Highlighting a New Potential Target in Stroke: Immunoreceptor NKG2D.

HomeJournal of the American Heart AssociationVol. 12, No. 12Highlighting a New Potential Target in Stroke: Immunoreceptor NKG2D Open AccessEditorialPDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citations ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toOpen AccessEditorialPDF/EPUBHighlighting a New Potential Target in Stroke: Immunoreceptor NKG2D Josef Anrather and Kimberly Marks Josef AnratherJosef Anrather *Correspondence to: Josef Anrather, VMD, Weill Cornell Medical College, Feil Family Brain and Mind Research Institute, 407 East 61st Street, RR 409, New York, NY 10065. Email: E-mail Address: [email protected] https://orcid.org/0000-0002-0468-9162 , Feil Family Brain and Mind Research Institute, Weill Cornell Medicine, , New York, , NY, , USA, Search for more papers by this author and Kimberly MarksKimberly Marks https://orcid.org/0009-0002-0363-7747 , Feil Family Brain and Mind Research Institute, Weill Cornell Medicine, , New York, , NY, , USA, Search for more papers by this author Originally published10 Jun 2023https://doi.org/10.1161/JAHA.123.030482Journal of the American Heart Association. 2023;12:e030482This article is a commentary on the followingImpact of NKG2D Signaling on Natural Killer and T‐Cell Function in Cerebral IschemiaOther version(s) of this articleYou are viewing the most recent version of this article. Previous versions: June 11, 2023: Ahead of Print In the past decade, the field of ischemic stroke has placed a special emphasis on the role of immune cells and inflammation in the pathogenesis of stroke injury and repair.1 Ischemic stroke is now known to engage both the innate and adaptive immune system, 2 systems largely thought to have opposing goals, with different cell types engaged for different tasks. We now know there to be significant overlap in these cell types and their coordinated efforts following injury.2, 3 Although much focus has been placed on elucidating the roles of the innate immune system, particularly monocytes, neutrophils, and natural killer (NK) cells, in the proinflammatory response to ischemic brain injury, the mechanisms by which cellular components of the adaptive immune system, like γδ T cells and cytotoxic CD8+ T cells, affect stroke pathogenesis during the acute phase are less well understood.Of recent particular interest are NK cells, a class of innate lymphoid cells that exhibit early cytotoxic effects independent of antigen presentation. NK cells are recruited into the brain in a chemokine guided manner, augment inflammation through release of proinflammatory cytokines, and have been implicated in various neurological conditions including ischemic brain injury.4, 5 Although their specific role in stroke has not yet been fully elucidated, previous reports have shown NK cell infiltration in the peri‐infarct stroke tissue of patients with middle cerebral artery occlusion and NK cells colocalizing to ischemic neurons, with ischemic neurons particularly susceptible to NK‐cell mediated death.5 In other studies of experimental stroke, NK cells were found to be increased in the ischemic hemisphere as early as 3 hours after ischemia and remained elevated for several days following injury. Depletion of NK cells via antibodies ameliorated overall stroke outcome, infarct size, and neurological deficits in middle cerebral artery occlusion in mice.5, 6Cytotoxic CD8+ T cells contribute to brain inflammation during acute and chronic stages following stroke. CD8+ T cells were found after experimental stroke up to 30 days after induction of ischemia, and depletion of CD8+ T cells starting at day 10 after stroke improved long‐term motor deficits.7 CD8+ T cells might also participate in the pathophysiology of acute stroke. A study identified the Fas "ligand"‐ 3‐phosphoinositide dependent protein kinase 1 pathway regulating the cytotoxic activity of CD8+ T cells, and it was shown that pharmacological inhibition or genetic deletion of components of this signaling pathway in CD8+ T cells reduced ischemic brain injury and neurobehavioral deficits at 3 days after stroke.8 However, the cellular targets and molecular mechanism of Fas "ligand"‐ 3‐phosphoinositide dependent protein kinase 1 activation have yet to be determined.In this issue of the Journal of the American Heart Association (JAHA), David and colleagues highlight the importance of cytotoxic lymphocytes like NK and CD8+ T cells and notably their shared expression of activating immunoreceptor natural killer group 2, member D (NKG2D), offering some mechanistic insight into a potential new target to study inflammation following stroke injury.9 David et al offer novel cell‐specific insights into this receptor and its specific implications in stroke. Of note, blocking NKG2D has been previously shown to attenuate ischemia–reperfusion injury in a cardiac transplant model and renal injury.10, 11 NKG2D is a C‐type, lectin‐like receptor found on NK cells and on activated CD8+ T cells, γδ T cells, macrophages, and some CD4+ T cells.12, 13 David and colleagues now show that blocking the NKG2D cascade prophylactically (1 hour before stroke) and therapeutically (1 hour after stroke) improved stroke outcomes at 24 hours. At 3 days, only the prophylactic group conferred additional improvement from NKG2D blockade, suggesting that the signaling cascade occurs early in the course of ischemic brain injury.9Immunodeficient NRG mice, lacking NK, T, and B cells, treated with NK cells either blocked by αNKG2D antibody or isotype control showed a decrease in infarct volume when using αNKG2D antibody‐treated cells, although this reduction did not reach statistical significance. Using adoptive transfer of mature CD8+ T cells, either blocked with αNKG2D antibody or isotype control, into recombinant activating gene 1 mice lacking mature T and B cells, a significant decrease in infarct volume was found in mice treated with αNKG2D cells, although not to levels of recombinant activating gene 1 controls, suggesting that although NKG2D may play a notable role in CD8+ T cell cytotoxicity following ischemia, other T‐cell mechanisms are independently at play. Although γδ T‐cell deficient mice were still amendable to the protection of αNKG2D treatment, this effect was less pronounced in cluster differentiation gene 1 mice lacking natural killer T cells, suggesting that NKG2D may also play a role in regulating the cytotoxic of this immune cell type. Importantly, when reconstituting recombinant activating gene 1 mice with CD8+ T cells from T‐cell receptor (TCR) transgenic mice expressing ovalbumin restricted TCR (OT‐1 mice) that were treated with αNKG2D, infarct volumes decreased, suggesting that in the setting of stroke, activation of CD8+ T cells via the NKG2D receptor is independent of TCR engagement. Using a degranulation assay as a proxy for cytolytic function, the investigators found diminished cytolytic activity in CD8+ T cells undergoing NKG2D blockade, whereas doing so in NK cells resulted in no changes.NKG2D ligands are sparsely expressed under homeostatic conditions and are upregulated following DNA damage and stress.14 The current study found that the high‐affinity ligand UL16 binding protein 1 and MULT1 (Mouse UL16 binding protein‐like transcript 1)15 was upregulated after cerebral ischemia. Using immunohistochemistry, several ULBP family members were localized to neurons, astrocytes, and microglia after experimental ischemia. Importantly, NKG2D blockage reduced the expression of the ULBP1 ligand in the brain. However, further studies are needed to determine whether this is a direct effect of blocking ligand‐receptor interactions or whether it is a consequence of the neuroprotection afforded by αNKG2D treatment.NKG2D is expressed on the surface of all NK and some T cells and is dependent on adapter proteins to transmit signals.16 Previous reports and data from this article have indicated that NKG2D alone cannot be independently responsible for enhancing cytotoxic function in these cells. Adaptor molecules DAP10 in CD8+ T cells and DAP10/DAP12 in NK cells interact with NKG2D.13 This differential expression could explain the different NKG2D‐dependent outcomes seen in both cell types. For example, in T cells, engagement of NKG2D costimulates T‐cell function as an amplifier to TCR‐mediated signaling while providing both activating and costimulatory signals in NK cells.16 In CD8+ T cells, the suggested major outcomes of NKG2D/DAP10 signaling are increased cytotoxicity via TCR signaling enhancement, although this article indicates this to not be the case given that NKG2D blockage in OT‐1 CD8+ T cells similarly resulted in a decreased infarct volume, suggesting a TCR‐independent mechanism. Whether other signals, such as IL‐15 (interleukin‐15), which has been shown to increase antigen‐independent cytotoxicity of certain CD8+ T cells,17 were altered in the current model remains to be established. Given that IL‐15 signaling contributes to ischemic brain injury through engagement of NK and CD8+ T cells,18 cooperativity of NKG2D and IL‐15 signaling to enhance NK and CD8+ T cell cytotoxicity could be explored in future studies. Enhanced T‐cell survival via Bcl‐2 upregulation has also been suggested, which may in the end contribute to a greater proportion of circulating and brain infiltrating cytotoxic T cells, thus enhancing overall neuroinflammation and neurotoxicity14 (Figure).Download figureDownload PowerPointFigure . Potential mechanism of NKG2D‐mediated CD8+ T‐cell cytotoxicity in stroke.The NKG2D ligands ULBP1 (MULT1), ULBP3, and ULBP4 are upregulated after cerebral ischemia in neurons, microglia, and astrocytes, respectively. ULBP presented to CD8+ T cells signals via NKG2D and DAP10, enhancing the PI3K (phosphoinositide 3‐kinase) pathway, leading to upregulation of Bcl‐2 and increasing CD8+ T‐cell survival. IL‐15 cytokine release and IL‐15 receptor signaling via DAP10 and NKG2D also potentiates the PI3K pathway, offering a potential TCR‐independent path for stimulating cytotoxicity in CD8+ T cells following ischemic brain injury. Bcl‐2 indicates B‐cell CLL/lymphoma 2; DAP10, DNAX‐activation protein 10; IL‐15, interleukin‐15; IL‐15R, interleukin‐15 receptor; NKG2D alias MULT1, killer cell lectin like receptor K1; and ULBP, UL16 binding protein.Other recent studies have shown pharmacological inhibition of NK cell activity during the acute stages of cerebral ischemia resulting in reduced interferon‐γ production and NKG2D expression by NK cells in the ischemic brain.19 The study suggested that STAT3 (signal transducer and activator of transcription 3) might regulate NKG2D expression, which should be also investigated in CD8+ T cells to gain further insight into the molecular mechanisms underlying NKG2D expression after stroke. Finally, to further validate the importance of this signaling molecule in ischemic brain injury, it would be prudent to replicate these pharmacological studies in NKG2D‐deficient mice, which have been studied in the context of NK cells and tumorigenesis.20 In summary, the work of David and colleagues identifies NKG2D signaling in CD8+ T cells as a relevant pathomechanism in ischemic brain injury, and additional studies should be pursued to better understand how precisely these mechanisms contribute to stroke outcome.DisclosuresNone.Footnotes*Correspondence to: Josef Anrather, VMD, Weill Cornell Medical College, Feil Family Brain and Mind Research Institute, 407 East 61st Street, RR 409, New York, NY 10065. Email: [email protected]cornell.eduSee Article by David et al.This article was sent to Neel S. Singhal, MD, PhD, Associate Editor, for editorial decision and final disposition.For Disclosures, see page 3.References1 Iadecola C, Anrather J. 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Published on behalf of the American Heart Association, Inc., by Wiley BlackwellThis is an open access article under the terms of the Creative Commons Attribution‐NonCommercial‐NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non‐commercial and no modifications or adaptations are made.https://doi.org/10.1161/JAHA.123.030482PMID: 37301754 Manuscript receivedMay 4, 2023Manuscript acceptedMay 5, 2023Originally publishedJune 10, 2023 KeywordsNKG2DEditorialsNK cellsstrokeCD8+ T cellsPDF download SubjectsIschemic Stroke