Th1 cells promote neurite outgrowth from cortical neurons via a mechanism dependent on semaphorins

https://doi.org/10.1016/j.bbrc.2010年10月02日9 Get rights and content

Abstract

The roles of T lymphocytes in the central nervous system (CNS) are diverse; their roles in the injured CNS have been reported to be both detrimental and advantageous. Hence, an investigation of the effects of specific subsets of T cells on neurons may provide an insight into the interaction between the nervous system and the immune system. In the present study, we demonstrate that a specific subset of T lymphocytes enhanced neurite outgrowth in vitro. When cultured T helper type 1 (Th1) cells were co-cultured with cortical neurons, neurite outgrowth from neurons was enhanced; however, the same was not observed when Th2 or naïve T cells were used. We observed that the promotion of neurite outgrowth by Th1 cells was completely inhibited by anti-interferon γ (IFN-γ) neutralizing antibody, but that IFN-γ did not directly promote neurite growth. Furthermore, experiments using knockout mice revealed that semaphorin 4A (Sema4A) but not Sema7A was required for the effect produced by Th1 cells. These results demonstrate that Sema4A and IFN-γ expressed in Th1 cells play a critical role in enhancing neurite outgrowth from cortical neurons.

Research highlights

► Interaction of helper T cells and neurons. ► Th1 cells promote neurite growth. ► Sema4A and IFN-γ play critical roles in inducing neurite growth.

Introduction

Immune reactions after central nervous system (CNS) trauma have been thought to be harmful for axonal regeneration and functional recovery [1], [2], [3]. However, during the last decade, it has been reported that the transfer of autoimmune T cells or active immunization with T cells promoted functional recovery after CNS injury [4], [5], [6], [7], [8], [9]. In contrast, others reported that T lymphocytes caused axonal damage after the CNS injury [3], [10], [11], [12]. It could be assumed that these contradictory results may be explained by distinct roles of subsets of T cells such as T helper type 1 (Th1) or Th2 cells.
Subsets of helper T cells have been suggested to be involved in the etiology of CNS diseases. Interferon γ (IFN-γ)-producing Th1 cells and interleukin (IL)-17-producing helper T (Th17) cells are associated with the onset and progression of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis [13], [14]. Interleukin-4-producing Th2 cells were considered to promote functional recovery after the CNS injury [15]. However, the role of each subset of T cells had not been explored in depth, and whether a specific subset of T cells was beneficial to CNS injury remained elusive. Contradictory reports on the role of T cells in vivo after a CNS injury may be because of the distinct roles of helper T cell-subsets. Hence, it is important to dissect out the precise effects of each T cell subset on neurons. For this purpose, we performed a neurite outgrowth assay using cortical neurons in vitro because this assay is considered to reveal the ability of neurons to regenerate axons in vivo. We report that Th1 cells but not Th2 cells enhanced neurite outgrowth from embryonic cortical neurons. Furthermore, we explored the molecular mechanism underlying the promotion of neurite outgrowth by Th1 cells. We investigated whether IFN-γ, which is mainly secreted by Th1 cells or semaphorins (axon guidance molecules as well as immunomodulators expressed on immune cells) [16], were required for the effects produced by Th1 cells. Our results demonstrated that Sema4A but not Sema7A was required for the effects produced by Th1 cells. Further, IFN-γ acting on Th1 cells was also required for producing these effects.

Section snippets

Mice

C57BL/6 mice were purchased from Charles River. Sema7A -/- mice on the C57BL/6 background were used [17]. Sema4A -/- mice were generated as described previously [18]. All mice used in this study were housed in specific pathogen-free conditions. All the experimental procedures were approved by the Institutional Committees of Chiba University and Osaka University.

Differentiation of CD4+ T cells

Spleens were collected from C57BL/6j female mice, and single-cell suspensions were prepared by mechanical disruption in RPMI-growth

Cultured Th1 cells enhance neurite outgrowth

In order to assess the functional interaction between neurons in the CNS and T cells, we performed a neurite growth assay. We examined neurite outgrowth from cortical neurons in the cerebral cortex of mice on embryonic day 15 and 16 for 24 hours. The effect of helper T cells on the neurons was examined by co-culture. For this experiment, CD4+ naïve helper T cells isolated ex vivo from mice spleens between postnatal weeks 7 and 9 were differentiated into Th1 or Th2 cells. First, we confirmed

Discussion

There have been controversies concerning the efficacy of adoptive transfer of T lymphocytes during CNS trauma [15]. Critically, no experiments focusing on specific T cell subsets with regard to these conflicting results have been reported. In the present study, we investigated the effects of specific T cell subsets on neurite outgrowth to evaluate neuronal regeneration in vitro. Unexpectedly, Th1 cells, which were suggested to be detrimental during CNS injury [15] and pathogenic during EAE [13]

Acknowledgments

We thank Dr. T. Nakayama, Dr. M. Yamashita, and Dr. M. Kuwahara at Chiba University for their technical advice on T cell-culture. This work was supported by a research grant from a Grant-in-Aid (S) from Ministry of Health, Labour and Welfare.

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