group were not able to maintain the lymphoid, granulocytic and megakaryocytic-erythroid differentiation capacity compared to the MSCs of the control group. In this way, the comprehension of the role of MSCs on the regulation of the hematopoietic cells, in protein malnutrition states, is for the first time showed. Therefore, we infer that hematopoietic alterations caused by protein malnutrition are due to multifactorial alterations and, at least in part, the MSCs' contribution to hematological impairment., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Ionizing radiation (IR) is an important medical tool. Despite the effects associated with high-dose radiation during or after treatment, as well as in accidental exposures, the direct or indirect effect of low-dose IR in cells remain poorly documented. IR can affect the tissue microenvironment, including mesenchymal stem cells (MSCs), which have high regenerative and immunomodulatory capacities. This study aimed to investigate the effect of low-dose IR in association with the inflammatory stimuli of TNF-α on the immunomodulatory capacity of MSCs. MSCs were irradiated with a low-dose IR, stimulated with TNF-α, and cultivated in a bystander system with murine spleen cells. The results showed that TNF-R1 is expressed in MSCs and is not affected, even in irradiated MSCs. However, irradiated MSCs produced reduced amounts of IL-6 and increased amounts of IL-10. The levels of PGE 2 and NO • in MSCs were also increased when stimulated with TNF-α. Furthermore, conditioned media from irradiated MSCs reduced the proliferation of bystander lymphocytes and reduced the metabolic activity of macrophages. In addition, conditioned media from irradiated MSCs modulated the profile of cytokines in bystander spleen cells (lymphocytes and macrophages), reducing inflammatory and increasing anti-inflammatory cytokines, also increasing Treg cells. In conclusion, low-dose IR in association with an inflammatory stimulus affects the immunomodulatory properties of MSCs. In this way, the immunosuppressive capability of MSCs can be explored for several disease treatments where IR usually part of the context of the treatment. However, a complete understanding of the mechanisms underlying these interactions need further investigation. Graphical Abstract.

Background & Aims: Magnesium (Mg 2+ ) is able to modulate the differentiation and proliferation of cells. Mg 2+ restriction can trigger neutrophilia, but the processes that result in this change have yet to be investigated and are not fully understood. Hematopoiesis is a complex process that is regulated by many factors, including cytokines and growth factors, and is strongly influenced by nutrient availability. In this context, our objective was to investigate the impact of the short-term restriction of dietary Mg 2+ on bone marrow hematopoietic and peripheral blood cells, especially in processes related to granulocyte differentiation and proliferation., Methods: Male C57BL/6 mice were fed a Mg 2+ restricted diet (50 mg Mg 2+ /kg diet) for 4 weeks. Cell blood count and bone marrow cell count were evaluated. Bone marrow cells were also characterized by flow cytometry. Gene expression and cytokine production were evaluated, and a colony-forming cell assay related to granulocyte differentiation and proliferation was performed., Results: Short-term dietary restriction of Mg 2+ resulted in peripheral neutrophilia associated with an increased number of granulocytic precursors in the bone marrow. Additionally, Mg 2+ restriction resulted in an increased number of granulocytic colonies formed in vitro. Moreover, the Mg 2+ restricted group showed increased expression of CSF3 and CEBPα genes as well as increased production of G-CSF in association with increased expression of STAT3 protein., Conclusion: Short-term dietary restriction of Mg 2+ induces granulopoiesis by increasing G-CSF production and activating the CEBPα and STAT-3 pathways, resulting in neutrophilia in peripheral blood., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest., (Copyright © 2021 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.)

The perivascular adipose tissue (PVAT) differs from other fat depots and exerts a paracrine action on the vasculature. The spleen has an important role in the immune response, and it was observed to have either a protective role or a contribution to obesity-related diseases. However, the relation between spleen and PVAT is elusive in obesity. We investigated the role of spleen in the inflammatory profile of the mesenteric PVAT (mPVAT) from mice fed a high-fat diet (HFD) for 16 weeks. Male C57Bl/6 mice were sham-operated or splenectomized (SPX) and fed a HFD for 16 weeks. mPVAT morphology was evaluated by hematoxylin and eosin staining, infiltrated immune cells were evaluated by flow cytometry, inflammatory cytokines were evaluated by ELISA and the splenic cell chemotaxis mediated by mPVAT was evaluated using a transwell assay. In SPX mice, HFD induced adipocyte hypertrophy and increased immune cell infiltration and proinflammatory cytokine levels in mPVAT. However, none of these effects were observed in mPVAT from sham-operated mice. Spleen from HFD fed mice presented reduced total leukocytes and increased inflammatory markers when compared to the spleen from control mice. Chemotaxis of spleen cells mediated by mPVAT of HFD fed mice was reduced in relation to standard diet fed mice. The spleen protects mPVAT against the effects of 16-week HFD. This information was missing, and it is important because PVAT is different from other fat depots and data cannot be extrapolated from any type of adipose tissue to PVAT.

Although branched-chain amino acids (BCAA) are commonly used as a strategy to recover nutritional status of critically ill patients, recent findings on their role as immunonutrients have been associated with unfavorable outcomes, especially in obese patients. The present study aimed to explore the effects of different BCAA supplementation protocols in the inflammatory response of LPS-stimulated RAW 264.7 macrophages. Cell cultures were divided into five groups, with and without BCAA supplementation, (2 mmol/L of each amino acid). Then, cell cultures followed three different treatment protocols, consisting of a pretreatment (PT), an acute treatment (AT), and a chronic treatment (CT) with BCAA and LPS stimulation (1 µg/mL). Cell viability was analyzed by MTT assay, NO production was assessed by the Griess reaction and IL-6, IL-10, TNF-α and PGE2 synthesis, was evaluated by ELISA. BCAA significantly increased cell viability in AT and CT protocols, and NO and IL-10 synthesis in all treatment protocols. IL-6 synthesis was only increased in PT and CT protocols. TNF-α and PGE2 synthesis were not altered in any of the protocols and groups. BCAA supplementation was able to increase both pro and anti-inflammatory mediators synthesis by RAW 264.7 macrophages, which was influenced by the protocol applied. Moreover, these parameters were significantly increased by isoleucine supplementation, highlighting a potential research field for future studies.

Purpose: Considering the COVID-19 pandemic, vitamin D is a target of research and speculation. Lockdown or home isolation reduces sunlight exposition and increases the risk of vitamin D deficiency. Special attention is needed for older people at risk of both severe forms of COVID-19 and vitamin D deficiency. This review aims to highlight the association of vitamin D and COVID-19 in two instances, the direct influence of vitamin D on the immune system, and the indirect risks for other vitamin D deficiency-related diseases, such as musculoskeletal properties in older persons., Methods: We performed a narrative review., Results: Whether vitamin D deficiency is associated with COVID-19 poor prognosis, and if vitamin D supplementation may improve the post-infection outcomes is still unclear. In any case, the pandemic generates indirect burden, such as the sequence: home isolation, low sunlight exposition, vitamin D deficiency, and fragility fractures., Conclusion: Therefore, it is time to debate how to optimize vitamin D status in older people, especially during the COVID-19 pandemic., Competing Interests: Conflict of interestThe authors declare that they have no conflict of interest., (© The Author(s), under exclusive licence to Springer Nature Switzerland AG part of Springer Nature 2021.)

In the last decade, mesenchymal stem cells (MSCs) have been gaining attention due their ability to influence the function of other cells as well as modulate the inflammatory response. This occurs via their immunomodulatory functions,  acting through direct cell-cell interaction or by releasing a broad spectrum of bioactive factors such as cytokines and growth factors. In addition, prostaglandins are arachidonic acid metabolites that play a key role in the generation and modulation of the inflammatory response. Among the bioactive prostaglandins, PGF 2α  is able to stimulate cell proliferation as well as act to inhibit progenitor cell differentiation, but no information about this prostaglandin's action on the immunoregulatory function of MSCs has been reported. In this study we evaluate important aspects of the influence of PGF 2α analog (17-phenyl-trinor PGF 2α ), which is a potent prostaglandin FP receptor agonist, on some mechanisms that control the main functions of MSCs. C3H10T1/2, a mesenchymal stem cell linage, was stimulated with PGF 2α under inflammatory conditions trigged by LPS in order to investigate PGF 2α inflammatory parameters as well as its ability to immunoregulate macrophages and lymphocytes. PGF 2α has the ability to increase proliferation tax without altering the cell viability of LPS-stimulated MSCs, while also diminishing the phosphorylation of NFκB transcription factor leading to attenuation of IL-1β and GM-CSF production. Additionally, MSC-s conditioned media from cells stimulated with PGF 2α was able to increase the lymphocytes' IL-10 production. Overall, this study implied that PGF 2α are able to modify some properties of MSCs., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Probiotic supplementation arises as playing an immune-stimulatory role. High-intensity and -volume exercise can inhibit immune cell function, which threatens athletic performance and recovery. We hypothesized that 30 days of probiotic supplementation could stabilize the immune system of athletes preventing immune suppression after a marathon race. Twenty-seven male marathonists were double-blinded randomly into probiotic (Bifidobacterium-animalis-subsp.-Lactis (10 × 10 9 ) and Lactobacillus-Acidophilus (10 × 10 9 ) + 5 g of maltodextrin) and placebo (5 g of maltodextrin) group. They received 30 sachets and supplemented 1 portion/day during 30 days before the race. Blood were collected 30 days before (rest), 1 day before (pre), 1 h after (post) and 5 days after the race (recovery). Both chronic and acute exercise modulated a different T lymphocyte population (CD3 + CD4 - CD8 - T-cells), increasing pre-race, decreasing post and returning to rest values at the recovery. The total number of CD8 T cell and the memory subsets statistically decreased only in the placebo group post-race. Pro-inflammatory cytokine production by stimulated lymphocytes decreased in the probiotic group after the supplementation period. 30 days of probiotic supplementation maintained CD8 T cell and effector memory cell population and played an immunomodulatory role in stimulated lymphocytes. Both, training and marathon modulated a non-classical lymphocyte population regardless of probiotic supplementation.

Objectives: The essential branched chain amino acids (BCAAs) valine, leucine, and isoleucine, are widely studied because of their effects on immunity and metabolism. Mesenchymal stem cells (MSCs) are a type of cell also studied due to their immunomodulatory properties. Since both BCAAs and MSCs have immunomodulatory capacity, the objective of this study was to evaluate the influence of BCAAs on some immunomodulatory aspects of MSCs., Methods: MSCs were cultivated in BCAA-supplemented media to evaluate metabolic activity, including cell cycle, proliferative nuclear cell antigen, peroxisome proliferator-activated receptor gamma coactivator 1-alpha, avian myelocytomatosis viral oncogene homolog, peroxisome proliferator activated receptor gamma, nuclear factor kappa B (NFкB), and signal transducers and activators of transcription 3 (STAT-3) expression. Additionally, some inflammatory mediators' synthesis, such as interleukin (IL) 1-beta, IL-10, granulocyte-macrophage colony-stimulating factor, transforming growth factor beta, nitric oxide, and prostaglandin E 2 , were also evaluated., Results: Supplementation with BCAA led not only to increased MSC proliferation with more cells in the S, G2, and M cycle phases, but also to increased metabolic activity. BCAA supplementation also altered the immunomodulatory capacity of MSCs by decreasing the p-NFкB/NFкB and increasing the p-STAT-3/STAT-3 gene expression ratios, in addition to increasing synthesis of the antiinflammatory mediators transforming growth factor beta and prostaglandin E 2 . Finally, MSCs cultivated in BCAA-supplemented media was shown to decrease the IL-6 and tumor necrosis factor alpha production by macrophages., Conclusions: BCAA supplementation affected some immunoregulatory aspects of MSCs., 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 © 2020 Elsevier Inc. All rights reserved.)

Magnesium (Mg 2+ ) is an essential mineral for the functioning and maintenance of the body. Disturbances in Mg 2+ intracellular homeostasis result in cell-membrane modification, an increase in oxidative stress, alteration in the proliferation mechanism, differentiation, and apoptosis. Mg 2+ deficiency often results in inflammation, with activation of inflammatory pathways and increased production of proinflammatory cytokines by immune cells. Immune cells and others that make up the blood system are from hematopoietic tissue in the bone marrow. The hematopoietic tissue is a tissue with high indices of renovation, and Mg 2+ has a pivotal role in the cell replication process, as well as DNA and RNA synthesis. However, the impact of the intra- and extracellular disturbance of Mg 2+ homeostasis on the hematopoietic tissue is little explored. This review deals specifically with the physiological requirements of Mg 2+ on hematopoiesis, showing various studies related to the physiological requirements and the effects of deficiency or excess of this mineral on the hematopoiesis regulation, as well as on the specific process of erythropoiesis, granulopoiesis, lymphopoiesis, and thrombopoiesis. The literature selected includes studies in vitro, in animal models, and in humans, giving details about the impact that alterations of Mg 2+ homeostasis can have on hematopoietic cells and hematopoietic tissue.

In March 2020, WHO declared a pandemic state due to SARS-CoV-2 having spread. TaqMan-based real-time RT-qPCR is currently the gold standard for COVID-19 diagnosis. However, it is a high-cost assay, inaccessible for the majority of laboratories around the world, making it difficult to diagnose on a large scale. The objective of this study was to standardize lower cost molecular methods for SARS-CoV-2 identification. E gene primers previously determined for TaqMan assays by Colman et al. (2020) were adapted in SYBR Green assay and RT-PCR conventional. The cross-reactivity test was performed with 17 positive samples for other respiratory viruses, and the sensibility test was performed with 8 dilutions (10 based) of SARS-CoV-2 isolated and 63 SARS-CoV-2-positive samples. The SYBR Green assays and conventional RT-PCR have not shown amplification of the 17 respiratory samples positives for other viruses. The SYBR Green-based assay was able to detect all 8 dilutions of the isolate. The conventional PCR detected until 10 7 dilution, both assays detected the majority of the 63 samples, 98.42% of positivity in SYBR Green, and 93% in conventional PCR. The average Ct variation between SYBR Green and TaqMan was 1.92 and the highest Ct detected by conventional PCR was 35.98. Both of the proposed assays are less sensitive than the current gold standard; however, our data shows a low sensibility variation, suggesting that these methods could be used by laboratories as a lower cost molecular method for SARS-CoV-2 diagnosis.

Hypercholesterolemia, also called high cholesterol, is a form of hyperlipidemia, which may be a consequence of diet, obesity or diabetes. In addition, increased levels of low-density lipoprotein (LDL) and reduced levels of high-density lipoprotein (HDL) cholesterol are associated with a higher risk of atherosclerosis and coronary heart disease. Thus, controlling cholesterol levels is commonly necessary, and fibrates have been used as lipid-lowering drugs. Gemfibrozil is a fibrate that acts via peroxisome proliferator-activated receptor alpha to promote changes in lipid metabolism and decrease serum triglyceride levels. However, anemia and leukopenia are known side effects of gemfibrozil. Considering that gemfibrozil may lead to anemia and that gemfibrozil acts via peroxisome proliferator-activated receptor alpha, we treated wild-type and peroxisome proliferator-activated receptor alpha-knockout mice with gemfibrozil for four consecutive days. Gemfibrozil treatment led to anemia seven days after the first administration of the drug; we found reduced levels of hemoglobin, as well as red blood cells, white blood cells and a reduced percentage of hematocrits. PPAR-alpha-knockout mice were capable of reversing all of those reduced parameters induced by gemfibrozil treatment. Erythropoietin levels were increased in the serum of gemfibrozil-treated animals, and we also observed an increased expression of hypoxia-inducible factor-2 alpha ( HIF-2α ) and erythropoietin in renal tissue, while PPAR-alpha knockout mice treated with gemfibrozil did not present increased levels of serum erythropoietin or tissue HIF-2α and erythropoietin mRNA levels in the kidneys. We analyzed bone marrow and found that gemfibrozil reduced erythrocytes and hematopoietic stem cells in wild-type mice but not in PPAR-alpha-knockout mice, while increased colony-forming units were observed only in wild-type mice treated with gemfibrozil. Here, we show for the first time that gemfibrozil treatment leads to anemia and leukopenia via peroxisome proliferator-activated receptor alpha in mice.

NRAS-mutations arise in 15-20% of all melanomas and are associated with aggressive disease and poor prognosis. Besides, the treatment for NRAS-mutant melanoma are not very efficient and is currently limited to immune checkpoints inhibitors or aggressive chemotherapy. 4-nerolidylcathecol (4-NC), a natural product extracted from Pothomorphe umbellata, induces apoptosis in melanoma cells by ROS production, DNA damage and increased p53 expression, in addition to inhibiting invasion in reconstructed skin. Moreover, 4-NC showed cytotoxicity in BRAF/MEKi-resistant and naive melanoma cells by Endoplasmic Reticulum (ER) stress induction in vitro. We evaluated the in vivo efficacy and the systemic toxicity of 4-NC in a NRAS-mutant melanoma model. 4-NC was able to significantly suppress tumor growth 4-fold compared to controls. Cleaved PARP and p53 expression were increased indicating cell death. As a proof of concept, MMP-2 and MMP-14 gene expression were decreased, demonstrating a possible role of 4-NC in melanoma invasion inhibition. Toxicological analysis indicated minor changes in the liver and bone marrow, but this toxicity was very mild when compared to other proteasome inhibitors and ER stress inductors already described. Our data indicate that 4-NC can counteract melanoma growth in vivo with minor adverse effects, suggesting further investigation as a potential NRAS-mutant melanoma treatment., 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 © 2020 Elsevier Ltd. All rights reserved.)

Background & Aims: Protein malnutrition (PM) affects hematopoiesis leading to bone marrow (BM) hypoplasia and arrests hematopoietic stem cells (HSC) in G 0 /G 1 cell cycle phases, which cause anemia and leukopenia. Hematopoiesis is mainly regulated by BM niches where endothelial cells (EC) present a key regulatory role. Thus, our objective is to evaluate whether PM affects the modulatory capacity of EC on hematopoiesis., Methods: C57BL/6 male mice received for 5 weeks a normal protein diet (12% casein) or a low protein diet (2% casein). MSC were isolated and differentiated in vitro into EC and the synthesis of SCF, Ang-1, CXCL-12, IL-11, TGF-β and G-CSF were evaluated. The HSC and hematopoietic progenitors were quantified and the EC capacity to modulate the hematopoietic system was also evaluated. Moreover, the ability of PM bone marrow to support hematopoieisis was assessed by proliferation of infused leukemic myelo-monoblasts cells., Results: PM decreases HSC and hematopoietic progenitor pool and promotes cell cycle arrest and a lower proliferation rate of leukemic myelo-monoblasts. PM also committed hematopoietic regulatory characteristics from EC, resulting in the modification of both cell cycle pattern and hematopoietic differentiation., Conclusion: BM microenvironment is compromised in PM, and since PM disturbs EC, it becomes one of the factors responsible for the hematopoietic cell cycle arrest and impairment of HSC differentiation., (Copyright © 2019 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.)

The demand for animal protein for human consumption has been risen exponentially. Modern animal production practices are associated with the regular use of antibiotics, potentially increasing the emerging multi-resistant bacteria, which may have a negative impact on public health. In poultry production, substances capable of maximizing the animals' performance and displaying an antimicrobial activity against pathogens are very well desirable features. Probiotic can be an efficient solution for such a task. In the present work, lactic acid bacteria (LAB) were isolated from chicken cecum and screened for their antagonistic effect towards many pathogens. Their capacity of producing the B-complex vitamins folate and riboflavin were also evaluated. From 314 isolates, three (C43, C175 and C195) produced Bacteriocin-Like Inhibitory Substances (BLIS) against Staphylococcus aureus (inhibition zones of 18.9, 21.5, 19.5 mm, respectively) and also inhibited the growth of Salmonella Heidelberg. The isolate C43 was identified as Enterococcus faecium, while C173 and C195 were both identified as Lactococcus lactis subsp. lactis. Moreover, the isolates L. lactis subsp. lactis strains C173 and C195 demonstrated high potential to be used as probiotic in poultry feed, in addition to their advantage of producing folate (58.0 and 595.5 ng/mL, respectively) and riboflavin (223.3 and 175.0 ng/mL, respectively).

Objective: For every 100 random children diagnosed with autism, at least 20 have morphological abnormalities, often associated with syndromes. Brazil does not have a standardized and validated instrument for morphological physical examination. This study aimed to translate into Brazilian Portuguese and culturally adapt the clinical signs described in the Autism Dysmorphology Measure, as well as validate the instrument in a sample of children with autism., Methods: The original instrument was translated, culturally adapted, and published in full, following traditional procedures for translation, back-translation, and terminology adaptation according to the Nomina Anatomica. The sample included 62 children from a published multicenter study, with intelligence quotient between 50-69, of both genders, with chronological age between 3-6 years. Two clinical geneticists performed the morphological physical examination, which consisted of investigating 82 characteristics assessing 12 body areas. We used Cohen's Kappa coefficient to evaluate the agreement between the two observers., Results: The final version of the instrument - translated into Brazilian Portuguese and culturally adapted - showed high agreement between the two observers., Conclusions: The translated instrument meets all international criteria, and minor anomalies and their clinical descriptions were standardized and are recognizable for physicians not specialized in genetics.

Objective: It is well known that protein malnutrition (PM) states can affect hematopoiesis, leading to severe leukopenia and reduced number of granulocytes, which act as the first line of defense, and are important to the innate immune response. The aim of this study was to elucidate some of the mechanisms involved in the impairment of granulopoiesis in PM., Methods: Male C57BL/6 mice were submitted to PM with a low-protein diet containing 2% protein. Control mice were fed a 12% protein-containing diet. Bone marrow histology and the percentage of granulocytic progenitors were evaluated after in vivo granulocyte-colony stimulating factor (G-CSF) stimulus. Cell proliferation, STAT3 signaling, and the expression of G-CSF receptor were evaluated in hematopoietic progenitor cells., Results: Malnourished animals presented with leukopenia associated with reduced number of granulocytes and reduced percentage of granulocytic progenitors; however, no differences were observed in the regulatory granulopoietic cytokine G-CSF. Additionally, the malnourished group presented with impaired response to in vivo G-CSF stimulus compared with control animals. PM was implicated in decreased ability of c-Kit + cells to differentiate into myeloid progenitor cells and downregulated STAT3 signaling. Furthermore, the malnourished group exhibited reduced expression of G-CSF receptor on granule-monocytic progenitors. This reduced expression was not completely reversible with G-CSF treatment., Conclusions: This study implies that PM promotes intrinsic alterations to hematopoietic precursors, which result in hematologic changes, mainly neutropenia, observed in peripheral blood in PM states., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Annexin A1 (AnxA1) modulates neutrophil life span and bone marrow/blood cell trafficking thorough activation of formyl-peptide receptors (FPRs). Here, we investigated the effect of exogenous AnxA1 on haematopoiesis in the mouse. Treatment of C57BL/6 mice with recombinant AnxA1 (rAnxA1) reduced the granulocyte-macrophage progenitor (GMP) population in the bone marrow, enhanced the number of mature granulocytes Gr-1 + Mac-1 + in the bone marrow as well as peripheral granulocytic neutrophils and increased expression of mitotic cyclin B1 on hematopoietic stem cells (HSCs)/progenitor cells (Lin - Sca-1 + c-Kit + : LSK). These effects were abolished by simultaneous treatment with Boc-2, an FPR pan-antagonist. In in vitro studies, rAnxA1 reduced both HSC (LSKCD90 low FLK-2 - ) and GMP populations while enhancing mature cells (Gr1 + Mac1 + ). Moreover, rAnxA1 induced LSK cell proliferation (Ki67 + ), increasing the percentage of cells in the S/G2/M cell cycle phases and reducing Notch-1 expression. Simultaneous treatment with WRW4, a selective FPR2 antagonist, reversed the in vitro effects elicited by rAnxA1. Treatment of LSK cells with rAnxA1 led to phosphorylation of PCLγ2, PKC, RAS, MEK, and ERK1/2 with increased expression of NFAT2. In long-term bone marrow cultures, rAnxA1 did not alter the percentage of LSK cells but enhanced the Gr-1 + Mac-1 + population; treatment with a PLC (U73122), but not with a PKC (GF109203), inhibitor reduced rAnxA1-induced phosphorylation of ERK1/2 and Elk1. Therefore, we identify here rAnxA1 as an inducer of HSC/progenitor cell differentiation, favouring differentiation of the myeloid/granulocytic lineage, via Ca 2+ /MAPK signalling transduction pathways., Competing Interests: Conflict of interestThe authors declare that they have no conflict of interest.

Background: Phelan-McDermid syndrome (PMS) is a rare genetic disorder characterized by global developmental delay, intellectual disability (ID), autism spectrum disorder (ASD), and mild dysmorphisms associated with several comorbidities caused by SHANK3 loss-of-function mutations. Although SHANK3 haploinsufficiency has been associated with the major neurological symptoms of PMS, it cannot explain the clinical variability seen among individuals. Our goals were to characterize a Brazilian cohort of PMS individuals, explore the genotype-phenotype correlation underlying this syndrome, and describe an atypical individual with mild phenotype., Methodology: A total of 34 PMS individuals were clinically and genetically evaluated. Data were obtained by a questionnaire answered by parents, and dysmorphic features were assessed via photographic evaluation. We analyzed 22q13.3 deletions and other potentially pathogenic copy number variants (CNVs) and also performed genotype-phenotype correlation analysis to determine whether comorbidities, speech status, and ASD correlate to deletion size. Finally, a Brazilian cohort of 829 ASD individuals and another independent cohort of 2297 ID individuals was used to determine the frequency of PMS in these disorders., Results: Our data showed that 21% (6/29) of the PMS individuals presented an additional rare CNV, which may contribute to clinical variability in PMS. Increased pain tolerance (80%), hypotonia (85%), and sparse eyebrows (80%) were prominent clinical features. An atypical case diagnosed with PMS at 18 years old and IQ within the normal range is here described. Among Brazilian ASD or ID individuals referred to CNV analyses, the frequency of 22q13.3 deletion was 0.6% (5/829) and 0.61% (15/2297), respectively. Finally, renal abnormalities, lymphedema, and language impairment were found to be positively associated with deletion sizes, and the minimum deletion to cause these abnormalities is here suggested., Conclusions: This is the first work describing a cohort of Brazilian individuals with PMS. Our results confirm the impact of 22q13 deletions on ASD and several comorbidities, such as hypotonia. The estimation of a minimal deletion size for developing lymphedema and renal problem can assist prediction of prognosis in PMS individuals, particularly those diagnosed in early infancy. We also identified one atypical individual carrying SHANK3 deletion, suggesting that resilience to such mutations occurs. This case expands the clinical spectrum of variability in PMS and opens perspectives to identify protective mechanisms that can minimize the severity of this condition.

Glutamine (GLN) is the most abundant free amino acid in the body, and is considered as a conditionally essential amino acid under stress conditions, acting as an important modulator of the immune response. We here investigated the role of exogenous GLN treatment on leukocyte migration after the onset of endotoxemia and the intracellular mechanisms of GLN actions on neutrophils. Two in vivo models of endotoxemia caused by lipopolysaccharide of Escherichia coli (LPS) injection were carried out in male outbred Balb/C mice 2-3 months old, as follow: (1) LPS (50 μg/kg) was intravenously injected 1 h prior to intravenous injection of GLN (0.75 mg/kg) and samples were collected 2 h later to investigate the role of GLN on the acute lung inflammation; (2) LPS (1 mg/kg) was intraperitoneally injected 1 h prior to intravenous injection of GLN (0.75 mg/kg) and samples were collected 18 h later to measure the effects of GLN on local and later phases of inflammation in the peritoneum. Results showed that GLN administration reduced the number of neutrophils in the inflamed lungs, partially recovery of the reduced number of leukocytes in the blood; reduced adhesion molecules on lung endothelium and on circulating neutrophils. Moreover, GLN treatment diminished the number of neutrophils, levels of chemotactic cytokine CXCL2 in the inflamed peritoneum, and neutrophils collected from the peritoneum of GLN-treated mice presented lower levels of Rho, Rac, and JNK. Together, our data show novel mechanisms involved in the actions of GLN on neutrophils migration.

Prader-Willi syndrome (PWS) and recurrent 15q13.3 microdeletion syndrome can be caused by genomic rearrangements in the complex 15q11q13 chromosomal region. Here, we describe the first female child with PWS and 15q13.3 microdeletion syndrome resulting from an unusual 10.7-Mb deletion from 15pter to 15q13.3 due to an unbalanced de novo 15;19 translocation. The patient presents with hypotonia, microcephaly, developmental delay with lack of speech, intellectual disability, happy demeanor, clinodactyly of the 4th and 5th fingers, and dysmorphic facial features discordant for PWS and consistent with an atypical phenotype., (© 2019 S. Karger AG, Basel.)

Micronutrients are indispensable for adequate metabolism, such as biochemical function and cell production. The production of blood cells is named haematopoiesis and this process is highly consuming due to the rapid turnover of the haematopoietic system and consequent demand for nutrients. It is well established that micronutrients are relevant to blood cell production, although some of the mechanisms of how micronutrients modulate haematopoiesis remain unknown. The aim of the present review is to summarise the effect of Fe, Mn, Ca, Mg, Na, K, Co, iodine, P, Se, Cu, Li and Zn on haematopoiesis. This review deals specifically with the physiological requirements of selected micronutrients to haematopoiesis, showing various studies related to the physiological requirements, deficiency or excess of these minerals on haematopoiesis. The literature selected includes studies in animal models and human subjects. In circumstances where these minerals have not been studied for a given condition, no information was used. All the selected minerals have an important role in haematopoiesis by influencing the quality and quantity of blood cell production. In addition, it is highly recommended that the established nutrition recommendations for these minerals be followed, because cases of excess or deficient mineral intake can affect the haematopoiesis process.

The immune system is essential for the control and elimination of infections, and macrophages are cells that act as important players in orchestrating the various parts of the inflammatory/immune response. Amino acids play important role in mediating functionality of the inflammatory response, especially mediating macrophages functions and cytokines production. We investigated the influence of glutamine, taurine and their association on the modulation of inflammatory pathway markers in macrophages. The RAW 264.7 macrophage cell line was cultivated in the presence of glutamine and taurine and proliferation rates, cell viability, cell cycle phases, IL-1α, IL-6, IL-10 and TNF-α as well as H 2 O 2 production and the expression of the transcription factor, NFκB, and its inhibitor, IκBα, were evaluated. Our results showed an increase in viable cells and increased proliferation rates of cells treated with glutamine concentrations over 2 mM, as well as cells treated with both glutamine and taurine. The cell cycle showed a higher percentage of cells in the phases S, G2 and M when they were treated with 2 or 10 mM glutamine, or with glutamine and taurine in cells stimulated with lipopolysaccharide. The pNFκB/NFκB showed reduced ratio expression when cells were treated with 10 mM of glutamine or with glutamine in association with taurine. These conditions also resulted in reduced TNF-α, IL-1α and H 2 O 2 production, and higher production of IL-10. These findings demonstrate that glutamine and taurine are able to modulate macrophages inflammatory pathways, and that taurine can potentiate the effects of glutamine, illustrating their immunomodulatory properties.

Magnesium (Mg 2+ ) is a mineral with the ability to influence cell proliferation and to modulate inflammatory/immune responses, due to its anti-inflammatory properties. In addition, mesenchymal stem cells (MSCs) modulate the function of all major immune cell populations. Knowing that, the current work aimed to investigate the effects of Mg 2+ enrichment, and its influence on the immunomodulatory capacity of MSCs. Murine C3H/10T1/2 MSCs were cultivated in media with different concentrations of Mg 2+ (0, 1, 3 and 5 mM), in order to evaluate the effects of Mg 2+ on MSC immunomodulatory properties, cell proliferation rates, expression of NFκB and STAT-3, production of IL-1β, IL-6, TGF-β, IL-10, PGE2 and NO, and TRPM7 expression. The results showed that TRPM7 is expressed in MSCs, but Mg 2+ , in the way that cells were cultivated, did not affect TRPM7 expression. Additionally, there was no difference in the intracellular concentration of Mg 2+ . Mg 2+ , especially at 5 mM, raised proliferation rates of MSCs, and modulated immune responses by decreasing levels of IL-1β and IL-6, and by increasing levels of IL-10 and PGE2 in cells stimulated with LPS or TNF-α. In addition, MSCs cultured in 5 mM Mg 2+ expressed lower levels of pNFκB/NFκB and higher levels of pSTAT-3/STAT-3. Furthermore, conditioned media from MSCs reduced lymphocyte and macrophage proliferation, but Mg 2+ did not affect this parameter. In addition, conditioned media from MSCs cultured at 5 mM of Mg 2+ modulated the production profile of cytokines, especially of IL-1β and IL-6 in macrophages. In conclusion, Mg 2+ is able to modulate some immunoregulatory properties of MSCs., (Copyright © 2018. Published by Elsevier Inc.)