Expansion of brain T cells in homeostatic conditions in lymphopenic Rag2−/− mice

Abstract

The concept of the brain as an immune privileged organ is rapidly evolving in light of new findings outlining the sophisticated relationship between the central nervous and the immune systems. The role of T cells in brain development and function, as well as modulation of behavior has been demonstrated by an increasing number of studies. Moreover, recent studies have redefined the existence of a brain lymphatic system and the presence of T cells in specific brain structures, such as the meninges and choroid plexus. Nevertheless, much information is needed to further the understanding of brain T cells and their relationship with the central nervous system under non-inflammatory conditions. In the present study we employed the Rag2−/− mouse model of lymphocyte deficiency and reconstitution by adoptive transfer to study the temporal and anatomical expansion of T cells in the brain under homeostatic conditions. Lymphopenic Rag2−/− mice were reconstituted with 10 million lymphoid cells and studied at one, two and four weeks after transfer. Moreover, lymphoid cells and purified CD4+ and CD8+ T cells from transgenic GFP expressing mice were used to define the neuroanatomical localization of transferred cells. T cell numbers were very low in the brain of reconstituted mice up to one week after transfer and significantly increased by 2 weeks, reaching wild type values at 4 weeks after transfer. CD4+ T cells were the most abundant lymphocyte subtype found in the brain followed by CD8+ T cells and lastly B cells. Furthermore, proliferation studies showed that CD4+ T cells expand more rapidly than CD8+ T cells. Lymphoid cells localize abundantly in meningeal structures, choroid plexus, and circumventricular organs. Lymphocytes were also found in vascular and perivascular spaces and in the brain parenchyma across several regions of the brain, in particular in structures rich in white matter content. These results provide proof of concept that the brain meningeal system, as well as vascular and perivascular spaces, are homing sites of lymphocytes and suggest the possibility of a brain specific T cell subtype.

Contributions of the adaptive immune system to mood regulation: Mechanisms and pathways of neuroimmune interactions

“…A vivid in vivo demonstration of T cell residence in the brain following adoptive transfer of green fluorescent protein (GFP)-positive lymph node cells in Rag2 −/− mice was recently published (Song et al, 2016). As expected, several weeks after repopulation into a naïve mouse, fluorescent lymphocytes were found scattered within the meninges, choroid plexus, and the circumventricular organs.…”

“…As expected, several weeks after repopulation into a naïve mouse, fluorescent lymphocytes were found scattered within the meninges, choroid plexus, and the circumventricular organs. Labeled cells within brain parenchyma were scarce and were largely confined to perivascular spaces (Song et al, 2016). Importantly, lymphocytes were not reported in the hippocampal vascular niche that supports DG neurogenesis (Palmer et al, 2000) in this or any other study to date.…”

CD8+ T cells promote cytokine responses to stress

“…Expression of this cytokine in the brain has been shown in varied cell types, but more prominently in microglial and perivascular cells [38]. Recently, lymphocytes and T cells have been shown to circulate in a brain lymphatic system [39], and work from our group has shown that they proliferate in the brain of reconstituted Rag2 −/− mice with a significant degree of interaction with the microvasculature during this process [32]. Thus, it is possible that CD8 + T cells increase their interaction with the brain microvasculature in response to stress, stimulating the production of IL-1β by perivascular cells, similar to the model of stress-induced vascular inflammation in hypertension [28].…”

“…Lymphopenic Rag2 −/− mice were reconstituted by adoptive transfer with 8–10 million naive CD4 + (n = 23) or CD8 + (n = 21) T cells in a 1:1 donor to recipient transfer from age-matched WT mice. Donor cells were prepared as previously described [32] using the EasySep CD4 + or CD8 + purification kits (Stem Cell Technologies, Vancouver, BC, Canada) following manufacturer’s guidelines. Control Rag2 −/− mice (n = 18) received PBS only.…”

“…Single cell suspensions from the lymph nodes (LNs) and Spleen were processed as described [32]. Antibodies used in this study were: anti-CD3-eFluor 450 (eBioscience, 48–0032-82), anti-CD8-FITC (BD Biosciences, cat #553030), anti-CD8-PerCP-Cy5.5 (BD Biosciences, cat #561109), anti-CD4-APC (BD Biosciences, cat #553051), anti- CD4-PE (BD Biosciences, cat #553730), anti-NK1.1-PerCP-Cy5.5 (BD Biosciences, cat #561111), anti-CD44-APC (BD Biosciences, cat #559250), anti-CD44-PE (BD Biosciences, cat #553133), PE-rat anti mouse CD62L (BD Biosciences, cat #553151; eFluor450-rat anti mouse IFNγ (eBioscience, cat #48–7311-82), PE-rat anti-mouse IL-6 (BD Biosciences, cat #: 562050) and APC-rat anti-mouse TNF-α (BD Biosciences, cat #: 561062).…”