Distinct immune and transcriptomic profiles in dominant versus subordinate males in mouse social hierarchies

Highlights

• Dominant mice exhibit higher proportion of adaptive immune cells in blood.

• Subordinate mice exhibit higher proportions of innate immune cells in blood.

• Dominants show increased hepatic expression of genes involved in organic acid, carboxylic and lipid catabolic processes.

• Genes related to wound healing and DNA repair genes are upregulated in subordinates.

• Genes related to maintaining and signaling social status are upregulated in dominants.

Abstract

Social status is a critical factor determining health outcomes in human and nonhuman social species. In social hierarchies with reproductive skew, individuals compete to monopolize resources and increase mating opportunities. This can come at a significant energetic cost leading to trade-offs between different physiological systems. In particular, changes in energetic investment in the immune system can have significant short and long-term effects on fitness and health. We have previously found that dominant alpha male mice living in social hierarchies have increased metabolic demands related to territorial defense. In this study, we tested the hypothesis that high-ranking male mice favor adaptive immunity, while subordinate mice show higher investment in innate immunity. We housed 12 groups of 10 outbred CD-1 male mice in a social housing system. All formed linear social hierarchies and subordinate mice had higher concentrations of plasma corticosterone (CORT) than alpha males. This difference was heightened in highly despotic hierarchies. Using flow cytometry, we found that dominant status was associated with a significant shift in immunophenotypes towards favoring adaptive versus innate immunity. Using Tag-Seq to profile hepatic and splenic transcriptomes of alpha and subordinate males, we identified genes that regulate metabolic and immune defense pathways that are associated with status and/or CORT concentration. In the liver, dominant animals showed a relatively higher expression of specific genes involved in major urinary production and catabolic processes, whereas subordinate animals showed relatively higher expression of genes promoting biosynthetic processes, wound healing, and proinflammatory responses. In spleen, subordinate mice showed relatively higher expression of genes facilitating oxidative phosphorylation and DNA repair and CORT was negatively associated with genes involved in lymphocyte proliferation and activation. Together, our findings suggest that dominant and subordinate animals adaptively shift immune profiles and peripheral gene expression to match their contextual needs.

Introduction

Social status is a significant factor affecting immune functioning and health outcomes in human and non-human social animals (Goymann and Wingfield, 2004, Sapolsky, 2004, Snyder-Mackler et al., 2020). For animals living in social dominance hierarchies, social rank strongly drives variation in behavior including access to energetic resources, territory, and mates. For example, higher-ranking males tend to monopolize access to food and consume more than lower-ranking males; however, they also tend to expend more energy via aggression, territory defense and investing in reproductive behavior than lower-ranking males (Lee et al., 2018, Lee et al., 2017, O’Connell and Hofmann, 2011). In stable hierarchies, lower-ranked individuals tend to receive higher levels of aggression and experience higher levels of stress and circulating glucocorticoids (Milewski et al., 2022).

Although the precise mechanisms by which social status influences immune function are yet to be determined, social rank dependent variation in stress levels and energetic demands are believed to be important mediators (Snyder-Mackler et al., 2016, Snyder-Mackler et al., 2020). For example, high-ranking individuals that direct energy away from immune processes and prioritize immediate reproductive success or sexual signaling exhibit compromised long-term immune functioning and health (Mills et al., 2010, Mills et al., 2009, Sheldon and Verhulst, 1996, Zuk et al., 1995). Conversely, low-ranking individuals often demonstrate compromised immune systems and worse health outcomes, such as high mortality rates and slow wound healing, even without physical injury or with access to ample nutritional resources (Archie et al., 2012, Blanchard et al., 1985, Koolhaas et al., 1997). These outcomes are likely induced through low-ranking individuals experiencing chronic psychosocial stress characterized by a loss of control and a lack of predictability over their social environment (Sapolsky, 1994). In rodents, socially defeated animals exhibit prolonged elevation of proinflammatory cytokines, enhanced splenocyte proliferation, and slow wound healing (Archie et al., 2012, Bartolomucci et al., 2001, Engler et al., 2004, Hodes et al., 2014, McKim et al., 2016, Sapolsky, 2005). Similarly, low status female rhesus monkeys exhibit dysregulated hypothalamic-pituitary adrenal axis (HPA) activity (Tung et al., 2012) as well as an immune system biased towards a proinflammatory, antibacterial response and away from an antiviral response, the reverse of what is observed in high status females (Snyder-Mackler et al., 2016). In contrast, high status male baboons, that experience consistently high levels of aggression, show a heightened inflammatory response to bacterial infection (Lea et al., 2018). This indicates that an important consideration in understanding the relationship between social status, stress, immune functioning and health is that it can vary by sex, species, and the specific social dynamics of social groups (Beery et al., 2020, Habig et al., 2018, Habig and Archie, 2015, Snyder-Mackler et al., 2020).

In this study, our aim was to investigate how social status in male mice is associated with markers of immune functioning. We used an established social dominance paradigm in which cohorts of 10–12 male CD-1 mice live in a group housing system and reliably form stable linear hierarchies (So et al., 2015, Williamson et al., 2016). As a social hierarchy is established, each animal occupies a unique social rank and dramatically different behavioral phenotypes emerge between ranks. The highest ranking male (alpha) exhibits higher rates of aggression compared to all other individuals, is more active in patrolling territory and eats and drinks more frequently (Lee et al., 2018). Alpha males also face further metabolic energy demands as they increase their production of major urinary proteins (MUPs) in the liver, which are then deposited for scent-marks and used to signal their dominance status to other animals (Lee et al., 2017, Nelson et al., 2015). Ranked below the alpha are the sub-dominant individuals who initiate aggression towards relatively subordinate individuals despite repeatedly receiving high levels of aggression from relatively more dominant individuals. Subordinates are the lowest ranking individuals who rarely show aggression. While these status-related phenotypes are typically consistent across cohorts, there can be some variation in the degree of aggressiveness (i.e. despotism) exhibited by alphas. Highly despotic alpha males exert frequent aggression and inhibit aggressive behavior of all other group members, and notably, subordinate individuals living with highly despotic alpha males exhibit elevated basal plasma corticosterone (CORT) concentrations (Williamson et al., 2017).

By utilizing this group-housing paradigm, we are able to directly compare the immunophenotypes of dominant and subordinate individuals from the same social hierarchies. We hypothesized that the energetic demands of establishing and maintaining dominance in a hierarchy would reshape immune profiles. In line with the trade-offs model proposed by Lee (2006), dominant individuals redirect resources to maintaining status and away from investing in innate immune components, relying preferentially on adaptive immune components (Habig et al., 2018, Habig and Archie, 2015, Lee, 2006). Conversely, we predicted that subordinate individuals would invest preferentially in innate immune components that would facilitate recovery from injuries (Archie et al., 2012, Kiecolt-Glaser et al., 1995). To test these hypotheses, we measured immune cell composition from peripheral blood of male CD-1 mice before and two weeks after the formation of social hierarchies. Given the purported role of glucocorticoids in mediating the relationship between status and immune functioning, we assessed basal levels of corticosterone from peripheral blood prior to and following group housing. As a proxy measurement of long-term stress exposure (Lee et al., 2010a, Lee et al., 2011), we also measured Fkbp5 DNA methylation levels in blood. Lastly, we profiled splenic and hepatic transcriptomes in dominant and subordinate individuals. The liver, an important metabolic organ that regulates energy metabolism, is also critically involved in the growth and transport of immune cells in conjunction with the spleen. This approach allowed us to identify genes and potential gene pathways involved in energy metabolism and immune system functioning that are differentially expressed according to social status.