Application of Allo-aca in preparation of medicine for improving liver injury and concurrent neurological symptoms

Abstract

The invention provides application of Allo-aca in preparation of a medicine for improving liver injury and concurrent neurological symptoms, and belongs to the technical field of medicines. The Allo-aca is a leptin receptor antagonist, and the core action mechanism of the Allo-aca is that the Allo-aca is competitively combined with a leptin receptor to block a leptin signal transduction pathway, so that the leptin-dependent pathological process is intervened. Experimental verification shows that the Allo-aca can significantly reduce the levels of glutamic-pyruvic transaminase and glutamic oxalacetic transaminase in mouse serum induced by chronic stress + AFB1, relieve liver cell edema balloon-like degeneration and inflammatory cell infiltration, and realize liver protection; meanwhile, the incubation period of a mouse dark avoidance experiment can be prolonged, the success frequency of a shuttle experiment is increased, the center staying time of an open field and the staying time of an open arm of an elevated cross maze are prolonged, and cognitive impairment and anxiety mood are effectively improved. The invention provides a new treatment scheme for the chronic stress induced AFB1 hepatotoxicity aggravation effect.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of medicine, and particularly relates to application of Allo-aca in preparation of a medicine for improving liver injury and concurrent neurological symptoms. BACKGROUND

[0002] Currently, humans are continuously exposed to a variety of harmful factors in daily environments and professional scenarios. Among them, biological toxins such as Aflatoxin B1 (AFB1) widely polluting the environment have become an important threat to global public health. AFB1, as a highly toxic mycotoxin, is mainly produced by Aspergillus flavus and parasitic aspergillus, and often occurs in various foods such as moldy grains, nuts, etc. The hypothalamus is the core regulatory center of the brain, and its functions include neuroendocrine, metabolic homeostasis, autonomic nervous and behavior regulation, and other multidimensional physiological processes.

[0003] Aflatoxin (AFB1) exposure is a chemical threat commonly found in the environment, which can trigger adaptive defense responses in the liver. Organisms respond to exogenous toxin challenges through metabolic regulation to maintain liver homeostasis. However, persistent physiological disorders caused by chronic stress can reshape the stress response pattern of the liver. The long-term activation of the stress system causes the body to be in a state of continuous metabolic alert, which can induce profound histopathological changes. Because the liver bears the core detoxification responsibility and has high metabolic activity, it becomes a vulnerable target for stress and toxin combined invasion. Chronic stress can cause hypothalamic structural degeneration and functional impairment, further causing cognitive impairment and exacerbating AFB1-induced liver toxicity damage. Stress-induced liver failure is a disastrous outcome of the interaction between toxin exposure and chronic stress.

[0004] The high intensity and fast-paced nature of modern life and professional environment causes a significant proportion of the population to be in a suboptimal state for a long time, even showing chronic stress-like pathological characteristics. This state significantly damages the individual's quality of life and work efficiency, and can form a vicious cycle of self-reinforcement, ultimately posing a serious threat to physical and mental health. SUMMARY

[0005] Therefore, the purpose of the present application is to provide the application of Allo-aca in the preparation of a medicine for improving liver injury and concurrent neurological symptoms, and to provide a new intervention idea for improving the exacerbation effect of chronic stress-induced aflatoxin hepatotoxicity.

[0006] In order to achieve the above-mentioned purpose of the application, the present application provides the following technical solutions: The present application provides the application of Allo-aca in the preparation of a medicine for improving liver injury and concurrent neurological symptoms.

[0007] Preferably, the liver injury and concurrent neurological symptoms are caused by aflatoxin.

[0008] More preferably, the liver injury and concurrent neurological symptoms are chronic stress-induced exacerbation of aflatoxin hepatotoxicity.

[0009] Preferably, the Allo-aca competitively binds to the leptin receptor, blocks the leptin signaling pathway, and thus interferes with the leptin-dependent pathological process.

[0010] Preferably, the Allo-aca has a liver protection effect.

[0011] Preferably, the Allo-aca has an effect of relieving hepatocyte edema ballooning and inflammatory cell infiltration.

[0012] Preferably, the Allo-aca has an effect of improving cognitive impairment and anxiety.

[0013] Preferably, the drug comprises an oral agent or an injection agent.

[0014] More preferably, the drug further comprises a pharmaceutically acceptable carrier and / or excipient.

[0015] Compared with the prior art, the present application has the following beneficial effects: The present application first applies the leptin receptor antagonist Allo-aca to the intervention of chronic stress-induced exacerbation of AFB1 hepatotoxicity related diseases, blocks the leptin-dependent pathological process, and thus interferes with the synergistic damage effect of chronic stress and AFB1 from the source, providing a new action target and intervention idea for such complex interactive diseases.

[0016] Based on the mouse model experiment induced by chronic stress + AFB1, Allo-aca has multiple and clear protective effects on liver injury caused by the specific cause: on the one hand, Allo-aca can reverse hepatocyte damage from the biochemical level by significantly reducing serum ALT and AST levels, and restore liver metabolism and detoxification function; on the other hand, Allo-aca can effectively relieve hepatocyte edema ballooning and reduce inflammatory cell infiltration, thereby reducing liver inflammation from the histopathological level; at the same time, Allo-aca can also improve the body weight change and liver / body ratio of mice induced by chronic stress + AFB1, and restore the regulatory effect of liver on body metabolism from the overall physiological state level. In terms of cognitive function, by prolonging the latency period of the dark avoidance experiment and reducing the number of errors, the learning and memory ability of mice is improved; by increasing the number of successful shuttle experiments, the function of establishing and executing conditioned reflex of mice is improved, and cognitive impairment is effectively reversed; in terms of emotional regulation, by prolonging the central stay time in the open field experiment and increasing the open arm stay time in the elevated plus maze, the anxiety of mice is relieved, and the normal behavior pattern is restored. BRIEF DESCRIPTION OF DRAWINGS

[0017] Figure 1 Body weight changes of different groups of mice; Figure 2 Liver / body ratios of different groups of mice; Figure 3 ALT and AST levels of different groups of mice; Figure 4 HE staining results of liver tissues of different groups of mice; Figure 5 Dark avoidance test results of different groups of mice; Figure 6 Shuttle test results of different groups of mice; Figure 7 Open field test results of different groups of mice; Figure 8 Elevated plus maze test results of different groups of mice. DETAILED DESCRIPTION

[0018] The application provides an application of Allo-aca in preparation of a medicine for improving liver damage and concurrent neurological symptoms.

[0019] The Allo-aca is a leptin receptor antagonist, and the molecular formula is C 48 H 75 N 13 O 15 , the molecular weight is 1074.19 Da, and the sequence is {H-allo}-Thr-Glu-{Nva}-Val-Ala-Leu-Ser-Arg-{Aca}-NH2.

[0020] The action mechanism of Allo-aca is to intervene in a leptin-dependent pathological process by competitively binding to a leptin receptor and blocking a signal transduction pathway of leptin. The specific source of the Allo-aca is not specially limited in the application, and the Allo-aca can be directly obtained through a market channel. The chronic stress-induced aflatoxin hepatotoxicity exacerbation effect is improved by the leptin receptor (lepr) antagonist by reducing the activity of hepatocytes through high expression of the lepr gene.

[0021] In the application, the medicine preferably includes an oral agent or an injection agent, and the medicine further preferably includes a pharmaceutically acceptable carrier and / or an excipient.

[0022] The technical solutions provided by the application will be described in detail below in combination with the embodiments, but they should not be understood as limiting the protection scope of the application.

[0023] Example 1 1. Experimental materials (1) Experimental animals: SPF grade male C57BL / 6 mice, 6-8 weeks old, weighing 20-22 g; (2) Reagents: aflatoxin B1 (AFB1), Allo-aca, physiological saline, dimethyl sulfoxide, polyethylene glycol 300, Tween; (3) Instruments: automatic biochemical analyzer, tissue section machine, microscope.

[0024] 2. Experimental method (1) Mouse grouping and treatment Normal control group (Contral): mice were free to move, and each control group of mice was injected with the same volume of solvent 0.2 mL / time / day, and the same volume of solvent mixture without AFB1 was given by gavage, and the mice were weighed on an electronic balance every morning at 9:00; Chronic stress + AFB1 group (AFBI + CRS): each mouse was injected with the same volume of solvent 0.2 mL / time / day, chronic restraint stress modeling, AFB1 solution was given by gavage 0.2 mL / time / day, lasting for three weeks, CRS for 3 hours in the restraint tube every day from 2:00 to 5:00 in the second and third weeks, and the mice were weighed on an electronic balance every morning at 9:00.

[0025] Allo-aca intervention group (AFBI + CRS + Allo-aca): Allo-aca was injected intraperitoneally at 1 mg / kg / day and AFB1 solution was given by gavage 0.2 mL / time / day, lasting for three weeks, CRS for 3 hours in the restraint tube every day from 2:00 to 5:00 in the second and third weeks, and the mice were weighed on an electronic balance every morning at 9:00.

[0026] (2) Index detection After the experiment, the eyeball was taken out and the serum was separated, and the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the serum were detected by automatic biochemical analyzer. The abdominal cavity was quickly opened, the liver was completely peeled off, the gallbladder, connective tissue and attached blood vessels were removed, and the average value was obtained after measuring three times on a precision balance. The liver tissue was taken out, HE staining was performed, and the inflammatory infiltration of the liver tissue was observed under a microscope.

[0027] 3. Experimental results As shown in Figures 1-4 , the body weight of the AFB1 + CRS group of mice increased slowly compared with the Control group, and Allo-aca alleviated the effect of chronic stress + AFB1 on the body weight of mice Figure 1 , the liver / body ratio of mice was significantly increased after AFB1 + CRS treatment, and Allo-aca improved this effect Figure 2). Compared with the Control group, the serum ALT and AST levels of the AFB1+CRS group of mice were significantly increased, and the degree of liver tissue inflammation was increased. The serum ALT and AST levels of the Allo-aca intervention group (AFB1+CRS+Allo-aca) of mice were significantly lower than those of the AFB1+CRS group (P<0.05), and the degree of liver tissue inflammation was reduced, indicating that Allo-aca had a significant liver protection effect on chronic stress-induced aflatoxin hepatotoxicity Figure 3 ). The Allo-aca intervention group (AFB1+CRS+Allo-aca) also had a relieving effect on the hepatocyte edema and ballooning degeneration and inflammatory cell infiltration caused by chronic stress+AFB1 Figure 4 .

[0028] Example 2 1. Experimental materials The experimental animals and reagents were the same as in Example 1.

[0029] Instruments: dark avoidance experiment box, shuttle experiment box, elevated plus maze, open field.

[0030] 2. Experimental methods (1) Mouse grouping and treatment: same as Example 1.

[0031] (2) Behavioral detection Dark avoidance experiment: The dark avoidance experiment device consists of a bright room and a dark room, and the two rooms are connected by a small hole. The bottom of the dark room is paved with electrified metal grid, and the animal will be slightly shocked when it enters the dark room. Before the experiment, the animal is placed in the bright room for a period of time, and then the experiment is started, and the latency of the animal from the bright room to the dark room for the first time is recorded.

[0032] During the experiment, normal animals usually have a tendency to dark, but due to the fear of electric shock, they will try to avoid entering the dark room. If the animal enters the dark room quickly or enters it more often, it indicates that its learning and memory ability may be poor, and it cannot effectively remember the bad experience of electric shock; on the contrary, the latency is long and the number of entries is small, indicating that the animal has good learning and memory ability and can remember the danger of the dark room and actively avoid it.

[0033] Shuttle experiment: The shuttle experiment device is a shuttle box composed of two interconnected boxes, with a metal grid at the bottom that can be electrified by an electric shock controller. One day before the experiment, the animals are allowed to freely explore the shuttle box for a period of time to familiarize them with the environment, and then a conditioned stimulus (such as light, sound) is given. After a certain period of time (e.g., 5 seconds), the animals are subjected to electric shock by electrifying the bottom of the box. To avoid the electric shock, the animals will shuttle back and forth in the shuttle box. During the experiment, the number of shuttles and escape latency of the animals under each conditioned stimulus are recorded. As the number of experiments increases, normal animals will gradually learn to escape the electric shock by shuttling to a safe area before the shock, and the escape latency will gradually shorten and the number of shuttles will increase. If the animal's escape latency is prolonged and the number of shuttles is reduced, it suggests that its learning and memory ability may be impaired.

[0034] Open field experiment: The open field experiment of all mice was performed after 18:00 every day. Three days before the experiment, the mice to be tested were moved to the experimental waiting area to adapt to the new environment, and each mouse was comforted to reduce the stress response caused by the environment and human beings. The experimental field was illuminated at a brightness of 80-120 lux, with suitable humidity and a temperature of 24±0.5°C. When the formal experiment began, the mice were placed in the central region of the open field experiment device (a topless cubic plastic box with dimensions of 50cm x 50cm x 50cm), and the movement trajectory of the mice in the open field device was recorded by an animal behavior video acquisition system for 6 minutes. After each mouse was tested, it was placed in a new cage, and the feces, urine in the open field device were cleaned, and the residual odor was eliminated with 75% alcohol. After the device was completely dried, the next mouse experiment was performed. During the entire experiment, the experimenter remained quiet and recorded the entire experimental process outside the experimental area (the experimental area was hung with curtains to block it) to avoid the experimenter's own odor causing stress to the mice and affecting the experimental results. After all the experiments were completed, the mice were all moved back to their original cages for rest.

[0035] Elevated plus maze test: The elevated plus maze test of all mice was performed after 18:00 every day. The mice to be tested were moved to the experimental waiting area 3 days before the experiment to adapt to the new environment, and each mouse was comforted to reduce the stress caused by the environment and human. The experimental site was 80-120 lux, the humidity was suitable, and the temperature was maintained at 24±0.5°C. The cross-maze assembly was connected by two opposite open arms (50 cm long x 10 cm wide) and two opposite closed arms (50 cm long x 10 cm wide x 40 cm high) and a central area (10 cm x 10 cm). The material was made of high-grade medical organic board, the contact surface was not reflective, and it could be cleaned with alcohol to remove the smell. At the beginning of the formal experiment, the mouse was placed in the central area of the maze experiment device. The animal behavior video acquisition system recorded the mouse's activity within 6 minutes after the mouse was placed in the maze from the central grid to the closed arm. The observation indicators included: the number of times entering the open arm (must have two front paws entering the arm), the open arm stay time, the closed arm entering time, and the closed arm stay time. Calculate the open arm stay time ratio, open arm entering time ratio, and total entering time in the elevated plus maze. After the experiment was completed, the mouse was taken out, the two arms were cleaned, alcohol was sprayed to remove the smell, and finally the animal behavior software was used for data analysis. The number of times entering the open arm and the stay time were positively correlated with the positive emotions of the mouse. The more the number of times entering the open arm and the longer the stay time, the higher the positive emotions of the mouse. After each mouse was tested, it was placed in a new cage and the feces, urine in the open field device were cleaned, and the residual odor was eliminated with 75% alcohol. After the device was completely dried, the next mouse experiment was performed. During the entire experimental process, the experimenter remained quiet and recorded the entire experimental process outside the experimental area (the experimental area was hung with curtains to block) to avoid the odor of the experimenter affecting the mouse and affecting the experimental results. After all the experiments were completed, the mice were all moved back to their original cages for rest.

[0036] 3. Experimental results As shown in Figures 5-8 , the AFB1+CRS group of mice had a significantly shortened latency to hide, an increased number of errors, a significantly reduced number of successful shuttles, a shortened central stay time in the open field, and a reduced open arm stay time in the elevated plus maze, indicating that there were obvious memory impairment and anxiety. The AFB1+CRS+Allo-aca group of mice had a significantly prolonged latency to hide compared with the AFB1+CRS group (P<0.05), a reduced number of errors, a significantly increased number of successful shuttles, a prolonged central stay time, and a prolonged open arm stay time, indicating that Allo-aca could effectively improve the cognitive impairment and anxiety caused by the exacerbation of aflatoxin hepatotoxicity induced by chronic stress.

[0037] The above merely describes the preferred embodiments of the present application, and it should be pointed out that, for those skilled in the art, several improvements and refinements can be made without departing from the principles of the present application, and these improvements and refinements should also be considered as falling within the protection scope of the present application.

Claims

1. Application of Allo-aca in the preparation of drugs to improve liver injury and associated neurological symptoms.

2. The application according to claim 1, characterized in that, The liver damage and accompanying neurological symptoms were caused by aflatoxin.

3. The application according to claim 1 or 2, characterized in that, The liver damage and accompanying neurological symptoms are due to the aflatoxin-induced exacerbation effect of chronic stress.

4. The application according to claim 1, characterized in that, Allo-aca competitively binds to the leptin receptor, blocking the leptin signaling pathway and thus intervening in leptin-dependent pathological processes.

5. The application according to claim 1, characterized in that, Allo-aca has a liver-protective effect.

6. The application according to claim 1, characterized in that, Allo-aca has the effect of relieving hepatocellular edema and ballooning degeneration, as well as inflammatory cell infiltration.

7. The application according to claim 1, characterized in that, Allo-aca has the effect of improving cognitive impairment and anxiety.

8. The application according to claim 1, characterized in that, The drug may be taken orally or injected.

9. The application according to claim 8, characterized in that, The drug also includes pharmaceutically acceptable carriers and / or excipients.

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