Modulation of astrocyte phenotype in response to T-cell interaction
Graphical abstract
Introduction
Classically, the brain has been described as an immune privileged organ separated from the peripheral immune system. There is increasing evidence that the brain is not devoid of peripheral immune cells. Immune cells can enter the brain via the lymphatic vessels, interacting with neurons and glial cells (Lopes Pinheiro et al., 2016; Schachtele et al., 2014), playing vital roles in immune surveillance (Louveau et al., 2018; Louveau et al., 2015) and bi-directional communication between the immune system and central nervous system (CNS) (Engelhardt and Ransohoff, 2012; Zhao et al., 2018; Chabot et al., 1999; Gimsa et al., 2004).
T-cells have been identified as resident cells in the brains of humans and rodents (Smolders et al., 2018; Xie et al., 2015). Previous research from our laboratory has showed that experimental ischemic stroke increased levels of T-cells in the brain, which have close interaction with astrocytes in the peri-infarct region. Furthermore, these T-cells increased gene expression and produced pro- and anti-inflammatory cytokines in vivo, including interleukin-10 (IL-10) (Xie et al., 2019). Astrocytes which have demonstrated their ability to act as antigen presenting cells in the brain (Rostami et al., 2020), play critical roles in brain homeostasis and contribute to neuroimmune communication (Li et al., 2017; Macht, 2016; Nasr et al., 2019; Sun and Jakobs, 2012; Wu et al., 2005). Studies have demonstrated that IL-10 increased when microglia and T-cells were in direct contact. IL-10 levels in T-cells-microglia co-cultures were reduced in a concentration dependent manner when CTLA-4 Fc was added to the culture media (Chabot et al., 1999). T-cell IL-10 production was modulated when astrocytes and T-cell interacted and induced upregulation of CTLA-4 levels in T-cells after 24 h in co-culture (Gimsa et al., 2004). Therefore, we reasoned that astrocyte T-cell interaction may have an impact on astrocyte phenotype and metabolism.
IL-10 is recognized as an anti-inflammatory cytokine which regulates immune cells in the periphery, suppressing excessive inflammation (Ledeboer et al., 2002; Lobo-Silva et al., 2016). Recently IL-10 has been identified in its ability to balance pro-inflammatory immune response in the brain (Lobo-Silva et al., 2016; Norden et al., 2016). It has been previously reported that IL-10 is released from several cell types, including astrocytes and T-cells, when immune challenged as occurs during traumatic brain injury and experimental ischemic stroke (Rodney et al., 2018; Schroeter and Jander, 2005; Xie et al., 2019; Xin et al., 2011). Mounting evidence suggests that the role of IL-10 in neurons is to induce an anti-apoptosis response during brain injury (He et al., 2017; Lobo-Silva et al., 2016; Xin et al., 2011; Zhou et al., 2009). However little if any attention has been given to the potential regulatory role of IL-10 on astrocyte proliferation and metabolism. The goal of this investigation was to determine the interaction of T-cells and astrocytes and the involvement of IL-10 in in vitro conditions. Here, we provided novel evidence to support that astrocytes interact with T-cells via direct cell-to-cell contact, increasing IL-10 gene expression and production leading to alterations in astrocyte's metabolic phenotype.
Section snippets
Astrocyte cell-line and primary astrocyte cultures
The astrocyte cell-line was purchased from American Type Culture Collection (ATCC): C8S (Astrocyte type II clone) ATCC CRL-2535™ Mus musculus. Adult (3–4 months old) C57BL6 mice were purchased from Jackson Laboratory. Breeding pairs of C57BL6 mice were established from which neonatal C57BL6 mice (postnatal day 1–3) were obtained for primary astrocyte cultures. Primary astrocytes were prepared as previously described (Prah et al., 2019; Roy Choudhury et al., 2015) with the following
T-cells interact directly with C8S astrocytes to increase IL-10 production
C8S astrocyte cell-lines were cultured alone for 48 h and after which they were either given fresh media or co-cultured at a 1:1 ratio with pan T-cells in fresh media for an additional 48 h (Supplementary Fig. S1A and B). C8S astrocytes co-cultured with pan T-cells generated an approximately 10-fold increased IL-10 mRNA level compared to C8S astrocytes alone (Fig. 1A). The increased IL-10 gene expression was confirmed by the increase of IL-10 protein secreted into the supernatant which peaked
Discussion
Our investigation demonstrates that the IL-10 pathway is implicated in the interaction between T-cells and astrocytes and that this anti-inflammatory pathway plays an important role in directing astrocyte metabolism and proliferation. These results strongly support our previous research which reported that there were increased levels of IL-10 gene expression from T-cells harvested from post-ischemic mouse brain hemispheres compared to the contralateral non-stroked hemispheres (Xie et al., 2019)
Acknowledgements
This research was in part supported by National Institutes of Health grants R01NS088596 (SY), R01NS109583 (SY), T32 AG020494 (JH), and a grant (#RP17301) from the Cancer Prevention and Research Institute of Texas (JH). The authors declare no conflict of interest.
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