The extracellular vesicle of depressive patient-derived Escherichia fergusonii induces vagus nerve-mediated neuroinflammation in mice

Abstract

Background: Gut microbiota dysbiosis is closely associated with psychiatric disorders such as depression and anxiety (DA). In our preliminary study, fecal microbiota transplantation from volunteers with psychological stress and subclinical symptoms of depression (Vsd) induced DA-like behaviors in mice. Escherichia fergusonii (Esf) was found to be more abundant in the feces of Vsd compared to healthy volunteers. Therefore, we investigated the effect of Esf on DA-like behavior and neuroinflammation in mice with and without celiac vagotomy.

Methods and results: Orally gavaged Esf increased DA-like behaviors, tumor necrosis factor (TNF)-α, and toll-like receptor-4 (TLR4) expression, and NF-κB⁺Iba1⁺ and lipopolysaccharide (LPS)⁺Iba1⁺ cell populations, while decreasing serotonin, 5-HT_1A receptor, and brain-derived neurotrophic factor (BDNF) expression in the hippocampus and prefrontal cortex. However, celiac vagotomy attenuated Esf-induced DA-like behavior and neuroinflammation. Orally gavaged extracellular vesicle (EV) from Vsd feces (vfEV) or Esf culture (esEV) induced DA-like behavior and inflammation in hippocampus, prefrontal cortex and colon. However, celiac vagotomy attenuated vfEV- or esEV-induced DA-like behaviors and inflammation in the brain alone, while vfEV- or esEV-induced blood LPS and TNF-α levels, colonic TNF-α expression and NF-κB-positive cell number, and fecal LPS level were not. Although orally gavaged fluorescence isothiocyanate-labeled esEV was translocated into the blood and hippocampus, celiac vagotomy decreased its translocation into the hippocampus alone.

Conclusions: esEVs may be translocated into the brain via the vagus nerve and bloodstream, subsequently inducing TNF-α expression and suppressing serotonin, its receptor, and BDNF expression through the activation of TLR4-mediated NF-κB signaling, thereby contributing to DA pathogenesis.

Keywords: Enterococcus faecium; Escherichia fergusonii; Depression; Gut dysbiosis; Vagus nerve.

Fig. 1

Fecal microbiota transplantation (FMT) from Vsd caused DA-like behavior and neuroinflammation in mice. Effects of FVsd and FVh on OT (a) and track path (b) in the EPMT and IT in the TST (c). Effects on serotonin (d), BDNF (e), TNF-α (f), and IL-10 (g) expression and BDNF⁺NeuN⁺ and NF-κB⁺Iba1⁺ cell populations (h) in the hippocampus. Effects on IL-6 (i) and corticosterone (CORT) levels (j) in the blood. Effects on TNF-α (k) and IL-10 expression (l) and NF-κB⁺CD11c⁺ cell populations (m) in the colon. NC, vehicle in normal control mice; Vsd, fecal microbiota of Vsd; Vh, the fecal microbiota of Vh. Data values were indicated as mean ± SD (n = 7). *p < 0.05. **p < 0.01. ***p < 0.001. ns, no significant

Fig. 2

EV from FVsd (vfEV) caused DA-like behavior and neuroinflammation in mice. Effect on IT in the TST (a), OT (b) and track path (c) in the EPMT, and total travelled distance (TD, d), distance travelled in the center (DC, e), and time spent in central area (TC, f), and track path (g) in OFT. Effect on serotonin (h), BDNF (i), TNF-α (j), and IL-10 expression (k), and BDNF⁺NeuN⁺, NF-κB⁺Iba1⁺, and LPS⁺Iba1⁺ cell populations (l) in the hippocampus. NC, vehicle in normal control mice; Sham, vehicle; vEv, vfEV in mice; Vx, vehicle in mice with vagotomy; VvEv, vfEV in mice with vagotomy. Data values were indicated as mean ± SD (n = 7). *p < 0.05. **p < 0.01. ***p < 0.001. ****p < 0.0001. ns, no significant

Fig. 3

vfEV increased inflammation-related marker levels in the blood and colon of mice. Effect of efEV on IL-6 (a), corticosterone (CORT, b), and LPS levels (c) in the blood. Effect on myeloperoxidase (MPO, d), TNF-α (e), IL-6 (f), and IL-10 expression (g) and NF-κB⁺CD11c⁺ cell populations (h) in the colon. (i) Effect on fecal LPS level. NC, vehicle in normal control mice; Sham, vehicle; vEv, vfEV in mice; Vx, vehicle in mice with vagotomy; VvEv, vfEV in mice with vagotomy. Data values were indicated as mean ± SD (n = 7). *p < 0.05. **p < 0.01. ***p < 0.001. ****p < 0.0001. ns, no significant

Fig. 4

The fecal microbiota composition of Vsd and Vh. (a) The composition at the phylum level (a) and family level (b). The α-diversity (OTU richness, c) and β-diversity (principal coordinate analysis plot based on Jensen-Shannon analysis, d). The number of gut bacteria Enterobacteriaceae and Enterococacceae grown in DHL (e) agar plates (inoculating Vsd and Vh feces). The levels of E. fergusonii (f), assessed Vsd and Vh feces by qPCR. Fmci, the feces of volunteers with MCI; Fhv, the feces of healthy volunteers. Vsd, n = 6; Vh, n = 6. ^#p < 0.05 vs. Vh

Fig. 5

Effect of E. fergusonii on DA-like behavior and neuroinflammation in mice with or without celiac vagotomy. Effect on IT in the TST (a), OT (b) and track path (c) in the EPMT, and TD (d), DC (e), TC (f), and track path (g) in OFT. Effect on serotonin (h), BDNF (i), TNF-α (j), and IL-10 expression (k) and BDNF⁺NeuN⁺, NF-κB⁺Iba1⁺, and LPS⁺Iba1⁺ cell populations (l) in the hippocampus. Effect on BDNF (m), serotonin (n), TNF-α (o), 5-HT_1AR (p), and NF-κB (p) in the prefrontal cortex. NC, vehicle in normal control mice; sham, vehicle; Es, E. fergusonii in mice; Vx, vehicle in mice with vagotomy; VEs, E. fergusonii in mice with vagtomy. Data values were indicated as mean ± SD (n = 7). *p < 0.05. **p < 0.01. ***p < 0.001. ****p < 0.0001. ns, no significant

Fig. 6

Effects of E. fergusonii on DA-related biomarker levels in the blood and colon of mice with or without celiac vagotomy. Effects on IL-6 (a), corticosterone (CORT, b), and LPS levels (c) in the blood. Effects on myeloperoxidase (MPO, d), TNF-α (e), IL-6 (f), and IL-10 expression (g) and NF-κB⁺CD11c⁺ cell populations (h) in the colon. (i) Effects on fecal LPS level. NC, vehicle in normal control mice; sham, vehicle; Es, E. fergusonii in mice; Vx, vehicle in mice with vagotomy; VEs, E. fergusonii in mice with vagtomy. Data values were indicated as mean ± SD (n = 7). *p < 0.05. **p < 0.01. ****p < 0.0001. ns, no significant

Fig. 7

Effect of esEV on DA-like behavior and neuroinflammation in mice with or without celiac vagotomy. Effects on IT in the TST (a), OT (b) and track path (b) in the EPMT, and TD (d), DC (e), TC (f), and track path (g) in OFT. Effects on serotonin (h), BDNF (i), TNF-α (j), and IL-10 expression (k) and BDNF⁺NeuN⁺, NF-κB⁺Iba1⁺, and LPS⁺Iba1⁺ cell populations (l). Effect on BDNF (m), serotonin (n), TNF-α (o), 5-HT_1AR (p), and NF-κB (p) in the prefrontal cortex. NC, vehicle; eEv, esEV in mice; Vx, vehicle in mice with vagotomy; VeEv, esEV in mice with vagotomy. Data values were indicated as mean ± SD (n = 7). *p < 0.05. **p < 0.01. ***p < 0.001. ****p < 0.0001. ns, no significant

Fig. 8

Effect of esEV on inflammation-related marker levels in the blood and colon mice. Effects on IL-6 (a), corticosterone (CORT, b), and LPS levels (c) in the blood. Effects on myeloperoxidase (MPO, d), TNF-α (e), IL-6 (f), and IL-10 expression (g) and NF-κB⁺CD11c⁺ cell populations (h) in the colon. (i) Effect on fecal LPS level. NC, vehicle; eEv, esEV in mice; Vx, vehicle in mice with vagotomy; VeEv, esEV in mice with vagotomy. Data values were indicated as mean ± SD (n = 7). *p < 0.05. **p < 0.01. ***p < 0.001. ****p < 0.0001. ns, no significant

Fig. 9

Orally gavaged FITC-labeled esEV (FEV) was translocated into the blood (a), hippocampus CA1 and CA3 (b). FITC-labeled esEV and vehicle were orally gavaged. NC, vehicle in normal mice; FEv, FITC-labeled esEV in mice; Vx, vehicle in normal mice; VFEv, FITC-labeled esEV in mice with vagotomy.