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The same types of cells can assume diverse states with varying functionalities. Effective cell therapy can be achieved by specifically driving a desirable cell state, which requires the elucidation of key transcription factors (TFs). Here, we integrated epigenomic and transcriptomic data at the systems level to identify TFs that define different CD8+T cell states in an unbiased manner. These TF profiles can be used for cell state programming that aims to maximize the therapeutic potential of T cells. For example, T cells can be programmed to avoid a terminal exhaustion state (TexTerm), a dysfunctional T cell state that is often found in tumors or chronic infections. However, TexTermexhibits high similarity with the beneficial tissue-resident memory T states (TRM) in terms of their locations and transcription profiles. Our bioinformatic analysis predictedZscan20, a novel TF, to be uniquely active in TexTerm. Consistently,Zscan20knock-out thwarted the differentiation of TexTermin vivo, but not that of TRM. Furthermore, perturbation ofZscan20programs T cells into an effector-like state that confers superior tumor and virus control and synergizes with immune checkpoint therapy. We also identifiedJdp2andNfil3as powerful TexTermdrivers. In short, our multiomics-based approach discovered novel TFs that enhance anti-tumor immunity, and enable highly effective cell state programming.One sentence summaryMultiomics atlas enables the systematic identification of cell-state specifying transcription factors for therapeutic cell state programming.

Catabolic responses of lean and fat energy stores are a component of the host response to infection. Cachexia is an extreme catabolic state characterized by unintentional weight loss and muscle loss, that can include fat loss. Whether cachexia plays any role in host defense or is a maladaptive consequence of host-pathogen interactions remains unknown. Traditionally studies have focused on understanding how inflammatory mediators and cells of the innate immune system contribute to the pathogenesis of cachexia and the depletion of energy stores. The cells of the adaptive immune system that regulate infection-induced cachexia remain elusive. In the present study, we examined the role of the adaptive immune response in cachexia pathogenesis using a murine model of the chronic parasitic infectionTrypanosoma brucei, the causative agent of sleeping sickness. We found that the cachectic response occurs in two phases with the first stage occurring early in the infection and involved a loss of body fat mass associated with anorexia, and the second stage occurring later in the infection and involved a sustained loss of fat mass that was accompanied by lean mass wasting. CD4+ T cells were necessary for the development of the sickness-induced anorexic response during stage 1 of the infection, which led to adipose triglyceride lipase dependent lipolysis in adipocytes and the resulting fat wasting. Adipose tissue wasting had no impact on host resistance defenses or survival of infection, both of which were antibody-mediated and independent ofCD4+ T cell responses. Our work reveals a new mechanism for infection induced cachexia involving CD4+ T cell regulation of host feeding behavior and an unexpected decoupling of adaptive immune mediated resistance from the cachectic response during infection.

In the tumor microenvironment, immune cells and tumor cells interact in a process called cancer immunoediting, giving rise to changes in gene expression, metabolism, mutational burden, and cellularity in the tumor. This SnapShot compares endogenous versus therapy-induced cancer immunoediting and outlines the molecular and cellular characteristics of interactions that result in complete tumor regression versus tumor escape and progression. To view this SnapShot, open or download the PDF.

SummaryThe limited efficacy of immunotherapies against glioblastoma illustrates the urgent need to better understand the interactions between the central nervous system and the immune system. Here, we showed that a protective response to αCTLA-4 therapy depended on a mutualistic relationship between microglia and CD4+ T cells. Suppression of gliomas by CD4+ T cells did not require tumor-intrinsic MHC-II expression, but rather was dependent on the selective expression of MHC-II and antigen presentation by local microglia that in turn, sustained CD4+ T cell tumoricidal effector functions. CD4+ T cell secretion of IFNγ made the glioma cells vulnerable to enhanced tumor surveillance and phagocytosis by microglia via the AXL/MER tyrosine kinase receptors that were necessary for tumor suppression. This work illustrates a novel partnership between CD4+ T cells and microglia that unleashes the tumoricidal properties of microglia that can be harnessed to improve immunotherapies for glioblastoma.

Summary Metabolic reprogramming is a hallmark of T cell activation and function. As our understanding of T cell metabolism increases, so does our appreciation of its inherent complexity. The metabolic heterogeneity of T cells that reside in different locations, such as lymphoid and non-lymphoid tissues, presents a challenge to developing therapies that exploit metabolic vulnerabilities. The roots of metabolic heterogeneity are only beginning to be understood. Here, we propose four factors that contribute to the adaptation of T cells to their dynamic tissue environment: (1) functional status of T cells, (2) local factors unique to the tissue niche, (3) type of inflammation, and (4) time spent in a specific tissue. We review emerging concepts about tissue-specific metabolic reprogramming in T cells with particular attention to explain how such metabolic properties are used as an adaptation mechanism. Adaptation of immune cells to the local microenvironment is critical for their persistence and function. Here, Varanasi et al. review the role and types of metabolic adaptation acquired by T cells in tissues and how these adaptations might differ between tissue type, disease state, and functionality of a T cell.

Light chain–associated amyloidosis is characterized by the extracellular deposition of amyloid fibrils in abdominothoracic organs, skin, soft tissue, and peripheral nerves. Phagocytic cells of the innate immune system appear to be ineffective at clearing the material; however, human light chain amyloid extract, injected subcutaneously into mice, is rapidly cleared in a process that requires neutrophil activity. To better elucidate the phagocytosis of light chain fibrils, a potential method of cell-mediated dissolution, amyloid-like fibrils were labeled with the pH-sensitive dye pHrodo red and a near infrared fluorophore. After injecting this material subcutaneously in mice, optical imaging was used to quantitatively monitor phagocytosis and dissolution of fibrils concurrently. Histologic evaluation of the residual fibril masses revealed the presence of CD68(+), F4/80(+), ionized calcium binding adaptor molecule 1(−) macrophages containing Congo red–stained fibrils as well as neutrophil-associated proteins with no evidence of intact neutrophils. These data suggest an early infiltration of neutrophils, followed by extensive phagocytosis of the light chain fibrils by macrophages, leading to dissolution of the mass. Optical imaging of this novel murine model, coupled with histologic evaluation, can be used to study the cellular mechanisms underlying dissolution of synthetic amyloid-like fibrils and human amyloid extracts. In addition, it may serve as a test bed to evaluate investigational opsonizing agents that might serve as therapeutic agents for light chain–associated amyloidosis.

Hepatocellular carcinoma (HCC) is an emerging health crisis due to rapidly increasing incidence rates and an overall 5-year survival rate of 18%. Consequently, there is an urgent need for new therapies and better preventive strategies. Aging and obesity are two of the biggest risk factors for HCC, and with ever-increasing aging and obese populations, uncovering how aging and obesity drive the increased risk for HCC has become imperative. Aging and obesity are well known to be characterized by immune dysregulation leading to persistent inflammation, and a dysregulated immune microenvironment is also a common theme shared by multiple hallmarks of HCC. In this study, we aim to assess the immune microenvironment of aged and obese mouse livers by performing spatial phenotyping of distinct immune signatures which can potentially predispose hepatocytes to HCC. To address this, we used Akoya’s ultra-high multiplexed CODEX (CO-Detection by indEXing) imaging system to interrogate 31 different biomarkers in liver samples at the single cell level. Specifically, we compared the liver immune microenvironment of mice within 2 different treatment groups (old mice on a high fat diet (HFD) and young mice on a normal diet (ND)) to the immune landscape of liver tumor tissue. Our goal is to identify distinct immune phenotypes present in both, liver tumor tissues and livers of HFD mice, compared to ND livers - to better understand how immune dysregulation can predispose an aged and obese liver to HCC. Citation Format: Adarsh Rajesh, Aaron Havas, Siva Karthik Varanasi, Katrina Evans, Bassem B. Cheikh, Clemens Duerrschmid, Najiba Mammadova, Peter Adams, Susan Kaech. a [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2531.

Glioblastoma multiforme (GBM) is the deadliest form of brain cancer, for which there is currently no effective treatment. Despite success of immunotherapies such as checkpoint inhibitors (CPI) in treating some cancers, less than 8% of GBM patients responded to anti-PD-1 therapy. Thus, we need to understand what mechanisms underlie tumor resistance to immunotherapies. In our GBM mouse model, we have found that anti-CTLA-4 treatment greatly improved survival and suppressed tumor growth, but surprisingly, its efficacy was lost in mice that were depleted of CD4+ but not CD8+ T cells, suggesting that CD4+ T cells are critical for protection against GBM. This matches clinical data showing decreased populations of CD4+ T cells in patients with adverse clinical outcomes such as high-grade GBM. Our preliminary results from flow cytometry and single-cell RNA sequencing (sc-RNAseq) also suggested enhanced cytokine production (IFNγ, TNFα, IL-2) in CD4+ T cells after anti-CTLA-4 treatment. Similarly, we found that microglia, the local sentinels of the central nervous system, also play a significant role in tumor control, as indicated by faster GBM growth and reduced number of tumor-infiltrating CD4+ T cells when mice are depleted of microglia by a CSF1R inhibitor (PLX3397). Additionally, microglia in GBM may serve as potential antigen-presenting cells, as shown by their high MHC class II (MHC-II) expression. Therefore, we postulate that microglia are necessary to sustain anti-tumor immunity by CD4+ T cells. Importantly, through a more detailed exploration of how CD4+ T cells and microglia interact to impact tumor control, our study will provide new insights for developing therapeutic strategies against “difficult-to-treat” cancers such as GBM.

SummaryT cell metabolic fitness plays a pivotal role in anti-tumor immunity and metabolic deregulation causes T cell dysfunction (i.e., ‘exhaustion’) in cancer. We identify that the scavenger receptor CD36 limits anti-tumor CD8+T cell effector functions through lipid peroxidation. In murine tumors, oxidized phospholipids (OxPLs) were highly abundant and CD8+TILs increased uptake and accumulation of lipids and lipid peroxidation. Functionally ‘exhausted’ CD8+TILs substantially increased CD36 expression and CD36-deficient CD8+TILs had more robust anti-tumor activity and cytokine production than wild-type cells. We further show that CD36 promotes uptake of oxidized low-density lipoproteins (OxLDL) and induces lipid peroxidation in CD8+TILs, and OxLDL inhibits CD8+T cell functions in a CD36-dependent manner. Moreover, glutathione peroxidase 4 (GPX4) over-expression lowers lipid peroxidation and restores functionalities in CD8+TILs. These results define a key role for an oxidized lipid-CD36 axis in promoting intratumoral CD8+T cell dysfunction.

This report deals with the possible mechanism by which IL-18 can contribute to the control and resolution of inflammatory lesions in the cornea caused by herpes simplex virus infection. Our results demonstrate that the expression of the IL-18R by both regulatory T cells (Treg) and effector T cells was a pivotal event that influenced lesion pathogenesis. The engagement of IL-18R on Treg with its cytokine ligand resulted in Amphiregulin expression a molecule associated with tissue repair. In support of this scheme of events, lesion severity became more severe in animals unable to express the IL-18R because of gene knockout and was reduced in severity when IL-18 was overexpressed in the cornea. These changes in lesion severity correlated with the frequency and number of both Treg and Teff that expressed Amphiregulin. Additional experiments indicated that IL-12 and IL-18 acted synergistically to enhance Amphiregulin expression in Treg, an event partly dependent on P38 MAPK activity. Finally, sub-conjunctival administration of Amphiregulin resulted in resolution of both developing and developed lesions. Thus, overall our results imply that IL-18 may participate in controlling the severity of SK and contribute to tissue repair by converting both Treg and effector T cells into those that produce Amphiregulin.

The outcome of virus infections depends on multiple factors. This review deals with the role of host metabolism as one such factor. We describe how different cells in the immune system employ differential metabolic pathways and how this relates to the outcome of virus infections. We also discuss how nutritional and metabolic diseases can influence the nature of viral pathogenesis as well as how targeted therapies against metabolic processes can impact on the outcome of virus infections. The case is also made for metabolic profiling as a potential tool to predict the outcome of a virus infection and to guide therapies that enhance host resistance.

Summary A common metabolic alteration in the tumor microenvironment (TME) is lipid accumulation, a feature associated with immune dysfunction. Here, we examined how CD8+ tumor infiltrating lymphocytes (TILs) respond to lipids within the TME. We found elevated concentrations of several classes of lipids in the TME and accumulation of these in CD8+ TILs. Lipid accumulation was associated with increased expression of CD36, a scavenger receptor for oxidized lipids, on CD8+ TILs, which also correlated with progressive T cell dysfunction. Cd36−/− T cells retained effector functions in the TME, as compared to WT counterparts. Mechanistically, CD36 promoted uptake of oxidized low-density lipoproteins (OxLDL) into T cells, and this induced lipid peroxidation and downstream activation of p38 kinase. Inhibition of p38 restored effector T cell functions in vitro, and resolution of lipid peroxidation by overexpression of glutathione peroxidase 4 restored functionalities in CD8+ TILs in vivo. Thus, an oxidized lipid-CD36 axis promotes intratumoral CD8+ T cell dysfunction and serves as a therapeutic avenue for immunotherapies.

This report deals with physiological changes and their implication following ocular infection with HSV. This infection usually results in a blinding inflammatory reaction in the cornea, orchestrated mainly by proinflammatory CD4 T cells and constrained in severity by regulatory T cells. In the present report, we make the unexpected finding that blood glucose levels change significantly during the course of infection. Whereas levels remained normal during the early phase of infection when the virus was actively replicating in the cornea, they increased around 2-fold during the time when inflammatory responses to the virus was occurring. We could show that glucose levels influenced the extent of induction of the inflammatory T cell subset in vitro that mainly drives lesions, but not regulatory T cells. Additionally, if glucose utilization was limited in vivo as a consequence of therapy in the inflammatory phase with the drug 2-deoxy-glucose (2DG), lesions were diminished compared with untreated infected controls. In addition, lesions in 2DG-treated animals contained less proinflammatory effectors. Glucose metabolism also influenced the acute phase of infection when the replicating virus was present in the eye. Thus, therapy with 2DG to limit glucose utilization caused mice to become susceptible to the lethal effects of HSV infection, with the virus spreading to the brain causing encephalitis. Taken together, our results indicate that glucose metabolism changed during the course of HSV infection and that modulating glucose levels can influence the outcome of infection, being detrimental or beneficial according to the stage of viral pathogenesis.

Our knowledge of T cell metabolism relies primarily on studies performed in vitro that may not fully recapitulate physiological conditions in vivo. In this issue of Immunity, Ma et al. find that the in vivo environment dictates the metabolic phenotype of effector CD8+ T cells—particularly their glucose utilization.

Ocular infection with herpes simplex virus (HSV) can result in a chronic immune inflammatory lesion that is a significant cause of human blindness. A key to controlling stromal keratitis (SK) lesion severity is to identify cellular and molecular events responsible for tissue damage and to counteract them. One potentially useful approach to achieve such therapy is Retinoic Acid (RA). Here we show that RA therapy reduces the severity of SK by having inhibitory effects on the T effector subtypes responsible for orchestrating SK. RA also served to stabilize the function of regulatory T cell (Treg) which counteract inflammatory cell activity. The Treg stabilizing effect was demonstrated by in vitro studies where RA was shown to retain Foxp3 expression when exposed to proinflammatory conditions such as IL-12 and IL-6+TGF-β. in vivo studies revealed that RA exerted its stabilizing effects by downregulating IL-6R expression on Treg after HSV-1 infection and this helped to control the progression of SK. Since the therapy was effective when used both early and after the initiation of lesions, it may represent a valuable means of therapy when used alone or along with additional therapies.

Herpes simplex 1 infection of the eye can cause blindness with lesions in the corneal stroma largely attributable to inflammatory events that include components of both adaptive and innate immunity. Several innate immune responses are triggered by herpes simplex 1, but it is unclear how such innate events relate to the subsequent development of stromal keratitis. In this study, we compared the outcome of herpes simplex 1 ocular infection in mice unable to express NLRP3 because of gene knockout (NLRP3−/−) to that of wild-type mice. The NLRP3−/− mice developed more-severe and earlier stromal keratitis lesions and had higher angiogenesis scores than did infected wild-type animals. In addition, NLRP3−/− mice generated an increased early immune response with heightened chemokines and cytokines, including interleukin-1β and interleukin-18, and elevated recruitment of neutrophils. Increased numbers of CD4+ T cells were seen at later stages of the disease in NLRP3−/− animals. Reduction in neutrophils prevented early onset of the disease in NLRP3−/− animals and lowered levels of bioactive interleukin-1β but did not lower bioactive interleukin-18. In conclusion, our results indicate that NLRP3 has a regulatory and beneficial role in herpetic stromal keratitis pathogenesis.

Ocular infection with herpes simplex virus 1 can result in a chronic immunoinflammatory stromal keratitis (SK) lesion that is a significant cause of human blindness. A key to controlling SK lesion severity is to identify cellular and molecular events responsible for tissue damage and to manipulate them therapeutically. Potential targets for therapy are miRNAs, but these are minimally explored especially in responses to infection. Here, we demonstrated that Mir155 expression was up-regulated after ocular herpes simplex virus 1 infection, with the increased Mir155 expression occurring mainly in macrophages and CD4 + T cells and to a lesser extent in neutrophils. In vivo studies indicated that Mir155 knockout mice were more resistant to herpes SK with marked suppression of T helper cells type 1 and 17 responses both in the ocular lesions and the lymphoid organs. The reduced SK lesion severity was reflected by increased phosphatidylinositol-3,4,5-trisphosphate 5-phosphatase 1 and interferon-γ receptor α-chain levels in activated CD4 + T cells in the lymph nodes. Finally, in vivo silencing of miR-155 by the provision of antagomir-155 nanoparticles to herpes simplex virus 1–infected mice led to diminished SK lesions and corneal vascularization. In conclusion, our results indicate that miR-155 contributes to the pathogenesis of SK and represents a promising target to control SK severity.

Ocular infection with HSV causes a chronic T cell–mediated inflammatory lesion in the cornea. Lesion severity is affected by the balance of different CD4 T cell subsets, with greater severity occurring when the activity of regulatory T cells (Tregs) is compromised. In this study, fate-mapping mice were used to assess the stability of Treg function in ocular lesions. We show that cells that were once Foxp3+ functional Tregs may lose Foxp3 and become Th1 cells that could contribute to lesion expression. The instability primarily occurred with IL-2Rlo Tregs and was shown, in part, to be the consequence of exposure to IL-12. Lastly, in vitro–generated induced Tregs (iTregs) were shown to be highly plastic and capable of inducing stromal keratitis when adoptively transferred into Rag1−/− mice, with 95% of iTregs converting into ex-Tregs in the cornea. This plasticity of iTregs could be prevented when they were generated in the presence of vitamin C and retinoic acid. Importantly, adoptive transfer of these stabilized iTregs to HSV-1–infected mice prevented the development of stromal keratitis lesions more effectively than did control iTregs. Our results demonstrate that CD25lo Treg and iTreg instability occurs during a viral immunoinflammatory lesion and that its control may help to avoid lesion chronicity.

Summary Immunotherapy has revolutionized cancer treatment, yet most patients do not respond. Here, we investigated mechanisms of response by profiling the proteome of clinical samples from advanced stage melanoma patients undergoing either tumor infiltrating lymphocyte (TIL)-based or anti- programmed death 1 (PD1) immunotherapy. Using high-resolution mass spectrometry, we quantified over 10,300 proteins in total and ∼4,500 proteins across most samples in each dataset. Statistical analyses revealed higher oxidative phosphorylation and lipid metabolism in responders than in non-responders in both treatments. To elucidate the effects of the metabolic state on the immune response, we examined melanoma cells upon metabolic perturbations or CRISPR-Cas9 knockouts. These experiments indicated lipid metabolism as a regulatory mechanism that increases melanoma immunogenicity by elevating antigen presentation, thereby increasing sensitivity to T cell mediated killing both in vitro and in vivo. Altogether, our proteomic analyses revealed association between the melanoma metabolic state and the response to immunotherapy, which can be the basis for future improvement of therapeutic response.

Stromal keratitis (SK) is a chronic immunopathological lesion of the eye, caused by HSV-1 infection, and a common cause of vision impairment in humans. The inflammatory lesions in the cornea are primarily caused by neutrophils with the active participation of CD4+ T cells. Therefore, the targeting of these immune cell types and their products represents a potentially valuable form of therapy to reduce the severity of disease. Resolvin D1 (RvD1) and its epimer aspirin-triggered RvD1 (AT-RvD1) are lipid mediators derived from docosahexaenoic acid (DHA) and were shown to promote resolution in several inflammatory disease models. In this report, we examined whether AT-RvD1 administration, begun before infection or at a later stage after ocular infection of mice with HSV-1, could control the severity of SK lesions. Treatment with AT-RvD1 significantly diminished the extent of corneal neovascularization and the severity of SK lesions. AT-RvD1-treated mice had fewer numbers of inflammatory cells that included neutrophils as well as Th1 and Th17 cells in the infected cornea. The mechanisms by which AT-RvD1 acts appear to be multiple. These include inhibitory effects on proinflammatory mediators, such as IL-1β, IL-6, IL-12, CXCL1, MCP-1, MIP-2, vascular endothelial growth factor (VEGF)-A, matrix metalloproteinase 9 (MMP-9), and proinflammatory miRNA, such as miR-155, miR-132, and miR-223, which are involved in SK pathogenesis and corneal neovascularization. In addition, AT-RvD1 attenuated STAT1, which plays an important role in Th1 cell differentiation and IFN-γ expression. These findings demonstrate that AT-RvD1 treatment could represent a useful strategy for the management of virus-induced immunopathological lesions.

Ocular infection with herpes simplex virus 1 (HSV-1) sets off an inflammatory reaction in the cornea which leads to both virus clearance and chronic lesions that are orchestrated by CD4 T cells. Approaches that enhance the function of regulatory T cells (Treg) and dampen effector T cells can be effective to limit stromal keratitis (SK) lesion severity. In this report, we explore the novel approach of inhibiting DNA methyltransferase activity using 5-azacytidine (Aza; a cytosine analog) to limit HSV-1-induced ocular lesions. We show that therapy begun after infection when virus was no longer actively replicating resulted in a pronounced reduction in lesion severity, with markedly diminished numbers of T cells and nonlymphoid inflammatory cells, along with reduced cytokine mediators. The remaining inflammatory reactions had a change in the ratio of CD4 Foxp3 + Treg to effector Th1 CD4 T cells in ocular lesions and lymphoid tissues, with Treg becoming predominant over the effectors. In addition, compared to those from control mice, Treg from Aza-treated mice showed more suppressor activity in vitro and expressed higher levels of activation molecules. Additionally, cells induced in vitro in the presence of Aza showed epigenetic differences in the Treg-specific demethylated region (TSDR) of Foxp3 and were more stable when exposed to inflammatory cytokines. Our results show that therapy with Aza is an effective means of controlling a virus-induced inflammatory reaction and may act mainly by the effects on Treg. IMPORTANCE HSV-1 infection has been shown to initiate an inflammatory reaction in the cornea that leads to tissue damage and loss of vision. The inflammatory reaction is orchestrated by gamma interferon (IFN-γ)-secreting Th1 cells, and regulatory T cells play a protective role. Hence, novel therapeutics that can rebalance the ratio of regulatory T cells to effectors are a relevant issue. This study opens up a new avenue in treating HSV-induced SK lesions by increasing the stability and function of regulatory T cells using the DNA methyltransferase inhibitor 5-azacytidine (Aza). Aza increased the function of regulatory T cells, leading to enhanced suppressive activity and diminished lesions. Hence, therapy with Aza, which acts mainly by its effects on Treg, can be an effective means to control virus-induced inflammatory lesions.

Ocular infection with Herpes Simplex Virus type 1 (HSV-1) sets off an inflammatory reaction in the cornea which leads to both virus clearance as well as chronic lesions that are orchestrated by CD4 T cells. Effector CD4+ T cells rely on glycolysis for inflammatory effector functions and Treg rely on mitochondrial metabolism. How these processes contribute to Herpes stromal keratitis (HSK) is unclear. We show that glycolysis is elevated in CD4+ T cells from mice with HSK as compared to non-immunized controls. In addition, glucose uptake by CD4 T cells at the lesion site and draining lymph nodes were higher in animals with HSK. In vitro, T cells activation led to enhanced glycolysis, which was inhibited by inhibitor 2-deoxy-D-glucose (2DG). Th1 generated in vitro displayed enhanced glycolysis rates compared to Treg cells. Hence, inhibition of glycolysis by 2DG inhibited the differentiation and function of interferon-γ (IFN-γ) producing Th1 cells, largely un-affecting Treg cells. In vivo, treatment of animals with 2DG normalized T cell metabolism rebalancing the Treg to Th1 ratio and limited the SK lesion progression. Furthermore, animals infected with HSV-1 displayed enhanced blood glucose levels at time points when the inflammation at draining lymph nodes and lesion site are at its peak, which may both be the cause and effect of the inflammation. These results suggest that normalization of T cell metabolism through inhibition of glycolysis is a promising therapeutic venue for HSK.

Ocular infection with herpes simplex virus 1 (HSV-1) can result in a chronic immuno-inflammatory lesion that is a significant cause of human blindness. IFN-γ producing CD4 + T cells are generally considered the main orchestrators, and lesions are more severe if the regulatory T cell (Treg) response is compromised. Tregs have been shown to lose FoxP3 (Treg lineage factor) and adopt alternate lineage fates in a changing cytokine environment. Moreover, little is known about the stability of Treg cells in an ongoing inflammatory reaction such as is the case of SK. In this study-using fate mapping mice, we were able to demonstrate that the population of ex-Tregs increased at the site of infection that is the cornea and also at the secondary lymphoid organs after HSV-1 infection. In vivo studies showed that these ex-Tregs acquired the Th1 and Th17 phenotype, which may play a destructive role in stromal keratitis. We also demonstrate in vivo that CD25lo subset of Tregs is less suppressive and more prone to conversion than CD25hi Tregs. Additionally, in-vitro studies suggested that Tregs in the presence of IL-12 converted into ex-Tregs, however this plasticity was prevented when Tregs were generated in the presence of DNMT inhibitor (Azacytidine). Thus, plasticity of Tregs might represent a problem during SK, which needs to be controlled with appropriate therapeutic procedures. Overall, our results indicate that Tregs lose FoxP3 expression in ocular inflammatory setting and drugs that reverse Treg plasticity could be useful to restore Treg function and to inhibit ocular immunopathology.