ABSTRACT
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.
Subject(s)
Insulin-Like Growth Factor I , Thymocytes , Mice , Animals , Insulin-Like Growth Factor I/pharmacology , Coculture Techniques , Mice, Inbred C57BL , Thymus Gland/metabolism , Cell Differentiation , CD4-Positive T-Lymphocytes/metabolism , Stromal Cells , DNA-Binding Proteins/metabolism , Transcription Factors/metabolismABSTRACT
OBJECTIVE: The aim of this study is to investigate the effects of topical growth hormone (GH) treatment on skin wound healing in mice. MATERIALS AND METHODS: An excisional wound healing model was established on male Swiss mice, and wound healing ability was evaluated by macroscopic and histologic analyses of mice treated with topical 10-8 M and 10-7 M of GH versus the mice receiving ve- hicle alone. Wound tissues were collected on post treatment days 3, 7, and 14. Skin fragments were subjected to hematoxylin and eo- sin and Masson's trichrome staining for morphological analyses. The expression of type I collagen and platelet endothelial cell adhesion molecule 1 (CD31) was detected by immunohistochemical analysis. RESULTS: Topical treatment with GH resulted in faster wound closure rates at all time points analyzed versus those observed in the control group (day 3: 18.3 ± 3.1 vs. 44.4 ± 7.4, 43.6 ± 0.6; day 7: 41.7 ± 6.3 vs. 73.8 ± 6.6, 71.3 ± 5.8; day 12: 94.3 ± 3.9 vs. 100 ± 0, 100 ± 0). Histological analysis of the wound on post treatment day 3 revealed a more diffused in ltration of in ammatory cells in the group treated with GH. After day 7, GH-treated animals began form- ing granulation tissue, and there was an increase in in ammatory cell in ltration. The GH signi cantly increased the expression of type I collagen (day 7: 57.4 ± 4.0 vs. 120.2 ± 9.7, 79.3 ± 7.9; day 14: 218.2 ± 10.4 vs. 301.5 ± 9.1, 235.0 ± 7.5) as well as the number of blood vessels (day 7: 10.0 ± 2.4 vs. 15.3 ± 2.0, 10.1 ± 2.2; day 14: 3.2 ± 0.8 vs. 5.6 ± 2.0, 6.2 ± 2.2) in the injured area. CONCLUSIONS: The GH accelerates the closure of skin wounds by resolving the in- ammatory phase faster, accelerating reepithelialization and collagen deposition, and stimulating angiogenesis.
Subject(s)
Granulation Tissue/drug effects , Granulation Tissue/pathology , Growth Hormone/pharmacology , Skin/drug effects , Skin/pathology , Wound Healing/drug effects , Wound Healing/physiology , Administration, Topical , Animals , Cell Proliferation/drug effects , Cell Proliferation/physiology , Collagen/metabolism , Disease Models, Animal , Immunohistochemistry , Male , Mice , Neovascularization, Physiologic/drug effects , Skin/injuriesABSTRACT
BACKGROUND: Several evidences indicate that hormones and neuropeptides function as immunomodulators. Among these, growth hormone (GH) is known to act on the thymic microenvironment, supporting its role in thymocyte differentiation. The aim of this study was to evaluate the effect of GH on human thymocytes and thymic epithelial cells (TEC) in the presence of laminin. RESULTS: GH increased thymocyte adhesion on BSA-coated and further on laminin-coated surfaces. The number of migrating cells in laminin-coated membrane was higher in GH-treated thymocyte group. In both results, VLA-6 expression on thymocytes was constant. Also, treatment with GH enhanced laminin production by TEC after 24 h in culture. However, VLA-6 integrin expression on TEC remained unchanged. Finally, TEC/thymocyte co-culture model demonstrated that GH elevated absolute number of double-negative (CD4(-)CD8(-)) and single-positive CD4(+) and CD8(+) thymocytes. A decrease in cell number was noted in double-positive (CD4(+)CD8(+)) thymocytes. CONCLUSIONS: The results of this study demonstrate that GH is capable of enhancing the migratory capacity of human thymocytes in the presence of laminin and promotes modulation of thymocyte subsets after co-culture with TEC.
Subject(s)
Epithelial Cells/drug effects , Growth Hormone/pharmacology , Laminin/biosynthesis , Thymocytes/drug effects , Thymus Gland/cytology , Analysis of Variance , CD4-Positive T-Lymphocytes , CD8-Positive T-Lymphocytes , Cell Adhesion/drug effects , Cell Differentiation/drug effects , Cell Movement/drug effects , Cells, Cultured , Child , Child, Preschool , Coculture Techniques , Flow Cytometry/methods , Humans , Immunohistochemistry , Infant , Infant, Newborn , Integrin alpha6beta1/analysis , Integrin alpha6beta1/metabolism , Laminin/drug effects , Reference Values , Thymus Gland/metabolism , Time FactorsABSTRACT
Therapeutic angiogenesis may be applied in medical conditions to promote stimulation of angiogenesis. Angiogenesis is a multistep process, which includes endothelial cell proliferation, migration, and tube formation, which is mediated by various angiogenic polypeptides. Thus, studies that elucidate the cellular mechanisms involved in these processes are necessary to develop novel therapeutic strategies. This study investigated the in vitro effects of the pro-angiogenic factors, insulin-like growth factor-1 (IGF-1) and/or chemokine (CC motif) ligand 2 (CCL2), on endothelial cells. Flow cytometry analysis showed that IGF-1 and CCL2 treatment did not interfere with IGF-1 receptor (IGF-1R) expression, but CCL2 treatment increased CCL2 receptor (CCR2) expression. Immunofluorescence analysis revealed that the IGF-1/CCL2 combination induced a greater increase in fibronectin deposition, but the treatments did not alter the expression of the fibronectin receptors, CD49e and CD44. The interaction of fibronectin with cytokines demonstrated that IGF-1/CCL2 promoted changes in intermediate F-actin remodeling that may result in increased endothelial cell adhesion and cell migration mediated by fibronectin. Furthermore, IGF-1/CCL2 stimulated endothelial cells, grown on fibronectin, to form capillary-like structures and intercellular lumina with greater luminal area. These data suggest that IGF-1/CCL2 combination and a fibronectin matrix may contribute to the angiogenesis process to stimulate adhesion, migration, and tube formation by endothelial cells as a result of F-actin remodeling.
