Elsevier

Brain, Behavior, and Immunity

Volume 87, July 2020, Pages 309-317
Brain, Behavior, and Immunity

Subcutaneous Mycobacterium vaccae promotes resilience in a mouse model of chronic psychosocial stress when administered prior to or during psychosocial stress

https://doi.org/10.1016/j.bbi.2019.12.018Get rights and content

Highlights

  • M. vaccae given s.c. prior to, but not during CSC promotes active stress coping.

  • M. vaccae given s.c. prior to and during CSC prevents CSC-induced general anxiety.

  • M. vaccae given s.c. prior to and during CSC prevents CSC-induced social anxiety.

Abstract

Chronic psychosocial stress is a risk factor for many mental disorders, including affective disorders, anxiety disorders, and trauma- and stressor-related disorders (i.e., posttraumatic stress disorder, PTSD). As these disorders are associated with an overreactive immune system and chronic low-grade inflammation, immunoregulatory approaches counterbalancing basal and/or stress-induced immune activation should be protective in this context. In support of this hypothesis, we recently demonstrated that repeated subcutaneous (s.c.) preimmunization with a heat-killed preparation of the immunoregulatory bacterium Mycobacterium vaccae (M. vaccae; National Collection of Type Culture (NCTC) 11659) promoted proactive stress coping and protected against stress-induced anxiety and intestinal pathology in a mouse model of chronic psychosocial stress. To induce development of a chronic anxiety-like state, the chronic subordinate colony housing (CSC) paradigm was used. Here we employed the CSC paradigm (start day 1) to confirm the stress-protective effects of repeated s.c. M. vaccae administrations prior to CSC exposure (days −21, −14, and −7) and to extend these findings to the stress-protective role of M. vaccae when administered repeatedly during CSC exposure (days 2, 8 and 15). As readouts we assessed the stress coping behavior on days 1, 8, and 15 and general and/or social anxiety-related behavior on days 19 (elevated plus-maze), 20 (open-field/novel object test), and day 21 (social preference/avoidance test) of CSC exposure. In line with our previous study, M. vaccae administered prior to CSC strongly promoted active stress coping and moderately reduced CSC-induced general and social anxiety. Although M. vaccae administered during CSC did not affect stress coping, this treatment protocol profoundly protected against CSC-induced general, and to a lesser extent social, anxiety. Taken together, these data broaden the framework for developing bioimmunoregulatory approaches, based on the administration of immunoregulatory microorganisms, for the prevention and/or treatment of affective disorders, anxiety disorders, and trauma- and stressor-related psychiatric disorders like PTSD.

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.

References (44)

  • T.W. Pace et al.

    Increased peripheral NF-kappaB pathway activity in women with childhood abuse-related posttraumatic stress disorder

    Brain. Behav. Immun.

    (2012)
  • I.C. Passos et al.

    Inflammatory markers in post-traumatic stress disorder: a systematic review, meta-analysis, and meta-regression

    Lancet Psychiat.

    (2015)
  • P. Pervanidou et al.

    Elevated morning serum interleukin (IL)-6 or evening salivary cortisol concentrations predict posttraumatic stress disorder in children and adolescents six months after a motor vehicle accident

    Psychoneuroendocrinology

    (2007)
  • S.O. Reber et al.

    Chronic subordinate colony housing paradigm: A mouse model for mechanisms of PTSD vulnerability, targeted prevention, and treatment-2016 Curt Richter Award Paper

    Psychoneuroendocrinology

    (2016)
  • D. Schmidt et al.

    Chronic psychosocial stress promotes systemic immune activation and the development of inflammatory Th cell responses

    Brain, Behav. Immun.

    (2010)
  • D.A. Slattery et al.

    Behavioural consequences of two chronic psychosocial stress paradigms: anxiety without depression

    Psychoneuroendocrinology

    (2012)
  • A. Sommershof et al.

    Substantial reduction of naive and regulatory T cells following traumatic stress

    Brain. Behav. Immun.

    (2009)
  • A.H. Veenema et al.

    Low inborn anxiety correlates with high intermale aggression: link to ACTH response and neuronal activation of the hypothalamic paraventricular nucleus

    Horm. Behav.

    (2007)
  • F.R. Walker et al.

    Coping with defeat: acute glucocorticoid and forebrain responses to social defeat vary with defeat episode behaviour

    Neuroscience

    (2009)
  • S.K. Wood et al.

    Resilience to the effects of social stress: evidence from clinical and preclinical studies on the role of coping strategies

    Neurobiol. Stress

    (2015)
  • S.K. Wood et al.

    Inflammatory factors mediate vulnerability to a social stress-induced depressive-like phenotype in passive coping rats

    Biol. Psychiat.

    (2015)
  • M.B. Young et al.

    A peripheral immune response to remembering trauma contributes to the maintenance of fear memory in mice

    Psychoneuroendocrinology

    (2018)
  • Cited by (18)

    • The PMN-MDSC – A key player in glucocorticoid resistance following combined physical and psychosocial trauma

      2023, Brain, Behavior, and Immunity
      Citation Excerpt :

      In Set 6 of ex-breeder mice (n = 6) whole blood leukocytes, PMN-MDSC-enriched PBMCs and neutrophil/PMN-MDSC-enriched WBCs were isolated and pooled, respectively, before myeloid subsets of whole blood leukocytes, PMN-MDSC-enriched PBMCs and neutrophil/PMN-MDSC-enriched WBCs were assessed in 5 (whole blood leukocytes), 6 (PMN-MDSC-enriched PBMCs) or 1 (neutrophil/PMN-MDSC-enriched WBCs) technical replicates using flow cytometry. The CSC paradigm was conducted as previously described (Amoroso et al., 2020; Amoroso et al., 2019; Langgartner et al., 2015; Reber et al., 2007; Reber et al., 2008). Briefly, on day 1 all experimental mice were assigned to either the CSC or the SHC group in a body weight-matched manner.

    • Effects of chronic psychosocial stress on ‘binge-like’ sucrose intake in mice

      2023, Progress in Neuro-Psychopharmacology and Biological Psychiatry
      Citation 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).

    View all citing articles on Scopus
    1

    These authors contributed equally.

    View full text