Virus-specific Memory CD8 Research Articles

Glucocorticoid-mediated Suppression of Effector Programming Assists the Memory Transition of Virus-specific CD8+ T Cells.

We demonstrate the role of signaling via the glucocorticoid receptor, NR3C1, in differentiation of CD8+ T cell memory. Pharmacological inhibition as well as the short hairpin RNA-mediated knockdown of the receptor hindered memory transition and limited the homeostatic turnover of the activated CD8+ T cells. Dexamethasone exposure of CD8+ T cells expanded during a resolving infection with influenza A virus or a γ-herpesvirus promoted conversion of effector cells into memory cells by modulating cellular metabolism and lowering the accumulation of reactive oxygen species. Reduced reactive oxygen species levels in the responding effector cells upregulated Bcl2 and enhanced survival. The generated virus-specific memory CD8+ T cells were efficiently recalled following challenge of animals with a secondary infection to control it better. The memory-enhancing effect was predominantly evident at low doses of dexamethasone. Therefore, controlled glucocorticoid signaling within the effector CD8+ T cells is crucial for optimal memory differentiation.

Immunological memory diversity in the human upper airway.

The upper airway is an important site of infection, butimmune memory in the human upper airway is poorly understood, with implications for COVID-19 and many other human diseases1-4. Here we demonstrate that nasal and nasopharyngeal swabs can be used to obtain insights into these challenging problems, and define distinct immune cell populations, including antigen-specific memory Bcells and T cells, in two adjacent anatomical sites in the upper airway. Upper airway immune cell populations seemed stable over time in healthy adults undergoing monthly swabs for more than 1 year, and prominent tissue resident memory T (TRM)cell and B (BRM) cellpopulations were defined. Unexpectedly, germinal centre cells were identified consistently in many nasopharyngeal swabs. In subjects with SARS-CoV-2 breakthrough infections, local virus-specific BRMcells, plasma cells and germinal centre B cells were identified, with evidence of local priming and an enrichment of IgA+ memory B cells in upper airway compartments compared with blood. Local plasma cell populations were identified with transcriptional profiles of longevity. Local virus-specific memory CD4+ TRM cellsand CD8+ TRM cellswere identified, with diverse additional virus-specific T cells. Age-dependent upper airway immunological shifts were observed. These findings provide new understanding of immune memory at a principal mucosal barrier tissue in humans.

Lung influenza virus-specific memory CD4 T cell location and optimal cytokine production are dependent on interactions with lung antigen-presenting cells

Influenza A virus (IAV) infection leads to the formation of mucosal memory CD4 T cells that can protect the host. An in-depth understanding of the signals that shape memory cell development is required for more effective vaccine design. We have examined the formation of memory CD4 T cells in the lung following IAV infection of mice, characterizing changes to the lung landscape and immune cell composition. IAV-specific CD4 T cells were found throughout the lung at both primary and memory time points. These cells were found near lung airways and in close contact with a range of immune cells including macrophages, dendritic cells, and B cells. Interactions between lung IAV-specific CD4 T cells and major histocompatibility complex (MHC)II+ cells during the primary immune response were important in shaping the subsequent memory pool. Treatment with an anti-MHCII blocking antibody increased the proportion of memory CD4 T cells found in lung airways but reduced interferon-γ expression by IAV-specific immunodominant memory CD4 T cells. The immunodominant CD4 T cells expressed higher levels of programmed death ligand 1 (PD1) than other IAV-specific CD4 T cells and PD1+ memory CD4 T cells were located further away from MHCII+ cells than their PD1-low counterparts. This distinction in location was lost in mice treated with anti-MHCII antibodies. These data suggest that sustained antigen presentation in the lung impacts the formation of memory CD4 T cells by regulating their cytokine production and location.

Myeloid derived suppressor cells potentiate virus-specific memory CD8+ T cell response

How therapeutically administered myeloid derived suppressor cells (MDSCs) modulate differentiation of virus-specific CD8+ T cell during virus infection was investigated. In vitro generated MDSCs from bone marrow precursors inhibited the expansion of stimulated CD8+ T cells but the effector cells in the recipients of MDSCs showed preferential memory transition during Influenza A virus (IAV) or an α− (Herpes Simplex Virus) as well as a γ- (murine herpesvirus 68) herpesvirus infection. Memory CD8+ T cells thus generated constituted a heterogenous population with a large fraction showing effector memory (CD62LloCCR7-) phenotype. Such cells could be efficiently recalled in the rechallenged animals and controlled the secondary infection better. Memory potentiating effects of MDSCs occurred irrespective of the clonality of the responding CD8+ T cells as well as the nature of infecting viruses. Compared to the MDSCs recipients, effector cells of MDSCs recipients showed higher expression of molecules known to drive cellular survival (IL-7R, Bcl2) and memory formation (Tcf7, Id3, eomesodermin). Therapeutically administered MDSCs not only mitigated the tissue damaging response during a resolving IAV infection but also promoted the differentiation of functional memory CD8+ T cells. Therefore, MDSCs therapy could be useful in managing virus-induced immunopathological reactions without compromising immunological memory.

Virus-Specific CD8+ Stem Memory T Lymphocytes Display Enhanced Anti-Virus Responses

Virus specific memory CD8+ T cells can be divided into three subsets, stem memory (T SCM), central (T CM) and effector (T EM), but their lineage relationships and their ability to persist and confer protective immunity against virus infection are not well understood. Our results show that virus-specific CD8+T SCM have a greater capacity than T CM and T EM to persist in vivo and in vitro and are more efficient in mediating protective immunity against CMV infection and EBV infection because of their increased self-renew potential whether in transcriptome or in epigenome. We firstly expanded virus-specific T cells containing CD8+TSCM and CD8+TCM cells in vitro by stimulating healthy human PBMCs with cytokines and CMV (pp65 and IE1) and EBV (LMP2a and EBNA1) virus peptides. Then we fully evaluated the phenotypic and functional differences of diverse virus-specific memory T subsets and found that CD8+TSCM cells are a subset of low exhaustion, high migration and high secretion of IL-2. However, the expression of IFN-γand TNF-a and CD107a were lower compared with TEM cells, although comparable to CD8+TCM cells. We then evaluated the self-renewal and differentiation capacity of CD8+TSCM cells, compared with TCM or TEM cells. The results showed after stimulated with a-CD3/CD2/CD28 for 5 days, that CD8+TSCM cells possess around 50% of CD45RA+CD62L+ subpopulation cells. Meanwhile, CD8+TSCM cells constantly keep a higher expression of CD45RA+ cells than TCM and TEM cells in different proliferation generations. Notably, around 20% of TCM cells,5% of TEM cells and 10% of TEFF cells were found, suggesting that TSCM cells can reconstitute the whole memory T subsets. By contrast, the TCM cells mainly retained a central memory phenotype or differentiated into effector memory T cells and the TEM cells maintained its original phenotype. We also used the stem cell index to evaluate the stemness of different subgroups, and the results confirmed that the stemness evaluation of TSCM cells were significantly higher than TCM and TEM subsets. In vivo rein-fusion of different virus-specific T cell into irradidated NPG mice also showed that CD8+TSCM cells in vivo can differentiate into all memory T subsets in vivo while TCM cells mainly retained TEM and TCM subgroups at 4W post T cell infusion in mouse liver,lung and spleen(Figure 1A-G). To explore whether the antiviral ability of TSCM cells lasted longer than that of TCM and TEM cells. We first evaluated the apoptosis of different T cell subsets stimulated by AD169 virus and found less apoptosis rate was lower than TCM and TEM cells, suggesting persistent virus infection leads to damage to T cells and CD8+TSCM cells have a strong ability to resist virus damage, which may potentially lead to stronger and longer lasting antiviral effects. Based on this, we also evaluated the apoptosis level of virus-infected MRC-5 cells to evaluate whether CD8+TSCM cells exert a stronger anti-virus effect. The result showed that AD-169 infected MRC-5 cells co-incubated with CD8+TSCM had a higher proportion of apoptotic level than CD8+TCM cells. We also used the 5day-long-term infection model in vitro to evaluate the effect of virus infection on T cell differentiation and evaluated the change of virus copy number after long-term co-culture of different cell subsets. We found that sorted CD8+TSCM system can significantly reduce the virus copy number compared with the control group, while there is no significant difference among TCM and TEM cells (Figure1H,). Further analysis into survival ability of T cell subpopulations found that the proportion of CD3+ T cells in TSCM group were significantly higher and the CD8+TSCM group was dominated by TCM and TEM cells after 5-day stimulation but still kept about 22 percentage of TSCM phenotype. (Figure1I). In summary, CD8+ TSCM cells have a sustainable ability against CMV than TCM and TEM. To examine the in vivo activity of virus-specific CD8+TSCM cells. We used Raji-HLA*A0201-pp65-luciferase cells to construct tumor infiltration mouse model. The result showed that in contrast to control mice, we observed a reduction in bioluminescence over three weeks in mice treated with CD8+TSCM cells. Although mice treated with CD8+TCM and CD8+TEM cells retained a lower tumor size in the early two weeks post adoptive T infusion, they showed a rapid tumor progresses and event lead to shorter survival compared with CD8+TSCM group, indicating a sustained anti-CMV effect of CD8+TSCM cells.

Single-cycle influenza virus vaccine generates lung CD8+ Trm that cross-react against viral variants and subvert virus escape mutants.

Influenza virus-specific tissue-resident memory (Trm) CD8+ T cells located along the respiratory tract provide cross-strain protection against a breadth of influenza viruses. We show that immunization with a single-cycle influenza virus vaccine candidate (S-FLU) results in the deposition of influenza virus nucleoprotein (NP)-specific CD8+ Trm along the respiratory tract that were more cross-reactive against viral variants and less likely to drive the development of cytotoxic T lymphocyte (CTL) escape mutants, as compared to the lung memory NP-specific CD8+ T cell pool established following influenza infection. This immune profile was linked to the limited inflammatory response evoked by S-FLU vaccination, which increased TCR repertoire diversity within the memory CD8+ T cell compartment. Cumulatively, this work shows that S-FLU vaccination evokes a clonally diverse, cross-reactive memory CD8+ T cell pool, which protects against severe disease without driving the virus to rapidly evolve and escape, and thus represents an attractive vaccine for use against rapidly mutating influenza viruses.

Radiation causes long term changes in virus-specific and virtual memory CD4 T cells

Abstract Threat of nuclear attack and accidents at nuclear power plants increase our risk of radiation exposure. Radiation effects on immune homeostasis are well-studied, but their effect on memory CD4 T cells is poorly defined. To explore that, we adoptively transferred naïve Thy1.1 TCR-Tg SMARTA CD4 T cells specific for LCMV-derived gp61 epitope into naïve Thy1.2 C57BL/6 mice before LCMV-Armstrong infection. At memory time point, infected recipient mice were subjected to 0Gy (non-radiated, control group) or 5Gy of total body radiation. We found that naïve CD4 T cells suffered significant numerical loss post radiation, but they numerically recovered in few weeks through increased proliferative expansion. However, radiated SMARTA and endogenous GP61-specific memory CD4 T cells sustained a prolonged loss in number that fail to recover even after 3 months. Radiation also leads to transient impairment in the ability of memory CD4 T cells to produce cytokines (IFNγ, TNFα) in response to cognate antigen stimulation, or to proliferate upon adoptive transfer into new hosts in response to secondary LCMV infection. Interestingly, the endogenous ‘effector-like’ CD4 T cell numbers recovered faster than SMARTA memory T cells. We hypothesized that virtual memory T cells (CD4 Tvm) are contributing to the recovery of this effector like CD4 T cells in the radiation mediated lymphoreplete conditions. Tetramer enrichment of CD4 T cells specific to the gp66 and 2W1S epitope showed that radiation increases the representation of gp66 specific CD4 Tvm in naïve mice. Together, these data suggest that radiation poses long-lasting defects to virus specific memory CD4 T cell responses, and it facilitates CD4 Tvm proliferation to overcome the lymphopenia.

Acute and chronic viral infection generate brain-resident memory T cells with distinct phenotypic, functional, spatial, and metabolic profiles

Abstract The rising incidence of post-viral neurological sequelae represents a substantial burden on global public health. Therefore, there is an urgent need to investigate alterations in brain immune composition after acute and chronic viral infection. While virus-specific tissue-resident memory (T RM) CD8 +T cells are observed in human health and neurological disease, these populations remain understudied in the brain. Here, we leveraged the comparative model system of lymphocytic choriomeningitis virus (LCMV) strain Armstrong vs. Clone 13 infection in mice to generate brain T RMwith quiescent and exhausted profiles respectively. We then interrogated the 1) phenotype, 2) functionality, 3) localization, and 4) metabolism of anti-viral T RMin the brain. Importantly, brain T RMexhibit higher expression of exhaustion markers (PD-1, LAG-3, TIM-3, TOX) compared to T circulating memory (T CIRCM) cells in the same host. Furthermore, brain T RMin chronically infected hosts are functionally impaired in cytokine production and cytolytic capacity. Anti-viral brain T RMseed distinct neuroanatomical sites, such as the cerebrospinal fluid, parenchyma, and choroid plexus, often adopting aggregate-like structures in solid compartments. Compared to T CIRCM, quiescent and exhausted brain T RMalso demonstrate higher expression of surface amino acid transporters with disparate amino acid uptake. These findings are contextualized by whole brain metabolomic profiling indicating global perturbations in amino acid metabolism post-acute and chronic viral infection. Altogether, these data highlight brain-intrinsic memory T cell signatures and offer broad insight into how brain T RMmay influence post-viral neurological outcomes. Supported by grants from the NIH (AI042767, AI114543, AI167847, 5T32GM007337) and the University of Iowa Post-Comprehensive Research Fellowship.

Effect of estrogen signaling in CD8 T cell mediated antiviral response

Abstract Estrogen, the major female sex hormone has been linked with immune response disparity between males and females. However, the exact mechanism by which estrogen impacts our ability to fight viral infection and development of immunologic memory remains unknown. CD8 T cells represent the primary protective mechanism against viruses and cancer. Here we aim to understand how CD8 T cell mediated antiviral immune response is regulated by estrogen. Systemic LCMV infection in ovariectomized female mice generated lower virus-specific memory CD8 T cells compared to sham surgery controls. To check if this reduction in memory response is CD8 T cell intrinsic, I generated a transgenic virus-specific estrogen receptor-α deficient CD8 mouse line (ESR1 KO). Using a co-adoptive transfer model, I compared the differentiation of ESR1 KO and control CD8 T cells in both systemic LCMV and local influenza infection. The lack of estrogen signaling resulted in reduced abundance of effector CD8 T cells in the context of both infections. ESR1 KO CD8 T cells also displayed reduced cytotoxic ability. In vitrostudies revealed that the CD8 T cell expansion was primarily impacted by the lack of estrogen signaling, whereas activation and cellular apoptosis remain unchanged. Altogether, this suggests a crucial role of sex steroid estrogen in initial effector CD8 T cell differentiation, that has significant ramification for generation of protective immune responses in pathological or physiological estrogen deficiency. Supported by grants from Searle Scholars Foundation, NIH 2P20GM109035, NIH P20GM121298, NIH T32HL134625, NIH T32GM136566

Virus-Specific Stem Cell Memory CD8+ T Cells May Indicate a Long-Term Protection against Evolving SARS-CoV-2

Immune memory to SARS-CoV-2 is key for establishing herd immunity and limiting the spread of the virus. The duration and qualities of T-cell-mediated protection in the settings of constantly evolving pathogens remain an open question. We conducted a cross-sectional study of SARS-CoV-2-specific CD4+ and CD8+ T-cell responses at several time points over 18 months (30-750 days) post mild/moderate infection with the aim to identify suitable methods and biomarkers for evaluation of long-term T-cell memory in peripheral blood. Included were 107 samples from 95 donors infected during the periods 03/2020-07/2021 and 09/2021-03/2022, coinciding with the prevalence of B.1.1.7 (alpha) and B.1.617.2 (delta) variants in Bulgaria. SARS-CoV-2-specific IFNγ+ T cells were measured in ELISpot in parallel with flow cytometry detection of AIM+ total and stem cell-like memory (TSCM) CD4+ and CD8+ T cells after in vitro stimulation with peptide pools corresponding to the original and delta variants. We show that, unlike IFNγ+ T cells, AIM+ virus-specific CD4+ and CD8+ TSCM are more adequate markers of T cell memory, even beyond 18 months post-infection. In the settings of circulating and evolving viruses, CD8+ TSCM is remarkably stable, back-differentiated into effectors, and delivers immediate protection, regardless of the initial priming strain.

Functional virus-specific memory CD8+ T cells survey glioblastoma

Abstract Glioblastoma multiforme (GBM) is among the most aggressive, treatment resistant cancers, and despite standard of care surgery, radiation and chemotherapy, is invariably fatal. GBM is marked by local and systemic immunosuppression, contributing to resistance to existing immunotherapies that have had success in other tumor types. Memory T cells specific for previous infections reside in tissues throughout the host, including the brain, and are capable of rapid and potent immune activation. Here, we show that virus-specific memory CD8+ T cells expressing tissue resident markers populate the mouse and human glioblastoma microenvironment. Reactivating virus-specific memory T cells through intra-tumoral delivery of adjuvant-free virus-derived peptide triggered local immune activation. This delivery translated to anti-neoplastic effects, which improved survival in a murine glioblastoma model. Our results indicate that virus-specific memory T cells are a significant part of the glioblastoma immune microenvironment and may be leveraged to promote anti-tumoral immunity. Supported by UMN SPORE Program Project Planning grant (DM, CCC), NCI 1R01CA238439 (DM), Humor to Fight the Tumor Foundation (JN), NCI 5P30CA023108-42 (PR)

Functional virus-specific memory T cells survey glioblastoma.

Glioblastoma multiforme (GBM) is among the most aggressive, treatment-resistant cancers, and despite standard of care surgery, radiation and chemotherapy, is invariably fatal. GBM is marked by local and systemic immunosuppression, contributing to resistance to existing immunotherapies that have had success in other tumor types. Memory T cells specific for previous infections reside in tissues throughout the host and are capable of rapid and potent immune activation. Here, we show that virus-specific memory CD8 + T cells expressing tissue-resident markers populate the mouse and human glioblastoma microenvironment. Reactivating virus-specific memory T cells through intratumoral delivery of adjuvant-free virus-derived peptide triggered local immune activation. This delivery translated to antineoplastic effects, which improved survival in a murine glioblastoma model. Our results indicate that virus-specific memory T cells are a significant part of the glioblastoma immune microenvironment and may be leveraged to promote anti-tumoral immunity.

The kinase complex mTORC2 promotes the longevity of virus-specific memory CD4+ T cells by preventing ferroptosis.

Antigen-specific memory CD4+ T cells can persist and confer rapid and efficient protection from microbial reinfection. However, the mechanisms underlying the long-term maintenance of the memory CD4+ T cell pool remain largely unknown. Here, using a mouse model of acute infection with lymphocytic choriomeningitis virus (LCMV), we found that the serine/threonine kinase complex mammalian target of rapamycin complex 2 (mTORC2) is critical for the long-term persistence of virus-specific memory CD4+ T cells. The perturbation of mTORC2 signaling at memory phase led to an enormous loss of virus-specific memory CD4+ T cells by a unique form of regulated cell death (RCD), ferroptosis. Mechanistically, mTORC2 inactivation resulted in the impaired phosphorylation of downstream AKT and GSK3β kinases, which induced aberrant mitochondrial reactive oxygen species (ROS) accumulation and ensuing ferroptosis-causative lipid peroxidation in virus-specific memory CD4+ T cells; furthermore, the disruption of this signaling cascade also inhibited glutathione peroxidase 4 (GPX4), a major scavenger of lipid peroxidation. Thus, the mTORC2-AKT-GSK3β axis functions as a key signaling hub to promote the longevity of virus-specific memory CD4+ T cells by preventing ferroptosis.

Phenotype of Memory HCV CD8+ T Cells in Chronic HCV

Abstract Background Hepatitis C virus (HCV) infection is a major cause of chronic liver disease, with approximately 71 million chronically infected individuals worldwide. After acute infection, a majority of healthy adults will develop persistent viremia. An infection with HCV activates the immune system to defend the host with a broad range of innate and adaptive immune responses. Cellular immune response shows antiviral immunity by means of virus specific CD8+ T cytotoxic lymphocytes and CD4+ T helper cells. After resolution of infection, a population of the virus-specific memory CD8+ T cells protects the host against reinfection. These subsets of memory cells differ with respect to their anatomical localization, their phenotype and their protective potential. Objectives The aim of the present study is to find out the phenotype pattern of HCV specific memory CD8+ T cells in chronic HCV patients in comparison to HCV exposed seronegative high-risk individuals. Patients and Methods This case control study included 60 individuals categorized into two main groups, (group I) included 20 chronic HCV patients and (group II= control group) included 40 HCV seronegative healthcare workers. Both groups were subjected to carboxyfluorescein diacetate succinimidyl ester (CFSE) proliferation assay in response to HCV specific peptides. Group II was divided into 2 subgroups according to proliferation index (cut-off<2) into, proliferation index +ve high-risk group (+PIHRG) (20 healthcare workers) and proliferation index -ve high-risk group (-PIHRG) (20 healthcare workers). Then flow cytometric analysis of HCV specific CD8+ T cells was done for the all subjects to determine the expression of CD127, CCR7, CD45RO and CD45RA on both proliferating and non-proliferating fractions of cells. Results We found that expression of CD45RA was significantly increased and CCR7 was significantly decreased on HCV CD8+ T cells in the chronic HCV group than in the high risk groups in response to different HCV peptides. Conclusion During persistent viremia in chronic HCV infection subthreshold signal levels established by HCV different proteins provide anergy of virus specific memory CD8+ T cells that are unable to differentiate into full effector cells upon a possible reencounter with the antigen. So, chronic HCV patients in our study showed predominance of less differentiated effector memory phenotype (CD45RA+, CCR7-) in peripheral blood in comparison to HCV exposed seronegative high-risk individuals who don’t show overt infection.

B cells promote CD8 Tcell primary and memory responses to subunit vaccines.

SUMMARYThe relationship between B cells and CD4 T cells has been carefully studied, revealing a collaborative effort in which B cells promote the activation, differentiation, and expansion of CD4 T cells while the so-called “helper” cells provide signals to B cells, influencing their class switching and fate. Interactions between B cells and CD8 T cells are not as well studied, although CD8 T cells exhibit an accelerated contraction after certain infections in B-cell-deficient mice. Here, we find that B cells significantly enhance primary CD8 T cell responses after vaccination. Moreover, memory CD8 numbers and function are impaired in B-cell-deficient animals, leading to increased susceptibility to bacterial challenge. We also show that interleukin-27 production by B cells contributes to their impact on primary, but not memory, CD8 responses. Better understanding of the interactions between CD8 T cells and B cells may aid in the design of more effective future vaccine strategies.

The purinergic receptor P2RX7 promotes the accumulation of effector CD4+ T cells in the lung parenchyma during hypervirulent mycobacterial infection

Abstract CD4+ T cells are crucial to control pulmonary tuberculosis, but also favor uncontrolled inflammation in severe disease. Effective CD4+ T cell response to tuberculosis is related to tissue location: Lung-resident CD4+ T cells display higher effector function in comparison to circulating cells. Among the factors favoring T cell residency in non-lymphoid tissues, extracellular ATP sensing via P2RX7 is crucial for the generation of virus-specific resident memory CD8+ T cells. Yet, its role in CD4+ T cell non-lymphoid tissue residency has not been defined. Moreover, mycobacterial infection induces high extracellular ATP levels due to lung tissue damage, which is increased in response to hypervirulent strains. Here, we used P2rx7 ablation models to test the role of P2RX7 in CD4+T cell function in tuberculosis. In response to the hypervirulent Mycobacterium bovis MP287/03 strain, lung-resident WT CD4+ T cells express higher P2RX7 levels compared to circulating CD4+ T cells. Compared to WT mice, P2RX7-deficient (P2rx7−/−) mice display reduced numbers of antigen-specific (I-AbESAT-64-17) and IFN-γ-producing CD4+ T cells in the lung parenchyma. Using cell adoptive transfers, while recipient CD4-deficient mice transferred with CD4+ WT T cells are protected, mice that received CD4+P2rx7−/− T cells develop severe tuberculosis. Mechanistically, CD4+P2rx7−/− T cells have poorer accumulation and IFN-γ production in the lung parenchyma; moreover, they show reduced proliferation in situ. This defect in proliferation is exclusively observed in the lung, with no differences observed in draining lymph nodes. Together, our results show that P2RX7 promote the accumulation of protective CD4+ T cells in the lung during severe tuberculosis.

SARS-COV-2 INFECTION: WHAT MAKES IT SO DIFFERENT?

[...]the infection can be transmitted by those who are asymptomatic or pre-symptomatic. [...]there is an activation of T cells in the first week of infection, with virusspecific memory CD4 and CD8 T cells reportedly peaking within 2 weeks, but remaining detectable at lower levels for 100 or more days of observation. The primary mechanism of action of Molnupiravir against RNA viruses is viral error catastrophe, a concept that is predicated on increasing the viral mutation rate beyond a biologically tolerable threshold, resulting in impairment of viral fitness and leading to viral extinction, determining a striking decrease in viral load in the nasopharyngeal cavity of the tested animals.

HIV-1 infection and the lack of viral control are associated with greater expression of interleukin-21 receptor on CD8+ T cells.

Interleukin-21 (IL-21) has been linked with the generation of virus-specific memory CD8+ T cells following acute infection with HIV-1 and reduced exhaustion of CD8+ T cells. IL-21 has also been implicated in the promotion of CD8+ T-cell effector functions during viral infection. Little is known about the expression of interleukin-21 receptor (IL-21R) during HIV-1 infection or its role in HIV-1-specific CD8+ T-cell maintenance and subsequent viral control. We compared levels of IL-21R expression on total and memory subsets of CD8+ T cells from HIV-1-negative and HIV-1-positive donors. We also measured IL-21R on antigen-specific CD8+ T cells in volunteers who were positive for HIV-1 and had cytomegalovirus-responding T cells. Finally, we quantified plasma IL-21 in treatment-naive HIV-1-positive individuals and compared this with IL-21R expression. IL-21R expression was significantly higher on CD8+ T cells (P = 0.0256), and on central memory (P = 0.0055) and effector memory (P = 0.0487) CD8+ T-cell subsets from HIV-1-positive individuals relative to HIV-1-negative individuals. For those infected with HIV-1, the levels of IL-21R expression on HIV-1-specific CD8+ T cells correlated significantly with visit viral load (r = 0.6667, P = 0.0152, n = 13) and inversely correlated with plasma IL-21 (r = -0.6273, P = 0.0440, n = 11). Lastly, CD8+ T cells from individuals with lower set point viral load who demonstrated better viral control had the lowest levels of IL-21R expression and highest levels of plasma IL-21. Our data demonstrates significant associations between IL-21R expression on peripheral CD8+ T cells and viral load, as well as disease trajectory. This suggests that the IL-21 receptor could be a novel marker of CD8+ T-cell dysfunction during HIV-1 infection.

Antigenic Restimulation of Virus-Specific Memory CD8+ T Cells Requires Days of Lytic Protein Accumulation for Maximal Cytotoxic Capacity.

In various infections or vaccinations of mice or humans, reports of the persistence and the requirements for restimulation of the cytotoxic mediators granzyme B (GrB) and perforin (PRF) in CD8+ T cells have yielded disparate results. In this study, we examined the kinetics of PRF and GrB mRNA and protein expression after stimulation and associated changes in cytotoxic capacity in virus-specific memory cells in detail. In patients with controlled HIV or cleared respiratory syncytial virus (RSV) or influenza virus infections, all virus-specific CD8+ T cells expressed low PRF levels without restimulation. Following stimulation, they displayed similarly delayed kinetics for lytic protein expression, with significant increases occurring by days 1 to 3 before peaking on days 4 to 6. These increases were strongly correlated with, but were not dependent upon, proliferation. Incremental changes in PRF and GrB percent expression and mean fluorescence intensity (MFI) were highly correlated with increases in HIV-specific cytotoxicity. mRNA levels in HIV-specific CD8+ T-cells exhibited delayed kinetics after stimulation as with protein expression, peaking on day 5. In contrast to GrB, PRF mRNA transcripts were little changed over 5 days of stimulation (94-fold versus 2.8-fold, respectively), consistent with posttranscriptional regulation. Changes in expression of some microRNAs, including miR-17, miR-150, and miR-155, suggested that microRNAs might play a significant role in regulation of PRF expression. Therefore, under conditions of extremely low or absent antigen levels, memory virus-specific CD8+ T cells require prolonged stimulation over days to achieve maximal lytic protein expression and cytotoxic capacity.IMPORTANCE Antigen-specific CD8+ T cells play a major role in controlling most virus infections, primarily by perforin (PRF)- and granzyme B (GrB)-mediated apoptosis. There is considerable controversy regarding whether PRF is constitutively expressed, rapidly increased similarly to a cytokine, or delayed in its expression with more prolonged stimulation in virus-specific memory CD8+ T cells. In this study, the degree of cytotoxic capacity of virus-specific memory CD8+ T cells was directly proportional to the content of lytic molecules, which required antigenic stimulation over several days for maximal levels. This appeared to be modulated by increases in GrB transcription and microRNA-mediated posttranscriptional regulation of PRF expression. Clarifying the requirements for maximal cytotoxic capacity is critical to understanding how viral clearance might be mediated by memory cells and what functions should be induced by vaccines and immunotherapies.

NK1.1+ innate lymphoid cells in salivary glands inhibit establishment of tissue-resident memory CD8+ Tcells in mice.

NK1.1+ cells found in salivary glands (SG) represent a unique cell population of innate lymphoid cells (ILC) with characteristics of both conventional NKcells and ILC1. Here, we demonstrate that these NK1.1+ cells limit the accumulation and differentiation of virus-specific tissue-resident memory CD8+ Tcells (TRM cells) in SG of mice infected with lymphocytic choriomeningitis virus (LCMV). The negative regulation of LCMV-specific CD8+ TRM cells by NK1.1+ cells in SG is independent of NKG2D, NKp46, TRAIL, and perforin. Moreover, analysis of NKp46iCre+ Eomesfl/fl mice revealed that Eomes-dependent conventional NKcells are dispensable for negative regulation. Since the SG are prone to autoimmune reactions, regulation of TRM cells by tissue-resident ILC may be particularly important to prevent immunopathology in this organ.