Multi-modal profiling of peripheral blood cells across the human lifespan reveals distinct immune cell signatures of aging and longevity

Abstract

Background: Age-related changes in immune cell composition and functionality are associated with multimorbidity and mortality. However, many centenarians delay the onset of aging-related disease suggesting the presence of elite immunity that remains highly functional at extreme old age.

Methods: To identify immune-specific patterns of aging and extreme human longevity, we analyzed novel single cell profiles from the peripheral blood mononuclear cells (PBMCs) of a random sample of 7 centenarians (mean age 106) and publicly available single cell RNA-sequencing (scRNA-seq) datasets that included an additional 7 centenarians as well as 52 people at younger ages (20-89 years).

Findings: The analysis confirmed known shifts in the ratio of lymphocytes to myeloid cells, and noncytotoxic to cytotoxic cell distributions with aging, but also identified significant shifts from CD4⁺ T cell to B cell populations in centenarians suggesting a history of exposure to natural and environmental immunogens. We validated several of these findings using flow cytometry analysis of the same samples. Our transcriptional analysis identified cell type signatures specific to exceptional longevity that included genes with age-related changes (e.g., increased expression of STK17A, a gene known to be involved in DNA damage response) as well as genes expressed uniquely in centenarians' PBMCs (e.g., S100A4, part of the S100 protein family studied in age-related disease and connected to longevity and metabolic regulation).

Interpretation: Collectively, these data suggest that centenarians harbor unique, highly functional immune systems that have successfully adapted to a history of insults allowing for the achievement of exceptional longevity.

Funding: TK, SM, PS, GM, SA, TP are supported by NIH-NIAUH2AG064704 and U19AG023122. MM and PS are supported by NIHNIA Pepper center: P30 AG031679-10. This project is supported by the Flow Cytometry Core Facility at BUSM. FCCF is funded by the NIH Instrumentation grant: S10 OD021587.

Keywords: Aging; Elite immunity; Extreme human longevity; Single cell RNA sequencing.

Fig. 1

The immune landscape of peripheral blood cells from subjects with extreme longevity at single cell resolution.A. UMAP embeddings of PBMCs from all 66 subjects representative of the human lifespan from the integrated scRNA-seq datasets (NECS, PNAS, and NATGEN, labelled by the identified immune cell subtypes. B. UMAP embeddings of PBMCs from all 66 subjects representative of the human lifespan from the integrated scRNA-seq datasets (NECS, PNAS, and NATGEN, labelled by the four age groups.

Fig. 2

Extreme longevity demonstrates shifts in immune cell repertoire compared to younger age groups.A. Bar chart of the relative proportions of the lymphocyte and myeloid immune cell subtypes for each sample across the integrated scRNA-seq datasets: Younger Age, Middle age, Older age, and EL. B. Boxplot of subject specific cell type diversity statistic of 12 immune cell subtypes, grouped by age groups (F-test p-value = 0.0001875). C. Bar chart of the estimated proportions of the lymphocyte and myeloid immune cell subtypes in each age group, grouped separately for males and females. The proportions were estimated using a Bayesian multinomial regression model, and the average 1833 cells per sample allowed us to estimate the smallest proportion 0.3% with 95% confidence. D. Heatmap of the age coefficient comparing Middle, Older, and EL age groups to the Younger age group (right) and heatmap of the sex coefficient for each cell type comparing Females compared to Males. We calculated the Z-statistic and p-value of significance for each coefficient, represented with: ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.

Fig. 3

Shift in the immune resilience strategy within lymphocyte and myeloid populations in centenarians. The average immune cell type proportions across the four age groups of the human lifespan along the hierarchy of peripheral immune compartments: PBMC (Myeloid v. Lymph), Myeloid (Mono v. DC), Lymph (NonCyt v. Cyt), DC (mDC v. pDC), Mono (M14 v. M16), NonCyt (CD4TC v. BC), Cyt (Cyt1 v. Cyt2), Cyt1 (gdTC v. NK), Cyt2 (cCD4TC v. cCD8TC), and CD4TC (nCD4TC v. mCD4TC) and BC (nBC v. mBC).

Fig. 4

Cell type gene expression changes demonstrate three patterns across the human lifespan.A. Table of the number of significant differentially expressed genes across aging comparisons: Middle v. Younger age, Older v. Younger age, EL v. Younger age based on fold change threshold of minimum 10% change and FDR less than 0.05. B. Heatmap of scaled average expression per sample of all significant genes across all cell types grouped by age group. C. Boxplots of expression levels of specific significant genes in particular cell types demonstrating changes in aging and EL (Aging-Related) with at least a nominal significance, D. changes only in EL (EL-Specific), and E. changes in aging not in EL (Aging-Specific). ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.