Nutrients, Vol. 15, Pages 139: Lactobacillus gasseri JM1 Isolated from Infant Feces Alleviates Colitis in Mice via Protecting the Intestinal Barrier

1. IntroductionThe gastrointestinal tract (GIT) is not only an important organ for digestion and absorption of nutrients, but it also protects the host from toxins and pathogens, thus maintaining environmental homeostasis. In addition, as the first line of defense against external threats, the intestinal barrier effectively separates the luminal contents from the host tissues, which is essential for the health of animals and humans [1,2]. It consists of a mucus layer secreted by goblet cells, a single layer of epithelial cells, and a mucosal lymphatic system containing complex immune cells [3]. When the intestinal barrier is damaged, it may lead to changes in intestinal permeability, causing intestinal diseases, such as inflammatory bowel disease (IBD), irritable bowel syndrome (IBS), food allergies, and obesity [4]. IBD is a disease of unknown origin characterized by chronic inflammation of the GIT, including Crohn’s disease (CD) and ulcerative colitis (UC), with clinical manifestations of persistent diarrhea, usually accompanied by blood and mucus [5]. Although the pathogenesis of IBD is still unclear, accumulated data have suggested that the disease is the result of a combination of genetic and environmental factors that could alter the balance of the intestinal environment and cause intestinal barrier dysfunction [6]. It has been demonstrated that all the tight junction proteins, mucous layer, and gut microbiota were altered [2,7,8,9]. Drug treatments, including aminosalicylates and steroids, are still commonly used for IBD, but long-term administration may cause side effects. New treatments are being researched, such as probiotics, prebiotics, and some microbial metabolites [10,11].Lactic acid bacteria (LAB), including several Lactobacillus gasseri (L. gasseri), are potential probiotics that can maintain gut homeostasis and contribute to the health of the host when given in sufficient amounts [12,13]. In addition, L. gasseri is one of the major species for early intestinal colonization, which is widely present in humans as a commensal bacterium [14]. Recent studies have also demonstrated that the use of LAB can alter the composition of gut microbiota, reduce intestinal damage, alleviate intestinal inflammation, ameliorate type 2 diabetes, and enhance immunomodulatory effects from in vitro and in vivo models [15,16,17,18,19]. Therefore, using probiotics to regulate gut microbiota is considered to be a way to promote health and treat certain diseases.Recent evidence suggested that different probiotics have a direct or indirect positive effect on animals in experimental models of IBD. In different mouse models, dextran sulfate sodium (DSS)-induced colitis has been widely used in the study of IBD because of its simplicity and many similarities with UC patients. In vivo studies have shown that probiotics such as probiotic mixture VSL#3, L. fermentum, L. reuteri, and L. rhamnosus improved the intestinal barrier by alleviating intestinal inflammation, decreasing intestinal barrier permeability, and enhancing mucosal integrity in mice with colitis [1,7,20,21]. Meanwhile, L. gasseri was used to evaluate the protective effect of the intestinal barrier in colitis, and the results showed that the levels of the inflammatory cytokines were restored and mucosal damage was avoided [14,19]. Although many promising results have been achieved with probiotics in alleviating DSS-induced colitis, how probiotics affect the GIT and their underlying mechanisms are unclear and need to be further elucidated.Previously, we conducted in vitro studies on the environmental tolerance, adhesion, and immunomodulatory activity of L. gasseri JM1 isolated from the feces of healthy infants. The results showed that L. gasseri JM1 could survive better in a high acid and high bile salt environment, better adhere to Caco-2 cells, and alleviate LPS-induced inflammation in Caco-2 cells [22]. Therefore, based on the previous study and combined with the current research on probiotics to improve the intestinal barrier, we proposed to continue to investigate the effects of L. gasseri JM1 on the regulation of intestinal mucosal damage, mucus layer characteristics, and gut microbiota in mice with colitis. 4. DiscussionIn recent years, the increasing incidence of IBD has made it one of the critical diseases affecting human health. The use of microorganisms to regulate colitis has become a research hotspot. Many studies have shown that probiotics can alleviate colitis in animal models or clinical IBD patients to varying degrees by enhancing the mucosal barrier, regulating immune response, and improving the composition of gut microbiota [41,42,43]. The results of this study showed the effect of L. gasseri JM1 on the intestinal mucosal injury, intestinal barrier, and intestinal metabolites in UC mice and explored the mechanism of its alleviation of UC.In this study, the acute UC model was induced with 3% DSS drinking water for one week. The DSS group began to show symptoms, such as weight loss, lethargy, loss of appetite and loose stools, on the second day of modeling followed by watery stools and gradual deterioration on the fifth day of model construction, indicating the successful establishment of the acute UC model. The degree of inflammation increased with the increase of modeling time, resulting in weight loss. However, the reason for the slight upward trend in body weight on the fourth day of modeling was tentatively speculated to be that the mice were also gavaged with L. gasseri JM1 at the initial stage of inflammation, and the inflammation continued to cause weight loss in mice, which gradually regained weight on the fourth day after adaptation [44]. No remission trend was observed in the 106 L. gasseri JM1 group, presumably due to inadequate dosage of probiotics. The higher the DAI index, the higher the degree of inflammation. With probiotic and drug interventions, the DAI index increased more slowly than that in the DSS group, and the symptoms of loose stools and bloody stools were relatively reduced. In addition, the length of the colon was an important indicator related to the degree of inflammation. We found the length of the colon was recovered and edema was improved to some extent with the intervention of L. gasseri JM1 and mesalazine. In addition, Yang et al. found that the colonic length was recovered in DSS-induced colitis after the intervention of Bifidobacterium breve CCFM683 [26], which was similar to the result of this study. Similarly, the pathological analysis of the colon showed consistent results that L. gasseri JM1 reversed the degree of intestinal mucosal injury in mice, indicating it was more effective at higher doses.Intestinal inflammation is the pathological manifestation of colitis caused by DSS. MPO has been shown to be a local mediator of tissue injury and resultant inflammation and can also enter the extracellular fluid to participate in circulation [34]. It was found that both L. gasseri JM1 and mesalazine could effectively reduce the activity of serum MPO, indicating that L. gasseri JM1 could reduce inflammatory cell infiltration and inflammatory response in the mucosa. MPO is also associated with oxidative stress levels, and the increase of it leads to oxidative stress in mice [45]. It was observed that the activity of MPO was decreased under the action of L. gasseri JM1, and oxidative stress in mice may be partially alleviated, but further studies are still needed. Mesalamine, a gold treatment for colitis, is also a free radical scavenger and antioxidant that is considered to be the most effective [46]. In this study, the effectiveness of mesalamine was also confirmed, and the comparison between L. gasseri JM1 and mesalamine indicated that L. gasseri JM1 had some alleviating effect.The regulation of LAB in the host immune system has always been one of the hot topics of research. Immune factors, including proinflammatory, anti-inflammatory, and growth factors, have important relationships with inflammation. For example, TNF-α is a critical proinflammatory cytokine causing intestinal inflammation and in the central position of many inflammatory cytokines, which can increase the secretion of inflammatory cytokines such as IL-1β and IL-6, leading to aggravation of intestinal mucosal damage [47]. In turn, the increased secretion of IL-6 further stimulates multiple targeted cells (APCs, T cells) to enhance the inflammatory response, and IL-1β is highly expressed in the inflammatory state, which is positively correlated with the degree of inflammation [7,35]. However, conversely, IL-10 has anti-inflammatory properties by inhibiting pro-inflammatory cytokines and chemokines with negative feedback regulation, thus acting as a downregulator of the inflammatory response. However, it was noteworthy that although the expression level of IL-10 in the DSS group was not significantly different from that in the control group, it also showed an upward trend, which may be related to the regulation of the nonspecific immune system activated by intestinal inflammation [9]. Similarly, we also found that the levels of TNF-α, IL-1β, and IL-6 were effectively reduced, and the level of IL-10 increased by L. gasseri JM1 intervention, indicating that L. gasseri JM1 exerted its immunomodulatory function to stimulate the organism to generate a cascade response and produce anti-inflammatory cytokines, which could play an immunomodulatory role in DSS-induced colitis.Intestinal barrier dysfunction with increased intestinal permeability is another characteristic symptom of IBD pathophysiology. We found similar results to those of Chen [23]. Specifically, goblet cells were stained purple by PAS staining, indicating that the intervention of L. gasseri JM1 significantly increased the number of goblet cells and the secretion of mucin, presumably because L. gasseri JM1 colonized and reproduced in the intestine, which stimulated the development of immune tissues and immune response to promote the growth of goblet cells. Furthermore, highly glycosylated mucins are classified into neutral and acidic mucins according to glycol-branched chain components [48]. Compared with neutral mucus, acidic mucins better protect against bacterial translocation because particularly sulfated mucins appear less degradable by bacterial glycosidases and host proteases [49]. L. gasseri JM1 increased the secretion of mucus and the proportion of acidic mucin, presumably due to the colonization and reproduction of the strain in the intestinal tract to exert a regulatory effect, resistance to the adhesion and migration of pathogenic bacteria, and recovery of the intestinal microenvironment. MUC2, the main component of colonic mucus, is predominantly expressed in a healthy colon. However, the expression of MUC2 was low in colitis due to the impaired goblet cells and reduced secretion of mucus [48,50]. Yang et al. found that the mRNA expression level of MUC2, which was significantly downregulated under DSS induction, could be restored to normal after the intervention of Bifidobacterium [26]. Correspondingly, L. gasseri JM1 and mesalamine improved the damage degree of goblet cells and increased the number of goblet cells, MUC2 gene expression, and mucin secretion as well, which ultimately promoted the formation of the tight mucus layer and thus alleviated colitis by resisting to the invasion of bacteria and other antigens. These were consistent with the staining results of PAS and AB-PAS.Tight junctions between intestinal epithelial cells are the most important mode of attachment and are a highly diversified structure composed of transmembrane and cytoplasmic proteins [38]. The Claudins family of proteins is an essential component of tight junctions. Claudins are a quad transmembrane protein with functions as a “fence” and “barrier”, which form tight junction chains by polymerization within the plasma membrane and secondary assembly with Claudins attached to the cell to cross the extracellular space and produce paracellular closure [38,51]. Occludin was the first important transmembrane protein to be identified. If it is damaged, it would lead to an increase in the permeability of paracellular to macromolecules. In addition, ZO-1 plays a role in connecting multiple types of tight junction proteins to maintain the integrity of the tight junction complex. Dou et al. found that L. casei ATCC 393 and its metabolites increased the expression levels of occludin, ZO-1, and Claudin-1 [52], and Din et al. found Bifidobacterium bifidum ATCC 29,521 also could increase the expression levels of ZO-1, MUC-2, and Claudin-3 [53]. In addition, it has been shown that DSS-induced colitis in mice was accompanied by an increase in the pore-forming protein Claudin-2 [54]. We found few relevant studies with simultaneous changes in Claudin-2 and Claudin-3, and the results showed that the extremely significant increase of Claudin-2 in the DSS group might increase the intercellular pores while the decrease of Claudin-3 would reduce the intercellular sealing ability. After intervention with L. gasseri JM1 and mesalazine, this phenomenon was alleviated in mice with colitis, and the mRNA expression levels of both Occludin and ZO-1 were effectively increased, which prevented the destruction of the intestinal barrier to maintain barrier integrity.It is well-known that physiological disturbances induced by IBD are bound to cause changes in gut microbiota diversity, composition, and structure [55]. It was found by detailed analysis of taxonomic composition that the abundance of Proteobacteria at the phylum level, Enterobacteriaceae, Peptostreptococcaceae, and Turicibacteraceae at the family level, and Shigella and Turicibacter at the genus level all increased significantly, which were the result of gut microbiota dysregulation after DSS. It was worth noting that the increased abundance of Proteobacteria, including many pathogenic bacteria, could exacerbate the microecological imbalances in the intestine [56]. The intestinal pathogen Shigella was a major factor in diarrhea in children under five years of age [57]. Studies found that the abundance of intestinal microorganisms Shigella was significantly higher in UC patients than in healthy individuals [58]. Turicibacter significantly increased in this study, which was consistent with Munyaka’s results [59]. However, there are few studies on Turicibacter at present, and its association with inflammation needs to be further investigated. When L. gasseri JM1 intervened, the abundance of Bacteroides, Mucispirillum, Oscillospira, Clostridium, Parabacteroides, Alistipes, and Ruminococcus increased on the basis of a reduction in the pathogenicity-associated flora mentioned above. Among them, Clostridiaceae belonging to Firmicutes is a kind of human-friendly bacteria, which are involved in various metabolic processes in the host. In particular, Clostridium has been shown in several studies to be a butyric acid-producing bacterium that can alleviate the condition of colitis in mice, induce Treg cell differentiation, and reduce inflammation [60]. It has been suggested that Mucispirillum was a resident bacterium in the intestinal mucosa and could also be associated with immunity, as manifested by the generation of Treg cells and IgA. However, there was still much controversy regarding the function of Mucispirillum, and follow-up studies would be needed. Moreover, Federica and others found that the content of Oscillospira was proportional to health, and the abundance of this genus decreased when the body was inflamed [61]. Additionally, Ruminococcus, which belongs to Ruminococcaceae, was a crucial group of gut microbiota in UC that could decompose cellulose, especially fermenting resistant starch to produce metabolites [62]. In this study, it was observed that the abundance of Ruminococcus increased, suggesting that it may play a role in alleviating inflammation through the production of SCFAs.Microbial-derived SCFAs, which are metabolites of gut microbiota, are beneficial to the host and have important functional effects, such as maintaining the water–electrolyte balance, reducing intestinal infections, lowering pH, and improving intestinal function [63]. Combined with the analysis of gut microbiota, it was found that the contents of SCFAs in the intestine significantly reduced in the DSS group while they increased dose-dependently after the L. gasseri JM1 intervention. This may be due to the fact that L. gasseri JM1 exerted its probiotic regulatory effect in the intestine, regulating the composition of gut microbiota, decreasing the abundance of harmful bacteria, and increasing the abundance of acetic and propionic acid-producing Bacteroides and Ruminococcus as well as butyric acid-producing Clostridium and Lactobacillus. This result was consistent with Bian, but unfortunately, F. prausnitzii, the typical butyric acid-producing strain, was not found in this study, and further research is still necessary [64].

留言 (0)

沒有登入
gif