Intrathymic somatotropic circuitry: consequences upon thymus involution.

Growth hormone (GH) is a classic pituitary-derived hormone crucial to body growth and metabolism. In the pituitary gland, GH production is stimulated by GH-releasing hormone and inhibited by somatostatin. GH secretion can also be induced by other peptides, such as ghrelin, which interacts with receptors present in somatotropic cells. It is well established that GH acts directly on target cells or indirectly by stimulating the production of insulin-like growth factors (IGFs), particularly IGF-1. Notably, such somatotropic circuitry is also involved in the development and function of immune cells and organs, including the thymus. Interestingly, GH, IGF-1, ghrelin, and somatostatin are expressed in the thymus in the lymphoid and microenvironmental compartments, where they stimulate the secretion of soluble factors and extracellular matrix molecules involved in the general process of intrathymic T-cell development. Clinical trials in which GH was used to treat immunocompromised patients successfully recovered thymic function. Additionally, there is evidence that the reduction in the function of the somatotropic axis is associated with age-related thymus atrophy. Treatment with GH, IGF-1 or ghrelin can restore thymopoiesis of old animals, thus in keeping with a clinical study showing that treatment with GH, associated with metformin and dehydroepiandrosterone, could induce thymus regeneration in healthy aged individuals. In conclusion, the molecules of the somatotrophic axis can be envisioned as potential therapeutic targets for thymus regeneration in age-related or pathological thymus involution.

IGF-1 increases survival of CD4+ lineage in a 2D model of thymocyte/thymic stromal cell co-culture.

Insulin-like growth factor-1 (IGF-1), in addition to its classic effects on cell proliferation and organism growth, has pleiotropic actions on the immune system, particularly on the thymus. Thus, the objective of this study was to evaluate the influence of IGF-1 on molecules involved in the survival of thymocytes in vitro using a co-culture system with thymic stromal cells obtained from C57BL/6 mice. The obtained thymic stroma has contained thymic epithelial cells, macrophages, dendritic cells, fibroblasts, and preserved the expression of the major histocompatibility complex (MHC) molecules. Fresh thymocytes were added to these cultures and the co-culture were treated daily with IGF-1 (100 ng/mL) for 3 days. In this scheme, the viability of the thymocytes was about 70%, either in the control (non-treated cells) or in the IGF-1-treated cultures. It was found that IGF-1 was able to increase the percentage of thymocytes from the CD4+ single-positive (SP) subset. This result was accompanied by an increase in the MHC II expression on thymic stromal cells and an augment on the interleukin-7 receptor (CD127) on the surface of the CD4 SP thymocytes after treatment with IGF-1. Finally, IGF-1 treatment increased the expression of the ThPOK encoding gene Zbtb7b, which is involved in the differentiation of CD4+ SP thymocytes. Our study demonstrates the participation of IGF-1 in the thymocyte/thymic stroma interactions, especially in the extended survival of the CD4+ lineage in the thymus.

Growth Hormone Stimulates Murine Macrophage Migration during Aging.

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 Responsiveness of Thymic Stromal Cells to semaphorin-3A.

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.

CXCL12-driven thymocyte migration is increased by thymic epithelial cells treated with prolactin in vitro.

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.