Pedalling toward a deeper understanding of exercise effects on immune function

Optimal immune function is essential for prolonged health and homeostasis, largely through the utilization of inflammation in order to regulate complex cellular processes. Inflammation is broadly classified as either pathogen-associated or damage-associated (i.e. sterile). Prolonged sterile inflammation is implicated in cardiovascular disease (CVD), prioritizing strategies to mitigate dysregulated inflammation in order to enhance long-term cardiovascular health. Exercise is touted as a critical lifestyle behaviour for promoting cardiovascular health due in part to its anti-inflammatory effects. Acute exercise stimulates a transient increase in circulating anti-inflammatory cytokines and leukocytes, whereupon the circulating leukocytes can patrol the vasculature or migrate into peripheral tissues. This repeated delivery of circulatory immune cells assists in the facilitation of tissue repair and resolution of inflammation. However, exercise of strenuous intensity and heavy volume has been shown to induce transient immunosuppression (Nieman & Wentz, 2019) and acute hyporesponsiveness to anti-inflammatory cytokine stimulation in populations with CVD risk (Barry et al., 2018). The mechanisms underlying these divergent immune responses are largely unknown and require further investigation in order to properly assess the anti-inflammatory effects of exercise. In a study recently published in The Journal of Physiology, Islam et al. (2024) sought to determine how variations in exercise intensity and pattern influence the cellular activity of anti-inflammatory cytokines by analysing (i) the fluctuation of circulating cytokine concentrations and leukocyte subtype counts, (ii) the functional ability of interleukin (IL)-10 and IL-6 to inhibit tumour necrosis factor-α (TNF-α), and (iii) the distinctions in monocyte polarization. They hypothesized that although exercise would increase circulating anti-inflammatory cytokines (IL-10, IL-6) and leukocytes, the intracellular signalling of IL-10 and IL-6 and subsequent inhibition of pro-inflammatory cytokines would be blunted Sixteen healthy, young and recreationally active adults (50% females) were recruited to participate in a crossover-model exercise study. Using a Latin-square design, participants completed the following conditions: (i) non-exercise control (CTL), (ii) moderate continuous exercise (MCE; cycling at 70% of lactate threshold), (iii) heavy continuous exercise (HCE; cycling at 10% of the difference between peak oxygen uptake ( V ̇ O 2 peak ${{\dot{V}}_{{{{\mathrm{O}}}_{\mathrm{2}}}{\mathrm{peak}}}}$ ) and lactate threshold), and (iv) heavy interval exercise (HIE; 1-min cycling intervals at HCE paired with 1 min of stationary rest), where all exercise conditions were matched for estimated energy expenditure. Venous blood samples were collected at baseline (PRE), as well as immediately post (POST), 30-min post (POST30), and 90-min post (POST90) exercise. Exercise elicited alterations in both circulating immune cells and cytokines. An acute increase in leukocytes was observed, as total white blood cell count, lymphocytes and monocytes were elevated at POST in each exercise condition compared to CTL POST, and neutrophils were increased at both HCE and HIE POST compared to CTL POST. Multiplex assay analysis revealed increases in IL-6 at HIE POST and both HCE and HIE POST30, compared to given time points within the CTL condition, while IL-10 elevations were observed in HCE POST30, compared to HCE PRE, MCE POST30 and HIE POST30. Notably, MCE had no impact on IL-6 or IL-10 at any time point as compared to given time points within the CTL condition. Additionally, TNF-α increased at HCE POST compared to all other POST conditions, while it decreased at MCE POST30 compared to HCE POST30. Overall, TNF-α, IL-6 and IL-10 increased in the brief (≤30-min POST) period following heavy-intensity exercise in a pattern-dependent manner. IL-10 and muscle-derived IL-6 are anti-inflammatory cytokines involved in the regulation of TNF-α signalling via the transcription factor signal transducer and activator of transcription 3 (STAT3). Phosphorylation of STAT3 (pSTAT3) exerts an anti-inflammatory effect on downstream targets of TNF signalling through inhibition of the transcription factor nuclear factor κB (NF-κB) and subsequent suppression of inflammatory cytokine production (Antonia et al., 2022). Islam et al. (2024) utilized flow cytometry to assess the expression of IL-6 and IL-10 receptors and pSTAT3 in CD14+ (classical) monocytes and CD4+ lymphocytes (helper T cells) following ex vivo stimulation with IL-6 and IL-10 at PRE, POST and POST90. While no changes in IL-6- or IL-10-mediated pSTAT3 were observed in CTL or MCE for either cell type, heavy-intensity exercise appeared to dampen pSTAT3. Specifically, IL-6 and IL-10-driven pSTAT3 was blunted in both HCE and HIE POST and POST90 compared to both HCE and HIE PRE, suggesting an impaired ability of IL-6 and IL-10 to reduce inflammatory signalling from circulating immune cells following heavy exercise. Indeed, whole blood cultures were collected at POST and co-stimulated with lipopolysaccharide (LPS) (to induce leukocytic TNF-α secretion) and IL-6 (LPS+IL-6) or IL-10 (LPS+IL-10) to provide an index of the anti-inflammatory cytokine's ability to inhibit TNF-α production. TNF-α from cultured supernatants was measured by ELISA. MCE and HIE had no impact on TNF-α secretion. However, HCE elicited increases in TNF-α that both low (1 ng/ml) and high (10 ng/ml) doses of IL-6 were unable to significantly inhibit. Conversely, while the low dose of IL-10 neared significance, only the high dose of IL-10 was effective at significantly blunting TNF-α, demonstrating secretion levels similar to those observed during the CTL condition. Though the precise explanation for the ability of IL-6 and IL-10 to inhibit LPS-induced TNF-α secretion requires further study, these findings suggest that IL-10 may be more effective at downregulating TNF-driven inflammation. In addition to cellular activation, the authors were interested in whether exercise could influence the polarization of monocytes toward a pro-inflammatory (M1-like) or anti-inflammatory (M2-like) phenotype. Classical monocytes were isolated from whole blood at POST and underwent 24-h incubation with either LPS + interferon γ (IFNγ) or IL-10 to promote M1-like or M2-like phenotypes, respectively. TNF-α was measured by ELISA and cells were stained for CD80 (M1-like) and CD163 (M2-like) expression and examined by flow cytometry. Other monocytes were stimulated with LPS or LPS + IL-10 to determine the effects of the phenotype polarization on TNF-α secretion and IL-10 inhibition. Flow cytometry analysis revealed a decrease in CD80 expression following HIE, while MCE and HCE had no effect. Additionally, the changes in CD163 expression following exercise did not reach statistical significance, and monocyte polarization did not affect IL-10 inhibition of TNF-α. Overall, HIE decreased M1-like polarization of monocytes, and even though heavier exercise-intensity trended towards M2-like polarization, it was not significant, suggesting that the ability of IL-10 to blunt TNF-α was unaffected by monocyte phenotype polarization. The elaborate study design implemented by Islam et al. (2024) highlights the importance of extending assessment of inflammation status beyond measurement of circulating cytokines alone. Cytokine actions are typically pleiotropic, so neglecting to connect changes in their concentrations to downstream cellular-level effects provides incomplete information about functional outcomes. By applying exercise protocols of differing intensity and pattern while still matching for overall energy expenditure, the findings of this study enhance our understanding of the short-term impact of exercise variables on immune function. However, the authors acknowledge that their findings may not be generalizable to less active or fit individuals, and that the sample size was insufficient to determine sex differences. Future studies that reflect the lower physical activity and fitness of the general population, and are adequately powered to determine sex differences are needed to understand the intracellular effects of different exercise protocols on immune function. It was also suggested that further research is warranted on the mechanisms by which exercise affects monocyte subset polarization. Blanks et al. (2020) attempted to address this gap by investigating the effects of moderate-intensity (60% V ̇ O 2 peak ${{\dot{V}}_{{{{\mathrm{O}}}_{\mathrm{2}}}{\mathrm{peak}}}}$ ) cycling on monocyte subset surface expression of CCR2, the receptor for monocyte chemoattractant protein-1 (MCP-1), and macrophage polarization. They observed a decrease in CCR2 expression on intermediate monocytes following exercise in their high physical activity group, along with a decrease in CD206 (M2) expression on macrophages in their low physical activity group. These findings, especially the differential outcomes between the physical activity groups, reinforce the needs asserted by Islam et al. (2024) to extend their recent work to other monocyte subsets. Additionally, while the current study measured TNF-α production, their study design could be replicated to examine the production and downstream effects of other inflammatory cytokines in the context of health and disease. The nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3 (NLRP3) inflammasome, a multi-protein sensing complex in the innate immune system, has received recent attention for its role in CVD. The NLRP3 inflammasome contributes to sterile inflammation via the production of pro-inflammatory cytokines IL-1β and IL-18 and is activated by, among other signals, NF-κB (Toldo et al., 2022). Given the aforementioned effects of exercise on pSTAT3, which helps regulate NF-κB activation, the role of exercise in modifying NLRP3 inflammasome activity for improved cardiovascular health warrants investigation (Fig. 1). In conclusion, the recent work by Islam et al. (2024) provides valuable insight into the impact of exercise intensity and pattern on anti-inflammatory activity and phenotype polarization of monocytes, and the importance of measuring both when trying to determine the effects of exercise on inflammation. This study provides a foundation for exciting avenues for future research in exercise immunology. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article. The authors declare no competing interests. All authors have read and approved the final version of this manuscript and agree to be accountable for all aspects of the work to ensure that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. All persons designated as authors qualify for authorship, and all those who qualify for authorship are listed. No funding was received for this work. The authors would like to thank their mentor, Dr R. Lee Franco, for his support and review of the manuscript.