Gradual Heat Exposure Triggers Distinct Lung Immune Shifts in Mice

A single gradual rise in ambient temperature from 20°C to 38°C over eight hours set off measurable immune changes in the lungs of mice while leaving most circulating inflammatory signals largely untouched, according to a new study published in Physiological Reports. The results suggest that the pace of heating matters: gradual exposure activated a selective pulmonary inflammatory program instead of the broad cytokine storm typically seen after sudden heat stress.

The research team exposed adult C57Bl/6 mice—24 females and 20 males, all 10 weeks old—to a carefully controlled temperature ramp inside an incubator. The temperature rose steadily from a morning minimum to a mid-to-late-afternoon maximum, approximating the diurnal temperature rise at the start of a heatwave. The 38°C ceiling falls within the range of maximum ambient temperatures recorded during extreme heat events, though the researchers chose that limit based on animal care concerns. Control mice and a pair-fed group, which received the same amount of food and water that heat-exposed mice consumed during the exposure window, stayed on a benchtop at 23°C in the same room.

This gradual paradigm fills an important gap in heat research. Most animal studies of heat stress use sudden exposure, which mimics abrupt transitions such as moving from an air-conditioned building into outdoor heat. Few studies have addressed gradual temperature increases that better reflect the real-world exposure profiles encountered by farmers, athletes, and military personnel working outdoors. The distinction is not trivial: mice, which are poor evaporative coolers that do not rely on whole-body sweating, respond to heat by increasing heat dissipation, reducing metabolic heat production, and raising core temperature, so findings in mice can inform but cannot directly equate to human physiological responses.

To disentangle heat effects from reduced food intake, the researchers included the pair-fed group as a critical control. Animals had been acclimated for three weeks under controlled conditions before exposure, with bedding mixed twice weekly to synchronize their microbiomes. Females were estrus-synchronized to eliminate estrous cycling as a confounding factor. After the eight-hour session, mice were euthanized immediately, and the team collected serum, lung tissue, and cecal contents for a battery of analyses spanning cytokine measurements, RNA sequencing, microbiome profiling, and serum metabolomics.

The cytokine results revealed a striking sex-specific pattern. In lung tissue, heat exposure significantly affected only 3 of 23 measured cytokines and chemokines. In males, IL-17α levels were significantly lower in heat-exposed animals than in pair-fed animals, while females showed no significant IL-17α differences between groups. In females, the chemokines MIP-1α and MIP-1β were significantly elevated after heat exposure compared with both control and pair-fed groups, but neither was significantly altered in males. In serum, IL-1α was significantly decreased in heat-exposed and pair-fed females compared to controls but unaffected in males. IL-12(p40) was significantly elevated in heat-exposed females compared to pair-fed animals, and RANTES levels were significantly increased in heat-exposed males compared to both controls and pair-fed animals.

The researchers interpreted this modest signature—3 of 23 serum cytokines altered—as evidence of a limited rather than systemic inflammatory response at the sampled time point. The chemokines that did change, including MIP-1α, MIP-1β, and RANTES, belong to the CC chemokine subfamily known for roles in T cell adhesion and migration. Elevated MIP-1α and MIP-1β in the lungs of females suggest increased immune cell recruitment and activation, consistent with an inflammatory response during heat exposure. The decrease in lung IL-17α in males may reflect impaired mucosal defense or a regulatory dampening of IL-17-driven tissue stress, since IL-17α plays a crucial role in maintaining mucosal immunity. Whether repeated insults, as occur during multi-day heatwaves, would allow incomplete resolution of inflammation to lead to sustained low-grade inflammation remains an open question.

The lung transcriptome told a richer story. Although principal component analysis did not reveal a separation of gene expression profiles by experimental group, differential expression analysis identified significant changes. In females, heat exposure upregulated 90 genes and downregulated 31 compared to controls; in males, 105 genes were upregulated and 48 downregulated. Pathway analysis showed that heat exposure primarily affected biological processes related to immune activation. Two immune-related processes, Immune System Process and Lymphocyte Activation, were significantly altered in both heat-exposed females and males compared to controls. Females additionally showed alterations in Immunoglobulin-Mediated Immune Response, B Cell-Mediated Immunity, and Immune Response, while males showed changes in Leukocyte Activation, Cell Activation, and B Cell Activation.

Four genes were upregulated in both male and female heat-exposed mice compared to controls: Fcer2a, Cd19, Ighd, and Gapt. These genes govern B-cell activation, immunoglobulin production, and inflammatory signaling. Shared effects across both the control and pair-fed comparisons—seven shared differentially expressed genes in females, including Cd19, Cd37, Fcrl1, and Tbx21, and seven in males, including Cd79a, Egr1, Egr3, and Fcer2a—likely indicate that the transcriptomic effects were mediated by heat rather than reduced feed intake. Compared with pair-fed animals, heat exposure upregulated 91 genes and downregulated 46 in females, and upregulated 207 genes and downregulated 80 in males. No shared differentially expressed genes were detected between males and females in that comparison, despite overlapping biological processes.

The serum metabolome and cecal microbiome proved remarkably resistant to gradual heating. Metabolomics revealed only small, non-significant shifts in endogenous metabolites in both sexes. Random forest analysis identified candidate discriminating metabolites—N-acetylglutamate, pseudouridine, and N-acetylalanine in females, and N-acetyl-phenylalanine and ornithine in males—but none differed significantly by ANOVA. KEGG enrichment analysis indicated that the Glycine, Serine, and Threonine Metabolism pathway was affected in both sexes, with a significant increase in threonine and a decrease in cystathionine due to heat exposure compared to pair-fed in females only. Similarly, alpha and beta diversity of the cecal microbiome did not change in either sex, and univariate testing found no statistically significant differences in the relative abundances of 20 taxa flagged by random forest analysis. This contrasts with earlier work showing that abrupt temperature changes alter gut microbiome composition and function.

Exploratory network analysis integrating lung gene expression with serum metabolites and dual-compartment cytokines identified two upregulated genes, Plac8 and Ltb, present in the heat-exposed versus control networks of both sexes. In females, both genes were positively correlated with serum IL-3; in males, Plac8 was positively correlated with lung MIP-1α and RANTES, while Ltb was negatively correlated with isovalerylcarnitine. Plac8 is expressed in the lung, required for innate antibacterial defense, and modulates monocytes in sepsis, while Ltb encodes a subunit of the membrane lymphotoxin complex critical for developing and maintaining secondary lymphoid organs. Lung GM-CSF and serum IL-3, cytokines that activate innate immune responses and are not typically assessed in heat-exposure studies, also emerged as key network features warranting further investigation. The researchers caution that because core body temperature and behavioral changes could not be recorded during exposure without disrupting the controlled thermal environment, the internal thermal dose experienced by the mice is unknown. Their results therefore characterize the systemic and molecular response to a defined ambient temperature profile, not a quantified physiological heat strain, and cannot be translated into the physiological consequences of human heatwave exposure. Future work should clarify the temporal sequence of molecular events driving pulmonary effects and whether these responses persist, resolve, or change with varying durations and patterns of heat exposure.

Subject of Research: Effects of gradual ambient heat exposure on pulmonary and systemic immune and metabolic responses in mice

Article Title: Ramping up the heat: Induction of systemic and pulmonary immune responses and metabolic adaptations in mice

Article References: Dean, L. E., Adamcakova‐Dodd, A., & Lehmler, H.-J. (2026). Ramping up the heat: Induction of systemic and pulmonary immune responses and metabolic adaptations in mice. Physiological Reports, 14(21), Article e71164. https://doi.org/10.14814/phy2.71164

Image Credits: AI Generated

DOI: 10.14814/phy2.71164

Keywords: heat exposure, mice, lung transcriptomics, cytokines, heatwave, immune response, metabolomics, microbiome, B-cell activation, sex differences, Physiological Reports, systems biology