Microbiota plays a role in shaping the HPA-axis response to psychological stressors. To examine the role of microbiota in response to acute immune stressor, we stimulated the adaptive immune system by anti-CD3 antibody injection and investigated the expression of adrenal steroidogenic enzymes and profiling of plasma corticosteroids and their metabolites in specific pathogen-free (SPF) and germ-free (GF) mice. Using UHPLC-MS/MS, we showed that 4 hours after immune challenge the plasma levels of pregnenolone, progesterone, 11-deoxycorticosterone, corticosterone (CORT), 11-dehydroCORT and their 3α/β-, 5α-, and 20α-reduced metabolites were increased in SPF mice, but in their GF counterparts, only CORT was increased. Neither immune stress nor microbiota changed the mRNA and protein levels of enzymes of adrenal steroidogenesis. In contrast, immune stress resulted in downregulated expression of steroidogenic genes (Star, Cyp11a1, Hsd3b1, Hsd3b6) and upregulated expression of genes of the 3α-hydroxysteroid oxidoreductase pathway (Akr1c21, Dhrs9) in the testes of SPF mice. In the liver, immune stress downregulated the expression of genes encoding enzymes with 3β-hydroxysteroid dehydrogenase (HSD) (Hsd3b2, Hsd3b3, Hsd3b4, Hsd3b5), 3α-HSD (Akr1c14), 20α-HSD (Akr1c6, Hsd17b1, Hsd17b2) and 5α-reductase (Srd5a1) activities, except for Dhrs9, which was upregulated. In the colon, microbiota downregulated Cyp11a1 and modulated the response of Hsd11b1 and Hsd11b2 expression to immune stress. These data underline the role of microbiota in shaping the response to immune stressor. Microbiota modulates the stress-induced increase in C21 steroids, including those that are neuroactive that could play a role in alteration of HPA axis response to stress in GF animals.

• Klíčová slova
• anti-CD3, germ-free, gut microbiota, immune stress, mice, steroidogenic genes, steroids,
• MeSH
• enzym štěpící postranní řetězce cholesterolu genetika   metabolismus   MeSH
• kortikosteron metabolismus   MeSH
• mikrobiota * MeSH
• myši MeSH
• steroidy metabolismus   MeSH
• systém hypofýza - nadledviny metabolismus   MeSH
• systém hypotalamus-hypofýza * metabolismus   MeSH
• tandemová hmotnostní spektrometrie MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• enzym štěpící postranní řetězce cholesterolu MeSH
• kortikosteron MeSH
• steroidy MeSH

Glucocorticoids (GCs) are hormones that are released in response to stressors and exhibit many activities, including immunomodulatory and anti-inflammatory activities. They are primarily synthesized in the adrenal gland but are also produced in peripheral tissues via regeneration of adrenal 11-oxo metabolites or by de novo synthesis from cholesterol. The present study investigated the influence of the microbiota on de novo steroidogenesis and regeneration of corticosterone in the intestine of germ-free (GF) and specific pathogen-free mice challenged with a physical stressor (anti-CD3 antibody i.p. injection). In the small intestine, acute immune stress resulted in increased mRNA levels of the proinflammatory cytokines IL1β, IL6 and Tnfα and genes involved in de novo steroidogenesis (Stard3 and Cyp11a1), as well as in regeneration of active GCs from their 11-oxo metabolites (Hsd11b1). GF mice showed a generally reduced transcriptional response to immune stress, which was accompanied by decreased intestinal corticosterone production and reduced expression of the GC-sensitive marker Fkbp5. In contrast, the interaction between stress and the microbiota was not detected at the level of plasma corticosterone or the transcriptional response of adrenal steroidogenic enzymes. The results indicate a differential immune stress-induced intestinal response to proinflammatory stimuli and local corticosterone production driven by the gut microbiota.

• Klíčová slova
• 11β-hydroxysteroid dehydrogenase, anti-CD3 antibody, extra-adrenal steroidogenesis, glucocorticoids, intestine, microbiome,
• MeSH
• 11-beta-hydroxysteroiddehydrogenasy genetika   metabolismus   MeSH
• kortikosteron metabolismus   MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• myši MeSH
• steroidy metabolismus   MeSH
• střevní mikroflóra fyziologie   MeSH
• tandemová hmotnostní spektrometrie MeSH
• tenké střevo metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• 11-beta-hydroxysteroiddehydrogenasy MeSH
• kortikosteron MeSH
• steroidy MeSH

The gut microbiota play an important role in shaping brain functions and behavior, including the activity of the hypothalamus-pituitary-adrenocortical (HPA) axis. However, little is known about the effect of the microbiota on the distinct structures (hypothalamus, pituitary, and adrenals) of the HPA axis. In the present study, we analyzed the influence of the microbiota on acute restraint stress (ARS) response in the pituitary, adrenal gland, and intestine, an organ of extra-adrenal glucocorticoid synthesis. Using specific pathogen-free (SPF) and germ-free (GF) male BALB/c mice, we showed that the plasma corticosterone response to ARS was higher in GF than in SPF mice. In the pituitary, stress downregulated the expression of the gene encoding CRH receptor type 1 (Crhr1), upregulated the expression of the Fkbp5 gene regulating glucocorticoid receptor sensitivity and did not affect the expression of the proopiomelanocortin (Pomc) and glucocorticoid receptor (Gr) genes. In contrast, the microbiota downregulated the expression of pituitary Pomc and Crhr1 but had no effect on Fkbp5 and Gr. In the adrenals, the steroidogenic pathway was strongly stimulated by ARS at the level of the steroidogenic transcriptional regulator Sf-1, cholesterol transporter Star and Cyp11a1, the first enzyme of steroidogenic pathway. In contrast, the effect of the microbiota was significantly detected at the level of genes encoding steroidogenic enzymes but not at the level of Sf-1 and Star. Unlike adrenal Sf-1, the expression of the gene Lrh-1, which encodes the crucial transcriptional regulator of intestinal steroidogenesis, was modulated by the microbiota and ARS and this effect differed between the ileum and colon. The findings demonstrate that gut microbiota have an impact on the response of the pituitary, adrenals and intestine to ARS and that the interaction between stress and the microbiota during activation of glucocorticoid steroidogenesis differs between organs. The results suggest that downregulated expression of pituitary Pomc and Crhr1 in SPF animals might be an important factor in the exaggerated HPA response of GF mice to stress.

• Klíčová slova
• HPA axis, acute restraint stress, extra-adrenal glucocorticoid synthesis, germ-free, gut microbiota, intestine, mice,
• MeSH
• enzym štěpící postranní řetězce cholesterolu genetika   MeSH
• fosfoproteiny genetika   MeSH
• fyzické omezení * MeSH
• hypofýza metabolismus   MeSH
• ileum metabolismus   mikrobiologie   MeSH
• kolon metabolismus   mikrobiologie   MeSH
• kortikosteron krev   MeSH
• myši inbrední BALB C MeSH
• nadledviny metabolismus   MeSH
• pro-opiomelanokortin genetika   MeSH
• psychický stres krev   mikrobiologie   MeSH
• receptor CRF typu 1 MeSH
• receptory hormonu uvolňujícího kortikotropin genetika   MeSH
• regulace genové exprese MeSH
• steroidní akutní regulační protein MeSH
• steroidogenní faktor 1 genetika   MeSH
• střevní mikroflóra * MeSH
• systém hypofýza - nadledviny * MeSH
• systém hypotalamus-hypofýza * MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• enzym štěpící postranní řetězce cholesterolu MeSH
• fosfoproteiny MeSH
• kortikosteron MeSH
• pro-opiomelanokortin MeSH
• receptor CRF typu 1 MeSH
• receptory hormonu uvolňujícího kortikotropin MeSH
• steroidní akutní regulační protein MeSH
• steroidogenní faktor 1 MeSH