Subcutaneous Mycobacterium vaccae promotes resilience in a mouse model of chronic psychosocial stress when administered prior to or during psychosocial stress
Introduction
Chronic psychosocial stress is an acknowledged predisposing factor for several mental disorders, including affective disorders, anxiety disorders, and trauma- and stressor-related disorders such as posttraumatic stress disorder (PTSD) (Reber et al., 2016a). For instance, psychological trauma induced by natural disasters, traffic accidents, sexual violence, war, and terror, amongst others, increases the risk of developing PTSD (Glaesmer et al., 2014, Husarewycz et al., 2014, March et al., 2014). In addition, a systematic review, meta-analysis, and meta-regression study that included 20 studies, found that interleukin (IL)-6, IL-1β, tumor necrosis factor (TNF)-α, and interferon γ were higher in individuals with a diagnosis of PTSD than in healthy controls (Passos et al., 2015). Although further mechanistic investigations are critically required, trauma- and stressor-related disorders in general are frequently accompanied by an over-(re)active immune system (Lindqvist et al., 2014, Pace et al., 2006, Pace et al., 2012). As the extent of stress-induced immune activation has been shown to predict subsequent development of PTSD in humans (Eraly et al., 2014, Pervanidou et al., 2007), as well as to predict increased anxiety-like defensive behavior (Hodes et al., 2014), and a compromised fear extinction in mice (Young et al., 2018), an increased stress-induced immune activation might even be involved in disease pathogenesis (Gao et al., 2018, Lowry et al., 2016, Miller and Raison, 2016, Rohleder, 2014). Moreover, social stress-induced immune activation is more pronounced in healthy human participants at high risk for developing anxiety and affective disorders (Böbel et al., 2018, Breines et al., 2014, Carpenter et al., 2010, Derry et al., 2013). Considering that PTSD (Sommershof et al., 2009) and other mental disorders like major depression (Li et al., 2010) are often characterized by a decreased number of regulatory T cells (Treg), one possible mechanism underlying the exaggerated stress-induced immune (re)activity in individuals at high risk for mental disorders might be a failure of adequate immunoregulation.
In accordance with the above-described hypothesis that stress-induced immune activation facilitates the development of several psychiatric conditions, we have recently shown that promoting immunoregulation by repeated subcutaneous (s.c.) administrations of a heat-killed preparation of Mycobacterium vaccae (M. vaccae; National Collection of Type Cultures (NCTC) 11659), a microorganism with anti-inflammatory and immunoregulatory properties, efficiently reduced subordinate, flight, and avoiding behavioral responses to a dominant aggressor, indicating a shift towards more proactive stress coping in a murine model of chronic psychosocial stress (Reber et al., 2016b). Moreover, M. vaccae preimmunization via the s.c. route has anxiolytic effects and prevents stress-induced spontaneous colitis and exaggeration of chemically-induced colitis in a model of inflammatory bowel disease (Reber et al., 2016b). In line with these findings, repeated administrations of M. vaccae either prior to or during stressor exposure in the same mouse model protected against stress-induced aggravation of chemically-induced colitis when administered via the non-invasive intranasal route (Amoroso et al., 2019). These studies used the chronic subordinate colony housing (CSC) paradigm, which induces a chronic anxiety-like phenotype (Füchsl et al., 2013, Füchsl et al., 2014, Langgartner et al., 2015, Reber et al., 2016b). This animal model is based on the chronic subordination of four male mice by a dominant resident male conspecific and, compared with single-housed controls (SHC), results in a lack of social preference, increased anxiety-like defensive behavioral responses, increased alcohol consumption/preference, hyperactivity, spontaneous colitis, and exaggerated chemically induced colitis. Notably, CSC mice show a reduced number of Treg (Schmidt et al., 2010), which likely contributes to the increased inflammatory state. Supporting this hypothesis, depletion of Treg by administration of anti-CD25 antibody prevents the stress-protective effects of M. vaccae administered via the s.c. route prior to exposure to the CSC model (Reber et al., 2016b).
The aim of the present study was to confirm the stress-protective effects of repeated s.c. M. vaccae administrations prior to CSC exposure and to extend these findings to the effects of s.c. administrations of M. vaccae during CSC exposure. Therefore, individual mice received s.c. injections of M. vaccae either: 1) on days −21, −14, and −7 prior to CSC, or 2) on days 2, 8, and 15 during CSC and were tested on the elevated plus-maze (EPM, day 19), open-field/novel object test (OF/NO, day 20) and social preference/avoidance test (SPAT, day 21) for changes in general and/or social anxiety. Pro- and reactive coping behaviors were assessed on days 1, 8, and 15 during CSC.
Section snippets
Animals
Male C57BL/6N mice weighing 17–19 g (Experiment 1; prior-to-CSC protocol) or 19–21 g (Experiment 2; during-CSC protocol) were used as experimental mice and male CD-1 mice weighing 30–35 g were used as dominant aggressors. Mice in Experiment 1 arrived at ~4–5 weeks of age, whereas mice in Experiment 2 arrived at ~5–6 weeks of age. This allowed all mice to enter the stress procedure at approximately the same age. All mice were sourced from Charles River, Sulzfeld, Germany. Standard polycarbonate
Effects of repeated s.c. M. vaccae on individual behavioral stress coping during CSC
To assess if s.c. administration of M. vaccae affects the behavioral stress coping of individual CSC mice (pro-active vs. re-active coping) during CSC exposure, mice were videotaped in the first 1 h after colony formation on days 1, 8, and 15 (Fig. 2). A dominance index (DI; number of pro-active minus number of re-active behaviors) was calculated.
Experiment 1: Statistical analysis of the DI employing a LMM approach revealed a significant time and M. vaccae × time interaction effect (Fig. 2A; F
Discussion
In the present study we confirm our previous results showing that repeated s.c. administrations of a heat-killed preparation of the immunoregulatory bacterium M. vaccae induces a pronounced shift towards active stress coping and has moderate protective effects against stress-induced anxiety when administered prior to stressor exposure in a mouse model of chronic psychosocial stress (Reber et al., 2016b). Moreover, we extend these findings by showing that M. vaccae has potent anxiolytic and,
Funding
This study was funded by the Office of Naval Research Global (N00014-17-S-B001).
Authors contributions
SOR, DL, and CAL planned the study; MA and DL performed the experiments; MA and AB did the statistical analysis; MA, DL, and SOR wrote the manuscript.
Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Declaration of interest: MA, AB, DL, and SOR have nothing to declare. CAL serves on the Scientific Advisory Board of Immodulon Therapeutics.
Acknowledgements
The authors thank P. Hornischer and U. Binder for their technical assistance and help in performing the experiments. Furthermore, the authors would also like to thank Dr. S. Ott, E. Merkel and S. Hummel (local animal research center) for their excellent support in terms of animal husbandry.
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2023, Progress in Neuro-Psychopharmacology and Biological PsychiatryCitation Excerpt :Regarding the effect of stress on anxiety, the results showed that 2 weeks of CSC exposure resulted in exacerbated anxiety-like behavior that was associated to increased sucrose intake. This is consistent with previous studies using a 19-days chronic social stress model in mice from our laboratory (Bahi, 2013a, 2017a, 2017b; Bahi and Dreyer, 2020) and from others (Amoroso et al., 2020; Foertsch et al., 2017). However, and in contrast to CSC mice, no differences in anxiety-like behaviors were observed when the CSC model was extended to Wistar rats (Nyuyki et al., 2012).
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These authors contributed equally.