The Responsiveness of Thymic Stromal Cells to semaphorin-3A

ABSTRACT

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.

KEYWORDS:

• Thymic stromal cell
• semaphorin-3A
• neuropilin-1
• thymocyte
• cell adhesion

Acknowledgments

Authors thank the laboratory technicians Ana Rúbia Batista Ribeiro (Physics Institute of Federal University of Alagoas) for assistance in image acquisition on the Scanning Electron Microscope and Juliane Pereira da Silva for handling of flow cytometer.

Declaration of interest statement

The authors declare that they have no conflict of interest.

Additional information

Funding

The study was supported by Brazilian National Council for Scientific and Technological Development - CNPq (Nº. 408677/2016-3 and Nº. 304408/2018-2) and Foundation for Funding Research in the State of Alagoas – FAPEAL (Nº. 60030 001260/2017).