阅读说明：本技术 可降解色氨酸的植物乳杆菌与色氨酸混合物的应用 (Application of plant lactobacillus capable of degrading tryptophan and tryptophan mixture ) 是由 李柏良 王娜娜 李娜 闫芬芬 刘飞 王松 霍贵成 于 2020-05-28 设计创作，主要内容包括：本发明公开了可降解色氨酸的植物乳杆菌与色氨酸混合物的应用。具体公开了可降解色氨酸的植物乳杆菌与色氨酸混合物在降低结肠组织髓过氧化物酶MPO活性中的应用、在改善结肠炎免疫应答紊乱中的应用、在保护肠道屏障中的应用、在保护肠道粘膜中的应用以及在调节肠道微生物中的应用。本发明通过体外评估16株乳酸杆菌对色氨酸的降解能力,筛选出具有较高降解色氨酸能力的植物乳杆菌KLDS1.0386；植物乳杆菌KLDS1.0386生长性能好,具有较强的产酸能力和耐酸耐胆盐能力,展现了良好的益生功能以及预防结肠炎功能。(The invention discloses application of a mixture of lactobacillus plantarum capable of degrading tryptophan and tryptophan. Specifically discloses application of a mixture of lactobacillus plantarum capable of degrading tryptophan and tryptophan in reducing activity of colon tissue Myeloperoxidase (MPO), application in improving colitis immune response disorder, application in protecting intestinal tract barrier, application in protecting intestinal tract mucosa and application in regulating intestinal microorganisms. According to the invention, the degradation capability of 16 lactobacillus strains on tryptophan is evaluated in vitro, and lactobacillus plantarum KLDS1.0386 with higher tryptophan degradation capability is screened out; the lactobacillus plantarum KLDS1.0386 has good growth performance, stronger acid production capacity and acid and bile salt resistance capacity, and shows good probiotic function and colitis prevention function.)

1. Application of mixture of plant lactobacillus capable of degrading tryptophan and tryptophan in preventing colitis.

2. Use of a mixture of lactobacillus plantarum capable of degrading tryptophan according to claim 1 and tryptophan for reducing the activity of the colon tissue myeloperoxidase MPO.

3. The use of the mixture of lactobacillus plantarum capable of degrading tryptophan according to claim 1 for improving colitis immune response disorder.

4. The use according to claim 3, wherein ameliorating colitis immune response disorders comprises reducing the levels of the proinflammatory cytokines IL-6, IL-1 β, TNF- α and increasing the level of the anti-inflammatory cytokine IL-10.

5. Use of a mixture of lactobacillus plantarum capable of degrading tryptophan according to claim 1 with tryptophan for protecting the intestinal barrier and protecting the intestinal mucosa.

6. The use according to claim 5, wherein protecting the intestinal barrier comprises elevating the levels of the claudin Occupudin, ZO-1 and the mucins MUC1, MUC2 in colon tissue; the intestinal mucosa is protected, and the lactobacillus plantarum capable of degrading tryptophan directly metabolizes tryptophan in the intestinal tract to generate 3-indoleacetic acid IAA, and the 3-indoleacetic acid IAA is used as a ligand of an aromatic hydrocarbon receptor AHR to participate in regulation of an AHR signal path closely related to intestinal immunity.

7. The use of claim 6, wherein the mixture of Lactobacillus plantarum capable of degrading tryptophan and tryptophan activates AHR, induces IL-22/STAT3 pathway, increases the expression of IL-22 and STAT3, further induces the expression of the antimicrobial peptide REG3 γ, promotes tissue repair and regeneration, and thus protects intestinal mucosa.

8. Use of the mixture of lactobacillus plantarum capable of degrading tryptophan according to claim 1 and tryptophan for modulating intestinal microorganisms.

9. The use of claim 8, wherein modulating gut microbiota comprises decreasing the relative abundance of shigella, cladia and helicobacter species in the gut and increasing the relative abundance of prevotella UCG-001, akkermansia and murine bacillaceae.

10. The use according to any one of claims 1 to 9, wherein the lactobacillus plantarum capable of degrading tryptophan comprises lactobacillus plantarum KLDS 1.0386; the amount of the mixture is: every 10 th⁸-10⁹0.2-2mg tryptophan is contained in 0.2mL of CFU/mL lactobacillus plantarum suspension.

Technical Field

The invention relates to an application of a mixture of lactobacillus plantarum capable of degrading tryptophan and tryptophan, in particular to an application of a mixture of lactobacillus plantarum capable of degrading tryptophan and tryptophan in preventing colitis, and belongs to the technical field of medicines.

Background

Inflammatory Bowel Disease (IBD) is a chronic and relapsing autoimmune disease, including both Ulcerative Colitis (UC) and Crohn's Disease (CD) disease types. IBD has been considered as a western disease in the past, and has been mainly concentrated in developed countries such as north america, europe, australia, and new zealand, while in recent years, IBD has been rapidly increasing in incidence in emerging industrial countries such as asia and the middle east, and has been developed as a global disease. IBD is clinically manifested by diarrhea, hematochezia, weight loss and other symptoms, UC mainly affects the mucous membrane of the colon to cause bloody stools, CD can be sectionally distributed in the whole gastrointestinal tract to cause fistulas, which seriously affect the quality of life of people, and heavy economic pressure is brought to patients due to long treatment period and easy relapse. At present, the etiology and pathogenesis of IBD are not completely clear, but with the development of detection technology, more and more evidence shows that the dysbiosis of host intestinal tract causes congenital and adaptive immune response disorder, which in turn leads to intestinal inflammation of genetically susceptible hosts. The existing IBD treatment method mainly adopts drug treatment, although the effect is good, the method has large toxic and side effects and is not suitable for long-term use, so that the search for a safe and effective IBD treatment method is very important.

Such as: at present, the drugs for clinically treating IBD mainly comprise aminosalicylic acid drugs, steroid hormones drugs and immunosuppressant drugs. Currently, aminosalicylic acid mainly comprises 5-aminosalicylic acid and sulfasalazine, and is mainly used for treating mild and moderate IBD, and the main mechanism is that PPAR gamma (Peroxisome proliferator-activator gamma) is activated to reduce the secretion of proinflammatory factors, so that the activation of NF-kappa B inflammatory pathways is inhibited. The glucocorticoid is mainly used for treating moderate and severe IBD patients, can inhibit cell differentiation and activation, reduce the production of proinflammatory cytokines, and induce apoptosis of DC and T cells, has good anti-inflammatory effect, is easy to cause organism metabolic disorder and induce adverse reaction, cannot be used as a medicament for long-term maintenance treatment, and can only be used as an emergency medicament. For IBD patients who cannot be treated by amino salicylic acid medicines and glucocorticoid medicines, immunosuppressive medicines are important treatment medicines, mainly comprising azathioprine, 6-mercaptopurine and methotrexate; the action mechanism of the medicine is to reduce cell proliferation including lymphocytes and inhibit the expression of inflammatory genes, but specific adverse reactions such as nausea, fever, arthralgia, acute pancreatitis and the like can be generated. Treatment with thiopurines in IBD would triple the relative risk of malignant, lymphoma and non-melanoma skin cancers.

Tryptophan is an essential aromatic amino acid for human body, its chemical name is α -amino- β -indole propionic acid, and its molecular formula is C₁₁H₁₂N₂O₂The amino acid is the only amino acid containing indole structure, namely a bicyclic compound, and consists of a six-membered benzene ring and a five-membered nitrogen-containing pyrrole ring. It is not synthesized in human body, and needs to be obtained from diet, and many studies find that tryptophan plays an important role in maintaining the balance between intestinal microorganisms and intestinal mucosal immunity. However, the capability of decomposing and utilizing tryptophan by intestinal microorganisms is limited, researches show that escherichia coli, clostridium, bacteroides and streptococcus digestans have the function of degrading tryptophan, a strain of enterococcus capable of utilizing tryptophan is screened from pig colon by the marmei bud, but the enterococcus is a pathogenic bacterium with potential pathogenicity, and the safety of the strain to human bodies cannot be guaranteed.

Therefore, a highly effective and safe medicine for relieving colitis is urgently needed.

Disclosure of Invention

The invention aims to solve the technical problem of providing an application of a mixture of lactobacillus plantarum and tryptophan capable of degrading tryptophan.

In order to solve the technical problems, the technical scheme adopted by the invention is as follows:

application of mixture of plant lactobacillus capable of degrading tryptophan and tryptophan in preventing colitis.

Use of a mixture of a tryptophan-degrading Lactobacillus plantarum and tryptophan for reducing the activity of the colon tissue myeloperoxidase MPO.

Application of a mixture of lactobacillus plantarum capable of degrading tryptophan and tryptophan in improvement of colitis immune response disorder.

Improving colitis immune response disorder includes reducing the levels of proinflammatory cytokines IL-6, IL-1 beta, TNF-alpha, and increasing the level of anti-inflammatory cytokines IL-10.

Use of a mixture of lactobacillus plantarum capable of degrading tryptophan and tryptophan for protecting the intestinal barrier.

Protecting the intestinal barrier includes elevating the levels of the claudin Occludin, ZO-1 and mucins MUC1, MUC2 in colon tissue.

Application of mixture of plant lactobacillus capable of degrading tryptophan and tryptophan in protecting intestinal mucosa is provided.

The intestinal mucosa is protected, and the lactobacillus plantarum capable of degrading tryptophan directly metabolizes tryptophan in the intestinal tract to generate 3-indoleacetic acid IAA, and the 3-indoleacetic acid IAA is used as a ligand of an aromatic hydrocarbon receptor AHR to participate in regulation of an AHR signal path closely related to intestinal immunity.

The mixture of the lactobacillus plantarum and the tryptophan, which can degrade the tryptophan, activates an aromatic hydrocarbon receptor AHR and induces an IL-22/STAT3 pathway, increases the expression of IL-22 and STAT3, further induces the expression of the antimicrobial peptide REG3 gamma, promotes tissue repair and regeneration, and plays a role in protecting intestinal mucosa.

Application of a mixture of lactobacillus plantarum capable of degrading tryptophan and tryptophan in regulating intestinal microorganisms.

Modulating gut microbiology includes reducing the relative abundance of shigella, rhodobacter and helicobacter species in the gut, and increasing the relative abundance of prevotella UCG-001, Akkermansia (Akkermansia) and muribacteriaceae (muribacteriaceae).

The plant lactobacillus capable of degrading tryptophan comprises plantsLactobacillus KLDS 1.0386; the amount of the mixture is: every 10 th⁸-10⁹0.2-2mg tryptophan is contained in 0.2mL of CFU/mL lactobacillus plantarum suspension.

The lactic acid bacteria used in the invention are regarded as safe food-grade microorganisms as main sources of probiotics, so that a lactic acid bacteria strain capable of efficiently degrading tryptophan is screened out to develop an efficient and safe natural medicine for relieving colitis. Lactic acid bacteria, which are gram-positive bacteria, are usually isolated from fermented foods such as yogurt and kimchi and are the main sources of probiotics. In addition to the extensive use of lactic acid bacteria in food products, they have been shown to be effective in preventing or treating various diseases, such as immune-modulating, cholesterol-lowering, anti-tumor, etc. probiotic functions. The lactobacillus can be attached to the intestinal tract by adhesion, regulate the microbial structure of the intestinal tract, and activate the immunity of the organism, thereby playing a corresponding probiotic function. A large number of in vivo, in vitro and clinical researches prove that different species of lactic acid bacteria have the function of relieving intestinal inflammation, but the influence of the lactic acid bacteria and tryptophan on colitis is not clarified yet.

The invention has the following beneficial effects:

1. through in vitro evaluation of the degradation capability of 16 strains of lactobacillus to tryptophan, lactobacillus plantarum KLDS1.0386 with higher degradation capability to tryptophan is screened out.

2. The lactobacillus plantarum KLDS1.0386 has good growth performance, stronger acid production capacity and acid and bile salt resistance capacity, and shows good probiotic function.

3. Tryptophan and Lactobacillus plantarum KLDS1.0386 can relieve intestinal inflammation caused by DSS-induced colitis of mice to different degrees, but the mixed intervention effect of the Tryptophan and the Lactobacillus plantarum KLDS1.0386 is the best. The mixed group can obviously reduce DAI index, prevent colon from shortening, relieve histopathological injury and reduce MPO activity, and the mixed group can obviously reduce the content of proinflammatory factors IL-6, IL-1 beta and TNF-alpha and obviously improve the content of anti-inflammatory factors IL-10. In addition, the mixed group can also obviously improve the colon intestinal barrier injury of mice caused by DSS, and paul obviously regulates the gene expression quantity of the claudin Occludin and ZO-1 and the levels of mucin MUC1 and MUC2, thereby protecting the integrity of the intestinal barrier.

4. The mixed group of tryptophan and lactobacillus plantarum KLDS1.0386 is used for dry prognosis, the content of tryptophan metabolite IAA in liver, serum and colon tissues of a mouse is obviously increased, the IAA is used as an AHR ligand, the AHR is activated, an IL-22/STAT3 pathway is induced, secretion of IL-22 and expression of STAT3 are increased, expression of antimicrobial peptide REG3 gamma is induced, and enteritis injury caused by DSS induction is relieved.

5. Tryptophan and Lactobacillus plantarum KLDS1.0386 can be mixed to intervene to increase the diversity of intestinal flora of enteritis mice, reduce the relative abundance of firmicutes and proteobacteria, and increase the relative abundance of bacteroidetes. Furthermore, at the genus level, it is possible to significantly reduce the relative abundance of harmful bacteria such as Escherichia-Shigella, Alisipes, and Helicobacter in the intestinal tract and increase the relative abundance of beneficial bacteria such as Akkermansia, Prevotella UCG-001, and Murebacteriaceae, thereby preventing intestinal microbial disorders caused by DSS.

Drawings

FIG. 1 is a standard graph of tryptophan;

FIG. 2 is a graph showing the ability of Lactobacillus to degrade tryptophan; note: different lower case letters indicate that the difference of each group is significant (P <0.05), the same letter indicates that the difference is not significant (P > 0.05);

FIG. 3 is a graph of the growth of Lactobacillus plantarum KLDS 1.0386;

FIG. 4 is a graph of acid production performance of Lactobacillus plantarum KLDS 1.0386;

FIG. 5 is a graph of the effect of each treatment on colon tissue myeloperoxidase activity; note: different lower case letters indicate that each group is significantly different (P < 0.05);

FIG. 6 is a graph showing the effect of each treatment group on cytokines in mouse serum; note: A-D are respectively TNF-alpha, IL-1 beta, IL-6 and IL-10 content, and different lower case letters indicate that the difference of each group is obvious (P is less than 0.05);

FIG. 7 is a graph showing the effect of each treatment group on the expression level of the claudin gene; note: different lower case letters indicate that each group is significantly different (P < 0.05);

FIG. 8 is a graph showing the effect of each treatment group on the expression level of mucin gene; note: control: a control group; DSS: a model group; trp: a group of tryptophans; LAB: KLDS1.0386 group; mix: and (4) mixing the groups. (A) MUC1 gene expression level, (B) MUC2 gene expression level, different lower case letters indicate that each group is significantly different (P < 0.05);

FIG. 9 is a graph showing the effect of treatment on IAA levels in liver, serum and colon tissues; note: control: a control group; DSS: a model group; trp: a group of tryptophans; LAB: KLDS1.0386 group; mix: mixing groups;

FIG. 10 is a graph showing the effect of each treatment group on the expression level of an AHR signaling pathway-associated gene; note: (A) AHR gene expression level, (B) STAT3 gene expression level, (C) REG3 γ gene expression level, (D) IL-22 expression level, with different lower case letters indicating that the groups are significantly different (P < 0.05);

fig. 11 is a venn diagram and a sample cluster diagram.

Detailed Description

The present invention will be further described with reference to the accompanying drawings.