QseC inhibition as an antivirulence approach for colitis-associated bacteria.

Hosts and their microbes have established a sophisticated communication system over many millennia. Within mammalian hosts, this dynamic cross-talk is essential for maintaining intestinal homeostasis. In a genetically susceptible host, dysbiosis of the gut microbiome and dysregulated immune responses are central to the development of inflammatory bowel disease (IBD). Previous surveys of stool from the T-bet-/-Rag2-/- IBD mouse model revealed microbial features that discriminate between health and disease states. Enterobacteriaceae expansion and increased gene abundances for benzoate degradation, two-component systems, and bacterial motility proteins pointed to the potential involvement of a catecholamine-mediated bacterial signaling axis in colitis pathogenesis. Enterobacteriaceae sense and respond to microbiota-generated signals and host-derived catecholamines through the two-component quorum-sensing Escherichia coli regulators B and C (QseBC) system. On signal detection, QseC activates a cascade to induce virulence gene expression. Although a single pathogen has not been identified as a causative agent in IBD, adherent-invasive Escherichia coli (AIEC) have been implicated. Flagellar expression is necessary for the IBD-associated AIEC strain LF82 to establish colonization. Thus, we hypothesized that qseC inactivation could reduce LF82's virulence, and found that an absence of qseC leads to down-regulated flagellar expression and motility in vitro and reduced colonization in vivo. We extend these findings on the potential of QseC-based IBD therapeutics to three preclinical IBD models, wherein we observe that QseC blockade can effectively modulate colitogenic microbiotas to reduce intestinal inflammation. Collectively, our data support a role for QseC-mediated bacterial signaling in IBD pathogenesis and indicate that QseC inhibition may be a useful microbiota-targeted approach for disease management.

Protective Role of the Hepatic Vagus Nerve against Liver Metastasis in Mice.

OBJECTIVE(S): Although accumulating evidence has shown that the autonomic nervous system is involved in liver pathology, its role in regulating cancer development remains unclear. The purpose of this study was to elucidate its detailed mechanisms. METHODS: A mouse model of liver metastasis of colorectal cancer was used. To elucidate the potential mechanisms involved, we examined the effect of selective hepatic vagotomy on the survival rate and liver-to-body weight. We further evaluated the possible involvement of the hepatic sympathetic nerve fibers in this model. RESULTS: The mortality rate and the liver-to-body weight ratio after cancer inoculation were significantly higher in the vagotomized mice than in the sham-operated mice. The vagotomized mice exhibited a transient decrease in hepatic norepinephrine levels following cancer inoculation. Interestingly, the vagotomy-induced exacerbation of liver metastasis was attenuated by supplementary norepinephrine or phenylephrine, a selective α1-adrenoceptor agonist, but not by clonidine, a selective α2-adrenoceptor agonist. CONCLUSION: Collectively, these results suggest that the hepatic vagus nerve may play a protective role against liver metastasis. Hepatic sympathetic nerves may also be involved as a protective efferent loop, possibly acting through the α1-adrenoceptor.

Dietary supplementation with 1-kestose induces altered locomotor activity and increased striatal dopamine levels with a change in gut microbiota in male mice.

1-Kestose (KES), a dietary fiber and prebiotic carbohydrate, benefits various physiological functions. This study aimed to examine whether diets supplemented with KES over three consecutive generations could significantly affect some host physiological aspects, including behavioral phenotypes and gut microbial ecology. Mice that received KES-supplemented diets for three generations demonstrated increased activity compared with those fed diets lacking KES. Furthermore, the KES group showed increased striatal dopamine (DA) and serotonin (5-HT) levels. The observed increase in DA levels within the striatum was positively correlated with locomotor activity in the KES group but not in the control (CON) group. The α-diversities were significantly lower in the KES group compared to the CON group. The three-dimensional principal coordinate analysis revealed a substantial distinction between the KES and CON groups across each generation. At the genus level, most gut microbiota genera exhibited lower abundances in the KES group than in the CON group, except for Bifidobacteria and Akkermansia. Spearman's rank-order analysis indicated significant negative correlations between the striatal DA levels and α-diversity values. These findings suggest that prolonged supplementation with KES may stimulate increased locomotor activity along with elevated striatal DA levels, which are potentially associated with KES-induced alterations in the gut microbiota.

Critical role of gut microbiota in the production of biologically active, free catecholamines in the gut lumen of mice.

There is increasing interest in the bidirectional communication between the mammalian host and prokaryotic cells. Catecholamines (CA), candidate molecules for such communication, are presumed to play an important role in the gut lumen; however, available evidence is limited because of the lack of actual data about luminal CA. This study evaluated luminal CA levels in the gastrointestinal tract and elucidated the involvement of gut microbiota in the generation of luminal CA by comparing the findings among specific pathogen-free mice (SPF-M), germ-free mice (GF-M), and gnotobiotic mice. Substantial levels of free dopamine and norepinephrine were identified in the gut lumen of SPF-M. The free CA levels in the gut lumen were lower in GF-M than in SPF-M. The majority of CA was a biologically active, free form in SPF-M, whereas it was a biologically inactive, conjugated form in GF-M. The association of GF-M with either Clostridium species or SPF fecal flora, both of which have abundant ß-glucuronidase activity, resulted in the drastic elevation of free CA. The inoculation of E. coli strain into GF-M induced a substantial amount of free CA, but the inoculation of its mutant strain deficient in the ß-glucuronidase gene did not. The intraluminal administration of DA increased colonic water absorption in an in vivo ligated loop model of SPF-M, thus suggesting that luminal DA plays a role as a proabsorptive modulator of water transport in the colon. These results indicate that gut microbiota play a critical role in the generation of free CA in the gut lumen.

Dietary tryptophan, tyrosine, and phenylalanine depletion induce reduced food intake and behavioral alterations in mice.

Important precursors of monoaminergic neurotransmitters, dietary tryptophan (TRP), tyrosine, and phenylalanine (all referred to as TTP), play crucial roles in a wide range of behavioral and emotional functions. In the current study, we investigated whether diets devoid of TTP or diets deficient in TRP alone can affect body weight, behavioral characteristics, and gut microbiota, by comparing mice fed on these amino acids-depleted diets to mice fed on diets containing regular levels of amino acids. Both dietary TTP- and TRP-deprived animals showed a reduction in food intake and body weight. In behavioral analyses, the mice fed TTP-deprived diets were more active than mice fed diets containing regular levels of amino acids. The TRP-deprived group exhibited a reduction in serum TRP levels, concomitant with a decrease in serotonin and 5-hydroxyindoleacetic acid levels in some regions of the brain. The TTP-deprived group showed a reduction in TTP levels in the serum, concomitant with decreases in both phenylalanine and tyrosine levels in the hippocampus, as well as serotonin, norepinephrine, and dopamine concentrations in some regions of the brain. Regarding the effects of TRP or TTP deprivation on gut microbial ecology, the relative abundance of genus Roseburia was significantly reduced in the TTP-deprived group than in the dietary restriction control group. Interestingly, TTP was found even in the feces of mice fed TTP- and TRP-deficient diets, suggesting that TTP is produced by microbial or enzymatic digestion of the host-derived proteins. However, microbe generated TTP did not compensate for the systemic TTP deficiency induced by the lack of dietary TTP intake. Collectively, these results indicate that chronic dietary TTP deprivation induces decreased monoamines and their metabolites in a brain region-specific manner. The altered activities of the monoaminergic systems may contribute to increased locomotor activity.

Dietary delivery of acetate to the colon using acylated starches as a carrier exerts anxiolytic effects in mice.

Recently, short-chain fatty acids (SCFA) have been shown to play an important role in mediating the gut-brain interaction and thereby participate in the patho-physiological process of stress-related disorders. In the current study, we examined whether SCFA generated in the lower gut affects host metabolic and behavioral characteristics. To determine this, we used special diets containing acylated starches that can reach the colon without being absorbed in the upper gastrointestinal tract of male mice. The delivery of SCFA to the colon using this method induced a substantial increase in acetate, butyrate, and propionate in the cecum. Moreover, the diets containing acylated starches also decreased microbial diversity in the cecum, concomitant with a significant impact on microbial composition. In marble-burying (MB) tests, the mice that consumed diets containing acetylated starches showed a decrease in anxiety-like behavior compared with the mice that consumed diets containing either butyrylated or propionylated starches. Cecal acetate contents were significantly associated with anxiety-like behaviors when evaluated by elevated plus-maze and MB tests. Collectively, these results indicate that gut acetate elevation of a dietary origin may exert anxiolytic effects on behavioral phenotypes of the host.

The Gut Microbiome Derived From Anorexia Nervosa Patients Impairs Weight Gain and Behavioral Performance in Female Mice.

Anorexia nervosa (AN) results in gut dysbiosis, but whether the dysbiosis contributes to AN-specific pathologies such as poor weight gain and neuropsychiatric abnormalities remains unclear. To address this, germ-free mice were reconstituted with the microbiota of four patients with restricting-type AN (gAN mice) and four healthy control individuals (gHC mice). The effects of gut microbes on weight gain and behavioral characteristics were examined. Fecal microbial profiles in recipient gnotobiotic mice were clustered with those of the human donors. Compared with gHC mice, gAN mice showed a decrease in body weight gain, concomitant with reduced food intake. Food efficiency ratio (body weight gain/food intake) was also significantly lower in gAN mice than in gHC mice, suggesting that decreased appetite as well as the capacity to convert ingested food to unit of body substance may contribute to poor weight gain. Both anxiety-related behavior measured by open-field tests and compulsive behavior measured by a marble-burying test were increased only in gAN mice but not in gHC mice. Serotonin levels in the brain stem of gAN mice were lower than those in the brain stem of gHC mice. Moreover, the genus Bacteroides showed the highest correlation with the number of buried marbles among all genera identified. Administration of Bacteroides vulgatus reversed compulsive behavior but failed to exert any substantial effect on body weight. Collectively, these results indicate that AN-specific dysbiosis may contribute to both poor weight gain and mental disorders in patients with AN.

Regulation of gut luminal serotonin by commensal microbiota in mice.

Gut lumen serotonin (5-hydroxytryptamine: 5-HT) contributes to several gastrointestinal functions such as peristaltic reflexes. 5-HT is released from enterochromaffin (EC) cells in response to a number of stimuli, including signals from the gut microbiota. However, the specific mechanism by which the gut microbiota regulates 5-HT levels in the gut lumen has not yet been clarified. Our previous work with gnotobiotic mice showed that free catecholamines can be produced by the deconjugation of conjugated catecholamines; hence, we speculated that deconjugation by bacterial enzymes may be one of the mechanisms whereby gut microbes can produce free 5-HT in the gut lumen. In this study, we tested this hypothesis using germ-free (GF) mice and gnotobiotic mice recolonized with specific pathogen-free (SPF) fecal flora (EX-GF). The 5-HT levels in the lumens of the cecum and colon were significantly lower in the GF mice than in the EX-GF mice. Moreover, these levels were rapidly increased, within only 3 days after exposure to SPF microbiota. The majority of 5-HT was in an unconjugated, free form in the EX-GF mice, whereas approximately 50% of the 5-HT was found in the conjugated form in the GF mice. These results further support the current view that the gut microbiota plays a crucial role in promoting the production of biologically active, free 5-HT. The deconjugation of glucuronide-conjugated 5-HT by bacterial enzymes is likely one of the mechanisms contributing to free 5-HT production in the gut lumen.

[A case of curatively resected AFP producing gastric cancer that responded remarkably to 1 course of TS-1 and showed complete loss of multiple liver metastatic tumors].

A 73-year-old woman was admitted to our hospital for evaluation of hypochondralgia, and a thorough examination revealed an AFP producing gastric cancer with multiple liver metastases. One course of TS-1 100 mg/day for 4 weeks and discontinuation for 2 weeks was started from February, 2003. After 3 months, the level of AFP reduced remarkably from 53,700 ng/ml to the normal limit. The metastatic tumors in the liver showed regression, and after 14 months, CT scanning showed that the tumors had disappeared. Since the size of the original tumor showed no change, distal gastrectomy was performed, and curability A was achieved. We consider this rare case has significant value in terms of treatment of AFP producing gastric cancer with multiple liver metastases. We think the combination of surgery and chemotherapy such as TS-1 will lead to a better prognosis in such cases.

Chronic psychological stress exaggerates the compound 48/80-induced scratching behavior of mice.

Although accumulating clinical evidence has shown that psychological stress worsens cutaneous symptoms by exaggerating scratching behavior, how the stress affects the scratching is unclear. Therefore, we herein investigated this using an animal model of scratching. Male BALB/c mice were exposed to 1h water avoidance stress (WAS) for ten consecutive days. Twenty-four hours after the last stress session, the mice were injected into the back of the neck with a condensation product of N-methyl-p- methoxyphenethylamine with formaldehyde (compound 48/80), and their scratching behavior was then observed for 120min. Mast cell number in the skin and histamine and corticosterone levels in the plasma were examined. The scratching number was significantly higher in the chronic WAS group than in the control group. Both mast cell number in the skin and the peak histamine in the plasma after the compound 48/80 injection were also significantly higher in the chronic WAS group in comparison to the control group. Chronic WAS delayed the peak corticosterone plasma response to the compound 48/40 injection. These findings indicate that chronic WAS exacerbates the compound 48/80-induced scratching behavior of mice. Both the increased number of skin mast cells and delayed glucocorticoid reaction may be related to this exacerbation.