As the status of cats increased in the family, so did their health and welfare. Changes in the environment increase the risk of IBD in cats. Similarly, the number of people with IBD has increased over the years due to deviant lifestyles. Human IBD consists of 2 subtypes: Crohn's disease(CD) and Ulcerative colitis(UC) . Despite being recognized for decades, IBD remains a clinical challenge today. Currently, IBD treating with antibiotics or immunosuppressant drugs. However, unwanted side effects and poor efficacy are inherent problems associated with these drugs . As a result, many traditional plant-based therapies have to explore as alternatives. Moreover, the mechanism of Traditional Chinese Medicine on IBD has increasingly raised interest in recent years.
BBr, as one of the most representative and most studied natural alkaloids, has been shown to display numerous pharmacological activities . There has been a significant and increasing interest in exploring the IBD-preventive effects BBr during the last decades. Our results show that BBr reduces the expression of IL-1β, TNF-α, and IL-6 in serum, suggesting a reduced inflammatory response . What is more, we found that BBr maintains the homeostasis of the antioxidant system in the gut . Studies have shown that BBr can produce oxidized berberine (OBB) under the action of intestinal flora, and oxidized berberine can significantly reduce colon shortening and histological damage in IBD patients . Therefore, BBr may also interact with the intestinal flora of cats to generate oxidized berberine, which is anti-inflammatory, alleviates diarrhea symptoms of cats with colitis, and makes the intestinal microenvironment conducive to the colonization of good bacteria.
Abnormal homeostasis of intestinal flora is one necessary pathologic mechanism for IBD. Our study based on 16S rRNA sequence analysis verified that the diversity and composition of intestinal flora in cats with IBD altered. In contrast to previous studies, the diversity of bacteria in this study increased after DSS treatment. The reason may be that the diversity of cats is lower than that of mice and dogs and disrupts the homeostasis more quickly. It should note that the DSS group had a single and small amount of bacteria. It further explains that DSS treatment can promote the propagation of harmful bacteria and temporarily cause an increase in bacterial diversity. We treated IBD cats with BBr and found that their gut microbiota was roughly the same as healthy cats. Firmicutes and Bacteroidetes are the major bacterial phyla in the gastrointestinal tract, and the Firmicutes/Bacteroidetes (F/B) ratio associating with maintaining homeostasis. Our results are consistent with previous studies showing a reduced F/B ratio in IBD cats compared to the CON group. However, Firmicutes and Bacteroidetes were more abundant than in the CON group. The increase of other phyla may influence the F/B ratio.
Significant differences in microbiota composition exist among species[26, 27]. One should remember that changes in this ratio can cause ecological disorders, thereby leading to various pathologies. Here, we found that the F/B ratio increased after BBr treatment to that of healthy cats. Specific probiotics can restore the gut microbial balance by influencing the F/B ratio, as the genus Lactobacillus. Our results imply that BBr treatment dramatically increased the abundance of Lactobacillus, suggesting that certain probiotic strains can manage IBD. Nevertheless, the therapeutic effect of Lactobacillus is limited, and their proliferation can cause pathological damage[28, 29].
A dysbiotic microbiome can affect the host not only directly but also indirectly by altering metabolic processes. Linoleic acid, a component of many cat diets, is degraded less in IBD cats, suggesting an abnormal digestive process that leads to weight loss in IBD cats. Notably, abnormal linoleic acid metabolism increases toxoplasma susceptibility. The abundance of Clostridium spp. is significantly reduced in IBD cats, whose primary function is to deconjugate bile acids and promote fat absorption in the small intestine. However, certain beneficial bacteria (e.g., Blautia spp., Faecalibacterium spp.) that produce immune metabolites such as SCFA increased in IBD cats, caused by the body's defense mechanism. It also illustrates that the intestinal tracts of cats harbor a highly complex microbiota.
Enterotoxins produced by pathogenic bacteria can destroy villous effacement and dysfunction the mucosal barrier[25, 27]. The physical barrier of intestinal epithelial cells and tight junction proteins is part of the intestinal barrier. This study demonstrated that DSS increases intestinal paracellular permeability, alters tight junction proteins' expressions, and destroys colon tissue morphology, suggesting that DSS induces intestinal tight junction barrier dysfunction in vivo. In line with the results obtained in patients with IBD, disrupted the intestinal barrier in cats with 5%DSS. Here, BBr treatment could improve the harsh pathological environments.
Intestinal microflora dysbiosis induces inflammatory reactions through Toll-like receptors (TLRs) . When properly activated, Toll-like receptors protect the gut, but when overactivated, they can cause persistent inflammation that can lead to severe disease. TLR4, the best-characterized member of the toll-like receptors, is known to activate adaptive immune responses and upregulate the expression of inflammatory genes by activating the NF-kB pathway. Notably, more expression of TLR4 in the colon in healthy cats. It may explain why the protein level of TLR4 in the DSS group was lower than that in the other two groups. Hence, BBr can maintain intestinal function and repair the intestinal barrier by controlling TLR4/NF-kB pathway.
Dysfunction of autophagy is associated with the pathogenesis of Crohn's disease . Autophagy is an intracellular degradation pathway and an essential cell survival mechanism. The index of autophagy induction mainly includes microtubule-associated protein one light chain 3 (LC3), Atg7, P62, and Atg5. We found that BBr treatment activated autophagy in IBD cats, as evidenced by increased LC3, Atg5, and Atg7 expression and inhibited P62 expression. Studies have suggested a tight, inverse coupling of autophagy induction and MTORC activation. The components of MTORC include Rictor, Raptor, and GβL. We found that both autophagy and MTORC were activated in IBD cats with BBr treatment, suggesting that the cross-talk between autophagy and MTORC has contradictory results. A recent study has revealed autophagy-induced MTORC activation to avoid excessive myofibroblast accumulation . Long-term stimulation of intestinal mucosa with DSS can induce chronic inflammation, typically characterized by excessive myofibroblasts accumulation. They were reversing this direct interaction under BBr treatment further. Taken together, BBr activates autophagy and MTORC to work together to repair the intestinal barrier.
In conclusion, the principal finding of this study is that BBr could severely recover intestinal microbiome homeostasis, including composition and function. More importantly, BBr reduces inflammation by inhibiting the colon's TLR4/NF-KB signaling pathway. Furthermore, BBr activates MTORC and autophagy in the intestine to promote intestinal epithelial cell proliferation and maintain cell metabolism, restoring intestinal barrier function. These findings provide new insights into the mechanisms of BBr therapy for IBD, in which activated MTORC and autophagy may be involved. Indeed, understanding the composition of intestinal flora in cats is beneficial to finding targeted drugs to treat intestinal diseases in cats, which is of great significance for treating pet gastroenteropathy.