Neutrophil-derived catecholamines mediate negative stress effects on bone.

Mental traumatization is associated with long-bone growth retardation, osteoporosis and increased fracture risk. We revealed earlier that mental trauma disturbs cartilage-to-bone transition during bone growth and repair in mice. Trauma increased tyrosine hydroxylase-expressing neutrophils in bone marrow and fracture callus. Here we show that tyrosine hydroxylase expression in the fracture hematoma of patients correlates positively with acknowledged stress, depression, and pain scores as well as individual ratings of healing-impairment and pain-perception post-fracture. Moreover, mice lacking tyrosine hydroxylase in myeloid cells are protected from chronic psychosocial stress-induced disturbance of bone growth and healing. Chondrocyte-specific ß2-adrenoceptor-deficient mice are also protected from stress-induced bone growth retardation. In summary, our preclinical data identify locally secreted catecholamines in concert with ß2-adrenoceptor signalling in chondrocytes as mediators of negative stress effects on bone growth and repair. Given our clinical data, these mechanistic insights seem to be of strong translational relevance.

Intact GR dimerization is critical for restraining plasma ACTH levels during chronic psychosocial stress.

Male C57BL/6N mice exposed to the chronic subordinate colony housing (CSC; 19 days) paradigm, a preclinically validated model of chronic psychosocial stress, are characterized by unaffected basal morning plasma corticosterone (CORT) concentrations despite adrenal and pituitary hyperplasia and increased adrenocorticotropic hormone (ACTH) plasma concentrations, compared with single-housed control (SHC) mice. However, as CSC mice are still able to show an increased CORT secretion towards novel heterotypic stressors, these effects might reflect an adaptation rather than a functional breakdown of general hypothalamus-pituitary-adrenal (HPA) axis functionality. In the present study we used male mice of a genetically modified mouse line, to investigate whether genetically-driven ACTH overexpression compromises adaptational processes occurring at the level of the adrenals during CSC exposure. Experimental mice carried a point mutation in the DNA binding domain of the glucocorticoid (GC) receptor (GR), attenuating dimerization of GR (GRdim), resulting in a congenially compromised negative feedback inhibition at the level of the pituitary. In line with previous studies, CSC mice in both the wild type (WT; GR+/+) and GRdim group developed adrenal enlargement. Moreover, compared with respective SHC and WT mice, CSC GRdim mice show increased basal morning plasma ACTH and CORT concentrations. Quantitative polymerase chain reaction (qPCR) analysis revealed neither a genotype effect, nor a CSC effect on pituitary mRNA expression of the ACTH precursor proopiomelanocortin (POMC). Finally, CSC increased anxiety-related behavior, active coping and splenocyte in vitro (re)activity in both WT and GRdim mice, while a CSC-induced increase in adrenal lipid vesicles and splenic GC resistance was detectable only in WT mice. Of note, lipopolysaccharide (LPS)-stimulated splenocytes of GRdim mice were resistant to the inhibitory effects of CORT. Together our findings support the hypothesis that pituitary ACTH protein concentration is negatively controlled by GR dimerization under conditions of chronic psychosocial stress, while POMC gene transcription is not dependent on intact GR dimerization under both basal and chronic stress conditions. Finally, our data suggest that adrenal adaptations during chronic psychosocial stress (i.e., ACTH desensitization), aiming at the prevention of prolonged hypercorticism, are protective only to a certain threshold of plasma ACTH levels.

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

Stress-associated somatic and psychiatric disorders are often linked to non-resolving low-grade inflammation, which is promoted at least in part by glucocorticoid (GC) resistance of distinct immune cell subpopulations. While the monocyte/macrophage compartment was in the focus of many clinical and preclinical studies, the role of myeloid-derived suppressor cells (MDSCs) in stress-associated pathologies and GC resistance is less understood. As GC resistance is a clear risk factor for posttraumatic complications in patients on intensive care, the exact interplay of physical and psychosocial traumatization in the development of GC resistance needs to be further clarified. In the current study we employ the chronic subordinate colony housing (CSC) paradigm, a well-characterized mouse model of chronic psychosocial stress, to study the role of myeloid cells, in particular of MDSCs, in innate immune activation and GC resistance following combined psychosocial and physical (e.g., bite wounds) trauma. Our findings support the hypothesis that stress-induced neutrophils, polymorphonuclear (PMN)-MDSCs and monocytes/monocyte-like (MO)-MDSCs get primed and activated locally in the bone marrow as determined by toll-like receptor (TLR)2 upregulation and increased basal and lipopolysaccharide (LPS)-induced in vitro cell viability. These primed and activated myeloid cells emigrate into the peripheral circulation and subsequently, if CSC is accompanied by significant bite wounding, accumulate in the spleen. Here, PMN-MDSCs and monocytes/MO-MDSCs upregulate TLR4 expression, which exclusively in PMN-MDSCs promotes NF-κB hyperactivation upon LPS-stimulation, thereby exceeding the anti-inflammatory capacities of GCs and resulting in GC resistance.

Changes in Functional Glucocorticoid Sensitivity of Isolated Splenocytes Induced by Chronic Psychosocial Stress - A Time Course Study.

Chronic psychosocial stress is a risk factor for the development of numerous disorders, of which most are associated with chronic low-grade inflammation. Given the immunosuppressive effects of glucocorticoids (GC), one underlying mechanism might be the development of stress-induced GC resistance in certain immune cell subpopulations. In line with this hypothesis, male mice exposed to the chronic subordinate colony housing (CSC, 19 days) model develop GC resistance of in vitro lipopolysaccharide (LPS)-stimulated splenocytes, splenomegaly and an increased percentage of splenic CD11b+ cells. Here male C57BL/6N mice were euthanized at different days during CSC, and following 30 days of single housing after stressor termination to assess when CSC-induced splenic GC resistance starts to develop and whether this is a transient effect. Moreover, splenic CD11b, GC receptor (GR) and/or macrophage migration inhibiting factor (MIF) protein levels were quantified at respective days. While mild forms of CSC-induced GC resistance, increased splenic CD11b expression and/or splenomegaly were detectable on days 8 and 9 of CSC, more severe forms took until days 15 and 16 to develop, but normalized almost completely within 30 days following stressor termination (day 51). In contrast, splenic GR expression was decreased in CSC versus single-housed control (SHC) mice at all days assessed. While MIF expression was increased on days 15 and 16 of CSC, it was decreased in CSC versus SHC mice on day 20 despite persisting splenomegaly, increased CD11b expression and functional GC resistance. In summary, our data indicate that GC resistance and CD11b+ cell-mediated splenomegaly develop gradually and in parallel over time during CSC exposure and are transient in nature. Moreover, while we can exclude that CSC-induced reduction in splenic GR expression is sufficient to induce functional GC resistance, the role of MIF in CD11b+ cell-mediated splenomegaly and GC resistance requires further investigation.