Monocyte response to mitochondrial DAMPs in elderly subjects: a possible contribution to chronic inflammation and atherosclerosis
Authors: Zanini G , Sinigaglia G , Lopez Domenech S , Selleri V , Micheloni G , Malerba M , Coppi F , Alboni S , Nasi M , Pinti M , Mattioli A V 
Published: October 2024

Abstract

Background
Aging is an important risk factor for atherosclerosis and persists as an independent contributor when all other known factors are controlled. Circulating free mitochondrial DNA (cf-mtDNA) can activate immune cells, including monocytes, that promote plaque formation and produce proinflammatory cytokines [1]. Several receptors, including TLR9, AIM2, NLRP3, cGAS can bind cf-mtDNA and activate downstream pathways, leading to the activation of proinflammatory genes [2]. We previously showed that mtDNA increases with age, and at the highest concentrations observed in plasma, can enhance the pro-inflammatory effect of bacterial PAMPs [3]. Nevertheless, it is unknown if the pathways triggered by mtDNA are functionally preserved in elderly, and if it can synergize with oxidized low-density lipoproteins (ox-LDL), which are known to interact with TLR4 and TLR2, in concert with scavenger receptors to regulate the inflammatory microenvironment in atherosclerosis.

Purpose
This study aims to understand if and how the cf-mtDNA activates monocytes triggering an inflammatory response, if monocyte from young and old donors present a different response to mtDNA if age-related difference in the functional activity of these cells.

Methods
We selected 10 old subjects (74.4 ± 2.4 y.o.; 2M:8F) without a history of cardiovascular or peripheral arterial diseases, 4 ultranonagenarians (98.3 ± 4.7 y.o.; 1M:3F) and 9 sex-matched young controls (28.4 ± 7.3 y.o.; 0M:9F). Monocytes were isolated from blood samples, and inflammatory response to mtDNA alone or in combination with LPS was assessed, by analyzing TLR9 expression, NF-kB activation and the release in the supernatants of a panel of proinflammatory cytokines.

Results
Monocyte expression of the mtDNA receptor TLR9 showed a slight reduction with aging, and is similar in old subjects and ultranonagenarians. mtDNA was internalized only in cells pre-treated with LPS and co-localized with TLR9. When internalization occurred, monocytes treated with mtDNA+LPS, released pro-inflammatory cytokines at higher levels than cells treated with LPS alone. Monocytes from elderly subjects showed a partial impairment in mtDNA uptake, and a lower capability to activate TLR9 downstream pathway. However, the capability to secrete pro-inflammatory cytokines (TNF-α, IL-1β, IL-6) was not compromised, and NF-kB translocation into the nucleus after treatment with mtDNA is preserved, suggesting that compensatory mechanisms allowed mtDNA-driven activation of monocytes in elderly people.

Conclusions
The capability of mtDNA to promote an inflammatory response is preserved in elderly people and is still present in ultranonagenarians subjects. As mtDNA plasma levels increase with aging, mtDNA likely act as proinflammatory agents in aged people and can contribute to maintain inflammation in the atherosclerotic plaques. Further studies will clarify if mtDNA can act synergistically with ox-LDL to promote monocyte activation.