Glucocorticoid modulates oxidative and thermogenic function of rat brown adipose tissue and human brown adipocytes.

Chronic and excessive glucocorticoid (GC) exposure can cause Cushing's syndrome, resulting in fat accumulation in selected body areas. Particularly in the brown adipose tissue (BAT), GC acts negatively, resulting in whitening of the tissue. We hypothesized that dysregulation of microRNAs by GC could be an additional mechanism to explain its negative actions in BAT. Male Wistar rats were divided into two groups: (1) Control sham and (2) GC group that was administered dexamethasone 6.25 mg/200 µL via osmotic pump implantation over 28 days. After this period, the animals were euthanized and BAT tissue was properly stored. Human fat cells treated with dexamethasone were used to translate the experimental results found in animals to human biology. GC-treated rat BAT presented with large lipid droplets, severely impaired thermogenic activation, and reduced glucose uptake measured by 18F-FDG PET/CT. GC exposure induced a reduction in the mitochondrial OXPHOS system and oxygen consumption. MicroRNA profiling of BAT revealed five top-regulated microRNAs and among them miR-21-5p was the most significantly upregulated in GC-treated rats compared to the control group. Although upregulation of miR-21-5p in the tissue, differentiated primary brown adipocytes from GC-treated rats had decreased miR-21-5p levels compared to the control group. To translate these results to the clinic, human brown adipocytes were treated with dexamethasone and miR-21-5p inhibitor. In human brown cells, inhibition of miR-21-5p increased brown adipocyte differentiation and prevented GC-induced glucose uptake, resulting in a lower glycolysis rate. In conclusion, high-dose GC therapy significantly impacts brown adipose tissue function, with a notable association between glucose uptake and miR-21-5p.

Oral Squamous Cell Carcinoma Lipid Evaluation in Gingiva Tissue Stored in TRIzol via Shotgun Lipidomics and MALDI Mass Spectrometry Imaging.

Oral squamous cell carcinoma (OSCC) is the prevalent type of oral cavity cancer, requiring precise, accurate, and affordable diagnosis to identify the disease in early stages, Comprehending the differences in lipid profiles between healthy and cancerous tissues encompasses great relevance in identifying biomarker candidates and enhancing the odds of successful cancer treatment. Therefore, the present study evaluates the analytical performance of simultaneous mRNA and lipid extraction in gingiva tissue from healthy patients and patients diagnosed with OSCC preserved in TRIzol reagent. The data was analyzed by partial least-squares discriminant analysis (PLS-DA) and confirmed via matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI). The lipid extraction in TRIzol solution was linear in a range from 330 to 2000 ng mL-1, r2 > 0.99, intra and interday precision and accuracy <15%, and absolute recovery values ranging from 90 to 110%. The most important lipids for tumor classification were evaluated by MALDI-MSI, revealing that the lipids responsible for distinguishing the OSCC group are more prevalent in the cancerous tissue in contrast to the healthy group. The results exhibit the possibilities to do transcriptomic and lipidomic analyses in the same sample and point out important candidates related to the presence of OSCC.

Polyunsaturated fatty acids alter the formation of lipid droplets and eicosanoid production in Leishmania promastigotes.

BACKGROUND: The knowledge about eicosanoid metabolism and lipid droplet (LD) formation in the Leishmania is very limited and new approaches are needed to identify which bioactive molecules are produced of them. OBJECTIVES: Herein, we compared LDs and eicosanoids biogenesis in distinct Leishmania species which are etiologic agents of different clinical forms of leishmaniasis. METHODS: For this, promastigotes of Leishmania amazonensis, L. braziliensis and L. infantum were stimulated with polyunsaturated fatty acids (PUFA) and LD and eicosanoid production was evaluated. We also compared mutations in structural models of human-like cyclooxygenase-2 (GP63) and prostaglandin F synthase (PGFS) proteins, as well as the levels of these enzymes in parasite cell extracts. FINDINGS: PUFAs modulate the LD formation in L. braziliensis and L. infantum. Leishmania spp with equivalent tissue tropism had same protein mutations in GP63 and PGFS. No differences in GP63 production were observed among Leishmania spp, however PGFS production increased during the parasite differentiation. Stimulation with arachidonic acid resulted in elevated production of hydroxyeicosatetraenoic acids compared to prostaglandins. MAIN CONCLUSIONS: Our data suggest LD formation and eicosanoid production are distinctly modulated by PUFAS dependent of Leishmania species. In addition, eicosanoid-enzyme mutations are more similar between Leishmania species with same host tropism.

Multifaceted effect of caffeic acid against Streptococcus mutans infection: microbicidal and immunomodulatory agent in macrophages.

The aim of the present study was to investigate the effects of caffeic acid in the interface between the antimicrobial and anti-inflammatory function in macrophage response against S. mutans. S. mutans (108 cfu/mL) were incubated with caffeic acid to determinate the half-maximal inhibitory concentration (IC50) and macrophage cells were incubated with caffeic acid to determinate cell viability and toxicity. Anti-inflammatory effects were measured by nitrite accumulation, TNF-α and PGE2 production, and NF-kB phosphorylation, and S. mutans survival following internalization by macrophages was investigated. We found that caffeic acid presented antimicrobial activity against S. mutans (IC50 = 2.938 ± 0.1225 mM) without exerting cytotoxicity. Caffeic acid inhibited nitrite, TNF-α and PGE2 production by the NF-kB dependent pathway, indicating an immunomodulatory property. Caffeic acid also contributed to macrophage bacteria clearance activity. In summary, caffeic acid presented antimicrobial activity against S. mutans and anti-inflammatory effects in macrophages.

Photosensitizers attenuate LPS-induced inflammation: implications in dentistry and general health.

Antimicrobial photodynamic therapy (aPDT) is a complementary therapeutic modality for periodontal and endodontic diseases, in which Gram-negative bacteria are directly involved. Currently, there are few evidences regarding the effects of aPDT on bacterial components such as lipopolysaccharide (LPS) and it would represent a major step forward in the clinical use of this therapy. In this context, this study aimed to evaluate the efficacy of different photosensitizers (PSs) used in aPDT in LPS inhibition. Four PSs were used in this study: methylene blue (MB), toluidine blue (TBO), new methylene blue (NMB), and curcumin (CUR). Different approaches to evaluate LPS interaction with PSs were used, such as spectrophotometry, Limulus amebocyte lysate (LAL) test, functional assays using mouse macrophages, and an in vivo model of LPS injection. Spectrophotometry showed that LPS decreased the absorbance of all PSs used, indicating interactions between the two species. LAL assay revealed significant differences in LPS concentrations upon pre-incubation with the different PSs. Interestingly, the inflammatory potential of LPS decreased after previous treatment with the four PSs, resulting in decreased secretion of inflammatory cytokines by macrophages. In vivo, pre-incubating curcumin with LPS prevented animals from undergoing septic shock within the established time. Using relevant models to study the inflammatory activity of LPS, we found that all PSs used in this work decreased LPS-induced inflammation, with a more striking effect observed for NMB and curcumin. These data advance the understanding of the mechanisms of LPS inhibition by PSs.

Phospholipids modifications in human hepatoma cell lines (HepG2) exposed to silver and iron oxide nanoparticles.

Metallic nanoparticles such as silver (Ag NPs) and iron oxide (Fe3O4 NPs) nanoparticles are high production volume materials due to their applications in various consumer products, and in nanomedicine. However, their inherent toxicities to human cells remain a challenge. The present study was aimed at combining lipidomics data with common phenotypically-based toxicological assays to gain better understanding into cellular response to Ag NPs and Fe3O4 NPs exposure. HepG2 cells were exposed to different concentrations (3.125, 6.25, 12.5, 25, 50 and 100 µg/ml) of the nanoparticles for 24 h, after which they were assayed for toxic effects using toxicological assays like cytotoxicity, mutagenicity, apoptosis and oxidative stress. The cell membrane phospholipid profile of the cells was also performed using shotgun tandem mass spectrometry. The results showed that nanoparticles exposure resulted in concentration-dependent cytotoxicity as well as reduced cytokinesis-block proliferation index (CBPI). Also, there was an increase in the production of ROS and superoxide anions in exposed cells compared to the negative control. The lipidomics data revealed that nanoparticles exposure caused a modulation of the phospholipidome of the cells. A total of 155 lipid species were identified, out of which the fold changes of 23 were significant. The high number of differentially changed phosphatidylcholine species could be an indication that inflammation is one of the major mechanisms of toxicity of the nanoparticles to the cells.

The inhibition of 5-Lipoxygenase (5-LO) products leukotriene B4 (LTB4) and cysteinyl leukotrienes (cysLTs) modulates the inflammatory response and improves cutaneous wound healing.

To analyze the participation of the enzyme 5-lipoxygenase (5-LO) in skin repair, WT wounds were compared to those in 5-LO deficient mice (5-LO-/-), which presented faster closure and reduced inflammatory infiltrate in the skin, together with increased CD4 regulatory T cells markers in the draining lymph nodes. The 5-LO-/- wounds also had diminished TNF-α, CCL11, CCL7, CCL2, CXCL9, CCR1 and CXCR2 mRNA expression in the lesions, besides differential extracellular matrix remodeling. Furthermore, when cysteinyl leukotriene (cysLT) and leukotriene (LTB4) receptors were antagonized in WT mice, there was a remarkable reduction in TNF-α expression and faster skin healing, similarly to the findings in 5-LO-/- animals. Finally, our results suggested that 5-LO products, in special cysLT and LTB4, underline skin inflammation that follows skin injury and their neutralization may be an important strategy to improve cutaneous healing.

Hyaluronidase decreases neutrophils infiltration to the inflammatory site.

OBJECTIVE: To evaluate the in vivo anti-inflammatory potential of bovine hyaluronidase (HYAL) using two different models of acute inflammation. METHODS: Air pouches were produced in the dorsal subcutaneous of mice and injected with phosphate saline solution or HYAL. The antiinflammatory action of HYAL was evaluated in carrageenan (Cg)-inflamed air pouches. After 4 and 24 h the cellular influx, protein exudation, cytokines and lipid mediators were evaluated. The action of HYAL on the rolling and adhesion of leukocytes was investigated in the LPS-stimulated mesenteric microcirculation by intravital microscopic. RESULTS: Treatment with HYAL reduced the cellular influx and protein exudation in non-inflamed and inflamed air pouches. HYAL treatment of Cg-inflamed air pouch reduced the production of tumor necrosis factor-alpha (TNF-α), interleukin-8 (IL-8), leukotriene B4 (LTB4) and LTC4, whereas prostaglandins E2 (PGE2) and D2 (PGD2) concentrations were unchanged. Histological analyses showed that HYAL administration diminished cell infiltration in the air-pouch lining. In LPS-stimulated mesenteric microcirculation, HYAL usage decreased rolling and adhesion of leukocytes, but did not affect the blood vessels diameters. CONCLUSION: The results demonstrate that HYAL inhibited cellular recruitment, edema formation and pro-inflammatory mediators production, resulting in decreased adherence of leukocytes to blood vessels and tissue infiltration. Our data suggest that HYAL may be considered an effective candidate to ameliorate acute inflammation.

Erythropoietin Exacerbates Inflammation and Increases the Mortality of Histoplasma capsulatum-Infected Mice.

Erythropoietin (EPO) is a key hormone involved in red blood cell formation, but its effects on nonerythroid cells, such as macrophages, have not been described. Macrophages are key cells in controlling histoplasmosis, a fungal infection caused by Histoplasma capsulatum (Hc). Considering that little is known about EPO's role during fungal infections and its capacity to activate macrophages, in this study we investigated the impact of EPO pretreatment on the alveolar immune response during Hc infection. The consequence of EPO pretreatment on fungal infection was determined by evaluating animal survival, fungal burden, activation of bronchoalveolar macrophages, inflammatory mediator release, and lung inflammation. Pretreatment with EPO diminished mononuclear cell numbers, increased the recruitment of F4/80(+)/CD80(+) and F4/80(+)/CD86(+) cells to the bronchoalveolar space, induced higher production of IFN-γ, IL-6, MIP-1α, MCP-1, and LTB4, reduced PGE2 concentration, and did not affect fungal burden. As a consequence, we observed an increase in lung inflammation with extensive tissue damage that might account for augmented mouse mortality after infection. Our results demonstrate for the first time that EPO treatment has a deleterious impact on lung immune responses during fungal infection.

Naphthoquinone derivatives as potential immunomodulators: prospective for COVID-19 treatment.

Inflammation plays a crucial role in COVID-19, and when it becomes dysregulated, it can lead to severe outcomes, including death. Naphthoquinones, a class of cyclic organic compounds widely distributed in nature, have attracted significant interest due to their potential biological benefits. One such naphthoquinone is 3,5,8-trihydroxy-6-methoxy-2-(5-oxohexa-1,3-dienyl)-naphthanthene-1,4-dione (3,5,8-TMON), a compound produced by fungi. Despite its structural similarity to shikonin, limited research has been conducted to investigate its biological properties. Therefore, the objective of this study was to evaluate the effects of 3,5,8-TMON and its synthetic derivatives in the context of inflammation induced by lipopolysaccharide (LPS) and SARS-CoV-2 infection in vitro using cell cultures. 3,5,8-TMON was obtained by acid treatment of crude extracts of fermentation medium from Cordyceps sp., and two derivatives were accessed by reaction with phenylhydrazine under different conditions. The results revealed that the crude extract of the fungi (C. Ex) inhibited the activity of transcription factor NF-kB, as well as the production of nitric oxide (NO) and interleukin-6 (IL-6) when LPS induced it in RAW 264.7 cells. This inhibitory effect was observed at effective concentrations of 12.5 and 3.12 µg mL-1. In parallel, 3,5,8-TMON and the new derivatives 3 and 4 demonstrated the ability to decrease IL-6 production while increasing TNF, with a specific effect depending on the concentration. These concentration-dependent agonist and antagonist effects were observed in THP-1 cells. Furthermore, 3,5,8-TMON inhibited IL-6 production at concentrations of 12.5 and 3.12 µg mL-1 in Calu-3 cells during SARS-CoV-2 viral infection. These findings present promising opportunities for further research into the therapeutic potential of this class of naphthoquinone in the management of inflammation and viral infections.

Immunologic mediators profile in COVID-19 convalescence.

SARS-CoV-2 caused the pandemic situation experienced since the beginning of 2020, and many countries faced the rapid spread and severe form of the disease. Mechanisms of interaction between the virus and the host were observed during acute phase, but few data are available when related to immunity dynamics in convalescents. We conducted a longitudinal study, with 51 healthy donors and 62 COVID-19 convalescent patients, which these had a 2-month follow-up after symptoms recovery. Venous blood sample was obtained from all participants to measure blood count, subpopulations of monocytes, lymphocytes, natural killer cells and dendritic cells. Serum was used to measure cytokines, chemokines, growth factors, anti-N IgG and anti-S IgG/IgM antibodies. Statistic was performed by Kruskal-Wallis test, and linear regression with days post symptoms and antibody titers. All analysis had confidence interval of 95%. Less than 35% of convalescents were anti-S IgM+, while more than 80% were IgG+ in D30. Anti-N IgG decreased along time, with loss of seroreactivity of 13%. Eosinophil count played a distinct role on both antibodies during all study, and the convalescence was orchestrated by higher neutrophil-to-lymphocyte ratio and IL-15, but initial stages were marked by increase in myeloid DCs, B1 lymphocytes, inflammatory and patrolling monocytes, G-CSF and IL-2. Later convalescence seemed to change to cytotoxicity mediated by T lymphocytes, plasmacytoid DCs, VEGF, IL-9 and CXCL10. Anti-S IgG antibodies showed the longest perseverance and may be a better option for diagnosis. The inflammatory pattern is yet present on initial stage of convalescence, but quickly shifts to a reparative dynamic. Meanwhile eosinophils seem to play a role on anti-N levels in convalescence, although may not be the major causative agent. We must highlight the importance of immunological markers on acute clinical outcomes, but their comprehension to potentialize adaptive system must be explored to improve immunizations and further preventive policies.

Cannabidiol reduces lipopolysaccharide-induced nociception via endocannabinoid system activation.

Bacterial infections are often accompanied by fever and generalized muscle pain. However, the treatment of pain with an infectious aetiology has been overlooked. Thus, we investigated the impact of cannabidiol (CBD) in bacterial lipopolysaccharide (LPS)-induced nociception. Male Swiss mice received intrathecal (i.t.) LPS injection, and the nociceptive threshold was measured by the von Frey filaments test. Spinal involvement of the cannabinoid CB2 receptor, toll-like receptor 4 (TLR4), microglia and astrocytes were evaluated by i.t. administration of their respectively antagonists or inhibitors. Western blot, immunofluorescence, ELISA and liquid chromatography-mass spectrometry were used to assess Cannabinoid CB2 receptors and TLR4 spinal expression, proinflammatory cytokines and endocannabinoid levels. CBD was administered intraperitoneally at 10 mg/kg. The pharmacological assay demonstrated TLR4 participation in LPS-induced nociception. In addition, spinal TLR4 expression and proinflammatory cytokine levels were increased in this process. CBD treatment prevented LPS-induced nociception and TLR4 expression. AM630 reversed antinociception and reduced CBD-induced endocannabinoids up-regulation. Increased spinal expression of the cannabinoid CB2 receptor was also found in animals receiving LPS, which was accompanied by reduced TLR4 expression in CBD-treated mice. Taken together, our findings indicated that CBD is a potential treatment strategy to control LPS-induced pain by attenuating TLR4 activation via the endocannabinoid system.

Effects of Moderate-Intensity Training Under Cyclic Hypoxia on Cardiorespiratory Fitness and Hematological Parameters in People Recovered From COVID-19: The AEROBICOVID Study.

BACKGROUND: Recent studies have indicated that people who live at altitude have a lower incidence of coronavirus disease (COVID-19) and lesser severity in infection cases. HYPOTHESIS: Hypoxia exposure could lead to health benefits, and it could be used in the recovery process as an additional stimulus to physical training to improve cardiorespiratory fitness (CRF). STUDY DESIGN: Randomized controlled clinical trial. LEVEL OF EVIDENCE: Level 2. METHODS: The 43 participants, aged 30 to 69 years, were divided into control group (CG, n = 18) and 2 training groups: normoxia (NG, n = 9) and hypoxia (HG, n = 16). Before and after the intervention were evaluated the lactate threshold 2 (L2), peak oxygen uptake (VO2peak), and a blood sample was collected at rest to evaluate hematological adaptation. Both groups performed an 8-week moderate-intensity physical training on a bike. The HG were trained under normobaric hypoxic conditions (fractional inspired oxygen [FiO2] = 13.5%). RESULTS: The 8-week intervention promoted a similar improvement in CRF of people recovered from COVID-19 in the HG (L2 = 34.6%; VO2peak = 16.3%; VO2peak intensity = 24.6%) and NG (L2 = 42.6%; VO2peak = 16.7%; VO2peak intensity = 36.9%). Only the HG presented differences in hematological variables (erythropoietin = 191.7%; reticulocytes = -32.4%; off-score = 28.2%) in comparison with the baseline. CONCLUSION: The results of the present study provide evidence that moderate-intensity training in normoxia or hypoxia promoted similar benefits in CRF of people recovered from COVID-19. Furthermore, the hypoxia offered an additional stimulus to training promoting erythropoietin increase and hematological stimulation. CLINICAL RELEVANCE: The present exercise protocol can be used for the rehabilitation of people recovered from COVID-19, with persistent low CRF. In addition, this is the first study demonstrating that physical training combined with hypoxia, as well as improving CRF, promotes greater hematological stimulation in people recovered from COVID-19.

Anandamide reduces the migration of lymphocytes to the intestine by CB2 activation and reduces TNF-α in the target organs, protecting mice from graft-versus-host disease.

Graft-versus-host disease (GVHD) is a serious inflammatory illness that often occurs as a secondary complication of bone marrow transplantation. Current therapies have limited effectiveness and fail to achieve a balance between inflammation and the graft-versus-tumor effect. In this study, we investigate the effects of the endocannabinoid anandamide on the complex pathology of GVHD. We assess the effects of an irreversible inhibitor of fatty acid amine hydrolase or exogenous anandamide and find that they increase survival and reduce clinical signs in GVHD mice. In the intestine of GVHD mice, treatment with exogenous anandamide also leads to a reduction in the number of CD3+, CD3+CD4+, and CD3+CD8+ cells, which reduces the activation of CD3+CD4+ and CD3+CD8+ cells, as assessed by enhanced CD28 expression, a T cell co-stimulatory molecule. Exogenous AEA was also able to reduce TNF-α and increase IL-10 in the intestine of GVHD mice. In the liver, exogenous AEA reduces injury, TNF-α levels, and the number of CD3+CD8+ cells. Interestingly, anandamide reduces Mac-1α, which lowers the adhesion of transplanted cells in mesenteric veins. These effects are mimicked by JWH133-a CB2 selective agonist-and abolished by treatment with a CB2 antagonist. Furthermore, the effects caused by anandamide treatment on survival were related to the CB2 receptor, as the CB2 antagonist abolished it. This study shows the critical role of the CB2 receptor in the modulation of the inflammatory response of GVHD by treatment with anandamide, the most prominent endocannabinoid.

Febrile response induced by cecal ligation and puncture (CLP) in rats: involvement of prostaglandin E2 and cytokines.

The purpose of the present study was to better understand the events involved in the febrile response induced by cecal ligation and puncture (CLP), a complex infectious process. To this end, we conducted in vivo experiments in rats examining (1) fever development, (2) bacterial number in the infection focus and in blood, (3) peripheral and hypothalamic synthesis of cytokines, (4) hypothalamic and cerebrospinal fluid (CSF) synthesis of prostaglandin E(2) (PGE(2)), (5) the effect of anti-IL-6 antibody on fever, and (6) the effect of celecoxib on fever and hypothalamic synthesis of PGE(2) after CLP induction. We found that CLP promotes fever and animal death depending on the number of punctures. The peak of CLP-induced fever overlapped with the maximal increase in the number of bacteria in the infectious focus and blood, which occurred at 6 and 12 h. The peak of the febrile response also coincided with increased amounts of interleukin (IL)-1ß, IL-6 and IL-10 in the peritoneal exudate and serum; IL-6 in the hypothalamus and PGE(2) in the CSF and predominantly in the hypothalamus. Moreover, intracerebroventricularly injected anti-IL-6 antibody reduced the febrile response while celecoxib reduced the fever and PGE(2) amount in the hypothalamus induced by CLP. Tumor necrosis factor (TNF)-α peaked at 3 h at all sites studied. Conversely, IL-10 concentration decreased in the hypothalamus. These findings show that the peak of CLP-induced fever is accompanied by an increase of bacteria in peritoneal fluid (local infection) and blood; local synthesis of pyrogenic (IL-1ß, IL-6) and antipyretic (IL-10) cytokines and central production of IL-6 and PGE(2), suggesting that these last are the central mediators of this response.

Phospholipids modifications, genotoxic and anticholinesterase effects of pepper fruit (Dennettia tripetala G. Baker) extract in Swiss mice.

The toxicity of D. tripetala fruit extract to mice was investigated using data obtained from lipidomic analyses, comet and Acetylcholinesterase (AChE) assays. Mice (n = 8) were exposed for 30 days via oral gavage to vehicle (5% Tween 80) (negative control), D. tripetala extract (100, 200 and 400 mg/kg) and 40 mg/kg methyl methanesulfonate (MMS) (positive control). The profile of compounds in the fruit extract was analyzed using gas chromatography-mass spectrometry. Out of the total of 32 compounds identified, considerable amount of established insecticidal compounds such as 2-phenylnitroethane, cis-vaccenic acid, linalool and linoleic acid were detected. Fruit extract did not induce DNA damage relative to negative control. Percentage gain in body weights differed significantly across the four weeks. Significantly highest and lowest brain AChE activity was observed in animals exposed to 200 and 400 mg/kg D. tripetala, respectively. Fruit extract modulated the brain phospholipid profile due to significant fold changes of 48 lipid species out of the total of 280 lipid species. High number of differentially expressed phosphatidylcholine (PC) species and significant levels of phosphatidylethanolamine (PE) at 400 mg/kg suggests that activation of inflammation and methylation pathways are the most plausible mechanisms of D. tripetala toxicity to mouse brain tissue.

Bioactive Lipids as Chronic Myeloid Leukemia's Potential Biomarkers for Disease Progression and Response to Tyrosine Kinase Inhibitors.

Chronic myelogenous leukemia (CML) is a myeloproliferative neoplasm that expresses the Philadelphia chromosome and constitutively activated Bcr-Abl tyrosine kinase in hematopoietic progenitor cells. Bcr-Abl tyrosine-kinase inhibitors (TKI) do not definitively cure all CML patients. The efficacy of TKI is reduced in CML patients in the blastic phase-the most severe phase of the disease-and resistance to this drug has emerged. There is limited knowledge on the underlying mechanisms of disease progression and resistance to TKI beyond BCR-ABL1, as well as on the impact of TKI treatment and disease progression on the metabolome of CML patients. The present study reports the metabolomic profiles of CML patients at different phases of the disease treated with TKI. The plasma metabolites from CML patients were analyzed using liquid chromatography, mass spectrometry, and bioinformatics. Distinct metabolic patterns were identified for CML patients at different phases of the disease and for those who were resistant to TKI. The lipid metabolism in CML patients at advanced phases and TKI-resistant patients is reprogrammed, as detected by analysis of metabolomic data. CML patients who were responsive and resistant to TKI therapy exhibited distinct enriched pathways. In addition, ceramide levels were higher and sphingomyelin levels were lower in resistant patients compared with control and CML groups. Taken together, the results here reported established metabolic profiles of CML patients who progressed to advanced phases of the disease and failed to respond to TKI therapy as well as patients in remission. In the future, an expanded study on CML metabolomics may provide new potential prognostic markers for disease progression and response to therapy.

Histoplasma capsulatum cell wall {beta}-glucan induces lipid body formation through CD18, TLR2, and dectin-1 receptors: correlation with leukotriene B4 generation and role in HIV-1 infection.

Histoplasma capsulatum (Hc) is a facultative, intracellular parasite of worldwide significance. Infection with Hc produces a broad spectrum of diseases and may progress to a life-threatening systemic disease, particularly in individuals with HIV infection. Resolution of histoplasmosis is associated with the activation of cell-mediated immunity, and leukotriene B(4) plays an important role in this event. Lipid bodies (LBs) are increasingly being recognized as multifunctional organelles with roles in inflammation and infection. In this study, we investigated LB formation in histoplasmosis and its putative function in innate immunity. LB formation in leukocytes harvested from Hc-infected C57BL/6 mice peaks on day 2 postinfection and correlates with enhanced generation of lipid mediators, including leukotriene B(4) and PGE(2). Pretreatment of leukocytes with platelet-activating factor and BLT1 receptor antagonists showed that both lipid mediators are involved in cell signaling for LB formation. Alveolar leukocytes cultured with live or dead Hc also presented an increase in LB numbers. The yeast alkali-insoluble fraction 1, which contains mainly beta-glucan isolated from the Hc cell wall, induced a dose- and time-dependent increase in LB numbers, indicating that beta-glucan plays a signaling role in LB formation. In agreement with this hypothesis, beta-glucan-elicited LB formation was inhibited in leukocytes from 5-LO(-/-), CD18(low) and TLR2(-/-) mice, as well as in leukocytes pretreated with anti-Dectin-1 Ab. Interestingly, human monocytes from HIV-1-infected patients failed to produce LBs after beta-glucan stimulation. These results demonstrate that Hc induces LB formation, an event correlated with eicosanoid production, and suggest a role for these lipid-enriched organelles in host defense during fungal infection.

Gram-negative periodontal pathogens and bacterial endotoxin in metallic orthodontic brackets with or without an antimicrobial agent: an in-vivo study.

INTRODUCTION: High levels of periodontal pathogens can cause periodontal alterations. The presence of endotoxin might be responsible for the occurrence and progression of tissue inflammation and bone resorption. The aims of this study were to use checkerboard DNA-DNA hybridization and limulus amebocyte lysate assay to evaluate in metallic orthodontic brackets (1) the presence of 16 gram-negative periodontal pathogenic microorganisms of the orange complex and red complex +Treponema socranskii, (2) the amount of bacterial endotoxin, and (3) the efficacy of 0.12% chlorhexidine gluconate mouthwash in reducing bacterial contamination and endotoxin amount. METHODS: Thirty-three patients (ages, 11-33 years) under orthodontic treatment with fixed appliances had 3 new metallic brackets bonded to 3 different premolars. Sixteen patients used a 0.12% chlorhexidine gluconate mouthwash (Periogard, Colgate-Palmolive, São Bernardo do Campo, São Paulo, Brazil) (experimental group), and 17 patients used a placebo mouthwash (control group) twice a week. After 30 days, the brackets were removed, and the samples were obtained. The data were analyzed statistically by Mann-Whitney, Kruskal-Wallis, and Dunn tests (α = 0.05). RESULTS: The 0.12% chlorhexidine gluconate group accumulated significantly lower levels of microorganisms than did the placebo group (P = 0.01). When each microbial complex was analyzed separately, a statistically significant difference between the experimental and control groups was found for the orange complex (P = 0.04). A greater amount of bacterial endotoxin was detected in the 0.12% chlorhexidine gluconate group than in the control group (P = 0.02). CONCLUSIONS: The 0.12% chlorhexidine gluconate oral rinses can be useful to reduce the levels of gram-negative periodontal pathogenic microorganisms in patients with fixed orthodontic appliances. Considering the increased amount of bacterial endotoxin after chlorhexidine gluconate use, further research is necessary to develop clinical procedures or antimicrobial agents with action against bacterial endotoxin adhering to metallic brackets.

Effects of moderate-intensity intermittent hypoxic training on health outcomes of patients recovered from COVID-19: the AEROBICOVID study protocol for a randomized controlled trial.

BACKGROUND: Recent studies point to a lower number and reduced severity of cases in higher altitude cities with decreased oxygen concentration. Specific literature has shown several benefits of physical training, so, in this sense, physical training with hypoxic stimulus appears as an alternative that supports the conventional treatments of the COVID-19 patient's recovery. Thus, this study's primary aim is to analyze the effects of moderate-intensity intermittent hypoxic training on health outcomes in COVID-19 recovered patients. METHODS: A clinical trial controlled double-blind study was designed. Participants (30-69 years old) will be recruited among those with moderate to severe COVID-19 symptoms, approximately 30 days after recovery. They will be included in groups according to the training (T) and recovery (R) association with hypoxia (H) or normoxia (N): (a) TH:RH, (b) TN:RH, (c) TN:RN, and last (d) the control group. The 8-week exercise bike intervention will be carried out with a gradual load increase according to the established periods, three times a week in sets of 5 min, 90 to 100% of the anaerobic threshold (AT), and a 2.5-min break. Blood will be collected for genotyping. First, after 4 weeks (partial), after 8 weeks, and later, 4 weeks after the end of the physical training intervention, participants will perform assessments. The primary outcome is the maximum oxygen consumption (VO2peak). The secondary outcomes include lung function, inflammatory mediators, hematological, autonomic parameters, AT, body composition analysis, quality of life, mental health, anthropometric measurements, and physical fitness. The statistical analysis will be executed using the linear regression model with mixed effects at a 5% significance level. DISCUSSION: This study is designed to provide evidence to support the clinical benefits of moderate-intensity intermittent hypoxic training as a part of the treatment of patients recovered from COVID-19. It may also provide evidence on the efficacy and safety of intermittent hypoxic training in different health conditions. Lastly, this study presents an innovative strategy enabling up to 16 participants in the same training session. TRIAL REGISTRATION: ClinicalTrials.gov RBR-5d7hkv. Registered after the start of inclusion on 3 November 2020 with the Brazilian Clinical Trials Registry.