Application of eubacterium inerti in preparation of medicine for treating senescence cognitive impairment

Abstract

The invention discloses an application of Eubacterium inertus in preparation of a medicine for treating senescence cognitive impairment, the Eubacterium inertus can effectively inhibit the occurrence and development of senescence cognitive impairment, has an obvious treatment effect on senescence cognitive impairment, does not generate obvious side effects, acts on intestinal flora, and can be used for treating senescence cognitive impairment. The traditional Chinese medicine composition has a remarkable improvement effect on the spatial memory ability caused by aging, can greatly enhance the anti-oxidation and anti-inflammatory ability, and has a great application prospect in the aspect of treatment of senescence cognitive impairment.

Description

Technical Field

[0001] The invention relates to application of Eubacterium inermis in preparing medicine for treating senile cognitive impairment. Background Art

[0002] As the population ages, many problems such as organ dysfunction and increased disease risk caused by aging are becoming increasingly prominent. Understanding the mechanism of aging-related cognitive impairment and finding anti-aging methods are of great significance to improving the quality of life and preventing and treating related diseases. However, current treatment methods are limited and have many problems. Drug therapy has side effects, and gene therapy technology and ethical issues need to be resolved. Overall, research on aging-related cognitive impairment still faces many challenges, and it is necessary to continue to explore safer and more effective treatment approaches.

[0003] Eubacterium lentum is a Gram-positive bacterium belonging to the family Eubacteriaceae and the phylum Firmicutes. It is an important intestinal bacterium found in the colon of healthy people. It can produce short-chain fatty acids, promote intestinal integrity, inhibit proinflammatory cytokines, and enhance immune regulation. It is a probiotic with the potential to resist aging-related cognitive impairment. However, there are very few reports on the application of Eubacterium lentum in the treatment of aging-related cognitive impairment.

[0004] At present, the clinical treatment of cognitive impairment associated with age has the following defects: 1) Risk of side effects: For example, women who use estrogen for anti-aging treatment may increase the risk of breast cancer, endometrial cancer and other diseases. 2) Individual differences: Because each person's genes, age, gender, lifestyle, etc. are different, the symptoms of cognitive impairment associated with age and the response to treatment are different. 3) Lack of effective treatment methods: The mechanism of cognitive impairment associated with age is extremely complex, involving changes at multiple levels, and there is a lack of effective treatment options. 4) Lack of specificity: For example, for patients with premature aging due to genetic factors, conventional anti-aging cognitive impairment drugs may not be effective. 5) High technical difficulty: Gene therapy requires precise editing or regulation of specific genes, which is very difficult. 6) High cost: The cost of research and development and clinical application of gene therapy is very high, and most patients cannot afford it. Therefore, it is urgent to seek new methods and new ideas for the treatment of cognitive impairment associated with age. Summary of the invention

[0005] Purpose of the invention: The purpose of the present invention is to provide the use of Eubacterium inermis in the preparation of a drug for treating senile cognitive impairment; it can effectively inhibit the occurrence and development of senile cognitive impairment without producing obvious side effects.

[0006] Technical solution: The present invention discloses the use of Eubacterium inermis in the preparation of medicines for treating senile cognitive impairment.

[0007] Among them, the aging-related cognitive impairment is the cognitive impairment caused by brain aging. In the medical field, Eubacterium iners is a potential treatment for various problems caused by aging, especially the disease of aging-related cognitive impairment. It can improve cognitive impairment by producing short-chain fatty acids, promoting intestinal integrity, inhibiting proinflammatory cytokines, and enhancing immune regulation. In the D-galactose-induced aging mouse model, Eubacterium iners significantly improved the spatial memory ability of the aging mouse model, reflecting its therapeutic application effect on aging-related cognitive impairment in this specific model.

[0008] Among them, the manifestations of aging-related cognitive impairment are impaired learning and memory ability, decreased antioxidant capacity, enhanced inflammatory response, decreased total antioxidant capacity in the brain, and increased tumor necrosis factor-α levels. In the aging mouse model, with the advancement of the aging process and the appearance of cognitive impairment, the level of T-AOC (total antioxidant capacity) in the brain decreased and the level of TNF-α (tumor necrosis factor-α) increased, indicating a weakening of antioxidant defense capacity and aggravation of neuroinflammation. When Eubacterium inermis was used for intervention, the experimental results showed that it could significantly increase the level of T-AOC in the brain and reduce the level of TNF-α.

[0009] Among them, the inert Eubacterium acts on the intestinal flora, significantly improving the spatial memory ability caused by aging, and can greatly enhance its antioxidant and anti-inflammatory capabilities.

[0010] Among them, the inert Eubacterium promotes the production of short-chain fatty acids, thereby regulating the gut-brain axis, enhancing the antioxidant capacity of brain tissue, and reducing the release of pro-inflammatory factors by regulating immune cell activity, thereby alleviating neuroinflammation.

[0011] The present invention also discloses a pharmaceutical composition for treating senile cognitive impairment, comprising the above-mentioned inert eubacterium and a pharmaceutically acceptable carrier or excipient, and prepared into a pharmaceutically acceptable dosage form.

[0012] The pharmaceutically acceptable carrier or excipient includes one or more of solid, semisolid or liquid excipients.

[0013] Wherein, the pharmaceutically acceptable dosage forms include tablets, capsules, granules, injections, pills, syrups, powders or pastes.

[0014] Principle of the invention: The potential intervention effect of Eubacterium inertes in the treatment of aging-related cognitive impairment through intestinal flora is mainly achieved through its regulatory effect in intestinal flora. First, Eubacterium inertes can inhibit the production of proinflammatory cytokines and reduce inflammatory responses, and this effect can effectively alleviate inflammation in the brain through signal transmission of the gut-brain axis. The inventor has found through research that excessive inflammatory response is closely related to brain aging and cognitive impairment. Excessive proinflammatory factors will not only damage nerve cells and their synaptic connections, but also cause disorders of the neural network in the brain, thereby affecting cognitive function. Eubacterium inertes inhibits these harmful reactions by reducing the release of inflammatory factors, thereby providing protection for brain function. At the same time, Eubacterium inertes indirectly reduces the level of oxidative stress by regulating the intestinal environment and reducing the generation of free radicals. Oxidative stress is considered to be a key factor in brain aging. Eubacterium inertes reduces oxidative stress by reducing the excessive production of free radicals, thereby indirectly enhancing antioxidant capacity. This antioxidant effect can stabilize the cell membrane of neurons, protect neurons from oxidative damage, and provide better conditions for maintaining cognitive function.

[0015] These effects bring unexpected results. For example, behavioral experiments show that the spatial memory ability of aging mice is significantly improved, and the antioxidant and anti-inflammatory capacity in the brain of aging mice is greatly enhanced, providing new evidence and direction for the treatment of cognitive disorders in brain aging, and revealing the potential of Eubacterium inermis as an anti-aging and cognitive disorder intervention in the gut-brain axis pathway.

[0016] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages: Eubacterium inermis can effectively inhibit the occurrence and development of aging-related cognitive impairment, has a significant therapeutic effect on aging-related cognitive impairment, and does not produce obvious side effects. It acts on the intestinal flora and has a significant improvement effect on the spatial memory ability caused by aging, and can greatly enhance its antioxidant and anti-inflammatory capabilities. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 Schematic diagram of the Y-maze-spontaneous alternation experiment Y in Example 1, wherein A is the blank group, B is the model group, and C is the Eubacterium inerans + model group;

[0018] Figure 2 The results of the Y-maze spontaneous alternation experiment Y in Example 1, wherein A is the number of new arm times, B is the spontaneous alternation rate, and C is the ratio of time in the new arm;

[0019] Figure 3 Schematic diagram of the new object recognition experiment in Example 2, wherein A is a blank group, B is a model group, and C is an Eubacterium inermis + model group;

[0020] Figure 4is the result of the novel object recognition experiment in Example 2, where A is the exploration time, B is the exploration time ratio, and C is the preference index;

[0021] Figure 5 The experimental results of the antioxidant capacity of Eubacterium inermis in the brain;

[0022] Figure 6 These are the experimental results of the tumor necrosis factor level in the brain of Eubacterium inermis. DETAILED DESCRIPTION

[0023] The technical scheme of the present invention is further described below in conjunction with the examples. The test materials used in the examples can all be purchased through conventional channels.

[0024] Among them, SPF-grade male C57BL / 6J mice, 6 weeks old, weighing 18-20 g, were purchased from Zhejiang Weitonglihua Experimental Animal Technology Co., Ltd. The quality inspection unit was Beijing Weitonglihua Experimental Animal Technology Co., Ltd., and the animal production license number was: SCXK(Zhe)2019-0001.

[0025] Eubacterium lentum, ATCC29066, was purchased from Ningbo Mingzhou Biotechnology Co., Ltd.

[0026] Example 1

[0027] Construction of D-galactose-induced aging mouse model and behavioral experiments:

[0028] (1) Establishment of aging mouse model: Male C57BL / 6J mice were intraperitoneally injected with 1000 mg / kg / day of D-galactose (Sigma-Aldrich) for 75 days.

[0029] (2) Activation of Eubacterium inermis (ATCC29066): Break the vacuum tube with a grinding wheel on a clean bench, and then sterilize the broken hole with an alcohol lamp flame. Use the needle of a sterile syringe to loosen the compacted bacterial powder, and then transfer the bacterial powder to a sterile cryopreservation tube, and then divide the bacterial powder into two. Treat the mouth and lid of the cryopreservation tube with an alcohol lamp flame, then seal the cryopreservation tube with a sealing film and transfer it to an anaerobic box, then dissolve the bacterial powder with Medium 110 culture medium (GibcoTM, ThermoFisher, USA) and place it in an anaerobic bag for culture.

[0030] (3) Cultivation and cryopreservation of Eubacterium inermis (ATCC29066): Place the culture glass tube in a shaker and culture at 37°C. When obvious turbidity appears, use a syringe to draw an appropriate amount of bacterial solution and transfer it to a 50 ml centrifuge tube for continued culture until obvious turbidity appears again. Attach a sealing film to the centrifuge tube and centrifuge at 4000 rpm for 10 minutes. Retain an appropriate amount of culture medium according to the number of bacteria. After resuspending, take 1 ml of bacterial solution and add it to a 2.5 ml sterile cryopreservation tube that has been pre-filled with 1 ml of 80% glycerol.

[0031] (4) Colonization of Eubacterium inermis (ATCC29066): Take Eubacterium inermis in the logarithmic growth phase, dilute it 100 times, and calculate the OD value based on the OD value. 600 The number of live bacteria was calculated by the absorbance of 1000 nm. The bacterial solution was centrifuged at 8000 g and the precipitate was collected. According to the number of bacteria measured, the inert Eubacterium was diluted to 5×10 9 CFU / ml and stored at 4°C for subsequent oral administration to mice.

[0032] (5) Dosage grouping scheme: After one week of adaptive feeding, C57BL / 6J mice were randomly divided into groups, with 10 mice in each group. In the mouse model, the daily dose of the Eubacterium inermis was 10 9 CFU / 0.2mL / d (i.e., 10 9 Colony forming units), administered continuously for 15 days.

[0033] Blank group: intraperitoneal injection of 0.2 ml of normal saline for 75 consecutive days. Starting from the 61st day, after intraperitoneal injection of normal saline, 0.2 ml of sterile normal saline was gavaged for 15 consecutive days.

[0034] Model group: D-galactose (1000 mg / kg / day) was intraperitoneally injected for 75 consecutive days. Starting from the 61st day, after intraperitoneal injection of D-galactose, 0.2 ml of sterile saline was gavage for 15 consecutive days.

[0035] Eubacterium inermis + model group: D-galactose (1000 mg / kg / day) was intraperitoneally injected for 75 consecutive days. Starting from the 61st day, Eubacterium inermis suspension was gavaged for 10 9 CFU / 0.2mL / d for 15 consecutive days.

[0036] Conduct the Y-maze-spontaneous alternation experiment:

[0037] The mice were placed in a Y-maze and allowed to freely explore and adapt to the environment for a period of time. After the adaptation period, the mice were placed at the end of one arm of the Y-maze, and the order and number of times the mice entered each arm were recorded. Figure 1 and Figure 2 The A in the figure was significantly increased, indicating that the mice were able to remember the arms they had explored and were more inclined to enter the new arms. Figure 1 and Figure 2 The ratio of time in the new arm ( Figure 1 and Figure 2 The C in the figure was significantly increased, indicating that the mice may have stronger attention and exploration ability to the new environment. The Y-maze-spontaneous alternation experiment showed that Eubacterium iners significantly improved the spatial memory ability of aging mice.

[0038] Example 2

[0039] Similar to Example 1, a blank group, a model group, and a Eubacterium inerans + model group were used to conduct a new object recognition experiment:

[0040] The mice were placed in a container without objects. Two identical objects were placed in the container at a fixed position. The animals were placed with their backs facing the objects at equal distances. The data on their exploration of the objects was recorded within 5 minutes. One day later, one of the objects was replaced with a new object and the animals were placed in the container for observation for 5 minutes. Figure 3 and Figure 4 A in, exploration time ratio ( Figure 3 and Figure 4 B), preference index ( Figure 3 and Figure 4 The C in the samples was significantly increased, the mice spent more time and more times around the new objects, and showed more exploratory behavior towards the new objects. The novel object recognition experiment showed that Eubacterium inermis significantly improved the cognitive ability of aging mice.

[0041] Example 3

[0042] The T-AOC and TNF-α in the brain were measured using the Bio-Tech test kit. The samples were processed according to the instructions. After the sample addition reaction, the absorbance was read and the concentration was calculated.

[0043] The results show that Figure 5 and 6 As shown in the results, Eubacterium iners significantly increased the level of T-AOC in the brain of aged mice, significantly reduced the level of TNF-α in the brain, and enhanced the antioxidant and anti-inflammatory capacity of the brain of aged mice, providing evidence for the role of intestinal flora in the treatment of cognitive impairment in brain aging. After intervention with Eubacterium iners, the experimental results showed that it could significantly increase the level of T-AOC and reduce the level of TNF-α in the brain, indicating that Eubacterium iners may improve aging-related cognitive impairment by enhancing the antioxidant capacity of the brain and reducing inflammatory response.

[0044] Through the above-mentioned novel object recognition experiment and Y-maze-spontaneous alternation experiment to detect the learning and memory ability of mice, it can be seen that the spatial memory function of mice was impaired after D-galactose aging modeling, and the preference index, exploration times and time of mice for new objects increased significantly after inoculation of Eubacterium inermis. Therefore, it is fully demonstrated that Eubacterium inermis can improve cognitive impairment by producing short-chain fatty acids, promoting intestinal integrity, inhibiting proinflammatory cytokines and enhancing immune regulation. In the D-galactose-induced aging mouse model, Eubacterium inermis significantly improved the spatial memory ability of the aging mouse model, reflecting its effective inhibition of the development of aging-related cognitive impairment, and has a significant therapeutic effect on aging-related cognitive impairment. There were no obvious side effects during the experiment, such as gastrointestinal reactions, liver and kidney damage, mental abnormalities, etc., and it has great application prospects in the treatment of aging-related cognitive impairment.

Claims

1. Application of Eubacterium inermis in the preparation of drugs for treating aging-related cognitive impairment.

2. The use according to claim 1, characterized in that: The aging-related cognitive impairment is a cognitive impairment caused by brain aging.

3. The use according to claim 1, characterized in that: The manifestations of aging-related cognitive impairment are impaired learning and memory abilities, decreased antioxidant capacity, enhanced inflammatory response, reduced levels of total antioxidant capacity in the brain, and increased levels of tumor necrosis factor-α.

4. The use according to claim 1, characterized in that: The inert eubacterium acts on the intestinal flora, significantly improving the spatial memory ability caused by aging, and can greatly enhance its anti-oxidation and anti-inflammatory capabilities.

5. The use according to claim 4, characterized in that: Eubacterium inermis promotes the production of short-chain fatty acids, thereby regulating the gut-brain axis, enhancing the antioxidant capacity of brain tissue, and reducing the release of proinflammatory factors by regulating immune cell activity, thereby alleviating neuroinflammation.

6. A pharmaceutical composition for treating senile cognitive impairment, characterized in that: The invention comprises the inert eubacterium according to claim 1 and a pharmaceutically acceptable carrier or excipient to prepare a pharmaceutically acceptable dosage form.

7. The composition according to claim 6, characterized in that The pharmaceutically acceptable carrier or excipient includes one or more of solid, semisolid or liquid excipients.

8. The composition according to claim 6, characterized in that The pharmaceutically acceptable dosage forms include tablets, capsules, granules, injections, pills, syrups, powders or pastes.

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