G-protein-coupled receptors (GPCRs) are critical sensors affecting the state of eukaryotic cells. To get systematic insight into the GPCRome of microglia, we analyzed publicly available RNA-sequencing data of bulk and single cells obtained from human and mouse brains. We identified 17 rhodopsin and adhesion family GPCRs robustly expressed in microglia from human brains, including the homeostasis-associated genes CX3CR1, GPR34, GPR183, P2RY12, P2RY13, and ADGRG1. Expression of these microglial core genes was lost upon culture of isolated cells ex vivo but could be acquired by human induced pluripotent stem cell (iPSC)-derived microglial precursors transplanted into mouse brains. CXCR4 and PTGER4 were higher expressed in subcortical white matter compared to cortical grey matter microglia, and ADGRG1 was downregulated in microglia obtained from normal-appearing white and grey matter tissue of multiple sclerosis (MS) brains. Single-cell RNA sequencing of microglia from active lesions, obtained early during MS, revealed downregulation of homeostasis-associated GPCR genes and upregulation of CXCR4 expression in a small subset of MS-associated lesional microglia. Functional presence of low levels of CXCR4 on human microglia was confirmed using flow cytometry and transwell migration towards SDF-1. Microglia abundantly expressed the GPCR down-stream signaling mediator genes GNAI2 (α_i2), GNAS (α_s), and GNA13 (α₁₃), the latter particularly in white matter. Drugs against several microglia GPCRs are available to target microglia in brain diseases. In conclusion, transcriptome profiling allowed us to identify expression of GPCRs that may contribute to brain (patho)physiology and have diagnostic and therapeutic potential in human microglia.

Circulating and tissue-resident T cells collaborate in the protection of tissues against harmful infections and malignant transformation but also can instigate autoimmune reactions. Similar roles for T cells in the brain have been less evident due to the compartmentized organization of the central nervous system (CNS). In recent years, beneficial as well as occasional, detrimental effects of T-cell-targeting drugs in people with early multiple sclerosis (MS) have increased interest in T cells patrolling the CNS. Next to studies focusing on T cells in the cerebrospinal fluid, phenotypic characteristics of T cells located in the perivascular space and the meninges as well as in the parenchyma in MS lesions have been reported. We here summarize the current knowledge about T cells infiltrating the healthy and MS brain and argue that understanding the dynamics of physiological CNS surveillance by T cells is likely to improve the understanding of pathological conditions, such as MS.

The human brain is populated by perivascular T cells with a tissue-resident memory T (T_RM)-cell phenotype, which in multiple sclerosis (MS) associate with lesions. We investigated the transcriptional and functional profile of freshly isolated T cells from white and gray matter. RNA sequencing of CD8⁺ and CD4⁺ CD69⁺ T cells revealed T_RM-cell signatures. Notably, gene expression hardly differed between lesional and normal-appearing white matter T cells in MS brains. Genes up-regulated in brain T_RM cells were MS4A1 (CD20) and SPP1 (osteopontin, OPN). OPN is also abundantly expressed by microglia and has been shown to inhibit T cell activity. In line with their parenchymal localization and the increased presence of OPN in active MS lesions, we noticed a reduced production of inflammatory cytokines IL-2, TNF, and IFNγ by lesion-derived CD8⁺ and CD4⁺ T cells ex vivo. Our study reports traits of brain T_RM cells and reveals their tight control in MS lesions.

Preventing T cell migration to the central nervous system (CNS) has remarkable therapeutic effects in relapsing-remitting multiple sclerosis (RRMS) but is poorly effective against the progressive form (PMS). Disability progression in PMS likely results from an interplay between smoldering local inflammation and neurodegeneration. The mechanisms sustaining the chronicity of PMS are poorly understood. Here, we investigated the potential role of tissue-resident memory CD4 ⁺ T cells (CD4 ⁺ Trm cells) in sustaining chronic CNS autoimmunity. We showed that CD4 ⁺ Trm cells were present in the CNS of mice with chronic experimental autoimmune encephalomyelitis (EAE) and in brain tissues from persons with PMS. Using flow cytometry and immunohistofluorescence analysis, we revealed the presence of bona fide CD4 ⁺ Trm cells expressing characteristic Trm cell surface markers, including CD69, CXCR6, P2RX7, and CD49a, in the CNS of mice with EAE and in the brains of persons with PMS. These T cells also expressed the transcription factor Hobit in mice with chronic EAE. Single-cell transcriptomic analysis uncovered the transcriptional heterogeneity and inflammatory potential of CD4 ⁺ Trm cells, and, accordingly, these cells localized within CNS inflammatory lesions of mice with EAE and persons with PMS. Last, either genetic or pharmacological depletion of CD4 ⁺ Trm cells combined with antibody-mediated depletion of the recirculating CD4 ⁺ T cell compartment alleviated neurological signs during the chronic phase of EAE. Our results indicate that CD4 ⁺ Trm cells contribute to maintain a chronic inflammatory state in the CNS and suggest that therapeutic strategies for PMS should consider targeting the CNS-resident T cell compartment.

Oxysterols, or cholesterol oxidation products, are naturally occurring lipids which regulate the physiology of cells, including those of the immune system. In contrast to effects that are mediated through nuclear receptors or by epigenetic mechanism, which take tens of minutes to occur, changes in the activities of cell-surface receptors caused by oxysterols can be extremely rapid, often taking place within subsecond timescales. Such cell-surface receptor effects of oxysterols allow for the regulation of fast cellular processes, such as motility, secretion and endocytosis. These cellular processes play critical roles in both the innate and adaptive immune systems. This review will survey the two broad classes of cell-surface receptors for oxysterols (G-protein coupled receptors (GPCRs) and ion channels), the mechanisms by which cholesterol oxidation products act on them, and their presence and functions in the different cell types of the immune system. Overall, this review will highlight the potential of oxysterols, synthetic derivatives and their receptors for physiological and therapeutic modulation of the immune system.

T cell surveillance is mandatory for maintaining central nervous system (CNS) homeostasis, while aberrant accumulation is linked to neuroinflammation. To explore the residency programs acquired by T cells through different anatomical locations of the human brain, we isolated CD8⁺ and CD4⁺ T cells from CNS border compartments (choroid plexus and leptomeninges), intrathecal compartments (cerebrospinal fluid [CSF] and subcortical white matter [WM]), and paired peripheral blood of brain donors. Flow cytometry revealed a shared effector memory phenotype across CNS compartments that was partially induced in circulating T cells interacting with brain endothelium in vitro. Intrathecal T cells expressed full tissue-residency traits, yet T cells from WM, compared to CSF, showed reduced expression of migratory, co-stimulatory, and recent activation markers despite a similar cytokine response upon ex vivo activation. This work demonstrates the versatility of T cell phenotypes across CNS compartments and provides insight into the programs regulating their recruitment and maintenance within the CNS.