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Background The thymus is responsible for thymocyte differentiation into immunocompetent T lymphocytes. Different cell types in the thymic microenvironment actively cooperate in this process, interacting with the developing thymocytes through soluble factors, extracellular matrix (ECM) molecules, and receptors. In addition, this microenvironment can be influenced by several factors, such as semaphorin-3A (Sema3A), which is a multifunctional protein involved in cell migration. We evaluated the Sema3A effects on the cellular parameters and functional features of thymic stromal cells. Methods Thymic stromal cells were obtained by enzymatic digestion of the murine thymus. These cells were treated with Sema3A and evaluated as follows: cell morphology by scanning electron microscope, F-actin cytoskeleton and deposition of ECM molecules by fluorescence microscopy, and adhesion assays with freshly obtained thymocytes. Results The obtained thymic stroma was composed of 67% of thymic epithelial cells (TECs), and 90% of the TECs were positive for the Sema3A receptor neuropilin-1. These cells secreted CXCL12, IL-7 and extended thymocyte survival. Sema3A changed the morphology of thymic stromal cells and promoted F-actin reorganization. In addition, the fibronectin fibers were reoriented, and the laminin production was increased in Sema3A-treated thymic stromal cells. In the adhesion assays, there was an increase in the number of adhered thymocytes when thymic stromal cells were pretreated with Sema3A. Conclusion Our data strongly suggest the active participation of Sema3A in thymic physiology, highlighting its role as an immunomodulatory molecule. This may provide important knowledge for understanding the interactions of thymic cells.

Bowdichia virgilioides Kunth (Fabaceae) is a tree species used in folk medicine against inflammatory disorders, and it is still unclear whether consumption of the B. virgilioides stem bark extract has any effect on immune cells. Thus, this study aimed to evaluate the effects of B. virgilioides stem bark extract (AEBv) on the functions of the thymus and B lymphocytes. The AEBv was obtained by decoction and administrated orally once a day in male mice for 7 days. After treatment, thymi were obtained for morphology analysis, relative weight, cellularity and deposition of laminin. Thymocytes obtained from control and treated animals were submitted to adhesion and migration assays followed by immunophenotyping. Also, mesenteric lymph nodes were removed, and the lymphoid cells were obtained to measure B cell proliferation and cytokine production. Mice treated with AEBv and their controls were also immunized with sheep red blood cells (SRBC), and 8 days post immunization the sera were collected to perform antibody titration using hemagglutination assays. AEBv administration decreased thymus weight and cellularity and increased the laminin deposition without any significant effect on the expression of the laminin receptor VLA-6 in the thymus microenvironment. Additionally, thymocytes of AEBv-treated mice showed reduced adhesion and migration on laminin substrates, and the B lymphocytes showed reduced proliferation, enhanced IL-10 secretion, decreased TNF-α production, and attenuated anti-SRBC antibody production. These findings showed that AEBv intake might cause modulatory effects on the functions of thymocytes and B-lymphocytes, contributing to the treatment of immune diseases. Key words: Thymocytes, cell migration, cell adhesion, immunosuppression, B cell.

Background: Age-related impairments in macrophage functions have important consequences for the health of the elderly population. The aging process is also accompanied by a reduction in several hormones, including growth hormone (GH). Previous studies have shown that this hormone can affect macrophage activity in young individuals; however, the biological effects of GH stimulation on macrophages during aging have not yet been elucidated. Objective: The aim of this work was to investigate the in vitro effects of GH on peritoneal macrophages from aged mice. Methods: Peritoneal macrophages isolated from young (4 months-old) and old (12-15 months-old) mice were treated in vitro with 100 ng/mL of GH for 24 hours. After treatment, cells were analysed for cell morphology, reactive oxygen species (ROS) production, expression of integrins, cell adhesion to extracellular matrix molecules, and migration in transwell chambers. Results: Although GH-treated cells from old mice exhibited decreased ROS production, we did not observe the effects of GH on macrophage morphology or macrophage phagocytic activity in young and old mice-derived cell cultures. Macrophages from old mice had increased adhesion to laminin and fibronectin substrates, as did cells obtained from young mice treated with GH, but no change was observed in the expression of integrin receptors. Furthermore, cells from old mice exhibited increased migration compared to young mice and a significant increase in macrophage migration was observed under GH stimulation. Conclusion: Our results showed that GH can interfere with the motility of macrophages from old mice, advancing our understanding of the interactions between the immune and neuroendocrine systems during aging.

The prolactin hormone (PRL), in addition to its known effects on breast development and lactation, exerts effects on the immune system, including pleiotropic effects on the thymus. The aim of this study was to evaluate the influence of PRL on the epithelial compartment of the thymus. Thymic epithelial cells (TECs) (2BH4 cells) and fresh thymocytes were used. Immunofluorescence assay revealed that PRL treatment (10 ng/ mL) increases the deposition of laminin and expression of the chemokine CXCL12 in 2BH4 cells. However, no change was observed in the deposition of fibronectin. Moreover, PRL altered F-actin polymerisation, allowing the formation of focal adhesion complexes in treated cells. When 2BH4 cells were pre-treated with PRL, thymocyte adhesion was not altered. However, in the cell migration assay, pre-treatment with PRL potentiated the chemotactic effect of CXCL12 on the migration of total, double-positive, CD4-positive, and CD8-positive thymocytes. Together, the results of this study demonstrate the effect of PRL on thymic epithelial cells, particularly on CXCL12-driven thymocyte migration, confirming that this hormone is a regulator of thymic physiology.

Growth hormone (GH), in addition to its classic actions on growth and metabolism in the body, exerts pleiotropic effects on the immune system, particularly on the thymus. The aim of this study was to evaluate the influence of GH on the interactions between mature thymocytes and the thymic endothelium involved in the migratory process. To this end, fresh thymocytes (C57BL/6 mice) and the thymic endothelial cell line (tEnd.1) were used. In the cell adhesion assay, the GH-treated thymocytes adhered more to tEnd.1 cells. Additionally, there was an improvement in the deposition of fibronectin by tEnd.1 cells when co-cultured with GH-pre-treated thymocytes. Furthermore, GH induced thymocyte F-actin polymerization. In the transendothelial migration assay, a large number of GH-treated thymocytes, mainly the CD4-CD8+ subset, migrated towards the endothelium under the stimulus of insulin-like growth factor 1. In conclusion, we demonstrated the positive actions of GH in thymocyte/thymic endothelium interactions, including transendothelial migration.

Growth hormone (GH) plays a physiological role in the immune system. In macrophages, GH enhances the production of hydrogen peroxide, superoxide anions, nitric oxide, cytokines, and chemokines, including interferon-γ and macrophage inflammatory protein-1α. However, some of the effects of GH stimulation on the biological functions of macrophages remain to be elucidated. Herein, we showed that in vivo GH treatment resulted in decreased expression of VLA-5 and VLA-6 integrins on the macrophage surface, accompanied by a reduction in macrophage adhesion to extracellular matrix (ECM) ligands, fibronectin, and laminin. Additionally, a decrease in macrophage adhesion to laminin was observed when the cells were treated in vitro with GH. In transwell migration assays, GH-treated macrophages showed increased migration after 6 h. Although in vitro GH treatment did not influence the phagocytic activity of macrophages, when the treatment was performed in vivo, peritoneal macrophages from GH-treated mice showed a higher percentage of phagocytosis and higher phagocytic capacity than cells from control animals. These results led us to analyse the role of insulin-like growth factor-1 (IGF-1), a GH stimulated factor, on macrophage phagocytosis. We observed an increase in phagocytic activity when J774 murine macrophages were treated with IGF-1 for 24 h. Our results revealed an important role for GH in resident macrophage migration and phagocytic activity. Specifically, we demonstrate that IGF-1 may be the GH stimulated factor that induces macrophage phagocytosis in vivo.

The generation of new blood vessels is a complex process mediated by a variety of growth factors, and the growth hormone (GH) has been shown to act as a proangiogenic factor. In fact, human GH deficiency or excess are associated with endothelial dysfunction. Moreover, mouse models have revealed the action of GH in both tissue repair and in the microvascular circulation of normal tissues. In this study, we investigated the in vitro effects of GH on endothelial cells. Using a murine endothelioma cell line (tEnd.1), we demonstrated that GH has a mitogenic effect. The hormone also affected the endothelial cellular morphology and augmented the deposition of the extracellular matrix molecules, laminin, and fibronectin, on tEnd.1 surface. GH could stimulate tEnd.1 cell fugetaxis, in transwell chambers migration assay, and increased the formation of capillary-like structures in Matrigel®-coated plates. Given the important role of angiogenesis during tissue injury, for example, at ischemic lesions, these findings shed light on therapeutic angiogenesis, particularly in pathologies where the cardiovascular system has been compromised.

Growth hormone (GH) plays a physiological role in the immune system. In macrophages, GH enhances the production of hydrogen peroxide, superoxide anions, nitric oxide, cytokines, and chemokines, including interferon-γ and macrophage inflammatory protein-1α. However, some of the effects of GH stimulation on the biological functions of macrophages remain to be elucidated. Herein, we showed that in vivo GH treatment resulted in decreased expression of VLA-5 and VLA-6 integrins on the macrophage surface, accompanied by a reduction in macrophage adhesion to extracellular matrix (ECM) ligands, fibronectin, and laminin. Additionally, a decrease in macrophage adhesion to laminin was observed when the cells were treated in vitro with GH. In transwell migration assays, GH-treated macrophages showed increased migration after 6 h. Although in vitro GH treatment did not influence the phagocytic activity of macrophages, when the treatment was performed in vivo, peritoneal macrophages from GH-treated mice showed a higher percentage of phagocytosis and higher phagocytic capacity than cells from control animals. These results led us to analyse the role of insulin-like growth factor-1 (IGF-1), a GH stimulated factor, on macrophage phagocytosis. We observed an increase in phagocytic activity when J774 murine macrophages were treated with IGF-1 for 24 h. Our results revealed an important role for GH in resident macrophage migration and phagocytic activity. Specifically, we demonstrate that IGF-1 may be the GH stimulated factor that induces macrophage phagocytosis in vivo.

Current therapeutic for the treatment of anxiety is associated with a wild variety of side effects. The traditional use of plant extract to health care can indicate an important source of new pharmaceuticals. Bowdichia virgilioides Kunth, Fabaceae, is a plant commonly employed in the Brazilian folk medicine to treat infl ammatory conditions. Nevertheless, despite its popular use there are no studies related to its possible neuropharmacological effect. Here, we investigated the possible anxiolytic effect of the extract of B. virgilioides after acute and sub-chronic treatment in mice. The aqueous extract from the stem barks of B. virgilioides (20, 200 or 400 mg/kg) was orally administered, and its anxiolytic effect was evaluated in the elevated plus maze, open-fi eld and rota-rod tests. Diazepam was employed as standard drug. The aqueous extract treatment was effective in inducing anxiolytic effects with single acute treatment, a phenomenon that remained after chronic treatment. However, no changes in spontaneous locomotor activity or myorelaxant effect after aqueous extract treatment. The extract was either safe with no deaths in mice treated orally with 1000 mg/kg. These fi ndings suggest that the aqueous extract from the stem barks of Bowdichia virgilioides has an acute and sub-chronic anxiolytic-like effect without compromising motor activity, demonstrating an advantage regarding to antidepressant drugs. Keywords: Bowdichia virgilioides, anxiety, elevated plus maze, open field, rota rod

Nanoparticles have extremely wide applications in the medical and biological fields. They are being used in biosensors, local drug delivery, diagnostics, and medical therapy. However, the potential effects of nanoparticles on target cell and tissue function, apart from cytotoxicity, are not completely understood. Thus, the aim of this study was to investigate the in vitro effects of silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs) on human fibroblasts with respect to their interaction with the extracellular matrix and in cell migration. Immunofluorescence analysis revealed that treatment with AgNPs or AuNPs decreased collagen and laminin production at all the concentrations tested (0.1, 1, and 10 μg/mL). Furthermore, cytofluorometric analysis showed that treatment with AgNPs reduced the percentage of cells expressing the collagen receptor very late antigen 2, α2β1 integrin (VLA-2) and the laminin receptor very late antigen 6, α6β1 integrin (VLA-6). In contrast, AuNP treatment increased and decreased the percentages of VLA-2-positive and VLA-6-positive cells, respectively, as compared to the findings for the controls. Analysis of cytoskeletal reorganization showed that treatment with both types of nanoparticles increased the formation of stress fibres and number of cell protrusions and impaired cell polarity. Fibroblasts exposed to different concentrations of AuNPs and AgNPs showed reduced migration through transwell chambers in the functional chemotaxis assay. These results demonstrated that metal nanoparticles may influence fibroblast function by negatively modulating the deposition of extracellular matrix molecules (ECM) and altering the expression of ECM receptors, cytoskeletal reorganization, and cell migration. Electronic supplementary material The online version of this article (doi:10.1186/s11671-017-1982-3) contains supplementary material, which is available to authorized users.

The generation of new blood vessels is a complex process mediated by a variety of growth factors, and the growth hormone (GH) has been shown to act as a proangiogenic factor. In fact, human GH deficiency or excess are associated with endothelial dysfunction. Moreover, mouse models have revealed the action of GH in both tissue repair and in the microvascular circulation of normal tissues. In this study, we investigated the in vitro effects of GH on endothelial cells. Using a murine endothelioma cell line (tEnd.1), we demonstrated that GH has a mitogenic effect. The hormone also affected the endothelial cellular morphology and augmented the deposition of the extracellular matrix molecules, laminin, and fibronectin, on tEnd.1 surface. GH could stimulate tEnd.1 cell fugetaxis, in transwell chambers migration assay, and increased the formation of capillary-like structures in Matrigel®-coated plates. Given the important role of angiogenesis during tissue injury, for example, at ischemic lesions, these findings shed light on therapeutic angiogenesis, particularly in pathologies where the cardiovascular system has been compromised.

Background Thymic involution is thought to be an important factor of age related immunodeficiency. Understanding the molecular mechanisms of human thymic senescence may lead to the discovery of novel therapeutic approaches aimed at the reestablishment of central and peripheral T cell repertoire. Results As an initial approach, here we report that the decline of human thymic FOXN1 transcription correlates with age, while other genes, DLL1, DLL4 and WNT4, essential for thymopoiesis, are constitutively transcribed. Using a human thymic epithelial cell line (hTEC), we show that FOXN1 expression is refractory to signals that induce FOXN1 transcription in primary 3D culture conditions and by stimulation of the canonical WNT signaling pathway. Blockage of FOXN1 induceability in the hTEC line may be mediated by an epigenetic mechanism, the CpG methylation of the FOXN1 gene. Conclusion We showed a suppression of FOXN1 transcription both in cultured human thymic epithelial cells and in the aging thymus. We hypothesize that the underlying mechanism may be associated with changes of the DNA methylation state of the FOXN1 gene. Electronic supplementary material The online version of this article (doi:10.1186/s12979-015-0045-9) contains supplementary material, which is available to authorized users.