Figure legend
Fig 1. IN OT improved the DSS-induced abnormal stress-related behavior of mice. During water avoidance stress exposure, DSS-induced IBD mice treated with intranasal oxytocin for 7 days showed (A) more activity, (B) less immobility and (C) more grooming behavior than untreated mice. IN atosiban reversed IN OT effects in IBD mice(A-C). Data are presented as means ± SEM. one-way ANOVA, n = 6. ***P<0.001 for main effect: DSS versus normal control; ###P < 0.001 for main effect: IN OT versus without IN OT in DSS-induced IBD mice;$$$P < 0.001 for main effect: IN atosiban versus without IN atosiban of DSS-induced IBD mice treated with IN OT.
Fig 2. Intranasal OT restored DSS-induced hippocampal change of gene expression. A. Representative Western blots of hippocampal nNOS, ERK, pERK and OTR.B. Statistical graph of pERK/ERK ratio in different groups. C. Statistical graph of OTR ratio in different groups. D. Statistical graph of nNOS ratio in different groups. E. Protein expression of NO in hippocampus between different groups. F. Protein expression of BDNF in hippocampus between different groups. Data are presented as means ± SEM. one-way ANOVA, n =3-6. **P<0.01 and ***P<0.001 for main effect: DSS versus normal control; #P<0.05,##P<0.01,###P < 0.001 and ####P<0.0001 for main effect: IN OT versus without IN OT in DSS-induced IBD mice;$P<0.05 and $$$P < 0.001 for main effect: IN atosiban versus without IN atosiban of DSS-induced IBD mice treated with IN OT.
Fig 3. IN OT ameliorated DSS-induced intestinal inflammation. weight(A), colon length (B), disease Activity Score (C), expression of TNF-α (D)and IL-1β (E), histological Score (F), colon macroscopic damage score (G) was assessed in different groups. H. Representative image of HE staining in different groups. Data are presented as means ± SEM. one-way ANOVA, n =6-10. **P<0.01 and ***P<0.001 for main effect: DSS versus normal control; #P<0.05 and ###P < 0.001 for main effect: IN OT versus without IN OT in DSS-induced IBD mice;$P<0.05,$$P<0.01 and $$$P < 0.001 for main effect: IN atosiban versus without IN atosiban of DSS-induced IBD mice treated with IN OT.
Fig 4. Intranasal OT reverted the changes induced by DSS in HPA axis and the sympathetic-adrenal medulla (SAM) axis. CRH in hypothalamus(A), ACTH(B), Cortisol(C), Catecholamine in serum(D), Oxytocin in hypothalamus(E), Oxytocin in serum(F), Ach(G) and TNF-α(H) in spleen were assessed in different groups. Data are presented as means ± SEM. one-way ANOVA, n =4-7. *P<0.05,**P<0.01 and ***P<0.001 for main effect: DSS versus normal control; #P<0.05,##P<0.01 and ###P < 0.001 for main effect: IN OT versus without IN OT in DSS-induced IBD mice; NS: Without statistically significant; $P<0.05,$$P<0.01 and $$$P < 0.001 for main effect: IN atosiban versus without IN atosiban of DSS-induced IBD mice treated with IN OT.
Fig 5. IN OT attenuated the DSS-induced changes in macrophage cell, NK cell, and Treg cell populations in the spleen. Percentages of M1 macrophage (A), M2 macrophage(B), CD3-NKp46+NK cell (C), CD4+ CD25+foxp3+Treg cell(D) in the spleen. Data are presented as means ± SEM. one-way ANOVA, n =4-6. **P<0.01 and ***P<0.001 for main effect: DSS versus normal control; #P<0.05 and ##P<0.01 for main effect: IN OT versus without IN OT of DSS-induced IBD mice;$P<0.05 and $$P<0.01 for main effect: IN atosiban versus without IN atosiban of DSS-induced IBD mice treated with IN OT.
Fig 6.Splenectomy abolished the protective effects of IN OT against DSS-induced abnormal stress-related behavior and intestinal inflammation. A-C. Protective effects of IN OT on DSS-induced abnormal stress-related behavior was abolished by splenectomy. weight(D), colon length (E), disease activity score (F), histological score (G), colon macroscopic damage score (H) was assessed in different groups. I. Representative image of HE staining in different groups. Data are presented as means ± SEM. two-way ANOVA, n =5. **P<0.01 and ***P<0.001 for main effect: DSS versus normal control; #P<0.05 and ##P<0.01 for main effect: IN OT versus without IN OT of DSS-induced IBD mice;$P<0.05 and $$P<0.01 for main effect: IN atosiban versus without IN atosiban of DSS-induced IBD mice treated with IN OT.
Fig 7. Possible mechanisms for IN OT improved DSS-induced abnormal stress-related behavior and intestinal inflammation.Intestinal inflammation “communicated” with brain through systemic communication, HPA axis and neural communication (vagal and sympathetic system), which induced psychological disorders. IN OT administration improved DSS-induced abnormal stress-related behavior and intestinal inflammation through HPA axis and cholinergic anti-inflammation pathway.