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  • EBI and its ligand s EBI

    2019-12-14

    EBI2 and its ligand(s) EBI2 was found in a screen of upregulated genes in human B cells upon infection with EBV [1]. EBI2 is a G-protein (Gαi type) coupled receptor [2] but as long as 18 years after its discovery the nature of its ligand remained undisclosed. With the help of transfected cell lines and EBI2 knockout animals, researcher from Novartis and J&J independently from each other discovered the ligands from extracts of sheep liver and porcine spleen respectively [3,4]. The description of the natural ligands of EBI2 advanced the understanding of the biology of the receptor enormously and led to the fast development of potent antagonists [5,6]. Both groups found 7α,25-OHC to be the most potent ligand in signaling and in induction of migration of transfected and primary immune cells. The second best ligand was 7α,27-OHC whereas 25-OHC showed only a minor residual activity [3,4]. Importantly, usage of cells from EBI2 deficient mice showed that the action of the ligands via EBI2 are non-redundant. Similarly, knockout of one of the major enzymes generating the ligand from cholesterol, CH25H, showed very similar phenotypes in terms of B cell localization in germinal centers as the EBI2 knockout mouse [3] (see below), proving that indeed these dihydroxycholesterols are the physiological relevant ligands of EBI2.
    Role of EBI2 in the immune system Even before the ligand was characterized, several high impact papers showed an important role of EBI2 in the germinal center reaction and B cell maturation, which is crucial for fast high affinity antibody production against foreign invaders [7,8]. After definition of the ligands, the cells expressing the ligand-generating enzymes were deciphered. It was shown that in lymphoid tissues so called lymphoid stromal cells express CH25H and CYP7B1 and the ligand degenerating enzyme HSD3B7 in a coordinated manner to localize B cells at the follicle perimeter. At this site at the border of the T cell zone, B cells receive T cell help for efficient antibody production [9]. An analysis of EBI2 expression on dendritic cells (DCs) showed that EBI2 is highly expressed on CD4+ conventional cDCs. In line with this mice deficient for EBI2 largely lack this subset of cDCs in the spleen. Furthermore, the localization of the residual cells is disturbed [10,11]. It was found that EBI2 positive DCs follow a ligand gradient to marginal zone bridging channels and most likely receive survival signals via the lymphotoxin β receptor at this place. Localization of DCs at these bridging channels enables efficient uptake of blood borne particulate MLCK inhibitor peptide 18 and therefore fast T and B cell responses against the latter. Another important role for EBI2 was demonstrated for T follicular helper cells (Tfh cells). Those cells have a specific role to help B cells in antibody production. It was shown that Tfh cells need to migrate towards activated DCs in the outer T zones to receive specific signals via ICOS/ICOS-L under IL-2 depleted conditions to differentiate into Tfh cells [12]. Very recently, an important role of EBI2 expression by innate lymphoid cells type 3 (ILC3) for development of inflammation in the gut in a mouse colitis model was demonstrated [13]. It was shown that stromal cells in the gut produce the EBI2 ligand, which leads to migration of colonic ILC3s into cryptopatches and isolated lymphoid follicles. This fostered colitis induced in RAG-1 deficient mice by injection of anti-CD40 antibodies [13].
    EBI2 in neuroinflammation The myelin sheet coating the axons of fast conducting neurons makes up largely for the so-called white matter of the CNS. The myelin sheet is basically a lipid-rich multilamellar membrane wrapped around the axon, which is needed to ensure fast saltatory electrical conduction. The myelin sheet contains very high concentrations of cholesterol, which is produced de novo inside the CNS. Due to the high amounts of those insulating sheets, the CNS is the organ containing the highest amount of cholesterol in the body [14]. In multiple sclerosis (MS) the myelin sheets are attacked by the patient\'s own immune system. T cells, B cells (with and without their antibodies) and macrophages play major parts in the pathology process leading to white matter lesions and neuronal cells death over time [[15], [16], [17]]. Cholesterol oxidation products in the serum or the cerebrospinal fluid (CSF) of MS patients (which would be predicted in an inflammatory destruction process of the myelin sheet) have been highly investigated [[18], [19], [20], [21], [22], [23], [24]] (Fig. 1). 24S-hydroxycholesterol is one of the major specific degradation products of cholesterol produced inside the CNS. In MS patients (as also in late stage neurodegenerative diseases) slightly less of this oxysterol is found in the plasma, which most likely reflects the loss of brain mass in these patients over time [19,25,26]. Conversely, increased levels of blood-derived 27-OHC can be found in the CSF of MS patients reflecting their blood brain barrier dysfunction [25,27]. A very recent systematic analysis of over 20 sterols in plasma and CSF revealed a decrease of 25-OHC levels in the plasma of relapsing-remitting MS patients but no significant changes of the EBI2 ligand 7α,25-OHC [24].