Impaired autophagy following ex vivo cooling of simulated hypothermic temperatures in peripheral blood mononuclear cells from young and older adults

Kelli E King; James J McCormick; Morgan K McManus; Kristina-Marie T Janetos; Nicholas Goulet; Glen P Kenny

doi:10.1016/j.jtherbio.2024.103831

Impaired autophagy following ex vivo cooling of simulated hypothermic temperatures in peripheral blood mononuclear cells from young and older adults

J Therm Biol. 2024 Mar 27:121:103831. doi: 10.1016/j.jtherbio.2024.103831. Online ahead of print.

Authors

Kelli E King¹, James J McCormick¹, Morgan K McManus¹, Kristina-Marie T Janetos¹, Nicholas Goulet², Glen P Kenny³

Affiliations

¹ Human and Environmental Physiology Research Unit, School of Human Kinetics, University of Ottawa, Ottawa, Canada.
² Human and Environmental Physiology Research Unit, School of Human Kinetics, University of Ottawa, Ottawa, Canada; Behavioural and Metabolic Research Unit, School of Human Kinetics, University of Ottawa, Ottawa, Ontario, Canada.
³ Human and Environmental Physiology Research Unit, School of Human Kinetics, University of Ottawa, Ottawa, Canada; Clinical Epidemiology Program, Ottawa Hospital Research Institute, Ottawa, Ontario, Canada. Electronic address: gkenny@uottawa.ca.

PMID: 38565070
DOI: 10.1016/j.jtherbio.2024.103831

Abstract

Hypothermia is a critical consequence of extreme cold exposure that increases the risk of cold-related injury and death in humans. While the initiation of cytoprotective mechanisms including the process of autophagy and the heat shock response (HSR) is crucial to cellular survival during periods of stress, age-related decrements in these systems may underlie cold-induced cellular vulnerability in older adults. Moreover, whether potential sex-related differences in autophagic regulation influence the human cold stress response remain unknown. We evaluated the effect of age and sex on mechanisms of cytoprotection (autophagy and the HSR) and cellular stress (apoptotic signaling and the acute inflammatory response) during ex vivo hypothermic cooling. Venous blood samples from 20 healthy young (10 females; mean [SD]: 22 [2] years) and 20 healthy older (10 females; 66 [5] years) adults were either isolated immediately (baseline) for peripheral blood mononuclear cells (PBMCs) or exposed to water bath temperatures maintained at 37, 35, 33, 31, or 4 °C for 90 min before PBMC isolation. Proteins associated with autophagy, apoptosis, the HSR, and inflammation were analyzed via Western blotting. Indicators of autophagic initiation and signaling (LC3, ULK1, and beclin-2) and the HSR (HSP90 and HSP70) increased when exposed to hypothermic temperatures in young and older adults (all p ≤ 0.007). Sex-related differences were only observed with autophagic initiation (ULK1; p = 0.015). However, despite increases in autophagic initiators ULK1 and beclin-2 (all p < 0.001), this was paralleled by autophagic dysfunction (increased p62) in all groups (all p < 0.001). Further, apoptotic (cleaved-caspase-3) and inflammatory (IL-6 and TNF-α) signaling increased in all groups (all p < 0.001). We demonstrated that exposure to hypothermic conditions is associated with autophagic dysfunction, irrespective of age or sex, although there may exist innate sex-related differences in cytoprotection in response to cold exposure as evidenced through altered autophagic initiation.

Keywords: Aging; Apoptosis; Cold; Hypothermia; Macroautophagy.