Skip to main content
NCBI home page
NIHPA Author Manuscripts logoLink to NIHPA Author Manuscripts
. Author manuscript; available in PMC: 2018 Jul 15.
Published in final edited form as: J Immunol. 2017 Jun 14;199(2):403–407. doi: 10.4049/jimmunol.1700625

The aging immune system reveals the biological impact of direct antigen presentation on CD8 T cell responses

Jennifer L Uhrlaub 1, Megan J Smithey 1, Janko Nikolich-Žugich 1,*
PMCID: PMC5520673  NIHMSID: NIHMS880210  PMID: 28615415

Abstract

The vertebrate immune system employs multiple, sometimes redundant, mechanisms to contain pathogenic microorganisms that are always evolving to evade host defenses. Thus, the cowpox virus (CPXV) uses genes encoding CPXV12 and CPXV203 to prevent direct MHCI presentation of viral peptides by infected cells. However, CD8 T cells are effectively primed against CPXV by cross-presentation of viral antigens in young mice. Old mice accumulate defects in both CD8 T cell activation and cross-presentation. Using a double-deletion mutant (Δ12Δ203) of CPXV, we show that direct priming of CD8 T cells in old mice yields superior recall responses, establishing a key contribution of this mechanism to host anti-poxvirus responses and enhancing our fundamental understanding of how viral manipulation of direct presentation impacts pathogenesis. This also provides a proof of principle that suboptimal CD8 T cell in old organisms can be optimized by manipulating antigen presentation, with implications for vaccine design.

Introduction

The immune system mobilizes a variety of innate and adaptive immune mechanisms to limit and eliminate infection. In youth, these mechanisms are both robust and overlapping, providing considerable redundancy in protecting against microbial infections. Evaluating the in vivo impact and limits of immune resource redundancy when confronted with microbial immune evasion has been difficult so far. In older age, many mechanisms of protective immunity exhibit defects, allowing us to use old mice as a model of suboptimal immunity, akin to a complex genetic hypomorph for adaptive or innate immunity.

Members of the Orthopoxvirus genus of the Poxviridae family are known to adversely affect individuals with vulnerable immune system, including older adults (1). Thus, vulnerability to wild-type (wt) ectromelia virus (ECTV) increases with age; anti-poxvirus-specific CD8 T cell responses are curtailed both in total number and function in ECTV-exposed old B6 mice (2), consistent with other models of viral and bacterial infections where CD8 T cell responses are impaired in old mice as compared to their adult counterparts (3–7). By contrast, 14–18 month old mice infected with poorly pathogenic orthopox viruses, such as vaccinia virus (VACV) and the mutant strain of ECTV (Δ166 ECTV) mounted CD8 T cell responses comparable to adult mice (2). The mechanistic basis for the improved CD8 responses in old mice to attenuated poxviruses remains incompletely understood (8).

Poxviruses utilize a diverse array of strategies to evade the immune system. At the present, it is not known whether and to what extent differences in the expression of viral immune evasion proteins play a role in increased susceptibility of older organisms to wild-type, but not attenuated, poxviruses (9). Multiple studies have mechanistically dissected Cowpox virus (CPXV) immune evasion (10–15). Two viral proteins, CPXV12 and CPXV203, down-regulate MHC Class I (MHCI) on the surface of infected cells. Consequently, antigen-specific CD8 T cells cannot recognize or exert their effector function on CPXV infected cells. Importantly, this evasion mechanism does not prevent the priming of a functional CD8 T cell response via cross-presentation(16, 17) (18). Indeed, C57BL/6 (B6) mice generate potent CD8 T cell responses to CPXV directed against a conserved immunodominant H-2Kb-restricted (Kb in the text) epitope B8R20–27 (B8R in the text). However, cross presentation has been shown to be less effective with aging (17–19). Thus, if direct presentation is blocked by the virus, and cross presentation is crippled with aging, then combined these two deficiencies may explain the reduced CD8 T cell responsiveness with aging (17–19). If this explanation is correct, then restoring of direct priming should improve CD8 T cell responses in old mice.

To test this hypothesis, we used a CPXV mutant lacking CPXV12 and CPXV203 (Δ12Δ203 CPXV) in old mice. We demonstrate that B8R-specific CD8 T cell responses to Δ12Δ203 CPXV are significantly improved in both abundance and function, as compared to those primed with wild-type CPXV (wt CPXV). Importantly, repairing direct priming with the mutant virus restored primary CD8 T cell responses in old mice to the same level as in adult mice responding to wt virus, and generated superior memory CD8 T cell responses upon recall in old mice even when compared to adult mice responding to wt CPXV, as judged by clearance of Listeria monocytogenes expressing the B8R epitope (Lm-B8R). This demonstrates that direct priming can induce strong effector and memory CD8 T cell responses, which helps explain the evolutionary pressure that lead to the generation of CPXV12 and 203 by the virus. Our approach highlights the power of using a vulnerable population with suboptimal immunity as a tool to dissect biological relevance of antimicrobial responses in the face of microbial immune evasion. We conclude that improving direct antigen presentation can be a powerful strategy to induce robust CD8 T cell responses even under conditions of suboptimal immunity (e.g. in the growing elderly segment of the population) and must be considered for vaccines where effective CD8 T cell memory is required to curtail or eliminate infection.

Materials and Methods

Ethics Statement

Mouse studies were carried out in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. Protocols were approved by the Institutional Animal Care and Use Committee at the University of Arizona (IACUC #08-102, PHS Assurance Number: A3248-01). Intranasal infections were performed under Ketamine/Xylazine anesthesia. Footpad injections were performed under isoflurane anesthesia. Euthanasia was performed by isoflurane overdose or cervical dislocation to preserve unobstructed analysis of cellular immunity (28).

Mouse experiments

Old (18–22 months) and adult (12–16 weeks) male C57BL/6 (B6) mice were purchased and/or obtained from the National Institute on Aging Rodent Resource via the Charles River Laboratories (Frederick, MD and Kingston, NY) and/or The Jackson Laboratory (Bar Harbor, ME). 10–16 weeks male RAG1-KO mice (strain #002216) and Ly5.1 congenic mice (strain #002014) were originally purchased from The Jackson Laboratory (Bar Harbor, ME). Mice anesthetized i.p. with ketamine and xylazine were infected intranasally (i.n.) with 1×106 pfu CPXV in 10uL sterile PBS administered to one nare. Mice anesthetized with isoflurane were infected subcutaneously in the footpad (f.p.) with 1×106 pfu of either wt or Δ12Δ203 CPXV in 50uL sterile PBS. Total T cells or CD8 T cells for adoptive transfer experiments were purified with Miltenyi Biotec, Inc. reagents and columns using negative magnetic selection. Purity was confirmed at >90% by FCM analysis.

Viruses and viral titer

Cowpox virus (CPXV) strain Brighton Red was obtained from the ATCC. The Δ12Δ203 CPXV mutant was kindly provided by Dr. Wayne Yokoyama (10). All viruses were propagated and titered on Vero cells. Titer was determined by plaque assay.

Flow cytometry and intracellular cytokine staining

Accutase-treated (eBioscience, San Diego, CA) spleen was disassociated over a 40µM cell strainer. Blood was hypotonically lysed to eliminate red blood cells. Lymphocytes were incubated overnight in B8R tetramer (NIH Tetramer Core Facility) and a saturating dose of mAbs including anti-CD3 and CD8α (eBioscience, San Diego, CA), stained with Live/Dead Aqua (Life Technologies, Grand Island, NY) and analyzed as below. Cells stimulated with B8R:Kb peptide (10−6M, 21st Century, Marlboro, MA) in the presence of brefeldin A for 5 hours were first stained for surface markers and viability as described above then fixed and permeabilized with the BD CytoFix/CytoPerm kit and stained for intracellular IFNγ, and TNFα (eBioscience, San Diego, CA) and Granzyme B (GzB, BD Bioscience, San Jose, CA). Samples were acquired using a BD LSR Fortessa cytometer (BD Bioscience, San Jose, CA) and analyzed by FlowJo software (Tree Star, Ashland, OR). Cell counts were extrapolated from either a CBC differential collected on a Hemavet LV (Drew Scientific, Waterbury, CT) instrument or by correction during acquisition with Count Beads (Life Technologies, Carlsbad, CA). The two counting methods were confirmed to be consistent.

Naïve CD8p number

Protocol adapted from (29). Spleen and visible lymph nodes (cervical, axillary, brachial, inguinal, and popliteal) were collected from naïve adult (12 weeks) and old (18 months) mice and treated with accutase (Ebioscience, San Diego, CA) for 30 min at 37°C followed by disassociation over a 40µM filter. Cells were washed in MACS buffer (PBS, 0.5% BSA, 2mM EDTA, pH 7.2) and enriched for total T cells by negative selection (Miltenyi Biotec, San Diego, CA; #130-095-130). Cells were transferred to FACS buffer (PBS + 2%FBS) and maintained cold (4°C) for all remaining steps. Enriched T cells were stained in the presence of Fc block for 1 hour with a saturating dose of mAbs for CD3, CD4, and CD8α (eBioscience, San Diego, CA) and Kb-B8R tetramers (NIH Tetramer Core Facility) labeled with two different fluorochromes. Cells were washed twice and stained in a saturating dose of anti-streptavidin PE antibody (BioLegend, San Diego, CA) for 30 min at 4°C. Cells were washed two more times and resuspended in 250uL of FACS buffer + count beads (Life Technologies, Carlsbad, CA). Upon acquisition with BD FACS Diva software, cells were first gated as a CD3+ histogram to the axis so that all CD3+ events and the count beads would be included. This was selected as the storage gate to minimize the size of the FCS file. The gating strategy for CD8p number is depicted in Supplemental Fig. 1. Final CD8p number was determined with count bead correction for abort rate per manufacturer protocol. CD8p from adult and old mice were determined alongside each other and reduced CD8p numbers with age was consistent with results from previous publications (19, 20). B8R-specific CD8p numbers from adult mice were consistent with the original publication (30).

In vivo bacterial clearance

Mice were intravenously infected with 105 CFU of Listeria monocytogenes expressing B8R (Lm-B8R) kindly provided by Dr. Sing Sing Way. 48 hours post-challenge spleens were harvested, weighed, and placed into sterile PBS. Tissues were homogenized mechanically using a Tissue-Tearor electric homogenizer (BioSpec Products, Bartlesville, OK). Serial dilutions were made in sterile PBS and plated onto BHI agar. Plates were incubated overnight at 37° C. The log10 CFU/g of tissue was calculated as: log10 [(CFU/dilution factor) × ((organ weight + homogenate volume)/organ weight)] (31).

Results

Reduced B8R-specific CD8 T cell responses to CPXV in old mice

Multiple models of infection have demonstrated an age-associated decrease in primary antigen-specific CD8 T cell responses (2–5, 8). We infected adult (12–16 weeks) and old (18–22 months) mice with wt CPXV and evaluated the B8R-specific CD8 T cell response (Fig. 1). Old mice exhibited reduced absolute numbers of, and diminished polyfunctional cytokine production amongst, the B8R-specific effector CD8 T cells on day 7–8 p.i (Fig. 1A, B). Moreover, as previously shown, many tetramer-specific CD8 T cells from old mice failed to produce IFNγ (Fig. 1C). Similar age-related differences were found with both the footpad (Fig. 1) and intranasal (not shown) administration of the virus.

FIGURE 1.

FIGURE 1

Open in a new tab

Curtailed antigen-specific CD8 T cell responses to CPXV in old mice. Adult (A; 12 weeks) and old (O; 18–22 months) B6 mice were inoculated with wt CPXV via i.n. route as described in Methods. (A) Total # of B8R-specific CD8 T cells from spleen on d7–8 p.i. (n=8 per group). Data are mean ± SEM. Statistical significance determined by unpaired student’s t-test. ** P< 0.01. (B) Frequency of IFNγ+ CD8 T cells from spleen when stimulated with 10−6M B8R:Kb peptide in the presence of BFA for 5 hours. IFNγ+ CD8 T cells also expressing TNFα and/or Granzyme B are shown as shaded bars within response. Data are represented as mean (n=8 per group). Statistical significance determined by unpaired student’s t-test. *** P< 0.001. (C) Ratio of IFNγ:Tetramer from A and B. Statistical significance determined by unpaired student’s t-test. *** P< 0.001.

Because CD8 T lymphocyte precursor (CD8p) numbers decline with age (19–21) we next asked if that reduction curtailed primary CD8 T cell response in old mice to CPXV. We confirmed that B8R-specific CD8p numbers were reduced with age (12-wk old =1603±124.2; 18-mo old = 810.0±115.9, Fig. 2A; FCM gating strategy, see Supplemental Fig. 1). Because the environment of the old mouse is reported to impair T cell responses, we compared the responsiveness of adult and old CD8 T cells in the adult environment. We transferred total adult or old splenic and lymph node (LN) T cells into Rag1−/− mice and challenged them with wt CPXV (Fig. 2B). Numbers of total T cells, CD8 T cells, B8R-specific CD8 T cells, and IFNγ producing CD8 T cells did not significantly differ with age on day 7 post infection (p.i.) with intranasally (i.n.)-administered CPXV (Fig. 2C–F). Therefore, a twofold reduction of B8R CD8 precursor numbers in 18-mo old mice was not sufficient to impair the response of old CD8 T cells in a young adult environment. This result further suggests that in the context of appropriate environmental cues, old CD8 T cells may be able to produce robust primary responses.

FIGURE 2.

FIGURE 2

Open in a new tab

Reduced numbers of B8R-specific precursors are not responsible for deficient antigen-specific CD8 T cell response in 18-mo old mice. (A) B8R-specific CD8p numbers from naïve adult (A; 12 weeks) and old (O; 18 months) mice (n=8–9 per group). Data are mean ± SEM. Statistical significance determined by unpaired student’s t-test. * P<0.05. (B) Experiment schematic for panels D–G. (C–F) Total T cells were isolated from spleen, popliteal, inguinal, brachial and cervical LN from adult (A; 12 weeks) or old (O; 18 months) mice into RAG1-KO mice followed by inoculation with WT CPXV via i.n. or f.p. route as described in Methods the day after transfer. Identical results were achieved from either route of infection. On d7 p.i. absolute number of (C) CD3+ T cells, (D) CD8+ T cells, (E) B8R:Kb+ CD8 T cells, (F) IFNγ+ CD8 T cells following stimulation with 10−6M B8R:Kb peptide in the presence of BFA for 5 hours (n=8 per group). Data are mean ± SEM. Statistical significance determined by unpaired student’s t-test. P=ns.

Antigen-specific CD8 T cell responses are improved by deletion of pathogen immune evasion mechanisms

CPXV12 and CPXV203 work together to prevent MHC I antigen presentation on the surface of infected cells, blocking their detection by CD8 T cells (22). While cross-presentation allows for priming a CD8 T cell response against CPXV in young mice, cross-presentation is impaired with age (23–25) (26). We hypothesized that restoring direct presentation during priming may improve CD8 T cell responses in old mice, and provide insight to how the basic biology of antigen presentation may impact immunity in a vulnerable population with suboptimal cellular immunity.

We confirmed that CPXV titers were equivalent in adult and old mice infected with either wt CPXV (wt CPXV) or CPXVΔ12Δ203 (Supplemental Fig. 2A). We next compared the CD8 T cell responses against CPXV and CPXVΔ12Δ203, both amongst the endogenous old or adult T cells, and, to control for possible intrinsic CD8 T cell defects in old mice, also amongst adult Ly5.1 congenic CD8 T cells transferred into adult and old mice (Fig. 3A). In adult mice, we confirmed (18) that there were no significant difference in the CD8 T cell response between the two viruses as measured by total number of CD8 T cells (Supplemental Fig. 2B), the number of B8R-specific CD8 T cells (Fig. 3B), or the frequency of IFNγ producing CD8 T cells (Fig. 3C). By contrast, old mice mounted an improved response (significantly higher number of B8R-specific and higher frequency of IFNγ producing CD8 T cells) in response to CPXVΔ12Δ203 (Fig. 3B–C). Total numbers of CD8 T cells remained comparable for both viruses (Supplemental Fig. 2B). The magnitude of the endogenous response in old mice remained 3–4 fold lower compared to adult mice.

FIGURE 3.

FIGURE 3

Open in a new tab

Deletion of genes evading direct presentation improves antigen-specific CD8 T cell responses. Ly5.1 congenic CD8 T cells (4×106 per recipient) were transferred into adult (A; 12 weeks) and old (O; 18–22 months) mice followed by immunization with either wt or Δ12Δ203 CPXV via f.p. route as described in Methods. (A) Experiment schematic for panels B–E. Endogenous (Ly5.2) and donor (Ly5.1) B8R-specific CD8 T cell responses were determined from spleen on d7 p.i. (n=8 recipients per group). (B) Total number of endogenous CD8 T cells that are B8R-specific from adult or old mice. (C) Frequency of endogenous CD8 T cells producing IFNγ from adult or old mice following stimulation with 10−6M B8R:Kb peptide in the presence of BFA for 5 hours. (D) Total number of donor Ly5.1+ CD8 T cells that are B8R-specific from adult or old mice. (E) Frequency of donor Ly5.1+ CD8 T cells producing IFNγ from adult or old mice following stimulation with 10−6M B8R:Kb peptide in the presence of BFA for 5 hours. Data are mean ± SEM. Statistical significance determined by unpaired student’s t-test. *P< 0.05; **P< 0.01, *** P< 0.001.

This pointed to additional age-related defects in the endogenous CD8 T cell responses, which we substantiated by examination of responding transferred adult CD8 T cells in the adult or old environments. We saw improved priming with CPXVΔ12Δ203 in both adult and old environments, as evidenced by an increased number of Ly5.1 congenic B8R-specific CD8 T cells and increased percentages of IFNγ+ cells responding to B8R peptide (Fig. 3D–E). Indeed, adult CD8 T cells primed with CPXVΔ12Δ203 in the old environment expanded to the same numbers like adult CD8 T cells responding to wt CPXV in the adult environment (Fig. 3D). The total number of donor Ly5.1 CD8 T cells recovered was comparable between groups (Supplemental Fig. 2C). Therefore, the restoration of direct Ag presentation improved primary CD8 T cell responses. Of note, previous studies examining CPXVΔ12Δ203 in young mice, where robust CD8 T cell responses exist in the absence of direct presentation, were unable to resolve the impact of immune evasion on CD8 T cell responses. The positive contribution of direct Ag presentation was below the limit of resolution for the assay, and became only fully evident when we examined a vulnerable old population with suboptimal immunity.

Superior generation of CD8 T cell memory in old mice following restoration of direct antigen presentation

Efficacious and protective long-term memory responses are key to immune defense. We, therefore, tested the efficacy of memory CD8 T cell responses elicited by CPXVΔ12Δ203. The acute CD8 T cell response measured in the blood on day 7 p.i. recapitulated previous data from the spleen, with superior responses to both infections in adult vs. old mice and an improved response in old mice to Δ12Δ203 as compared to wt CPXV (Fig. 4A). In the resting memory phase, at day 45 p.i., antigen-specific CD8 T cell frequencies were comparable between groups, albeit those in old mice immunized with the Δ12Δ203 trended higher (Fig. 4B). Most importantly, upon challenge with Listeria monocytogenes expressing the B8R epitope (Lm-B8R), mice vaccinated with CPXVΔ12Δ203 exhibited improved clearance as measured by decreased bacterial burden in the spleen (Fig. 4C). This clearance was similarly improved in both adult and old mice and CD8 memory T cell responses in old mice vaccinated with CPXVΔ12Δ203 were as potent as their adult counterparts in clearing the bacterium.

FIGURE 4.

FIGURE 4

Open in a new tab

Vaccination with a direct priming-competent vector produces a highly protective CD8 T cell memory response. Adult (A; 12 weeks) and old (O; 18 months) mice were infected with either wt or Δ12Δ203 CPXV via f.p. route as described in Methods. Frequency of B8R:Kb+ CD8 T cell responses on (A) d7 and (B) d45 p.i. (n=7–8 per group). Statistical significance determined by unpaired student’s t-test. **P< 0.01. At d45 p.i. immune (n=6–8 per group) and naïve (n=4 per age) mice were challenged with 1×106 cfu Listeria monocytogenes expressing the Kb-restricted B8R epitope. 2 days post-challenge bacterial burden in the spleen was determined as cfu/g. Data are mean ± SEM. Statistical significance determined on log transformed data by one-way ANOVA with Bonferroni post-test. *P< 0.05; **P< 0.01, *** P< 0.001.

Discussion

The data presented here underscore the significant contribution of direct antigen presentation to the priming of functional CD8 T cell memory, as one of the key mechanisms to long-term resistance to infections. Our results stress the extent to which these mechanisms function in a vulnerable population to provide superior CD8 T cell memory, over and above what would have been predicted based on primary responses. Specifically, by circumventing immune evasion mechanisms that down-regulate class I expression and prevent the participation of infected DC’s in CD8 T cell priming, we were able to generate strongly protective memory CD8 T cell responses against an intracellular bacterium.

These results confirm our prior data in the West Nile virus model (8), that in old mice weak primary CD8 T cell responses do not predict strength of CD8 T cell protection upon recall. They also underscore the importance of strong direct antigen presentation in the course of priming of functional CD8 T cell memory in a vulnerable population, and highlight the unappreciated interplay between immune evasion and of reduced immune resources in determining long-term immune protection.

Our studies also highlight the importance of using vulnerable populations with suboptimal immunity as a tool to dissect biological relevance of microbial immune evasion in the context of antimicrobial and anti-vaccine responses. Indeed, CD8 responses following direct antigen presentation were not significantly improved when assessed in adult mice that possess strong, competent, cross-presentation mechanisms (23–25). The contribution of direct presentation could only be appreciated following adoptive transfer of adult CD8 T cells into old mice, and by studies of memory populations – neither of which would have been undertaken in the absence of the improved endogenous response measured in old mice.

The above basic findings have substantial and immediate relevance to vaccine design. The contribution of poor CD8 T cell responses to increased susceptibility to infection with aging is pathogen specific (3, 4, 6, 27). Nonetheless, our study highlights that ensuring robust direct presentation is a promising vaccine-engineering strategy in vulnerable populations against strictly intracellular microbial pathogens that do not lend themselves to antibody neutralization. Such infections include, but are not limited, to Listeria and Salmonella and viruses whose control require robust CD8 T cell responses, including Respiratory Syncytial Virus.

Supplementary Material

1

Acknowledgments

Grant Support:

This work was supported by NIAID NIH HHS U54 AI081680.The UACC/ALR Cytometry Core Facility and the Cancer Center Support Grant (CCSG-CA 023074) supported our flow cytometry

Kb-restricted B8R tetramer obtained through the NIH Tetramer Core Facility.

Abbreviations

A

adult mouse, 3–4 mo old

CPXV

cowpox virus

Δ12Δ203 CPXV

CPXV lacking genes 12 and 203

O

old mouse, >18 months old

References

  • 1.Hanna W. Studies In Small-pox and Vaccination. John Wright, Marshall Simpkin, Kent Hamilton, Bristol, London: 1913. [Google Scholar]
  • 2.Fang M, Roscoe F, Sigal LJ. Age-dependent susceptibility to a viral disease due to decreased natural killer cell numbers and trafficking. J Exp Med. 2010;207:2369–2381. doi: 10.1084/jem.20100282. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Smithey MJ, Renkema KR, Rudd BD, Nikolich-Zugich J. Increased apoptosis, curtailed expansion and incomplete differentiation of CD8+ T cells combine to decrease clearance of L. monocytogenes in old mice. Eur J Immunol. 2011;41:1352–1364. doi: 10.1002/eji.201041141. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 4.Brien JD, Uhrlaub JL, Hirsch A, Wiley CA, Nikolich-Zugich J. Key role of T cell defects in age-related vulnerability to West Nile virus. J Exp Med. 2009;206:2735–2745. doi: 10.1084/jem.20090222. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Po JL, Gardner EM, Anaraki F, Katsikis PD, Murasko DM. Age-associated decrease in virus-specific CD8+ T lymphocytes during primary influenza infection. Mech Ageing Dev. 2002;123:1167–1181. doi: 10.1016/s0047-6374(02)00010-6. [DOI] [PubMed] [Google Scholar]
  • 6.Bhadra R, Moretto MM, Castillo JC, Petrovas C, Ferrando-Martinez S, Shokal U, Leal M, Koup RA, Eleftherianos I, Khan IA. Intrinsic TGF-beta signaling promotes age-dependent CD8+ T cell polyfunctionality attrition. J Clin Invest. 2014;124:2441–2455. doi: 10.1172/JCI70522. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 7.Yager EJ, Ahmed M, Lanzer K, Randall TD, Woodland DL, Blackman MA. Age-associated decline in T cell repertoire diversity leads to holes in the repertoire and impaired immunity to influenza virus. J Exp Med. 2008;205:711–723. doi: 10.1084/jem.20071140. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 8.Uhrlaub JL, Brien JD, Widman DG, Mason PW, Nikolich-Zugich J. Repeated in vivo stimulation of T and B cell responses in old mice generates protective immunity against lethal West Nile virus encephalitis. J Immunol. 2011;186:3882–3891. doi: 10.4049/jimmunol.1002799. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 9.Schuren AB, Costa AI, Wiertz EJ. Recent advances in viral evasion of the MHC Class I processing pathway. Curr Opin Immunol. 2016;40:43–50. doi: 10.1016/j.coi.2016.02.007. [DOI] [PubMed] [Google Scholar]
  • 10.Byun M, Verweij MC, Pickup DJ, Wiertz EJ, Hansen TH, Yokoyama WM. Two mechanistically distinct immune evasion proteins of cowpox virus combine to avoid antiviral CD8 T cells. Cell Host Microbe. 2009;6:422–432. doi: 10.1016/j.chom.2009.09.012. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.McCoy WHt, Wang X, Yokoyama WM, Hansen TH, Fremont DH. Cowpox virus employs a two-pronged strategy to outflank MHCI antigen presentation. Mol Immunol. 2013;55:156–158. doi: 10.1016/j.molimm.2012.11.011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Alzhanova D, Edwards DM, Hammarlund E, Scholz IG, Horst D, Wagner MJ, Upton C, Wiertz EJ, Slifka MK, Fruh K. Cowpox virus inhibits the transporter associated with antigen processing to evade T cell recognition. Cell Host Microbe. 2009;6:433–445. doi: 10.1016/j.chom.2009.09.013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Dasgupta A, Hammarlund E, Slifka MK, Fruh K. Cowpox virus evades CTL recognition and inhibits the intracellular transport of MHC class I molecules. J Immunol. 2007;178:1654–1661. doi: 10.4049/jimmunol.178.3.1654. [DOI] [PubMed] [Google Scholar]
  • 14.Luteijn RD, Hoelen H, Kruse E, van Leeuwen WF, Grootens J, Horst D, Koorengevel M, Drijfhout JW, Kremmer E, Fruh K, Neefjes JJ, Killian A, Lebbink RJ, Ressing ME, Wiertz EJ. Cowpox virus protein CPXV012 eludes CTLs by blocking ATP binding to TAP. J Immunol. 2014;193:1578–1589. doi: 10.4049/jimmunol.1400964. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 15.Byun M, Wang X, Pak M, Hansen TH, Yokoyama WM. Cowpox virus exploits the endoplasmic reticulum retention pathway to inhibit MHC class I transport to the cell surface. Cell Host Microbe. 2007;2:306–315. doi: 10.1016/j.chom.2007.09.002. [DOI] [PubMed] [Google Scholar]
  • 16.Bevan MJ. Cross-priming for a secondary cytotoxic response to minor H antigens with H-2 congenic cells which do not cross-react in the cytotoxic assay. J Exp Med. 1976;143:1283–1288. doi: 10.1084/jem.143.5.1283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 17.Bevan MJ. Cross-priming. Nat Immunol. 2006;7:363–365. doi: 10.1038/ni0406-363. [DOI] [PubMed] [Google Scholar]
  • 18.Gainey MD, Rivenbark JG, Cho H, Yang L, Yokoyama WM. Viral MHC class I inhibition evades CD8+ T-cell effector responses in vivo but not CD8+ T-cell priming. Proc Natl Acad Sci U S A. 2012;109:E3260–3267. doi: 10.1073/pnas.1217111109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 19.Rudd BD, Venturi V, Li G, Samadder P, Ertelt JM, Way SS, Davenport MP, Nikolich-Zugich J. Nonrandom attrition of the naive CD8+ T-cell pool with aging governed by T-cell receptor:pMHC interactions. Proc Natl Acad Sci U S A. 2011;108:13694–13699. doi: 10.1073/pnas.1107594108. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20.Smithey MJ, Li G, Venturi V, Davenport MP, Nikolich-Zugich J. Lifelong persistent viral infection alters the naive T cell pool, impairing CD8 T cell immunity in late life. J Immunol. 2012;189:5356–5366. doi: 10.4049/jimmunol.1201867. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.Decman V, Laidlaw BJ, Doering TA, Leng J, Ertl HC, Goldstein DR, Wherry EJ. Defective CD8 T cell responses in aged mice are due to quantitative and qualitative changes in virus-specific precursors. J Immunol. 2012;188:1933–1941. doi: 10.4049/jimmunol.1101098. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Gainey MD, Rivenbark JG, Cho H, Yang L, Yokoyama WM. Viral MHC class I inhibition evades CD8(+) T-cell effector responses in vivo but not CD8(+) T-cell priming. Proc Natl Acad Sci U S A. 2012;109:E3260–3267. doi: 10.1073/pnas.1217111109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 23.Chougnet CA, Thacker RI, Shehata HM, Hennies CM, Lehn MA, Lages CS, Janssen EM. Loss of Phagocytic and Antigen Cross-Presenting Capacity in Aging Dendritic Cells Is Associated with Mitochondrial Dysfunction. J Immunol. 2015;195:2624–2632. doi: 10.4049/jimmunol.1501006. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24.Zacca ER, Crespo MI, Acland RP, Roselli E, Nunez NG, Maccioni M, Maletto BA, Pistoresi-Palencia MC, Moron G. Aging Impairs the Ability of Conventional Dendritic Cells to Cross-Prime CD8+ T Cells upon Stimulation with a TLR7 Ligand. PLoS One. 2015;10:e0140672. doi: 10.1371/journal.pone.0140672. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25.Li G, Smithey MJ, Rudd BD, Nikolich-Zugich J. Age-associated alterations in CD8alpha+ dendritic cells impair CD8 T-cell expansion in response to an intracellular bacterium. Aging Cell. 2012;11:968–977. doi: 10.1111/j.1474-9726.2012.00867.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 26.Hickman HD, Takeda K, Skon CN, Murray FR, Hensley SE, Loomis J, Barber GN, Bennink JR, Yewdell JW. Direct priming of antiviral CD8+ T cells in the peripheral interfollicular region of lymph nodes. Nat Immunol. 2008;9:155–165. doi: 10.1038/ni1557. [DOI] [PubMed] [Google Scholar]
  • 27.Richner JM, Gmyrek GB, Govero J, Tu Y, van der Windt GJ, Metcalf TU, Haddad EK, Textor J, Miller MJ, Diamond MS. Age-Dependent Cell Trafficking Defects in Draining Lymph Nodes Impair Adaptive Immunity and Control of West Nile Virus Infection. PLoS Pathog. 2015;11:e1005027. doi: 10.1371/journal.ppat.1005027. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 28.Pecaut MJ, Smith AL, Jones TA, Gridley DS. Modification of immunologic and hematologic variables by method of CO2 euthanasia. Comp Med. 2000;50:595–602. [PubMed] [Google Scholar]
  • 29.Jenkins MK, Moon JJ. The role of naive T cell precursor frequency and recruitment in dictating immune response magnitude. J Immunol. 2012;188:4135–4140. doi: 10.4049/jimmunol.1102661. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 30.Hamilton SE, Schenkel JM, Akue AD, Jameson SC. IL-2 complex treatment can protect naive mice from bacterial and viral infection. J Immunol. 2010;185:6584–6590. doi: 10.4049/jimmunol.1001215. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31.Smithey MJ, Brandt S, Freitag NE, Higgins DE, Bouwer HG. Stimulation of enhanced CD8 T cell responses following immunization with a hyper-antigen secreting intracytosolic bacterial pathogen. J Immunol. 2008;180:3406–3416. doi: 10.4049/jimmunol.180.5.3406. [DOI] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

1
NIHMS880210-supplement-1.pdf (161.8KB, pdf)

RESOURCES