CN114452375A

CN114452375A - Application of rehabilitation new liquid and FGF-10 in synergistically regulating intestinal flora and improving immune system steady state

Info

Publication number: CN114452375A
Application number: CN202111659043.7A
Authority: CN
Inventors: 孔晓霞; 黄鹏; 张宏宇; 胡斐
Original assignee: Cixi Institute Of Biomedicine Wenzhou Medical University; Ruian Peoples Hospital
Current assignee: Cixi Institute Of Biomedicine Wenzhou Medical University; Ruian Peoples Hospital
Priority date: 2021-12-30
Filing date: 2021-12-30
Publication date: 2022-05-10

Abstract

The invention provides application of a novel rehabilitation liquid and FGF-10 in synergistically regulating intestinal flora and improving the steady state of an immune system, discloses the synergistic effect of the novel rehabilitation liquid and FGF-10 in treating intestinal inflammation for the first time, finds that FGF-10 can obviously improve the structure of intestinal flora after the novel rehabilitation liquid is enema, promotes the recovery of intestinal inflammation, has a synergistic promotion effect on both the steady state of the intestinal immune system and the repair of an intestinal mucosa barrier, and has a wide application prospect.

Description

Application of rehabilitation new liquid and FGF-10 in synergistically regulating intestinal flora and improving immune system steady state

Technical Field

The invention relates to the field of biomedicine, in particular to application of FGF-10 and a novel rehabilitation solution in regulation of intestinal flora and improvement of immune system homeostasis.

Background

Chronic enteritis, also called Inflammatory Bowel Disease (IBD), is an idiopathic chronic intestinal inflammation affecting the ileum, rectum and colon, mainly including Ulcerative Colitis (UC) and Crohn's Disease (CD). The clinical manifestations are chronic or recurrent abdominal pain, diarrhea and dyspepsia, and the serious one may have mucous stool or watery stool. IBD is considered to be a systemic disease that involves various extra-intestinal tissues and organs and causes extra-intestinal manifestations and general digestive and metabolic dysfunction.

Various kinds of microorganisms including bacteria, viruses, fungi, and the like live in human intestinal tissues. It is estimated that the number of bacteria in human intestinal tract is up to 100 trillion, which is 10 times of total cell number of human body, and the number of genes coded by them is 100 times of human body's own gene number, which has great influence on physiological function and nutrition intake. In recent years, researches show that the intestinal flora is an indispensable important part of human bodies, is one of important 'organs' for the human bodies, and has great influence on the growth and the life of the human bodies. Gut flora imbalance plays an important role in the development of IBD disease. In healthy individuals, 95% of the intestinal bacteria belong to the phylum Bacteroides and the subgroups of Clostridium sphaericum and Clostridium flexibilizatum flora, but the number of these flora is reduced in the intestine of IBD patients, in particular of the genera Bifidobacterium and Thelephora. In CD patients, the diversity of the intestinal flora decreases and the number of firmicutes, which is the major proportion, decreases considerably, while the number of bacteroides increases. In addition, the number of Propionibacterium belonging to the Clostridium family having an anti-inflammatory effect was also found to be reduced in CD patients. Intestinal flora transplantation experiments have proved that intestinal flora imbalance is one of the factors causing symptoms such as IBD. Studies have shown that intestinal flora from IBD patients are transplanted into germ-free mice, which exhibit IBD symptoms, and that the diversity and number ratio of intestinal flora in the mice is similar to that of IBD patients. Similarly, in clinical studies, IBD symptoms can be ameliorated and treated by transplantation of healthy individuals with intestinal fecal flora. For example, transplantation of fecal flora standardized to the midgut site into CD patients significantly increases patient weight and nutritional status, as well as IgM levels, with possible mechanisms of action being clostridium populations IV, XIVa and XVIII induce Treg cell differentiation and production by providing bacterial antigens and TGF- β microenvironments and bacterial metabolites Short Chain Fatty Acids (SCFAs), producing IL-10 with anti-inflammatory effects, etc., and increasing the diversity of intestinal flora. From the above research data, we conclude that intestinal flora plays an important role in maintaining normal physiological processes of the intestinal tract, and imbalance of intestinal flora can cause the occurrence and progression of IBD. Therefore, the improvement and treatment of IBD through the regulation of intestinal flora is of feasibility and research value.

The new rehabilitation liquid is an extract of dried periplaneta americana, and mainly comprises amino acids, small molecular polypeptides and nucleic acids. The existing rehabilitation new liquid is mainly used for clinically treating various ulcerative diseases, particularly skin ulcer. Pharmacological research shows that the new rehabilitation liquid can enhance immunity, reduce inflammation, promote epidermal cell growth and granulation tissue proliferation, accelerate the repair and regeneration of damaged tissues and further promote wound healing. FGF10 is a member of the fibroblast growth factor family and is involved in the development and maturation of various organs, and studies have shown that FGF10 may play an important role in human anorectal development. However, the therapeutic effect of FGF10 on IBD has not been studied and reported.

Disclosure of Invention

In order to fill the blank of the prior art, the invention provides the application of FGF-10 and a rehabilitation new liquid in regulating intestinal flora and improving intestinal homeostasis of an immune system. Specifically, the invention provides the following technical scheme:

in a first aspect of the invention, there is provided a composition for use in the treatment of intestinal inflammation, said composition comprising FGF-10 and a convalescent liquid.

In one embodiment, the composition is a liquid formulation.

In one embodiment, the composition is administered as an enema.

In another aspect of the invention, there is provided a use of the above composition for the preparation of a medicament for the regulation of intestinal flora.

In another aspect of the invention, there is provided a use of the above composition for the preparation of a medicament for improving gut immune system homeostasis.

In another aspect of the invention, there is provided a use of the above composition for the preparation of a medicament for repairing the intestinal mucosal barrier.

Compared with the prior art, the invention has the following beneficial technical effects:

the invention discloses the synergistic effect of the rehabilitation new liquid and FGF-10 in the treatment of intestinal inflammation for the first time, and surprisingly discovers that FGF-10 can obviously improve the structure of intestinal flora after the rehabilitation new liquid is clystered, promotes the recovery of intestinal inflammation, and has the synergistic promotion effect on the steady state of an intestinal immune system and the repair of an intestinal mucosa barrier.

Drawings

The accompanying drawings, which are included to provide a further understanding of the invention and are incorporated in and constitute a part of this specification, illustrate embodiments of the invention and together with the description serve to explain the principles of the invention and not to limit the invention. In the drawings:

FIG. 1 shows the number of Th cells and Treg cells of different groups of mice;

FIG. 2 short chain fatty acid content in feces of different groups of mice.

Detailed Description

The preferred embodiments of the present invention will be described in conjunction with the accompanying drawings, and it will be understood that they are described herein for the purpose of illustration and explanation and not limitation.

Example 1 method for constructing enteritis model of mouse

The mouse Inflammatory Bowel Disease (IBD) model was induced by Dextran Sulfate Sodium (DSS) as is conventional in the art, and was specifically operated as follows:

10 BALB/c mice with the age of 8 weeks are selected, the weight of the mice is 16-18 g, the number of the mice is half that of the mice, and 5% DSS is prepared by distilled water. After the mice freely drink 5% DSS for 7 days, the mice freely drink distilled water for 10 days, and 1 cycle is formed. The mice have diarrhea on the 4 th to 5 th days after the start of molding, the occult blood test is positive, and the naked eye bloody stool on the 5 th to 7 th days lasts until the 8 th to 9 th days. After 7 days after 1 cycle, 10 mice were randomly selected and sacrificed, and the pathological results are visible: the lesions developed continuously from the anus to the buccal end of the mouse, with the rectum being the most severe. Congestion and edema of mucous membrane at the affected part can lead to scattered erosion or ulcer. The mucosa and submucosa are predominantly infiltrated with neutrophils. The mouse IBD model is shown to be successfully modeled.

Example 2 evaluation of the synergistic therapeutic Effect of convalescent liquid and FGF-10 on a mouse model of IBD by DAI scoring

Taking 80 BALB/c mice with 8 weeks old, weighing 16-18 g and half each female and half, randomly dividing the mice into three groups:

normal control group: 20 animals, each half of male and female, are fed with normal feed and distilled water every day;

IBD model group: 20, male and female halves, under daily normal feed, were modeled according to the method of example 1;

IBD convalescence treatment group: 20 mice, each male and female, were modeled according to the method of example 1 under daily normal feed feeding, and mice were subjected to enema daily with rehabilitation new liquid (purchased from Sichuan good doctor Panxi pharmaceutical industry, LLC) supplemented with FGF-10 from day 5 of modeling, with the concentration of rehabilitation new liquid in the enema liquid being 6 mg/mL;

IBD convalescence + FGF-10 treatment group: 20 mice, each male and female, were modeled according to the method of example 1 under daily normal feed feeding, and mice were subjected to enema daily with rehabilitation new liquid (purchased from Sichuan good doctor Panxi pharmaceutical industry, LLC) added with FGF-10 from day 5 of modeling, the concentration of rehabilitation new liquid in the enema liquid was 6mg/mL, and the concentration of FGF-10 was 1 mg/mL;

from day 1 of modeling, mice were observed daily for hair color, mental state, activity, fecal behavior, occult blood, and condition of macroscopic bloody stool, Disease Activity Index (DAI) scores were made according to table 1, daily DAI values were calculated for each mouse based on the body mass reduction, fecal behavior, and occult blood condition scores, and the mean was calculated. To assess disease activity.

Table 1: DAI scoring criteria

Note: normal feces: forming excrement;

loosening feces: pasty, semi-formed feces that do not adhere to the anus;

sparse feces: can adhere to the anus and the anus to form a thin water sample.

The results of the experiments are shown in table 2, and the weight of the mice is reduced and bloody stool appears after the DSS is used for inducing the enteritis of the mice. After the treatment of enema of the refreshment liquid, the DAI index of the mouse can be obviously improved. FGF-10 is added into the novel rehabilitation liquid, so that the treatment effect of the novel rehabilitation liquid on IBD (infectious bursal disease) can be synergistically improved, the weight loss of mice is remarkably improved, and bloodless stools are avoided under naked eyes.

Table 2: DAI scores in mice of different treatment groups

Group of	n	DAI score
			Control group	20	0
IBD model group	20	3.33
			IBD model + convalescence liquid treatment group	20	1.32
IBD model + convalescent liquid + FGF-10 treatment group	20	0.96

Example 3 synergistic Regulation of intestinal flora in mice with enteritis by convalescent liquid and FGF-10

The total number of intestinal flora of 4 groups of mice treated differently, the levels of 2 intestinal probiotics (such as bifidobacterium and lactobacillus) and 6 intestinal pathogenic bacteria (such as escherichia coli, staphylococcus, veillonella, helicobacter pylori, streptococcus and pseudomonas aeruginosa) are detected by a real-time fluorescence quantitative PCR method. The results are shown in table 3, the number of intestinal pathogenic bacteria of the DSS-induced enteritis model mice is increased, and the number of probiotics is reduced; the number of pathogenic bacteria in the intestinal tract can be obviously reduced and the number of intestinal tract probiotics can be increased after the new rehabilitation liquid is treated, however, as can be seen from table 3, after the new rehabilitation liquid is clystered, although the number of pathogenic bacteria is obviously reduced, the effect is not obvious for the recovery of the intestinal tract probiotics, the inflammatory reaction is probably reduced because the new rehabilitation liquid plays more roles in clystering, and after FGF-10 is added, the inhibition effect on the intestinal tract pathogenic bacteria is further improved, the content of the probiotics in the intestinal tract is obviously improved, and the FGF-10 plays an important role in the recovery of the intestinal tract environment.

TABLE 3 comparison of the intestinal flora (log/g feces, x. + -. s)

Example 4 Effect of convalescing solution and FGF-10 on the improvement of intestinal immune System homeostasis in enteritis mice

In order to further study the role of the convalescent liquid and FGF-10 in intestinal immune homeostasis, the number of immune cells in colon tissues of each group of mice is subjected to flow detection on the modeling 10 Th day (namely, the treatment 5 Th day), the detection results of each group are shown in figure 1, the number of Th cells of DSS-induced enteritis mice is increased, the number of Treg cells is reduced, the convalescent liquid treatment can inhibit the increase of the Th cells and increase the number of the Treg cells, and the addition of FGF-10 in the convalescent liquid can synergistically increase the number of the immune cells in intestinal tracts.

Example 5 repair of mucosal barrier function of epithelial cells in mouse intestinal tract by the novel healing liquid

Short-chain fatty acids (SCFAS) are important for maintaining intestinal mucosal barrier. The short chain fatty acid is an organic carboxylic acid containing 1 to 6 carbon atoms, and includes acetic acid, propionic acid, butyric acid, lactic acid, isobutyric acid, isogenic acid, isocaproic acid, and the like. 90% of SCFAS is produced by indigestible carbohydrates in the intestine by bacterial fermentation in the colon, the rest is produced by metabolism such as dietary intake and proteins, and normal human intestinal tract can produce about 50-100 mmol of SCFA every day. Most of the SCFAS are absorbed in the intestine, and a small amount (about 5% to 10% of the total amount) of SCFAS can be detected in the stool sample, so the concentration of SCFAS in the stool can reflect the concentration of SCFAS produced in the intestine to some extent.

To investigate the effect of convalescing fluid on intestinal permeability of inflammatory bowel disease, we measured the short chain fatty acid content in each group of mouse feces by gas chromatography on day 10 of modeling (i.e., day 5 of treatment). The results are shown in fig. 2, the total short-chain fatty acid (SCFA) of DSS-induced enteritis mice is greatly reduced, the SCFA content can be significantly increased after the rehabilitation new liquid is treated, and the synergistic effect of repairing the intestinal mucosal barrier can be achieved by adding FGF-10 into the rehabilitation new liquid.

The results show that FGF-10 can remarkably enhance the treatment effect of the rehabilitation new liquid on IBD and promote the intestinal function recovery of an IBD model, and the two have synergistic technical effects.

It will be apparent to those skilled in the art that various changes and modifications may be made in the present invention without departing from the spirit and scope of the invention. Thus, if such modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include such modifications and variations.

Claims

1. A composition for treating intestinal inflammation, said composition comprising FGF-10 and a healing lotion.

2. The composition of claim 1, wherein the composition is a liquid formulation.

3. The composition according to claim 1 or 2, wherein the composition is administered by enema.

4. Use of a composition according to any one of claims 1 to 3 for the manufacture of a medicament for the regulation of intestinal flora.

5. Use of a composition according to any one of claims 1 to 3 for the manufacture of a medicament for improving gut immune system homeostasis.

6. Use of a composition according to any one of claims 1 to 3 for the preparation of a medicament for the repair of the intestinal mucosal barrier.