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. 2022 Dec 13;19(1):63.
doi: 10.1186/s12979-022-00321-9.

Phenotypic and functional alterations of monocyte subsets with aging

Yu Cao  1   2   3 , Yang Fan  1   2   3 , Fangyuan Li  4   5 , Yu Hao  4   5 , Yaxian Kong  1   2   3 , Chen Chen  4   5 , Xing Hao  6 , Dannuo Han  6 , Guoli Li  1   2   3 , Zengtao Wang  1   2   3 , Chuan Song  1   2   3 , Junyan Han  7   8 , Hui Zeng  9   10   11   12   13
Affiliations

Phenotypic and functional alterations of monocyte subsets with aging

Yu Cao et al. Immun Ageing. .

Abstract

Background: It has been widely accepted that monocytes are one of the central mediators contributing to inflammaging. However, it remains unclear whether aged monocytes, similar to aged T cells, have characteristics of hyperactivation and increased expression of co-inhibitory molecules.

Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from young (21-40 years old), middle-aged (41-60 years old), and older human subjects (> 60 years old). Flow cytometry was used to monitor changes in the expression of surface molecules of monocyte subsets and cytokine-producing capacity.

Results: We observed increased tumor necrosis factor-α: TNF-α and decreased interleukin-6 (IL-6) production in monocytes from older adults compared with young and middle-aged adults. Older adults had a greater percentage of intermediate and non-classical monocyte subsets, along with increased levels of the immune activation markers human leukocyte antigen-DR (HLA-DR), and adhesion molecules cluster of differentiation molecule 11b (CD11b) and L-selectin (CD62L). Furthermore, we observed increased C-C motif chemokine receptor 2 (CCR2) expression on classical monocytes and decreased C-X3-C motif chemokine receptor 1 (CX3CR1) expression on non-classical monocytes in older adult subjects. The expression of co-inhibitory receptors was reduced on monocyte subsets in older adults.

Conclusions: Circulating monocytes in older adults exhibit increased expression of activation, adhesion, and migration markers, but decreased expression of co-inhibitory molecules.

Keywords: Activation; Aging; Immunosenescence; Monocytes.

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Conflict of interest statement

none.

Figures

Fig. 1
Fig. 1
The cytokine profiles of total monocytes in healthy adults in different age groups. A. Comparison of the percentage of total monocytes among peripheral blood mononuclear cells (PBMCs) in young, middle-aged, and older adults via flow cytometry (young: 21–40 years, n = 42; middle-aged: 41–60 years, n = 34; older: > 60 years, n = 34). Data are shown as a box plot with medians (lines inside boxes), 25th, and 75th quartiles (limits of boxes). Whiskers indicate the range, and each dot represents one sample. P-values were obtained using a Kruskal–Wallis test, followed by post hoc analysis. B. Representative flow cytometry data of intracellular staining for TNF-α, IL-6, IL-10, GM-CSF, and IL-1β in total monocytes stimulated with LPS (100 ng/mL) for 3 h in vitro from young, middle-aged, and older adults (young: 21–40 years, n = 42; middle-aged: 41–60 years, n = 34; older: > 60 years, n = 34). C. Intracellular staining for the percentage of TNF-α+ and IL-6+ monocytes stimulated with LPS (100 ng/mL) for 3 h in vitro from young, middle-aged, and older adults (young: 21–40 years, n = 42; middle-aged: 41–60 years, n = 34; older: > 60 years, n = 34) upon in vitro LPS stimulation. Data are shown as box-plots with medians (lines inside boxes), 25th, and 75th quartiles (limits of boxes). Whiskers indicate the range, and each dot represents one sample. P-values were obtained by a Kruskal–Wallis rank test, followed by post hoc analysis
Fig. 2
Fig. 2
Percentage of peripheral monocyte subsets from young, middle-aged, and older adults. A. Ideograph and representative flow cytometry analyses of monocytes. According to the expression pattern of CD14 and CD16, human monocytes were divided into CD14lowCD16− (Mo0), CD14highCD16− (Mo1), CD14highCD16+ (Mo2), and CD14low CD16+ (Mo3) subsets. B. Comparison of the percentages of Mo0, Mo1, Mo2, and Mo3 subpopulations among all monocytes in young, middle-aged, and older adults were performed according to the expression pattern of CD14 and CD16 (young: 21–40 years, n = 42; middle-aged: 41–60 years, n = 34; older: > 60 years, n = 34). P-values were calculated using the non-parametric Kruskal–Wallis rank test, followed by post hoc analysis
Fig. 3
Fig. 3
The expression levels of HLA-DR, CD88, CD29, CD11b, and CD62L in monocyte subsets. A. The MFI of HLA-DR, CD11b, and CD62L from monocyte subsets (Mo0, Mo1, Mo2, and Mo3) among young, middle-aged, and older persons (young: 21–40 years, n = 23; middle-aged: 41–60 years, n = 15; older: > 60 years, n = 44). B. The Z-scores of HLA-DR, CD88, CD29, CD11b, and CD62L expression of monocyte subsets (Mo0, Mo1, Mo2, and Mo3) from young (black dots, n = 23), middle-aged (blue dots, n = 15) and older adults (red dots, n = 44) as a radar plot, and the values range from 0 to 1
Fig. 4
Fig. 4
The expression of CCR2 and CX3CR1 in monocyte subsets. A. The MFI of CCR2 and CX3CR1 from monocyte subsets in young participants was analyzed by flow cytometry. Histograms were created using FlowJo software. B. Boxplots showing the MFI of CCR2 and CX3CR1 from monocyte subsets in young, middle-aged, and older individuals (young: 21–40 years, n = 23; middle-aged: 41–60 years, n = 15; older: > 60 years, n = 44). P-values were calculated using the non-parametric Kruskal–Wallis rank test, followed by post hoc analysis
Fig. 5
Fig. 5
Percentage of monocyte subsets expressing co-inhibitory molecules decreased in older adults. A. Representative flow cytometry histograms display the expression of the co-inhibitory molecules 2B4, T-cell immunoglobulin domain and mucin domain 3 (TIM-3), CD200R, T-cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif (ITIM) domain (TIGIT), B and T lymphocyte attenuator (BTLA), CD160, programmed death-1 (PD-1), and lymphocyte-activation gene 3 (LAG-3) on monocyte subsets from young adults. B. The percentage of monocyte subsets expressing TIM-3, 2B4, CD200R, TIGIT, and BTLA from young, middle-aged, and older adults (young: 21–40 years, n = 42; middle-aged: 41–60 years, n = 34; older: > 60 years, n = 34). P-values were calculated using the non-parametric Kruskal–Wallis rank test, followed by post hoc analysis
Fig. 6
Fig. 6
Summary of function-related molecule expression profiles of monocyte subsets between younger and older adults. The different expression levels of function-related molecules (activation and adhesion molecules and receptors, chemokine receptors, and co-inhibitory molecules) in four monocyte subsets (Mo0, Mo1, Mo2, and Mo3) from older adults (> 60 years old) compared with younger adults (< 60 years old). Fold changes were calculated by the ratio of expression levels between older and young adults. Significant differences in the molecular expression within monocyte subsets from older adults compared with young adults are presented as dots (the dot size is -LogP). P-values were calculated using the Wilcoxon rank-sum test. Orange and blue dots represent increased and decreased values of monocyte subsets in older adults compared to young adults
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