Bacterial infection and LPS (Lipopolysaccharide) induced broiler liver injury model as well as construction method and application thereof

Abstract

The invention discloses a bacterial infection and LPS induced broiler chicken liver injury model and a construction method and application thereof, and the construction method comprises the following steps: administering 10-day-old broiler chicken with Escherichia coli E.coli O78 with a preset concentration in an intragastric administration manner; after broiler chickens are subjected to intragastric administration of escherichia coli for 18 hours, an LPS solution is injected into the intraperitoneal cavity of the broiler chickens, after the LPS solution is injected for 24 hours, the broiler chicken liver injury model is obtained, when the broiler chickens are 11 days old, the broiler chickens subjected to intragastric administration of escherichia coli are subjected to intraperitoneal injection of the LPS solution, and after the LPS solution is injected for 24 hours, the broiler chicken liver injury model is obtained. The broiler chicken liver injury model is constructed by intragastric administration of avian pathogenic escherichia coli (E.coli O78) and intraperitoneal injection of LPS (2 mg / kg), the pathology of the model is highly consistent with that of clinical broiler chicken bacterial liver injury, the liver has clinical typical lesions macroscopically, liver cells are degenerative and necrotic microscopically, and a molecular pathway activation mode is consistent with that of clinical application; the model constructed by the invention realizes specific liver injury, only the liver has specific injury, multiple organ lesions such as heart, peritoneum and the like do not exist, and the model availability is high.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of animal injury model construction, and particularly relates to a bacterial infection and LPS-induced broiler liver injury model and a construction method and application thereof. BACKGROUND

[0002] In recent years, with the rapid development of the breeding industry, more and more liver damage of livestock and poultry caused by incorrect use of drugs, feed mold, bacteria, viruses or parasites and environmental factors. Animal liver damage caused material metabolism disorder, biliary formation and excretion disorder, detoxification ability decreased and a series of pathological changes and corresponding syndromes, greatly affected the production performance of animals, and brought serious harm to the livestock and poultry breeding industry. Every year, a large number of poultry are sick and die due to bacterial infection. Through comprehensive observation of the pathological changes, it is found that many common bacterial diseases such as chicken white diarrhea, chicken typhoid, avian cholera and colibacillosis can cause liver damage in chickens to varying degrees. A suitable liver injury animal model is the basis for studying liver injury pathology, evaluating the effect and mechanism of liver-protecting drugs. At present, there are many liver injury animal models, and the commonly used single induction methods include lipopolysaccharide (LPS), CCl4, acetaminophen, enrofloxacin ethanol, etc.; the commonly used compound induction methods include LPS combined with cefalonium sodium or D-galactosamine (GalN), ethanol, CCl4 combined with concanavalin A, etc., but there is no report on the LPS combined with bacterial infection to induce the liver injury model of poultry.

[0003] The liver injury of livestock and poultry mainly includes drug-induced liver injury and immune-induced liver injury. The immune-induced liver injury refers to a type of liver injury mediated by immune response. Scholars such as Han Chao et al. established a stable immune liver injury animal model by intraperitoneal injection of LPS. LPS is mainly derived from gram-negative bacteria in the intestinal tract. Under normal circumstances, only a small amount of LPS is produced in the intestinal tract, and enters the liver through the portal vein, and is then rapidly and regularly absorbed by various scavenger cells in the liver. Generally, a small amount of LPS in the intestinal tract does not stimulate the liver to induce immune response, which is of great significance to avoid immune damage of the liver due to excessive inflammatory response. However, if the intestinal flora is unbalanced, a large number of pathogenic bacteria will be produced, and the level of LPS will be increased. As the first line of defense between intestinal bacteria and other parts of the body, the liver inevitably becomes the first target organ of LPS attack, causing liver damage. Studies have shown that the occurrence of many liver diseases is accompanied by an increase in the level of LPS, and the LPS produced during the development of the disease also causes secondary damage to the liver. At present, the most common poultry liver injury model is to induce liver injury in chickens by LPS alone, but LPS alone cannot fully reflect the bacterial liver injury of poultry.

[0004] Therefore, the prior art remains to be improved and developed. SUMMARY

[0005] In view of the deficiencies of the prior art described above, the purpose of the present application is to provide a bacterial infection and LPS-induced liver injury model of broilers, as well as a construction method and application thereof, aiming to solve the problem that the existing single cause manufacturing method of liver injury model cannot reproduce the clinical real process of the pathological basis laid by bacterial infection and the damage amplified by LPS.

[0006] The technical solution of the present application is as follows: A construction method of a bacterial infection and LPS-induced liver injury model of broilers, comprising the following steps: 10-day-old broilers are given E. coli O78 of a preset concentration by gavage; After the broilers are given E. coli by gavage for 18h, the broilers are intraperitoneally injected with LPS solution, and a liver injury model of broilers is obtained 24h after the injection of LPS solution; at 11 days old, the broilers given E. coli by gavage are intraperitoneally injected with LPS solution, and a liver injury model of broilers is obtained 24h after the injection of LPS solution.

[0007] The construction method of the bacterial infection and LPS-induced liver injury model of broilers, wherein the preset concentration of E. coli O78 is 10 6 -10 11 CFU / mL.

[0008] The construction method of the bacterial infection and LPS-induced liver injury model of broilers, wherein the LPS solution is composed of physiological saline and LPS dissolved in the physiological saline, and the concentration of the LPS solution is 2mg / kg.

[0009] The construction method of the bacterial infection and LPS-induced liver injury model of broilers, wherein the broilers are yellow-feathered broilers.

[0010] A bacterial infection and LPS-induced liver injury model of broilers is prepared by using the construction method of the bacterial infection and LPS-induced liver injury model of broilers.

[0011] The application of a bacterial infection and LPS-induced liver injury model of broilers, wherein the bacterial infection and LPS-induced liver injury model of broilers is used for drug screening, and the drug is used for treating liver injury of broilers.

[0012] Beneficial effects: This invention constructs a broiler liver injury model by gavage administration of pathogenic avian Escherichia coli (E. coli O78) and intraperitoneal injection of LPS (2 mg / kg). The pathology of the model is highly consistent with clinical bacterial liver injury in broilers. The liver macroscopically shows typical clinical lesions, and microscopically, hepatocytes show degeneration and necrosis. The molecular pathway activation pattern is consistent with clinical findings. The model constructed by this invention achieves specific liver injury, with only the liver showing specific damage and no lesions in multiple organs such as the heart and peritoneum. The model has high usability. Attached Figure Description

[0013] Figure 1 This is a flowchart of a method for constructing a bacterial infection and LPS-induced liver injury model in broilers according to the present invention.

[0014] Figure 2 The images show the clinical symptoms and feces of chicks in the normal control group in Example 1.

[0015] Figure 3 The images show the clinical symptoms and feces of chicks in the gavage group in Example 1.

[0016] Figure 4 The image shows the results of the observation of chicken liver tissue in the normal control group, the gavage group, and the intraperitoneal injection group in Example 1.

[0017] Figure 5 The image shows the results of HE staining microscopic observation of chicken liver tissue in the normal control group and the gavage group in Example 1.

[0018] Figure 6 The image shows the results of the epigenetic observation of chicken liver tissue in the normal control group, gavage group, single gavage group, and single LPS group in Example 2.

[0019] Figure 7 The image shows the results of HE staining microscopic observation of chicken liver tissue from the normal control group, gavage group, single gavage group, and single LPS group in Example 2. Detailed Implementation

[0020] This invention provides a method for constructing a bacterial infection and LPS-induced liver injury model in broilers. To make the objectives, technical solutions, and effects of this invention clearer and more explicit, the invention is further described in detail below. It should be understood that the specific embodiments described herein are only for explaining the invention and are not intended to limit the invention.

[0021] Current broiler liver injury models are mainly divided into three categories: chemically induced (such as carbon tetrachloride and florfenicol), single infection (such as simple E. coli infection), and single endotoxin (such as direct injection of LPS). All of them have significant defects, as shown in Table 1.

[0022] Table 1 Model Classification

[0023] As shown in Table 1, while the chemically induced model exhibits good reproducibility, it cannot simulate the most common pathological mechanisms of infectious liver injury in clinical practice, particularly lacking key features such as gut microbiota dysbiosis and immune cell infiltration. The single infection model suffers from uneven liver damage due to significant individual variability in bacterial colonization; the serum ALT coefficient of variation can exceed 45% within the same batch of models. The single LPS model, by directly triggering a systemic inflammatory response, often results in a mortality rate exceeding 50% in broilers, and its pathological characteristics do not match the progressive liver damage seen in clinical bacterial liver disease.

[0024] Clearly, traditional models generally sever the pathological link between the gut and the liver, while clinical studies have confirmed that bacterial liver disease in broilers is essentially a result of gut-liver axis dysfunction. As the primary colonization site for Gram-negative bacteria, the integrity of the gut's barrier function directly determines the concentration of endotoxins entering the bloodstream, thus affecting the degree of liver damage. While existing single-infection models can induce intestinal infection, they fail to assess the degree of barrier damage and therefore cannot establish a quantitative relationship between intestinal lesions and liver injury. The single LPS model completely skips the intestinal stage, directly inducing liver inflammation, resulting in a severe disconnect from the clinical pathological progression.

[0025] Based on this, the present invention provides a method for constructing a bacterial infection and LPS-induced liver injury model in broilers, such as... Figure 1 As shown, it includes the following steps: S10. Ten-day-old broilers were given a predetermined concentration of Escherichia coli O78 via gavage. S20. After 18 hours of gavage with Escherichia coli, the broilers were intraperitoneally injected with LPS solution. A broiler liver injury model was obtained 24 hours after the LPS solution injection. At 11 days of age, the broilers that had been gavaged with Escherichia coli were intraperitoneally injected with LPS solution. A broiler liver injury model was obtained 24 hours after the LPS solution injection.

[0026] Specifically, existing technologies mostly rely on single-cause modeling (such as simple LPS injection or single bacterial infection), which can only simulate a certain pathological stage of liver injury and cannot reproduce the complete pathological process of bacterial liver disease in broilers: intestinal infection → barrier disruption → endotoxin translocation → liver inflammation → tissue necrosis. The bacterial infection + LPS secondary stimulation dual-factor modeling strategy proposed in this invention is not a simple factor superposition, but rather a creatively designed three-level pathological chain based on the principle of pathological progression: Primary pathological initiation (intestinal colonization period, 10 days old): Pathogenic Escherichia coli (E. coli O78) was administered to broilers via gavage. Its fimbrial adhesin (FimA) and outer membrane protein (OmpA) specifically bind to the intestinal epithelial cells of broilers, triggering an innate immune response in the intestinal mucosa. At this time, the number of intestinal goblet cells increased by 23.6% ± 3.2%, and mucus secretion was increased, laying the infection foundation for subsequent damage and addressing the deficiency of traditional models in lacking a source of pathogen invasion. Secondary pathological amplification (barrier disruption period, 11 days old): 18 hours after oral administration of E. coli, the hemolysin (HlyA) secreted by E. coli O78 disrupted the tight junctions of intestinal epithelial cells, increasing intestinal permeability. The expression levels of tight junction proteins (ZO-1 and Occludin) decreased by 38.7%±3.8% and 41.2%±4.3%, respectively. At this time, intraperitoneal injection of LPS (2 mg / kg) could rapidly enter the bloodstream through the damaged barrier, avoiding the problem that a high dose (≥5 mg / kg) of LPS alone was required to break through the intact intestinal barrier, thus reducing the model mortality rate (from 52.3% in the single LPS model to 12.7%). Level III pathological diagnosis (liver injury stage, 12 days old): Entered blood LPS activates hepatic Kupffer cells through the TLR4-MyD88-NF-κB pathway, prompting them to release pro-inflammatory factors such as TNF-α (content reached 286.5pg / mL±25.3pg / mL, 2.1 times that of the single infection group) and IL-6 (318.7pg / mL±30.2pg / mL, 1.8 times that of the single LPS group), while inhibiting hepatocyte antioxidant enzymes, forming a synergistic effect of inflammatory storm and oxidative damage, which increases the rate of hepatocyte necrosis. The pathological characteristics of liver injury caused by E. coli infection in broilers are consistent with those of clinical E. coli infection in 92.6%, far exceeding the existing models.

[0027] Existing models generally neglect the correlation between strain pathogenicity, broiler age, and stimulus dose, resulting in poor model reproducibility (between-batch coefficient of variation > 35%). This invention, however, establishes a quantitative matching relationship among these three factors through extensive preliminary experiments: Creative strain selection: Traditional laboratory-preserved non-pathogenic Escherichia coli (such as DH5α) were discarded; clinically isolated E. coli O78 (serotype matching rate with prevalent strains in Chinese broiler farms reached 89.7%) was selected. The virulence genes (tsh, vat, iucD) carried by this strain were detected in 100% of cases, and its adhesion rate to broiler intestinal epithelial cells was 4.2 × 10⁻⁶. 5 The CFU / cell ratio is 3.7 times that of laboratory strains, ensuring infection efficiency; Precise age selection: Breaking away from the traditional practice of arbitrarily selecting chickens aged 7-14 days, we have determined that 10-day-old yellow-feathered broilers are the best model subjects. At this stage, the liver metabolic function of broilers has been initially improved, but the intestinal barrier has not yet been fully developed (the expression level of tight junction protein is only 58.2% of that of adult chickens). They are susceptible to bacterial invasion and can clearly show the pathological changes of liver damage. This avoids the problems of excessive liver damage in 7-day-old chicks (mortality rate > 25%) or insignificant damage in 14-day-old broilers (hepatocyte necrosis rate < 15%). Dosage gradient design: 10 dose gradient designs for E. coli O78 6 -10 11 Six concentration gradients of CFUs, combined with a fixed dose of 2 mg / kg LPS, form a low concentration (10 6 -10 8 CFUs) Mild damage (liver tissue score 1.70-2.33), moderate concentration (10 9 -10 10 CFUs) Moderate damage (2.87-3.13 points), high concentration (10) 11 A gradient model for severe CFU (4.07 points) injury can meet different research needs (such as research on mild inflammation mechanisms and drug screening for severe injury), while existing models can only achieve a single degree of injury (mostly moderate), limiting their application scenarios.

[0028] Furthermore, this invention does not inject LPS simultaneously with bacterial infection, but rather at an interval of 18 hours (bacteria are administered via gavage at 10 days of age, and LPS is injected at 11 days of age). This interval precisely corresponds to the pathological timeline of bacterial infection → intestinal barrier disruption → endotoxin entry into the bloodstream in clinical practice (clinical monitoring shows that the incidence of endotoxemia in broilers infected with E. coli reaches 78.3% within 24-18-36 hours). This makes the pathological process of the model closer to the real disease, while a single administration method (such as administering bacteria and LPS simultaneously) will lead to disordered pathological processes (endotoxemia occurs as early as 8 hours, which is inconsistent with clinical practice).

[0029] The present invention will be further explained and illustrated below through specific embodiments: Example 1 Yellow-feathered broiler chickens were placed in a temperature-controlled room (25℃) and subjected to a 12-hour light-dark cycle (light from 7:00 to 19:00). The chickens had free access to standard experimental feed and drinking water. Ten-day-old yellow-feathered broiler chickens were randomly divided into 13 groups: a normal control group, a gavage group (10... 6 CFUs, 10 7 CFUs, 10 8 CFUs, 10 9 CFUs, 10 10 CFUs, 10 11CFUs E. coli O78) + intraperitoneal injection of LPS (2 mg / kg), peritoneal injection group (10 6 CFUs, 10 7 CFUs, 10 8 CFUs, 10 9 CFUs, 10 10 CFUs, 10 11 CFUsE.coli O78) + intraperitoneal injection of LPS (2 mg / kg), 12 animals per group. In this example, the gavage groups were designated as: PO.6 + LPS, PO.7 + LPS, PO.8 + LPS, PO.9 + LPS, PO.10 + LPS, PO.11 + LPS; the intraperitoneal injection groups were designated as: IP.6 + LPS, IP.7 + LPS, IP.8 + LPS, IP.9 + LPS, IP.10 + LPS, IP.11 + LPS. Chickens in the gavage group were administered a series of concentrations of *E. coli* via gavage, while chickens in the intraperitoneal injection group were administered a series of concentrations of *E. coli* via intraperitoneal injection. Each chicken received 1 mL of the administered medication, while the normal control group received an equal volume of sterile saline. After 18-18 hours, chickens in both the gavage and intraperitoneal injection groups were intraperitoneally injected with 2 mg / kg LPS (LPS dissolved in saline). 24 hours after LPS injection, blood was collected from the jugular vein of the chickens, and the right liver lobe was fixed in 4% paraformaldehyde solution for histopathological observation. Fecal samples and clinical symptoms of the chicks were recorded during the experiment.

[0030] Figure 2 Images show the clinical symptoms and feces of chicks in the normal control group. Figure 3 Images of clinical symptoms and feces in chicks in the gavage group were collected. Figure 2 It can be seen that the chicks in the normal control group had no clinical symptoms, their feces were conical and light gray in color, and white urate was attached to the surface of the feces; while the chicks in the gavage group preferred to lie down and had no other obvious abnormal clinical symptoms, but some chicks had watery feces.

[0031] Furthermore, the epigenetic observation of chicken liver tissue in the normal control group, gavage group, and intraperitoneal injection group was performed, and the results are as follows: Figure 4 As shown in the figure. Compared with the normal control group, no significant changes were observed in the liver of PO.6-PO.9 in the gavage group after necropsy, and the liver of PO.10-PO.11 in the gavage group was whitish in color after necropsy. No changes were observed in the heart and peritoneum in any of the gavage groups. However, in the intraperitoneal injection group, fibrinous adhesions of the heart and liver, a brownish-green color of the liver, and peritonitis were observed after necropsy, indicating multi-organ damage. This suggests that the intraperitoneal injection group is not suitable for use as a poultry liver injury model.

[0032] Furthermore, HE staining and microscopic observation were performed on chicken liver tissue from the normal control group and the gavage group. The results are as follows: Figure 5 As shown. From Figure 5 It can be seen that the hepatocytes of the chicks in the normal control group were in a normal physiological state, with neatly arranged cell cords and clear structure, and no pathological changes such as inflammatory cell infiltration, cell necrosis, or degeneration were observed. However, in the gavage group (E. coli + LPS), the chicks showed hepatocyte vacuolar degeneration, hepatocyte necrosis, blood vessels filled with red blood cells, and inflammatory cell infiltration in the portal area.

[0033] Quantitative analysis of liver histological damage was performed, and the results are shown in Table 2.

[0034] Table 2 Liver Tissue Injury Score

[0035] Note: Data in the table are mean ± standard deviation; * indicates statistical difference compared with the control group (p < 0.05).

[0036] Combined with Table 2 and Figure 5 The data show that the liver tissue damage score in the control group (without E. coli + without LPS) was 0.77±0.25, and the hepatocytes were in a normal physiological state. Figure 5 The control group showed no pathological changes such as inflammatory cell infiltration and cell necrosis, proving that the liver of broilers was not damaged under normal feeding conditions, and providing a blank control benchmark for the model.

[0037] Low concentration group (PO.6-PO.8+LPS, 10) 6 -10 8 The CFUs / E. coli scores were 1.70±0.26, 1.83±0.29, and 2.33±0.58, respectively. Although these scores were higher than those in the control group, there was no statistically significant difference (p>0.05). The corresponding liver tissue showed only mild hepatocellular edema. Figure 5 The low-concentration group showed no significant necrosis, indicating that low-concentration E. coli combined with LPS could only induce mild, non-significant liver injury, which could not meet the needs of moderate to severe injury studies.

[0038] Medium and high concentration group (PO.9-PO.11+LPS, 10) 9 -10 11 The scores for CFUs / E. coli were significantly elevated, at 2.87±0.23, 3.13±0.23, and 4.07±0.12 (all marked with *, p<0.05), and continued to rise with increasing E. coli concentration: PO.9 group showed punctate hepatocyte necrosis, PO.10 group showed an increase in necrotic foci, and PO.11 group showed extensive inflammatory cell infiltration and hepatocyte fusion necrosis in the portal area.Figure 5 (Medium and high concentration groups) demonstrate that the higher the E. coli concentration, the more severe the liver tissue damage, achieving a controllable gradient of mild-moderate-severe damage and solving the problem of the single damage degree in existing models.

[0039] This embodiment also tested five core liver function indicators in the serum of chicks in the normal control group and the gavage group: AST (aspartate aminotransferase), ALT (alanine aminotransferase), ALP (alkaline phosphatase), T-Bil-V (total bilirubin), and D-Bil-V (direct bilirubin). The impact of the model on liver function was quantified from the dimensions of hepatocyte damage and bile excretion function. The results are shown in Table 3.

[0040] Table 3 Results of serum liver function enzyme activity tests in chicks

[0041] As shown in Table 3, the high concentration groups (PO.10 and PO.11) of hepatocyte damage markers (AST and ALT) exhibited significantly abnormal increases, reflecting hepatocyte necrosis. Specifically, for AST (mitochondrial damage marker), the control group had a value of 296.61±12.81 U / L, while the low concentration groups (PO.6-PO.8 + LPS) showed a slight increase (328.81-338.00 U / L) but without statistical significance. The medium-to-high concentration groups (PO.10-PO.11 + LPS) significantly increased to 357.76±20.68 and 361.53±32.88 U / L (p<0.05), representing an increase of 20.6%-21.9% compared to the control group. AST is mainly found in hepatocyte mitochondria, and its elevation demonstrates that E.coli+LPS has damaged the mitochondrial structure of hepatocytes, which completely corresponds to the pathological characteristics of hepatocyte necrosis (mitochondrial rupture) in the medium-to-high concentration groups in Table 2. Regarding ALT (a marker of cytoplasmic damage), the control group had a value of 3.54 ± 0.72 U / L, while the PO.10-PO.11 + LPS group showed a significant increase to 4.09 ± 0.42 and 4.10 ± 0.41 U / L (p < 0.05). Although the absolute increase in ALT was small, the ALT content in broiler livers is only 1 / 10 to 1 / 5 that of mammals, and this change is sufficient to reflect hepatocyte cytoplasmic damage. Figure 5 The changes in hepatocytes (vacuole degeneration and cytoplasmic structure destruction) were consistent with the changes in AST, jointly demonstrating that high concentrations of E. coli + LPS can lead to hepatocyte necrosis and liver function impairment.

[0042] Regarding ALP (a marker of bile duct function), the data from each group fluctuated between 6006.76 and 7378.39 U / L, with no significant difference (p > 0.05). ALP is mainly secreted by hepatobiliary epithelial cells, and its stability indicates that the damage in the model was concentrated in hepatocytes and did not disrupt the structure and function of the bile ducts. This is consistent with the absence of bile duct dilation and cholestasis observed during necropsy, proving that the model represents hepatocyte-specific damage and avoiding interference from bile duct injury in the evaluation of liver function.

[0043] For T-Bil-V (total bilirubin) and D-Bil-V (direct bilirubin) levels, the control group showed values ​​of 34.49±8.21 μmol / L and 1.26±0.30 μmol / L, respectively, with no significant differences among the groups (p>0.05). Bilirubin requires uptake, conjugation, and excretion by hepatocytes. The stability of these levels indicates that although hepatocytes showed necrosis, the remaining hepatocytes could still maintain basic bilirubin metabolism, further demonstrating that the model's damage level was controllable (not reaching the level of liver failure), making it suitable for screening hepatoprotective drugs (the drugs can exert their effects by repairing the remaining hepatocytes).

[0044] Example 2 Yellow-feathered broiler chickens were placed in a temperature-controlled room (25℃) and subjected to a 12-hour light-dark cycle (light from 7:00 to 19:00). The chickens had free access to standard experimental feed and drinking water. Ten-day-old yellow-feathered broiler chickens were randomly divided into four groups: a normal control group, a gavage group (10... 11 CFUs E. coli O78) + intraperitoneal injection of LPS (2 mg / kg), single gavage group (10 11 CFUsE.coli O78), single LPS group (intraperitoneal injection of LPS 2 mg / kg), 12 animals in each group. Chickens in the gavage group were administered LPS 2 mg / kg via gavage. 11 After 24-18 hours, broilers in the gavage group were intraperitoneally injected with 2 mg / kg LPS (LPS dissolved in physiological saline); the single gavage group was administered 10 mg / kg LPS via gavage at 10 days of age. 11 CFU concentration of E. coli; the LPS-only group received intraperitoneal injection of 2 mg / kg LPS directly at 11 days of age. 24 h later, blood was collected from the jugular vein of the broilers, and the right liver lobe was fixed with 4% paraformaldehyde solution for histopathological observation.

[0045] Similarly, the epigenetic observation of chicken liver tissue was performed in the normal control group, the gavage group (E. coli + LPS group), the single gavage group (E. coli group), and the single LPS group (LPS group). The results are as follows: Figure 6 As shown, from Figure 6It can be seen that no obvious abnormalities were observed in the livers of broilers in the normal group, the single gavage group, and the single LPS group after necropsy, while the livers of chicks in the gavage group were slightly brownish-green after necropsy.

[0046] Similarly, HE staining and microscopic observation were performed on chicken liver tissue from the normal control group, gavage group, single gavage group, and single LPS group. The results are as follows: Figure 7 As shown, from Figure 7 It can be seen that the hepatocytes of the normal control group chicks were in a normal physiological state, with neatly arranged cell cords and clear structure, and no pathological changes such as inflammatory cell infiltration, cell necrosis, or degeneration were observed. Their liver tissue damage score was 0.80±0.20. In the LPS-only group and the gavage-only group, the liver tissue of chicks was filled with erythrocytes and inflammatory cells, and their liver tissue damage scores were 2.23±0.25* and 2.57±0.40*, respectively, both significantly higher than the normal group. The gavage-only group showed inflammatory cell infiltration in the portal area of ​​the liver tissue, and its liver tissue damage score was 3.93±0.15 (higher than the LPS-only group or the E. coli group).

[0047] In this embodiment, five core liver function indicators—AST (aspartate aminotransferase), ALT (alanine aminotransferase), ALP (alkaline phosphatase), T-Bil-V (total bilirubin), and D-Bil-V (direct bilirubin)—were tested in the serum of chicks in the normal control group, the single gavage group, the single LPS group, and the gavage group. The results are shown in Table 4.

[0048] Table 4 Results of serum liver function enzyme activity tests in chicks

[0049] Compared with the normal control group, the AST levels in the single gavage group, the single LPS group, and the gavage group were significantly increased (p < 0.05). The AST level in the gavage group was further increased compared with the single gavage group and the single LPS group (p < 0.05). The changes in ALT level were consistent with those in AST, indicating that the gavage group had a more significant effect on liver injury than the single regimen.

[0050] It should be understood that the application of the present invention is not limited to the examples above. Those skilled in the art can make improvements or modifications based on the above description, and all such improvements and modifications should fall within the protection scope of the appended claims.

Claims

1. A method for constructing a bacterial infection and LPS-induced liver injury model in broilers, characterized in that, Including the following steps: Ten-day-old broilers were administered a pre-set concentration of Escherichia coli O78 via gavage. After 11 days and 18 hours of gavage with E. coli, the broilers that had been gavaged with E. coli were intraperitoneally injected with LPS solution. A broiler liver injury model was obtained 24 hours after the LPS solution injection.

2. The method for constructing a bacterial infection and LPS-induced liver injury model in broilers according to claim 1, characterized in that, The preset concentration of Escherichia coli O78 is 10. 6 -10 11 CFU / mL.

3. The method for constructing a bacterial infection and LPS-induced liver injury model in broilers according to claim 1, characterized in that, The LPS solution consists of physiological saline and LPS dissolved in physiological saline, and the concentration of the LPS solution is 2 mg / kg.

4. The method for constructing a bacterial infection and LPS-induced liver injury model in broilers according to claim 1, characterized in that, The broiler chickens mentioned are yellow-feathered broiler chickens.

5. A bacterial infection and LPS-induced liver injury model in broilers, prepared using the method described in any one of claims 1-4.

6. An application of a bacterial infection and LPS-induced liver injury model in broilers, characterized in that, The bacterial infection and LPS-induced liver injury model in broilers described in claim 5 is used for drug screening, and the drug is used to treat liver injury in broilers.

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