Construction method and application of atrophic gastritis animal model

Abstract

The invention discloses a construction method and application of an atrophic gastritis animal model. The atrophic gastritis model of the meriones unguiculatus is constructed through combination of helicobacter pylori infection and benzopyrene induction for the first time, and the obtained model proves that the helicobacter pylori infection causes physiological injury of the meriones unguiculatus and the helicobacter pylori infection and benzopyrene have a synergistic injury causing effect. Important reference is provided for a physiological injury mechanism caused by combined action of the two, and a starting point is provided for assisting in making an eradication treatment scheme to prevent occurrence and development of physiological injury. Meanwhile, the construction method is easy to operate, low in cost and stable in pathological change, and the constructed meriones unguiculatus atrophic gastritis model is high in success rate and has good application prospects.

Description

Technical Field

[0001] The present invention relates to the technical field of animal models, and in particular to a method for constructing an atrophic gastritis animal model and an application thereof. Background Art

[0002] Atrophic gastritis, also known as chronic atrophic gastritis, is a chronic digestive system disease characterized by atrophy of gastric mucosal epithelium and glands, decreased number, thinning of gastric mucosa, thickening of mucosal basal layer, or accompanied by pyloric gland metaplasia and intestinal gland metaplasia, or atypical hyperplasia. It is often manifested as upper abdominal pain, fullness, belching, loss of appetite, or weight loss, anemia, etc., without specificity. It is a multi-pathogenic disease or precancerous lesion.

[0003] The research and treatment of atrophic gastritis have attracted much attention. The construction of an animal model of atrophic gastritis is the key material for the basic and clinical research of the disease. A good animal model will provide direct in vivo evidence for the prevention and treatment of the disease. The existing animal models of atrophic gastritis are mainly constructed by chemical modeling, surgical modeling and biochemical immunology. Among them, the chemical modeling method uses chemical reagents to simulate high-risk factors to intervene in rats. The chemical factors selected are N-methyl-N'-nitro-N-nitrosoguanidine, ammonia, sodium deoxycholate, acid suppressants, non-steroidal anti-inflammatory drugs, ethanol, etc. The above factors cause repeated chronic damage and inflammatory stimulation, resulting in the destruction of the intrinsic glands of the stomach and atrophy. The surgical modeling method commonly uses gastrojejunostomy and spring pylorus implantation. The methods reported by the biochemical immunology method are: using the gastric mucosa of the same rat, physiological saline homogenate, and complete Freund's adjuvant in a 1:1 ratio to form an emulsion, subcutaneously injecting rats, combined with sodium deoxycholate and ethanol stimulation, successfully replicating the chronic atrophic gastritis model. H.pylori strain SS1 has also been isolated from clinical practice. This strain can effectively colonize the stomach of experimental animals for a long time and successfully produce chronic inflammation, mucosal atrophy, intestinal metaplasia and atypical hyperplasia in the stomach of experimental animals. Although there are many ways to model, single-factor modeling still has some defects, such as long cycle, unstable model, low success rate, low similarity between the induced animal model and the clinic, immune deficiency and high cost.

[0004] In view of this, the present invention is proposed. Summary of the invention

[0005] The purpose of the present invention is to provide a method for constructing an atrophic gastritis animal model and its application. The atrophic gastritis animal model obtained by the construction method of the present invention can improve the defects existing in the above-mentioned prior art.

[0006] The present invention is achieved in that:

[0007] In a first aspect, the present invention provides a method for constructing an atrophic gastritis animal model, which comprises treating an experimental animal with Helicobacter pylori and benzopyrene to obtain an atrophic gastritis animal model.

[0008] Helicobacter pylori (Hp) is a Gram-negative microaerobic spirochete that generally colonizes the gastric mucosa and, as a pathogenic factor, can cause diseases such as gastritis, peptic ulcer and gastric cancer. The present invention selects Hp infection because it is the initiating factor of the evolution process of chronic non-atrophic gastritis → atrophic gastritis → intestinal metaplasia → dysplasia → gastric cancer.

[0009] Benzopyrene (BaP) is a group of polycyclic aromatic hydrocarbons containing five benzene rings. Humans ingest it through contact with contaminated food and air. It does not react directly with human DNA, but forms adducts with DNA after being metabolically activated through the diol-epoxide pathway, o-quinone pathway and free radical cation pathway, causing damage to DNA. Severe damage can lead to gene mutations, inflammation, and even cancer.

[0010] In real life, Hp and BaP exposure at the same time are relatively common phenomena in human activities, and the cause of atrophic gastritis is complex. Studies have confirmed that it is the result of the combined action of multiple factors. Based on this, the inventors of the present invention adopted a comprehensive modeling method combining multiple factors to combine Hp infection with BaP induction, making it closer to the clinical pathogenesis, so as to simulate unhealthy human living habits.

[0011] In some embodiments, the above construction method includes: firstly infecting the experimental animals with Helicobacter pylori by gavage, then gavage benzopyrene to the experimental animals, and then feeding the treated experimental animals normally.

[0012] In some embodiments, the experimental animal comprises a gerbil.

[0013] An ideal animal model should be able to simulate the infection and pathological development process of humans in a natural state. Therefore, when constructing an animal model, the higher the grade of the selected animal is, the better. The use of sterile animals, immunodeficient animals, etc. to construct an infection animal model also has the disadvantages of harsh feeding conditions, high feeding costs, limited experimental operations, and difficult management. The present invention uses long-clawed gerbils as an Hp infection model, which has the advantages that: long-clawed gerbils rarely suffer from gastritis naturally; the amount of Hp required for infection is small, and the pathological histological changes are more obvious; the gastric lesions are most similar to those of humans, and atrophic gastritis, ulcers, intestinal metaplasia and gastric cancer may occur; the lifespan is longer than that of mice, suitable for long-term observation; the amount of gastric tissue is larger than that of mice, suitable for direct histological examination.

[0014] In some embodiments, Helicobacter pylori is administered intragastrically to experimental animals once a week for 5 consecutive weeks.

[0015] In some embodiments, the Helicobacter pylori is administered to the experimental animals by gavage at a concentration of 1×10 9 CFU / mL of Helicobacter pylori suspension, 0.5 mL / time gavage.

[0016] In some embodiments, benzopyrene is administered orally to experimental animals once a week for 4-5 consecutive weeks. More preferably, administration for 5 consecutive weeks has a reinforcing effect on establishing atrophic gastritis model in gerbils.

[0017] In some embodiments, the method of gavage of benzopyrene to experimental animals each time is: using a benzopyrene solution with a concentration of 5 mg / mL, 0.5 mL / time gavage.

[0018] During the experiment, the present invention not only verified the effects of Hp infection alone and BaP induction alone, but also verified the effects of combining Hp infection with BaP induction. The results showed that Hp infection has an enhancing effect on BaP induction. When the above-mentioned doses of Hp and BaP are combined, a long-clawed gerbil model of atrophic gastritis with a high infection rate, pathological changes similar to those of patients with chronic gastritis and high stability can be obtained.

[0019] In some embodiments, the normal feeding period is from week 11 to week 36.

[0020] In a second aspect, the present invention also provides an atrophic gastritis animal model obtained by the above-mentioned method for constructing an atrophic gastritis animal model.

[0021] In a third aspect, the present invention also provides the application of the above-mentioned atrophic gastritis animal model in the research of related diseases, including the research on the mechanism, drug screening and treatment methods of atrophic gastritis-related diseases.

[0022] The present invention has the following beneficial effects:

[0023] (1) The present invention is the first to construct an atrophic gastritis model of long-clawed gerbils by combining Hp infection with BaP induction. The obtained model proves that Hp infection causes physiological damage to long-clawed gerbils and that it has a synergistic damaging effect with BaP. This provides an important reference for further clarifying the mechanism of physiological damage caused by Hp infection and BaP induction, as well as the joint action of the two. It also provides a starting point for assisting in the formulation of eradication treatment plans to prevent the occurrence and development of physiological damage.

[0024] (2) The construction method of the present invention is simple to operate, low in cost, and has stable pathological changes. In addition, the success rate of the constructed gerbil atrophic gastritis model is high, and it has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings required for use in the embodiments are briefly introduced below. It should be understood that the following drawings only show certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other related drawings can be obtained based on these drawings without creative work.

[0026] Figure 1 This is a graph showing the weight trends of gerbils in different treatment groups;

[0027] Figure 2 The tailing of cells in different treatment groups under CASP analysis software, where A is the blank control (NC), B is the Hp group, C is the BaP group, and D is the Hp+BaP group;

[0028] Figure 3 is the expression level of inflammatory cytokines IL-1β, IL-6 and TNF-α in the serum of gerbils in different treatment groups;

[0029] Figure 4 The Warthin-Starry silver staining of the gastric mucosal tissue of long-clawed gerbils in the Hp group and the Hp+BaP group, where the left picture is the staining picture of the Hp group at 200×, and the right picture is the staining picture of the Hp+BaP group at 200×;

[0030] Figure 5 The 40× sections of the gastric antrum of gerbils in different treatment groups, where A is the blank control (NC), B is the Hp group, C is the BaP group, and D is the Hp+BaP group;

[0031] Figure 6 The 200× sections of the gastric antrum of gerbils in different treatment groups, where A is the blank control (NC), B is the Hp group, C is the BaP group, and D is the Hp+BaP group;

[0032] Figure 7 The relative abundance of intestinal flora of gerbils in different treatment groups at the phylum level;

[0033] Figure 8 The relative abundance of intestinal flora of long-clawed gerbils in different treatment groups at the genus level. DETAILED DESCRIPTION

[0034] In order to make the purpose, technical scheme and advantages of the embodiments of the present invention clearer, the technical scheme in the embodiments of the present invention will be described clearly and completely below. If the specific conditions are not specified in the embodiments, they are carried out according to conventional conditions or conditions recommended by the manufacturer. If the manufacturer of the reagents or instruments used is not specified, they are all conventional products that can be purchased commercially.

[0035] The features and performance of the present invention are further described in detail below in conjunction with the embodiments.

[0036] The Helicobacter pylori used in the present invention is an international standard strain of Helicobacter pylori (Sydney strain 1, SS1) purchased from Hangzhou Zhiyuan Institute of Laboratory Medicine; benzopyrene was purchased from Shanghai Yiboyuan Biotechnology Co., Ltd.; SPF-grade healthy male long-clawed gerbils (weight 50±10g) were purchased from the Animal Center of Hangzhou Medical College [production license number SCXK (Zhejiang) 2019-0002, use license number (Zhejiang) 2019-0011].

[0037] Example 1

[0038] This embodiment provides a method for constructing an atrophic gastritis animal model, and the specific operations are as follows:

[0039] S1. 1×10 9 CFU / mL bacterial count (turbidimetric method) Hp suspension was intragastrically perfused into SPF healthy male long-clawed gerbils once a week for 5 consecutive weeks, 0.5 mL / time.

[0040] S2. Dissolve BaP in salad oil at a BaP concentration of 5 mg / mL, and administer 0.5 mL / time by intragastric administration for 5 consecutive weeks, once a week.

[0041] S3. At the 36th week, blood was collected from the eyeballs for DNA damage experiments, serum inflammatory factor detection, and feces were collected for intestinal flora 16SrDNA sequencing analysis; after the 36th week, all long-clawed gerbils were fasted for 12 hours and anesthetized by injection of sodium pentobarbital and then killed. Urease and Warthin-Starry silver staining solutions were used to detect H p colonization, respectively; HE staining was used for diagnosis of gastric mucosal pathology of long-clawed gerbils.

[0042] Comparative Example 1

[0043] In this comparative example, only Hp infection was performed on the long-clawed gerbils. The operation method was as follows: 1×10 9 CFU / mL bacterial count (turbidimetric method) Hp suspension was intragastrically perfused into SPF healthy male long-clawed gerbils once a week for 5 consecutive weeks, 0.5 mL / time.

[0044] Comparative Example 2

[0045] The long-clawed gerbils of this comparative example were only induced with BaP, and the operation method was as follows: BaP was dissolved in salad oil, the BaP concentration was 5 mg / mL, 0.5 mL / time was gavaged, and the gavage was continued for 5 weeks, once a week.

[0046] Experimental example

[0047] The atrophic gastritis gerbil model constructed in Example 1 and Comparative Examples 1-2 and the blank control were subjected to status observation and body weight measurement, DNA damage experiment, serum inflammatory factor detection, 16SrDNA sequencing analysis, Hp colonization detection and gastric mucosal pathology diagnosis. The experimental operation is as follows:

[0048] 1. Observation of the state and weight measurement of gerbils

[0049] (1) The mental state of the gerbils was observed every day before, during, and after drug administration, and the differences in hair, mental state, and hunched back between the gerbils in different groups were photographed and recorded.

[0050] (2) Determination of body weight of gerbils: The body weight of gerbils was measured once before administration and once every 2 weeks after administration, and records were kept.

[0051] 2. DNA damage experiment

[0052] The DNA damage of long-clawed gerbils after administration was determined using the comet electrophoresis cell damage detection kit of Jiangsu Keygene Biotechnology Co., Ltd. and according to its instructions. The analysis was performed using CASP software to measure the comet length, tail length, tail distance, Olive tail distance, and DNA content in the comet tail. According to the proportion of DNA in the comet tail, the damage can be divided into 5 levels: Level 0: <5%, no damage, no tailing of cells; Level I: 5% to 20%, low damage, a small amount of tailing; Level II: 20%-40%, moderate damage, obvious tailing; Level III: 40% to 95%, high damage, severe tailing; Level IV: >95%, severe damage, fragmentation. Based on this, the degree of damage to cells in each experimental group was observed and graded.

[0053] 3. Detection of key related inflammatory factors - ELISA method

[0054] Blood was collected from the eyeballs of each group of long-clawed gerbils, and serum was separated. The inflammatory factors IL-1β, IL-6, and TNF-α were determined using the proteintech kits Mouse IL-1beta, Mouse IL-6, and Mouse TNF-alpha in strict accordance with the instructions.

[0055] 4. Examination of Hp infection and colonization in gastric tissue of gerbils

[0056] ① Rapid urease test: The gastric mucosa was obtained after the long-clawed gerbil was killed. If the indicator turned rose red within 30 minutes in the liquid urease reagent, it was positive; ② Warthin-Starry silver staining: The gastric mucosal tissue was obtained after the long-clawed gerbil was killed and fixed with 10% neutral formalin solution, and the procedure was performed according to the instructions of the Warthin-Starry silver staining solution kit purchased from Beijing Leagene Biotechnology Co. Ltd.

[0057] 5. Pathological analysis of gerbil tissue sections

[0058] The gastric tissue of the gerbil was fixed with 10% neutral formalin solution and then sampled. It was dehydrated by an automatic dehydrator and embedded in paraffin after being transparent and immersed in wax. Finally, it was cut into 3 μm thick sections, stained with HE and sealed with neutral gum to complete the preparation of glass slides for observation under an optical microscope.

[0059] 6. 16S rDNA sequencing analysis of gerbil feces

[0060] Fecal samples of each group of long-clawed gerbils were sent to Hangzhou Lianchuan Biotechnology Co., Ltd. for 16SrDNA sequencing analysis.

[0061] 7. Experimental results

[0062] (1) Observation of the condition and weight measurement of gerbils

[0063] like Figure 1 As shown in the figure, the NC group was healthy gerbils, and their weight increased steadily. The weight of the gerbils in the Hp group increased before 15 weeks, and in the BaP group before 28 weeks, but the increase was smaller than that in the control group, indicating that both Hp and BaP caused physiological damage to the gerbils. After 15 weeks, the weight of the Hp group did not increase, which was related to the fact that Hp colonized in the stomach of the gerbils at this time, causing chronic active gastritis. The weight of the Hp+BaP group increased slightly after 15 weeks, which may be because BaP initially removed some Hp, making the gastritis of Hp+BaP less severe than that caused by a single Hp, or it may be caused by the hormetic effect. The hormetic effect is an adaptive response of the organism, that is, chemical pollutants themselves or in combination with another substance can stimulate the body's active stress response. After 28 weeks, the weight of the Hp+BaP group decreased rapidly, which was related to the increasingly serious damage under the coordinated action of the two.

[0064] (2) DNA damage detection results of blood cells of gerbils

[0065] like Figure 2The cell morphology of the four groups of cells observed under a fluorescence microscope. The cells of NC were round, with clear and neat edges of the nucleus and no tailing. There was a small amount of tailing in the Hp and BaP groups. In the Hp+BaP group, the damaged DNA fragments moved out of the nucleus and showed a tail-shaped distribution, with obvious tailing, resembling a "comet".

[0066] Table 1 Cell damage in each group of gerbils

[0067]

[0068] Table 1 shows that compared with the NC group, the comet tail length, tail length, tail distance, Olive tail distance and comet tail DNA content of the Hp+BaP group were significantly different (P<0.01). Compared with the Hp group or BaP group, the above five indicators of the Hp+BaP group were also significantly different (P<0.01), while only the comet tail length, tail length and comet tail DNA content of the single Hp group and the single BaP group were significantly different from the NC group (P<0.01).

[0069] The degree of cell damage was graded according to the DNA content in the comet tail: the average DNA content in the comet tail of the Hp+BaP group exceeded 40%, which was highly damaged; the average DNA content in the comet tail of the Hp single group and the BaP single group was greater than 5% and less than 20%, which was low damage. HP infection causes cellular oxidative stress, which leads to DNA chain breakage, and then causes oxidative DNA damage. DNA damage plays an important role in the evolution of gastric tissue from gastritis to gastric cancer after HP infection. After benzopyrene enters the body, under the action of the CYP450 enzyme system, it undergoes a series of metabolic activation reactions to generate the ultimate carcinogen BPDF, which directly acts on DNA, causing DNA damage and releasing broken DNA fragments. Because the gerbils were strong and immune before oral administration in this study, the exposure to Hp and BaP was low, so the DNA damage in the single action group was not very serious, which was low damage, while the same exposure, the Hp and BaP synergistic group reached high damage, indicating that Hp and BaP have a synergistic effect at the cellular level.

[0070] (3) Expression levels of inflammatory cytokines IL-1β, IL-6, and TNF-α in gerbil serum

[0071] Cytokines / inflammatory mediators, cytotoxins, Hp enzymes and adhesion factors are involved in the inflammatory response during Hp infection. Cytokines can chemotactic and activate neutrophils, causing nonspecific inflammatory responses in the body and causing damage to the gastric and duodenal mucosa. TNF-α is a cytokine released by macrophages and monocytes after being stimulated by inflammatory factors. It is a pro-inflammatory protein that causes damage to the endothelial cells of the gastric mucosa, reduces blood flow, and thus causes gastric mucosal damage and gastritis. IL-1β is an inflammatory cytokine with a wide range of biological effects. It can stimulate various immune cells and inflammatory cells, promote the release of inflammatory proteins and inflammatory mediators, and has a chemotactic effect on neutrophils, macrophages and lymphocytes, enhancing the inflammatory response. IL-6 is a biologically active polypeptide substance with multiple functions. It can induce local aggregation of inflammatory cells, mainly neutrophils, causing a burst of granulocyte respiration, generating reactive oxygen species, and causing tissue inflammation. Activated neutrophils can secrete cytokines such as IL-1, IL-8, and TNF-α, forming a complex cell network that jointly participates in gastric mucosal inflammatory damage and causes gastritis.

[0072] like Figure 3 As shown, compared with the NC group, the expression level of IL-1β in the serum of the Hp+BaP group was significantly increased, with significant differences (P<0.05); compared with the BaP group, the expression level of IL-1β in the serum of the Hp+BaP group was significantly increased, with significant differences (P<0.05); compared with the NC group and the Hp group, the expression level of IL-6 in the serum of the Hp+BaP group was significantly increased, with extremely significant differences (P<0.01); compared with the NC group, the expression level of TNF-α in the serum of the Hp+BaP group was significantly increased, with significant differences (P<0.05).

[0073] Cytokines / inflammatory mediators, cytotoxins, Hp enzymes and adhesion factors are involved in the inflammatory response during Hp infection. Cytokines can chemotactic and activate neutrophils, causing nonspecific inflammatory responses in the body and causing damage to the gastric and duodenal mucosa. TNF-α is a cytokine released by macrophages and monocytes after being stimulated by inflammatory factors. It is a pro-inflammatory protein that causes damage to the endothelial cells of the gastric mucosa, reduces blood flow, and thus causes gastric mucosal damage and gastritis. IL-1β is an inflammatory cytokine with a wide range of biological effects. It can stimulate various immune cells and inflammatory cells, promote the release of inflammatory proteins and inflammatory mediators, and has a chemotactic effect on neutrophils, macrophages and lymphocytes, enhancing the inflammatory response. IL-6 is a biologically active polypeptide substance with multiple functions. It can induce local aggregation of inflammatory cells, mainly neutrophils, causing a burst of granulocyte respiration, generating reactive oxygen species, and causing tissue inflammation. Activated neutrophils can secrete cytokines such as IL-1, IL-8, and TNF-α, forming a complex cell network that jointly participates in gastric mucosal inflammatory damage and causes gastritis.

[0074] In this experiment, compared with the NC group, the expression levels of inflammatory cytokines TNF-α, IL-1β, and IL-6 in the serum of the Hp+BaP group were significantly increased (P<0.05), which is consistent with the conclusions of the general survival status of long-clawed gerbils and the number of gastritis cases in the subsequent gastric tissue pathological examination.

[0075] (4) Hp colonization

[0076] The Hp group and the Hp+BaP group were Hp colonization groups, with a total of 18 long-clawed gerbils, which were killed 36 weeks after the start of the experiment for Hp infection detection. All long-clawed gerbils were positive in the rapid urease test (development time: 0.5 hours, color: red to purple red), and typical short rod-shaped or seagull-shaped black particles (such as granules) were observed on the surface of the gastric mucosa and in the glandular cavity under a microscope after Warthin-Starry silver staining of gastric tissue sections. Figure 4 Both positive results are the gold standard for determining Hp infection. All long-clawed gerbils in the Hp group and the Hp+BaP group were successfully colonized with Hp.

[0077] The success of establishing an animal model of Hp infection and its application prospects are closely related to both the strain and the animal. Currently, the strains that are widely used and can be successfully colonized include Hp Sydney strain (HpSS1) and HpATCC43504 strain. The common characteristics of the above strains are that both the cytotoxicity-related gene (CagA) and the vacuolating toxin gene (VacA) are positive, and they are highly virulent strains. Therefore, selecting highly virulent strains to infect animals can increase the colonization rate.

[0078] Among many animal models, the long-clawed gerbil has a low probability of naturally developing gastritis, is susceptible to Hp, and can serve as a host for long-term stable colonization of Hp. The pathological changes in the gastric mucosa after infection are similar to those in humans after Hp infection. At the same time, the animal has a longer lifespan than mice, making it suitable for long-term observation; it is cheaper than primates and has a lower cost.

[0079] In this experiment, the international standard strain Hp SS1 was selected to infect long-clawed gerbils. The rapid urease test and Warthin-Starry silver staining test showed that Hp easily colonized in the stomach of long-clawed gerbils, and the colonization rate could reach 100%.

[0080] (5) Pathological examination of gastric tissue sections of different groups of gerbils

[0081] As shown in Table 2, the conditions of atrophic gastritis in the stomach of gerbils in different treatment groups. As can be seen from Table 2, at 36 weeks, the probability of atrophic gastritis in the Hp+BaP group was 100%.

[0082] Table 2 Construction success rate of different treatment groups

[0083]

[0084] Figure 5 These are 40× sections of the gastric antrum of different groups of gerbils. Figure 6 The 200× sections of the gastric antrum of long-clawed gerbils in different groups show no obvious abnormalities in the NC group, with neatly arranged gastric mucosal glands and intact structures, without atrophy or loss. In the Hp group, the epithelial glands of the gastric mucosal layer near the pylorus in the gastric antrum were atrophied and shortened, the number of major cells (chief cells, parietal cells and other cells) decreased, the number of glands in the lamina propria decreased, the interstitium was mildly infiltrated with inflammatory cells, and there was gastric mucosal loss. In the BaP group, the gastric antral glands were atrophied and decreased in number, with mild interstitial edema. In the Hp+BaP group, the gastric antral glands were atrophied and disordered, with interstitial edema and inflammatory cell infiltration in the lamina propria.

[0085] The prior art reports that the gastric mucosa was observed 1 week and 2 weeks after infection with Hp, and the main manifestations of acute injury were blood, erosion, and a large number of neutrophil infiltration. In this experiment, the lesions at each observation point were mainly gastric gland atrophy, decreased number, and chronic atrophic inflammation with inflammatory cell infiltration, and no acute gastritis was found.

[0086] From the perspective of tissue sections, there was no difference in the results between the Hp group and the BaP group, with 1 case of no difference, 7 cases of mild symptoms, and 1 case of moderate symptoms. However, compared with any single factor group, 9 cases of atrophic gastritis developed after the addition of BaP on the basis of Hp infection, and the model success rate was 100%, with 4 cases of mild symptoms and 5 cases of moderate symptoms. Not only did the number of cases increase, but the degree of atrophy also worsened, which was significantly different, as shown in Table 2.

[0087] The above results confirm that Hp infection has the effect of enhancing the action of BaP.

[0088] (6) Effects of Hp, BaP, and Hp+BaP exposure on the relative abundance of phyla and genus levels in the intestinal flora of gerbils

[0089] like Figure 7 As shown in the figure, at the phylum level, the intestinal flora includes Firmicutes, Bacteroidetes, Actinobacteriota, Patescibacteria, Desulfobacteria, Proteobacteria, and Verrucomicrobia. Compared with the NC group, the abundance of Firmicutes and Actinobacteriota in the Hp, BaP, and Hp+BaP groups was significantly lower, while the abundance of Bacteroidetes and Patescibacteria was significantly higher. This indicates that Hp, BaP, and Hp+BaP will change the composition of the intestinal flora of long-clawed gerbils and cause intestinal flora imbalance, which is one of the causes of physiological damage to the model and atrophic gastritis.

[0090] like Figure 8As shown in the figure, at the genus level, the intestinal flora included Lactobacillus, Candidatus_Saccharimonas, Muribaculaceae_unclassified, Allobaculum, Ligilactobacillus, HT002, Desulfovibrio, Limosilactobacillus, Monoglobus, Bifidobacterium, etc. Compared with the NC group, the relative abundance of Lactobacillus and Limosilactobacillus in the Hp, BaP, and Hp+BaP groups was significantly reduced (P<0.05). Both Lactobacillus and ligilactobacillus are probiotics and belong to lactic acid bacteria. They have similar functions, which can promote the digestion and absorption of nutrients, improve gastrointestinal function, enhance the body's immunity, and have anti-cancer and anti-tumor activities. The relative abundance of Allobaculum, Monoglobus, Candidatus_Saccharimonas, and Muribaculaceae_unclassified increased. In summary, exposure to Hp, BaP, and Hp+BaP caused changes in the species composition of the intestinal flora of long-clawed gerbils, and the abundance of probiotics among dominant bacteria was significantly reduced, while the abundance of harmful bacteria was increased.

[0091] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A method for constructing an atrophic gastritis animal model, characterized in that: The experimental animals were treated with Helicobacter pylori and benzopyrene to obtain an animal model of atrophic gastritis.

2. The construction method according to claim 1, characterized in that: The construction method comprises: firstly, infecting experimental animals with Helicobacter pylori by gavage, then gavage benzopyrene to the experimental animals, and then feeding the treated experimental animals normally.

3. The construction method according to claim 2, characterized in that: The experimental animals were intragastrically administered with Helicobacter pylori once a week for 5 consecutive weeks.

4. The construction method according to claim 3, characterized in that: The method of gavage of Helicobacter pylori to experimental animals each time is: using a concentration of 1×10 9 CFU / mL of Helicobacter pylori suspension, 0.5 mL / time gavage.

5. The construction method according to claim 4, characterized in that: The experimental animals are intragastrically administered with benzopyrene once a week for 4-5 consecutive weeks.

6. The construction method according to claim 5, characterized in that: The method of gavage of the experimental animals with benzopyrene each time is: using a benzopyrene solution with a concentration of 5 mg / mL, 0.5 mL / time gavage.

7. The construction method according to claim 6, characterized in that: The normal feeding period is from the 11th week to the 36th week.

8. The construction method according to claim 7, characterized in that: The experimental animals include gerbils.

9. An atrophic gastritis animal model obtained by the method for constructing an atrophic gastritis animal model according to any one of claims 1 to 8.

10. Use of the atrophic gastritis animal model according to claim 9 in the study of related diseases, characterized in that: The research includes studies on the mechanisms, drug screening and treatment methods of atrophic gastritis-related diseases.

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