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Abstract The tumor microenvironment (TME) harbors several cell types, such as tumor cells, immune cells, and non-immune cells. These cells communicate through several mechanisms, such as cell-cell contact, cytokines, chemokines, and extracellular vesicles (EVs). Tumor-derived vesicles are known to have the ability to modulate the immune response. Monocytes are a subset of circulating innate immune cells and play a crucial role in immune surveillance, being recruited to tissues where they differentiate into macrophages. In the context of tumors, it has been observed that tumor cells can attract monocytes to the TME and induce their differentiation into tumor-associated macrophages with a pro-tumor phenotype. Tumor–derived EVs have emerged as essential structures mediating this process. Through the transfer of specific molecules and signaling factors, tumor-derived EVs can shape the phenotype and function of monocytes, inducing the expression of cytokines and molecules by these cells, thus modulating the TME towards an immunosuppressive environment.

Background: Obesity poses a significant global health challenge, given its association with the excessive accumulation of adipose tissue (AT) and various systemic disruptions. Within the adipose microenvironment, expansion and enrichment with immune cells trigger the release of inflammatory mediators and growth factors, which can disrupt tissues, including bones. While obesity’s contribution to bone loss is well established, the direct impact of obese AT on osteoblast maturation remains uncertain. This study aimed to explore the influence of the secretomes from obese and lean AT on osteoblast differentiation and activity. Methods: SAOS-2 cells were exposed to the secretomes obtained by culturing human subcutaneous AT from individuals with obesity (OATS) or lean patients, and their effects on osteoblasts were evaluated. Results: In the presence of the OATS, mature osteoblasts underwent dedifferentiation, showing an increased proliferation accompanied by a morphological shift towards a mesenchymal phenotype, with detrimental effects on osteogenic markers and the calcification capacity. Concurrently, the OATS promoted the expression of mesenchymal and adipogenic markers, inducing the formation of cytoplasmic lipid droplets in SAOS-2 cells exposed to an adipogenic differentiation medium. Additionally, TGF-β1 emerged as a key mediator of these effects, as the OATS was enriched with this growth factor. Conclusions: Our findings demonstrate that obese subcutaneous AT promotes the dedifferentiation of osteoblasts and increases the adipogenic profile in these cells.

Obesity is an alarming disease that favors the upset of other illnesses and enhances mortality. It is spreading fast worldwide may affect more than 1 billion people by 2030. The imbalance between excessive food ingestion and less energy expenditure leads to pathological adipose tissue expansion, characterized by increased production of proinflammatory mediators with harmful interferences in the whole organism. Bone tissue is one of those target tissues in obesity. Bone is a mineralized connective tissue that is constantly renewed to maintain its mechanical properties. Osteoblasts are responsible for extracellular matrix synthesis, while osteoclasts resorb damaged bone, and the osteocytes have a regulatory role in this process, releasing growth factors and other proteins. A balanced activity among these actors is necessary for healthy bone remodeling. In obesity, several mechanisms may trigger incorrect remodeling, increasing bone resorption to the detriment of bone formation rates. Thus, excessive weight gain may represent higher bone fragility and fracture risk. This review highlights recent insights on the central mechanisms related to obesity-associated abnormal bone. Publications from the last ten years have shown that the main molecular mechanisms associated with obesity and bone loss involve: proinflammatory adipokines and osteokines production, oxidative stress, non-coding RNA interference, insulin resistance, and changes in gut microbiota. The data collection unveils new targets for prevention and putative therapeutic tools against unbalancing bone metabolism during obesity.

Immune system cells, including neutrophils, are recruited by the tumor microenvironment as a site of chronic inflammation and begin to favor tumor growth. Neutrophils present in the tumor site are called tumor-associated neutrophils (TAN) and can present two phenotypes: N1 (antitumor) or N2 (pro-tumor). Evidence shows the high capacity of immune system cells to interact with extracellular vesicles (Evs) released by tumor cells. Evs can modulate the phenotype of cells within the immune system, contributing to tumor development. Here, we investigated the role of MDA-MB-231-derived Evs upon the polarization of neutrophils towards an N2 phenotype and the underlying mechanisms. We observed that neutrophils treated with Evs released by MDA cells (MDA-Evs) had their half-life increased, increased their chemotactic capacity, and released higher levels of NETs and ROS than neutrophils treated with non-tumoral Evs. We also observed that neutrophils treated with MDA-Evs released increased IL-8, VEGF, MMP9, and increased expression of CD184, an N2-neutrophil marker. Finally, neutrophils treated with MDA-Evs increased tumor cell viability. Our results show that MDA-Evs induce an N2-like phenotype, and the blockage of phosphatidylserine by annexin-V may be an essential agent counter-regulating this effect.

The significant incidence of deforestation in South America culminates in the contact of humans with typical forests species. Among these species, one may highlight Lonomia obliqua caterpillar, which, when touched by humans, can poison them through their bristles. Therefore, better acknowledging the mechanisms involved in envenomation caused by Lonomia obliqua caterpillar bristle extract (LOCBE) may contribute to further treatments. Recently, we demonstrated that LOCBE induces a pro-inflammatory profile in endothelial cells; thus, we decided to investigate the effects of LOCBE on human polymorphonuclear neutrophils (PMN), which are the first leukocytes that migrate to the inflammatory focus. Our results showed that treatment with LOCBE induced PMN chemotaxis together with alterations in actin cytoskeleton and focal adhesion kinase (FAK) activation, favoring migration. Concurrently, LOCBE induced PMN adhesion to matrix proteins, such as collagen IV, fibronectin, and fibrinogen. Moreover, we observed that LOCBE attenuated PMN apoptosis and increased reactive oxygen species (ROS) production together with nuclear factor kB (NF-κB) activation—a redox-sensitive transcription factor—as well as interleukin (IL)-1β and IL-8 release. We call attention to the ROS-dependent effect of LOCBE on increased cell migration once an antioxidant treatment reverted it. In summary, we report that LOCBE activates PMN, inducing pro-inflammatory responses modulated by ROS.

The data provides information in support of the research article Moraes et al., Atherosclerosis 243(2) (2015) 477–485 [1]. Here we provide data behind the mechanisms involved in Angiotensin II (Ang II) effects on vascular smooth muscle cells (VSMC). Ang II-induced VSMC ROS production is modulated by alpha1beta1 integrin. Ang II also stimulates ROS production in VSMC via p47phox, a NOX2 subunit. Furthermore, Ang II effect on VSMC migration was also inhibited by NOX2 inhibitor. We showed that obtustatin, alpha1beta1 integrin blocker, inhibited Ang II effect on p47phox activation. Ang II effect on ROS production is also PI3K dependent. Finally we showed that NOX1 and Integrin-Linked-Kinase (ILK) are crucial to NOX2 activation. The research provides information about the sequential events of NOX1/alpha1beta1 integrin/ILK/NOX2 in Ang II effects on VSMC.

In obesity, the dysfunctional adipose tissue (AT) releases increased levels of proinflammatory adipokines such as TNFα, IL-6, and IL-1β and free fatty acids (FFAs), characterizing a chronic, low-grade inflammation. Whilst FFAs and proinflammatory adipokines are known to elicit an inflammatory response within AT, their relative influence upon preadipocytes, the precursors of mature adipocytes, is yet to be determined. Our results demonstrated that the conditioned medium (CM) derived from obese AT was rich in FFAs, which guided us to evaluate the role of TLR4 in the induction of inflammation in preadipocytes. We observed that CM derived from obese AT increased reactive oxygen species (ROS) levels and NF-ĸB nuclear translocation together with IL-6, TNFα, and IL-1β in 3T3-L1 cells in a TLR4-dependent manner. Furthermore, TLR4 signaling was involved in the increased expression of C/EBPα together with the release of leptin, adiponectin, and proinflammatory mediators, in response to the CM derived from obese AT. Our results suggest that obese AT milieu secretes lipokines, which act in a combined paracrine/autocrine manner, inducing inflammation in preadipocytes via TLR4 and ROS, thus creating a paracrine loop that facilitates the differentiation of adipocytes with a proinflammatory profile.

Hepcidin is a key hormone that induces the degradation of ferroportin (FPN), a protein that exports iron from reticuloendothelial macrophages and enterocytes. The aim of the present study was to experimentally evaluate if the obesity induced by a high-fat diet (HFD) modifies the expression of FPN in macrophages and enterocytes, thus altering the iron bioavailability. In order to directly examine changes associated with iron metabolism in vivo, C57BL/6J mice were fed either a control or a HFD. Serum leptin levels were evaluated. The hepcidin, divalent metal transporter-1 (DMT1), FPN and ferritin genes were analyzed by real-time polymerase chain reaction. The amount of iron present in both the liver and spleen was determined by flame atomic absorption spectrometry. Ferroportin localization within reticuloendothelial macrophages was observed by immunofluorescence microscopy. Obese animals were found to exhibit increased hepcidin gene expression, while iron accumulated in the spleen and liver. They also exhibited changes in the sublocation of splenic cellular FPN and a reduction in the FPN expression in the liver and the spleen, while no changes were observed in enterocytes. Possible explanations for the increased hepcidin expression observed in HFD animals may include: increased leptin levels, the liver iron accumulation or endoplasmic reticulum (ER) stress. Together, the results indicated that obesity promotes changes in iron bioavailability, since it altered the iron recycling function.

Envenomation caused by human contact with the caterpillar Lonomia is characterized by deleterious effects on coagulation and patency of blood vessels. The cellular effects induced by Lonomia obliqua venom highlights its capacity to activate endothelial cells, leading to a proinflammatory phenotype. Having more knowledge about the mechanisms involved in envenomation may contribute to better treatment. We aimed to evaluate the effects of Lonomia obliqua caterpillar bristle extract (LOCBE) on vascular smooth muscle cells (VSMC). We observed that LOCBE induced VSMC migration, which was preceded by alterations in actin cytoskeleton dynamics and Focal Adhesion Kinase activation. LOCBE also induced Extracellular Signal-Regulated Kinase (ERK) phosphorylation in VSMC, and the inhibition of this pathway impaired cell proliferation. Stimulation of VSMC with LOCBE triggered reactive oxygen species (ROS) production through the activation of NADPH oxidase. The rapid increase in these ROS further induced mitochondrial ROS production, however only NADPH oxidase-derived ROS were involved in ERK activation in VSMC. We that demonstrated the chemotactic and proliferative effects of LOCBE on VSMC were dependent on ROS production, mainly through NADPH oxidase. Together, the data show that Lonomia obliqua venom can interact with and activate VSMC. These effects rely on ROS production, suggesting new potential targets for treatment against vascular damage during envenomation.

Alveolar macrophages play a central role in the pathogenesis of chronic obstructive pulmonary disease (COPD). Monocytes are recruited from blood during inflammation and then mature into alveolar macrophages. The aim of this study was to investigate the effect of cigarette smoke (CS) at different times in lung macrophages and monocytes from blood and bone marrow in mice. Male mice (C57BL/6, n = 45) were divided into groups: control, CS 5 days, CS 14 days and CS 30 days. Five days’ CS exposure induced a pronounced influx of neutrophils and macrophages in the lung associated with increased levels of keratinocyte chemoattractant (KC), tumor necrosis factor-α (TNF-α), nitric oxide (NO) and matrix metalloproteinase (MMP)-12. After 14 days of CS exposure, neutrophil recruitment and cytokine production were greatly reduced. Moreover, chronic CS exposure led to increased recruitment of macrophages (with high expression of CD206), transforming growth factor-β (TGF-β) production as well as no detection of TNF-α, interleukin (IL)-6 and KC. CS can also change the monocyte phenotype in the blood and bone marrow, with an increase in Ly6Clow cells. These results show for the first time that CS can change not only macrophage polarization but also monocyte. These results suggest that continued recruitment of Ly6Clow monocytes may help the distinct renewing macrophage M2 population required for COPD progression.

We investigated the consequences of mild maternal malnutrition in rat dams, in terms of thymocyte responses and the putative role of leptin. The young progeny of dams submitted to protein deprivation (PD) during lactation showed at 30 days of age lower body and thymus weights, significant alterations in CD4/CD8-defined T cell subsets without modifications in total thymocyte number as well as in proliferative response. Despite, the rats from PD group did not present alterations in leptin circulating levels, the expression of leptin receptor ObRb was enhanced in their thymocytes. This change was accompanied by an increase in leptin signaling response of thymocytes from PD rats, with an increase in JAK2 and STAT3 phosphorylation after leptin stimulation. Thymocytes from PD rats also presented a decreased rate of spontaneous apoptosis when compared to controls. Accordingly, higher expression of anti-apoptotic protein Bcl-2, and lower of pro-apoptotic protein Bax, with no change of pro-apoptotic Bad, and higher pro-caspase 3 content were detected in PD thymocytes. Moreover, thymocytes from PD group exhibited a constitutive higher nuclear content of p65 NF-kB associated to a lower IkB content in the cytoplasm. Finally, although there was no change in ob gene expression in PD thymocytes, a higher mRNA expression for the Ob gene was observed in the thymic microenvironment from PD animals. Taken together, the results show that mild maternal protein deprivation during lactation affects thymic homeostasis, enhancing leptin activity, which in turn protects thymocytes from apoptosis in the young progeny, with possible consequences upon the immune response of these animals in adult life.

Human overnutrition has caused a rise in the prevalence of obesity in recent years. In addition to the deleterious effects of obesity during childhood, long-term effects in adulthood have been described as well. For instance, cardiovascular diseases and type 2 diabetes are among the diseases associated with a history of obesity. Altered insulin secretion and action have been described as important links between these diseases and obesity. Insulin acts as a unique anabolic hormone providing regulation of whole-body glucose homeostasis and peripheral tissue glucose uptake in tissues such as the heart. In this study, we examined insulin signaling in the heart of obese animals using an experimental model of inducing overweight adult animals by overnutrition in early life. In these animals, overfeeding during lactation was able to induce a significant increase in body weight starting at the 10th day of life, and this increased weight persisted until adulthood. Impairment in glucose tolerance, hyperinsulinemia, and an increased insulin/glucose ratio were also observed in these animals. Moreover, an increased heart weight/tibia length ratio was also observed, indicating an enlarged heart size. The overfed animals also had decreased insulin sensitivity in the heart, as confirmed by decreased insulin receptor (IR)-β and IR substrate-1 (Irs1) phosphorylation, increased phosphatase, non-receptor type 1 (Ptpn1)–IR-β association, decreased –Irs1-associated activity, and reduction in anti-phospho Akt1 phosphorylation. In conclusion, our findings showed that overnutrition during early life induced obesity and insulin resistance in the adult offspring, and further increased heart size and impaired cardiac insulin signaling, putatively due to an increase in Ptpn1 activity. [ABSTRACT FROM AUTHOR]

Recently, we demonstrated high serum leptin and 25(OH)D (calcidiol) in obese animals, with high C/EBP β and PPAR γ expression in adipose tissue. Since the role of vitamin D in adipogenesis remains controversial and hyperleptinemia is found in obesity, we asked if leptin could interfere in vitamin D action on adipocytes. Here, we studied the direct effect of these two hormones upon 3T3L1 preadipocytes incubated with or without 1,25(OH)2D (100 nM, 24 h) and with leptin (10 −7  M, 4 h later). RT-PCR (VDR and Cyp27b1/1α-hydroxylase ), western blotting (VDR, Cyp27b1/1α-hydroxylase, ObR-b, C/EBP β , PPAR γ and Bax content), a cell proliferation assay and an Annexin V-FITC binding assay were performed. Incubation with 1,25(OH)2D decreased Cyp27b1/1α-hydroxylase and VDR. Co-incubation of 1,25(OH)2D and leptin did not change Cyp27b1/1α-hydroxylase and had no additive effect upon the decreased VDR mRNA. Incubation with 1,25(OH)2D decreased C/EBP β and PPARγ. In the cell proliferation assay, 1,25(OH)2D decreased the number of 3T3L1 cells. No changes in OBR-b or apoptotic parameters (Bax and annexin-V) were observed. The 1,25(OH)2D decreased pro-adipogenic factors and proliferation of adipocytes. However, since it inhibits the conversion of 25(OH)D to 1,25(OH)2D and VDR mRNA long-term, it could decrease the vitamin D response in adipocytes, leading to greater adipogenesis. The co-incubation of both hormones, simulating what occurs in obesity, even neutralizing the effect on Cyp27b1/ 1α- hydroxylase , did not change the vitamin D sensitivity but decreased SOCS-3 and pSTAT-3. Thus, an excess of vitamin D and hyperleptinemia could decrease vitamin D sensitivity in adipocytes, contributing to increased adipogenesis. [ABSTRACT FROM AUTHOR]