Application of lactobacillus plantarum GUANKE in prevention and treatment of liver injury

Abstract

The invention relates to the technical field of medicines, and discloses application of lactobacillus plantarum GUANKE in prevention and treatment of liver injury. According to the invention, a liver injury mouse model is established through a ZEN infected mouse, and lactobacillus plantarum GUANKE powder is fed into the stomach of the liver injury mouse, so that compared with a model group, the serum, liver function, antioxidant enzyme activity and liver related inflammation indexes of the mouse fed with the lactobacillus plantarum GUANKE are correspondingly improved; and the effect of high-dose treatment of the lactobacillus plantarum GUANKE is more remarkable than that of low-dose treatment of the lactobacillus plantarum GUANKE. Moreover, the tissue pathological section of the liver shows that compared with the inflammation, necrosis and lipid degeneration of the mouse tissue of the model group, the inflammation, necrosis and lipid degeneration of the mouse tissue are relieved after the treatment of the lactobacillus plantarum GUANKE. In conclusion, the lactobacillus plantarum GUANKE has the beneficial effect of treating the liver injury.

Description

Technical Field

[0001] The invention relates to the technical field of medicine, and in particular to application of Lactobacillus plantarum GUANKE in preventing and treating liver damage. Background Art

[0002] Zearalenone (ZEN) is an estrogen-like mycotoxin found in food and feed that can cause oxidative damage and inflammation levels in the liver and kidneys, seriously endangering the safety of animal and human health. Zearalenone has strong heat resistance and can only be completely destroyed by treatment at up to 110°C for 1 hour. Zearalenone mainly contaminates grains such as corn, wheat, rice, barley, millet and oats. According to statistics, the positive detection rate of zearalenone in corn is 45%, and the positive detection rate of wheat is 20%. Therefore, in people's daily lives, it is necessary to prevent the body damage caused by zearalenone. In addition, such as liver damage, it can also occur in the overuse of daily medications such as acetaminophen. Liver damage and other situations occur from time to time, and finding effective drugs for the treatment of liver damage has become an urgent problem to be solved.

[0003] There have been many studies in the prior art on the treatment of liver damage. For example, the patent with application number CN2022101988323 discloses manganese-based nanozymes as ferroptosis inhibitors and their application in liver damage. The above patent applies manganese-based nanozyme preparations to mouse embryonic fibroblasts and living mice, and finds that manganese-based nanozyme preparations, as ferroptosis inhibitors, can significantly inhibit cell ferroptosis induced by RSL3 and Erastin, and have a long-term and stable inhibitory effect. It also has a mitigating effect on drug-induced liver damage in mice induced by acetaminophen (APAP). However, the manganese-based nanozymes in this patent have limited mitigating effects on liver damage.

[0004] In order to provide more practical solutions for the treatment and relief of liver damage, more effective therapeutic drugs are still needed. Summary of the invention

[0005] In order to solve the above technical problems, the present invention provides the use of Lactobacillus plantarum GUANKE in preventing and treating liver damage.

[0006] The specific technical scheme of the present invention is: In one aspect, the present invention provides use of Lactobacillus plantarum GUANKE in preparing a medicament for preventing and / or treating liver damage.

[0007] Lactobacillus plantarum GUANKE, the strain preservation number is CGMCC NO.21720, the preservation date is January 22, 2021, the preservation classification name is Lactobacillus Plantarum GUANKE, and the preservation unit is the General Microbiology Center of China Microbiological Culture Collection Administration.

[0008] Researchers of the present invention establish liver injury mouse model by ZEN infection mice, and gavage plant lactobacillus GUANKE bacterial powder to liver injury mice, find that compared with the model group, the serum, liver function, antioxidant enzyme activity and liver-related inflammation index of the mice gavaged with plant lactobacillus GUANKE have been improved accordingly, and the effect of plant lactobacillus GUANKE high-dose treatment is more significant than the effect of plant lactobacillus GUANKE low-dose treatment. In addition, the histopathological sections of the liver show that inflammation, necrosis and lipid degeneration of mouse tissues are reduced after plant lactobacillus GUANKE treatment compared to inflammation, necrosis and lipid degeneration of mouse tissues in the model group. Show in summary, plant lactobacillus GUANKE has a beneficial effect in treating liver injury.

[0009] As a preferred embodiment of the above application, the dosage form of the drug includes granules, ordinary tablets, dispersible tablets, effervescent tablets, orally disintegrating tablets, lozenges, chewable tablets, capsules, soft capsules, microcapsules, pills, powders, pellets, sustained-release preparations, controlled-release preparations, oral liquid preparations and injections.

[0010] As a preferred embodiment of the above application, the liver injury is acute liver injury.

[0011] Acute liver injury refers to a type of acute inflammatory liver disease caused by improper medication, infection, ischemia-reperfusion, idiosyncratic reactions, etc., among which improper medication is more common. Based on the beneficial effects of the above-mentioned Lactobacillus plantarum GUANKE in treating liver injury, Lactobacillus plantarum GUANKE can be used to alleviate acute liver injury caused by drugs.

[0012] As a preferred embodiment of the above application, the liver damage is caused by zearalenone.

[0013] In another aspect, the present invention provides a composition for treating liver damage, wherein the active ingredient of the composition is Lactobacillus plantarum GUANKE.

[0014] As a preferred embodiment of the above composition, the liver injury is acute liver injury.

[0015] As a preferred embodiment of the above composition, the liver damage is caused by zearalenone.

[0016] Preferably, the above composition comprises one or more pharmaceutically acceptable excipients.

[0017] As a preferred embodiment of the above composition, the composition is prepared into granules, ordinary tablets, dispersible tablets, effervescent tablets, orally disintegrating tablets, lozenges, chewable tablets, capsules, soft capsules, microcapsules, pills, powders, pellets, sustained-release preparations, controlled-release preparations, oral liquid preparations or injections.

[0018] Compared with the prior art, the present invention has the following technical effects: 1. A mouse model of liver injury was established by infecting mice with ZEN, and mice with liver injury were gavaged with Lactobacillus plantarum GUANKE powder. It was found that compared with the model group, the serum, liver function, antioxidant enzyme activity and liver-related inflammatory indicators of mice gavaged with Lactobacillus plantarum GUANKE were improved accordingly, and the effect of high-dose treatment with Lactobacillus plantarum GUANKE was more significant than that of low-dose treatment with Lactobacillus plantarum GUANKE. In addition, histopathological sections of the liver showed that compared with the inflammation, necrosis and lipid degeneration of mouse tissues in the model group, Lactobacillus plantarum GUANKE treatment reduced the inflammation, necrosis and lipid degeneration of mouse tissues. In summary, Lactobacillus plantarum GUANKE has a beneficial effect in the treatment of liver injury.

[0019] 2. At the same time, as a probiotic, Lactobacillus plantarum GUANKE has less side effects on the body. Experiments have shown that compared with the blank control group, the body weight and liver weight of mice gavaged with GUANKE did not change, which shows that GUANKE bacteria is harmless to mice. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 The statistical results of body weight measurement of mice in 5 experimental groups; Figure 2 The statistical results of liver weights of mice in the five experimental groups; Figure 3 The results of ALT concentration test in the serum of mice in the five experimental groups are shown in Figure 2. Figure 4 The results of ALP concentration test in the serum of mice in the five experimental groups are shown in Figure 2. Figure 5 The results of the detection of AST concentration in the serum of mice in the five experimental groups; Figure 6 The results of the detection of GSH activity in the liver of mice in the five experimental groups; Figure 7 The results of the detection of CAT activity in the liver of mice in the five experimental groups; Figure 8 The results of the detection of SOD activity in the liver of mice in the five experimental groups; Fig. 9The results of the detection of MDA content in the liver of mice in the five experimental groups; Fig.10 The results are for the detection of TNF-α content in the liver of mice in the five experimental groups; Fig.11 The results of the detection of IL-1β content in the liver of mice in the five experimental groups; Fig.12 The results of the detection of IL-6 content in the liver of mice in the five experimental groups; Fig.13 The results of the detection of IL-10 content in the liver of mice in the five experimental groups; Fig.14 HE staining images of the livers of mice in the five experimental groups. DETAILED DESCRIPTION

[0021] The present invention will be further described below in conjunction with embodiments. Those of ordinary skill in the art will be able to implement the present invention based on these descriptions. In addition, the embodiments of the present invention involved in the following description are generally only embodiments of a part of the present invention, rather than all embodiments. Therefore, based on the embodiments in the present invention, all other embodiments obtained by those of ordinary skill in the art without making creative work should belong to the scope of protection of the present invention.

[0022] The plant lactobacillus GUANKE used in the embodiment of the present invention has a strain deposit number of CGMCC NO.21720, a deposit date of January 22, 2021, and a deposit classification named Lactobacillus Plantarum GUANKE. The deposit unit is the General Microbiological Center of the China Microbiological Culture Collection Administration Committee, and the address is the Institute of Microbiology, Chinese Academy of Sciences, No. 3, No. 1, Beichen West Road, Chaoyang District, Beijing, Postal Code: 100101. This strain is a public strain. The above-mentioned plant lactobacillus GUANKE is fermented and freeze-dried to obtain GUANKE powder.

[0023] Example 1 Establishment of mouse model and drug administration (1) Experimental preparation Before the experiment, 50 newly purchased 7-week-old BALB / C female mice were fed for 7 days of adaptation. Then, according to the random principle, these mice were randomly divided into 5 experimental groups, with 5 mice in each group. The experimental groups included Control group, GUANKE group, ZEN group, ZEN+LG group, and ZEN+HG group.

[0024] (2) Model establishment and drug administration The total drug administration of the 5 experimental groups is as follows: Control group: The animals were fed with the same amount of saline solution as the other groups every day during the experiment, for a total of 28 days; GUANKE group: GUANKE powder was administered intragastrically every day during the experiment, with a dosage of 1×10 10 CFU / mouse, total administration for 28 days; ZEN group: During the experiment, ZEN was administered intragastrically every day at a dose of 40 mg / kg. Four hours after oral administration of ZEN, PBS was administered intragastrically again at the same volume as that of the ZEN+LG group and ZEN+HG group. The administration lasted for 28 days in total. ZEN+LG group: ZEN was administered orally every day during the experiment at a dose of 40 mg / kg, and GUANKE (administered at a dose of 1×10 6 CFU / mouse), and the drug was administered for 28 days; ZEN+HG group: ZEN was administered orally every day during the experiment at a dose of 40 mg / kg, and GUANKE (administered at a dose of 1×10 10 CFU / mouse), and the drug was administered for 28 days in total.

[0025] The specific experimental process is: Day 1 to 27: During the 28-day test period, the animals were provided with adequate food and water and were free to eat. The mental state, behavior, diet, weight changes and deaths of mice in each group were observed and recorded every day. The mice in the five experimental groups were dosed daily according to the above-mentioned dosing conditions.

[0026] Day 28: After fasting for 24 hours, the mice in each group were weighed, the eyeballs were removed and blood was collected, and the serum was separated after standing at room temperature for 4 hours. The mice were killed by dislocating the neck, the liver was removed and rinsed with PBS, wiped dry and weighed, and the liver organ index was calculated, and the liver surface was checked for organic lesions. The left lobe of the liver was prepared for routine histopathological testing, and the right lobe of the liver was used for molecular level related index testing.

[0027] Example 2 Histopathological study 1. Weight and organ indicators On the 28th day, the weight of mice in the five experimental groups was measured. Figure 1 , the statistical results of mouse liver weight are shown in Figure 2 The body weight and liver weight of mice can directly reflect the health status of mice.

[0028] The results of weighing the body weight and liver weight of the mice showed that compared with the Control group, the body weight and liver weight of the mice in the GUANKE group did not change; compared with the Control group, the body weight and liver weight of the mice in the ZEN group decreased by 28% (P<0.01) and 42.3% (P<0.01), respectively; compared with the ZEN group, the body weight and liver weight of the mice in the ZEN+LG group increased by 12.7% (P<0.05) and 22.2% (P<0.05), respectively; compared with the ZEN group, the body weight and liver weight of the mice in the ZEN+HG group increased by 27% (P<0.01) and 34.1% (P<0.05) compared with the ZEN group; compared with the ZEN+LG group, the body weight and liver weight of the mice in the ZEN+HG group increased by 11.3% (P<0.01) and 0.097% (P<0.05), respectively. The above results indicate that GUANKE bacteria are harmless to mice, while ZEN is harmful to mice. ZEN reduces the body weight and liver weight of mice, while GUANKE bacteria treatment reverses this damage. Within a certain range, the effect increases with the increase of GUANKE bacteria dose.

[0029] 2. Serological indicators On the 28th day, the ALT, ALP and AST concentrations in the serum of mice were detected. The ALT, ALP and AST concentrations of the mice in the five experimental groups were shown in Table 1. Figure 3 , Figure 4 , Figure 5 . ALT, ALP and AST are key transaminases in liver cells and are important indicators of liver damage. In a healthy state, the activities of ALT, ALP and AST in the blood are relatively low. However, once the liver is damaged, ALT, ALP and AST in the cells are released into the blood, causing the activities of ALT, ALP and AST in the blood to increase.

[0030] The results of ALT, ALP and AST concentrations in mice showed that compared with the Control group, there was no change in ALT, ALP and AST in the serum of mice in the GUANKE group; ALT, ALP and AST in the serum of mice in the ZEN group were increased by 46% (P<0.01), 65.7% (P<0.01) and 38.3% (P<0.01), respectively. Compared with the ZEN group, ALT, ALP and AST in the serum of mice in the ZEN+LG group were decreased by 26.5 (P<0.05), 17% (P<0.05) and 20% (P<0.05), respectively; compared with the ZEN group, ALT, ALP and AST in the serum of mice in the ZEN+HG group were decreased by 34.5% (P<0.01), 46.9% (P<0.01) and 28.3% (P<0.01), respectively. Compared with the ZEN+LG group, the ALT, ALP and AST in the serum of mice in the ZEN+HG group were down-regulated by 10.9% (P<0.05), 36% (P<0.01) and 15.7% (P<0.05), respectively. The above results show that ZEN can cause liver damage in mice, and GUANKE bacteria can treat liver damage caused by ZEN, and the effect is better with the increase of dose within a certain range.

[0031] 3. Oxidative stress indicators On the 28th day, the activities of GSH, CAT, SOD and the content of MDA in the liver of mice were detected. The results of the activities of GSH, CAT, SOD and the content of MDA were shown in Table 1. Figure 6 , Figure 7 , Figure 8 , Fig. 9 .

[0032] From the test results of GSH, CAT, SOD activities and MDA content in mouse liver, it can be seen that compared with the Control group, there was no difference in the activities of GSH, CAT, SOD and MDA content in the liver of mice in the GUANKE group; compared with the Control group, the activities of GSH, CAT and SOD in the liver of mice in the ZEN group were downregulated by 41% (P<0.01), 53.1% (P<0.01), and 19.1% (P<0.01), and the content of MDA was upregulated by 43.4% (P<0.01). Compared with the ZEN group, the activities of GSH, CAT and SOD in the liver of mice in the ZEN+LG group were increased by 19.8%, 42% (P<0.05), and 8.1%, and the content of MDA was decreased by 9.2% (P<0.05); compared with the ZEN group, the activities of GSH, CAT and SOD in the liver of mice in the ZEN+HG group were increased by 38.4% (P<0.01), 55% (P<0.01), and 18% (P<0.05), and the content of MDA was decreased by 34.8% (P<0.01). Compared with the ZEN+LG group, the activities of GSH, CAT and SOD in the liver of mice in the ZEN+HG group were increased by 26.2% (P<0.05), 22.3% (P<0.05), and 11%, and the content of MDA was decreased by 28.2% (P<0.05). The above results show that ZEN treatment increases free radicals in the liver of mice and generates lipid peroxidation products. GUANKE bacteria reduces the oxidative stress damage caused by ZEN to the liver, and the effect is better with the increase of dose within a certain range.

[0033] 4. Inflammatory factor indicators On the 28th day, the levels of TNF-α, IL-1β, IL-6, and IL-10 in the liver of mice were detected. The results of TNF-α, IL-1β, IL-6, and IL-10 content detection were shown in Table 1. Fig.10 , Fig.11 , Fig.12 , Fig.13 .

[0034] From the detection results of IL-10, TNF-α, IL-1β and IL-6 contents in mouse liver, it can be seen that compared with those in the Control group, there was no difference in the levels of IL-10, TNF-α, IL-1β and IL-6 in the liver of mice in the GUANKE group; compared with the Control group, the levels of TNF-α, IL-1β and IL-6 in the liver of mice in the ZEN group were increased by 76.2% (P<0.01), 96.2% (P<0.01) and 207.4% (P<0.01), and the level of IL-10 was decreased by 47% (P<0.01). Compared with the ZEN group, the levels of TNF-α, IL-1β and IL-6 in the liver of the mice in the ZEN+LG group were downregulated by 36.9% (P<0.05), 38.3% (P<0.05), and 36.9%, and the level of IL-10 was upregulated by 19.5%; compared with the ZEN group, the levels of TNF-α, IL-1β and IL-6 in the liver of the mice in the ZEN+HG group were downregulated by 62.8% (P<0.01), 79.7% (P<0.01), and 153% (P<0.01), and the level of IL-10 was upregulated by 37.4% (P<0.01). Compared with the ZEN+LG group, the levels of TNF-α, IL-1β and IL-6 in the liver of the mice in the ZEN+HG group were downregulated by 18.9%, 29.8%, and 85% (P<0.05), and the level of IL-10 was upregulated by 22.2% (P<0.05). The above results show that ZEN treatment increased the level of pro-inflammatory factors and reduced the level of anti-inflammatory factors in the liver of mice. GUANKE bacteria reduced the inflammatory damage caused by ZEN to the liver, and the effect was better with the increase of dose within a certain range.

[0035] 5. Pathological indicators On the 28th day, the left lobe of the liver was removed from the mice in each experimental group, fixed with 10% formalin, embedded, sliced, and stained with hematoxylin-eosin (HE). The damage of the liver was then observed under an electron microscope. The HE staining images of the left lobe of the liver of the mice in each group are shown in Fig.14 HE staining of liver sections can most intuitively reveal the morphology of tissue damage. By observing HE staining of liver sections, the damage to the liver can be clearly observed.

[0036] Depend on Fig.14It can be seen from the liver sections of the Control group and the GUANKE group that the boundaries of liver cells are clear, the structure of liver cells is intact (black arrow), the structure of liver sinusoids is intact, and there is no inflammatory infiltration; while in the liver sections of the ZEN group, the boundaries of liver cells are unclear (black arrow), liver cells have multiple focal necrosis, a large number of neutrophils infiltrate (red arrow), most liver cell nuclei undergo self-fusion (blue arrow), and the liver is severely damaged; in the ZEN+LG group, the boundaries of liver cells are unclear (black arrow), liver cells have partial focal necrosis, some neutrophils infiltrate (red arrow), liver cells are slightly edematous, some liver cell nuclei undergo self-fusion (blue arrow), and the liver is moderately damaged; in the ZEN+HG group, the boundaries of liver cells are clearer (black arrow), most of the liver sinusoids are visible, a small amount of neutrophils infiltrate (red arrow), and a small amount of stem cell nucleus self-fusion (blue arrow) can be seen. The above results show that ZEN can cause liver damage in mice, and GUANKE bacteria can treat liver damage caused by ZEN, and the effect is better with the increase of dose within a certain range.

[0037] The raw materials and equipment used in the present invention, unless otherwise specified, are all commonly used raw materials and equipment in the art; the methods used in the present invention, unless otherwise specified, are all conventional methods in the art.

[0038] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any way. Any simple modification, change and equivalent transformation made to the above embodiment based on the technical essence of the present invention still falls within the protection scope of the technical solution of the present invention.

Claims

1. Application of Lactobacillus plantarum GUANKE in the preparation of a medicament for preventing and / or treating liver damage.

2. The use according to claim 1, characterized in that: The dosage forms of the drug include granules, ordinary tablets, dispersible tablets, effervescent tablets, orally disintegrating tablets, lozenges, chewable tablets, capsules, soft capsules, microcapsules, pills, powders, drop pills, sustained-release preparations, controlled-release preparations, oral liquid preparations and injections.

3. The use according to claim 1, characterized in that: The liver injury is acute liver injury.

4. The use according to claim 1, characterized in that: The liver damage is caused by zearalenone.

5. A composition for treating liver damage, characterized in that: The active ingredient of the composition is Lactobacillus plantarum GUANKE.

6. The composition according to claim 5, characterized in that: The liver injury is acute liver injury.

7. The composition according to claim 5, characterized in that: The liver damage is caused by zearalenone.

8. The composition according to claim 5, characterized in that: The composition includes one or more pharmaceutically acceptable excipients.

9. The composition according to claim 5 or 8, characterized in that: The composition is prepared into granules, ordinary tablets, dispersible tablets, effervescent tablets, orally disintegrating tablets, lozenges, chewable tablets, capsules, soft capsules, microcapsules, pills, powders, drop pills, sustained-release preparations, controlled-release preparations, oral liquid preparations or injections.

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