NADPH oxidase deficiency underlies dysfunction of aged CD8+ Tregs
Zhenke Wen, Yasuhiro Shimojima, Tsuyoshi Shirai, Yinyin Li, Jihang Ju, Zhen Yang, Lu Tian, Jörg J. Goronzy, Cornelia M. Weyand
Zhenke Wen, Yasuhiro Shimojima, Tsuyoshi Shirai, Yinyin Li, Jihang Ju, Zhen Yang, Lu Tian, Jörg J. Goronzy, Cornelia M. Weyand
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Research Article Aging Immunology

NADPH oxidase deficiency underlies dysfunction of aged CD8+ Tregs

Abstract

Immune aging results in progressive loss of both protective immunity and T cell–mediated suppression, thereby conferring susceptibility to a combination of immunodeficiency and chronic inflammatory disease. Here, we determined that older individuals fail to generate immunosuppressive CD8+CCR7+ Tregs, a defect that is even more pronounced in the age-related vasculitic syndrome giant cell arteritis. In young, healthy individuals, CD8+CCR7+ Tregs are localized in T cell zones of secondary lymphoid organs, suppress activation and expansion of CD4 T cells by inhibiting the phosphorylation of membrane-proximal signaling molecules, and effectively inhibit proliferative expansion of CD4 T cells in vitro and in vivo. We identified deficiency of NADPH oxidase 2 (NOX2) as the molecular underpinning of CD8 Treg failure in the older individuals and in patients with giant cell arteritis. CD8 Tregs suppress by releasing exosomes that carry preassembled NOX2 membrane clusters and are taken up by CD4 T cells. Overexpression of NOX2 in aged CD8 Tregs promptly restored suppressive function. Together, our data support NOX2 as a critical component of the suppressive machinery of CD8 Tregs and suggest that repairing NOX2 deficiency in these cells may protect older individuals from tissue-destructive inflammatory disease, such as large-vessel vasculitis.

Authors

Zhenke Wen, Yasuhiro Shimojima, Tsuyoshi Shirai, Yinyin Li, Jihang Ju, Zhen Yang, Lu Tian, Jörg J. Goronzy, Cornelia M. Weyand

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Figure 3

The suppressive function of CD8 Tregs is deficient in older individuals.

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The suppressive function of CD8 Tregs is deficient in older individuals....
(A) CD8 Tregs induced from young (age < 30 yrs) and older (age > 60 yrs) healthy individuals were tested for their ability to suppress pZAP70 in naive CD4 T cells 10 minutes after stimulation. Results are shown as mean ± SD MFI from 9 independent experiments. (B) CD8 Tregs generated from young and older subjects were mixed with CD4 T cells at increasing ratios. pZAP70 in CD4 T cells was analyzed by flow cytometry. Results are shown as mean ± SD from 4 independent experiments. (C) CD8+CD39+CD26– Tregs were sorted from peripheral blood and immediately assessed for their suppressive activity by mixing them at increasing ratios with CD4 T cells. pZAP70 in CD4 T cells was measured after 10 minutes stimulation. Results are shown as mean ± SD from 6 independent experiments. (D and E) NSG mice were reconstituted with naive CD4 T cells, monocytes, and CD8 Tregs generated from young or older donors. Nine days later, expansion of CD4 T cells was quantified by enumerating the (D) frequency and (E) number of human CD4 T cells in the murine spleen. Results from 7 independent experiments are shown as box-and-whisker plots. Boxes represent the 25th and 75th percentile, and lines within the boxes are medians. The 10th and 90th percentiles are shown as whiskers. (F–H) CD4 T cells were labeled with CFSE prior to the transfer. Proliferation of CD4 T cells was assessed by flow cytometric analysis of CFSE dilution in splenocytes harvested after 9 days. (F) A representative image is shown, and results from 6 independent experiments are summarized as box-and-whisker plots of (G) division index and (H) proliferation index. Unpaired 2-tailed Student’s t test was used for comparisons.