Application of ackermann mucilaginosus and lactic acid in preparation of medicine for improving 5-hydroxytryptamine level of organism

Abstract

The invention provides application of combination of ackermann mucilaginosus and lactic acid in preparation of a medicine for improving the 5-hydroxytryptamine level of an organism. The invention further provides a pharmaceutical composition for improving the 5-hydroxytryptamine level of a body. The pharmaceutical composition contains ackermann mucilaginosus and lactic acid. According to the application, 16s rRNA sequencing and metabonomics means are applied, a scheme for improving the 5-hydroxytryptamine level of an organism through combination of ackermann mucilaginosus and lactic acid is screened and proposed for the first time, animal experiments prove that the scheme is used for improving the 5-hydroxytryptamine level, and a brand new treatment strategy is provided for 5-hydroxytryptamine deficiency related diseases.

Description

Technical Field

[0001] The invention belongs to the field of biomedicine and relates to Akkermansia muciniphila, and specifically to the use of Akkermansia muciniphila and lactic acid in combination in the preparation of a drug for increasing the level of 5-hydroxytryptamine in an organism. Background Art

[0002] In recent years, 5-HT (5-hydroxytryptamine), as a key neurotransmitter, has attracted widespread attention for its role in regulating emotions, sleep and intestinal function. 5-HT deficiency is closely related to a variety of mental and psychological diseases such as depression and anxiety, and is also associated with diseases such as intestinal dysfunction and immune disorders. However, traditional methods of increasing 5-HT mainly rely on drug intervention, such as selective 5-HT reuptake inhibitors, whose use is often accompanied by adverse reactions, and the effects vary from person to person. In recent years, studies have found that intestinal flora plays a key role in regulating 5-HT levels, especially certain specific probiotics and metabolites can significantly affect the 5-HT metabolic pathway, thereby increasing the synthesis and circulation levels of 5-HT. Summary of the invention

[0003] In view of the above technical problems in the prior art, the present invention provides the use of Akkermansia muciniphila and lactic acid in the preparation of a drug for increasing the level of 5-hydroxytryptamine in the body. The use is to solve the technical problem that the drugs in the prior art have adverse reactions and poor effects on increasing 5-HT.

[0004] The present invention provides the use of Akkermansia muciniphila and lactic acid in the preparation of a medicine for increasing the level of 5-hydroxytryptamine in an organism.

[0005] Furthermore, the amount of Akkermansia muciniphila is 2 x 10 8 CFU / d, the lactic acid dosage is 750mg / kg / d.

[0006] The present invention also provides the use of Akkermansia muciniphila and lactic acid in the preparation of a medicine for treating or improving diseases related to 5-hydroxytryptamine deficiency.

[0007] Furthermore, the amount of Akkermansia muciniphila is 2 x 10 8 CFU / d, the lactic acid dosage is 750mg / kg / d.

[0008] The present invention also provides a pharmaceutical composition for increasing the level of 5-hydroxytryptamine in the body, comprising Akkermansia muciniphila and lactic acid.

[0009] The present invention also provides a pharmaceutical composition for treating or improving diseases related to 5-hydroxytryptamine deficiency, characterized in that it contains Akkermansia muciniphila and lactic acid.

[0010] Furthermore, the pharmaceutical composition also contains a carrier and / or excipient acceptable in pharmaceuticals or the food industry.

[0011] Furthermore, the dosage form of the pharmaceutical composition is tablets, capsules, granules, pills, ointments, solutions, suspensions, emulsions, creams, sprays, drops, patches or tube feeding preparations.

[0012] The present invention screens anxiety-related microorganisms based on fecal 16S rRNA sequencing, and comprehensively screens out Akkermansia muciniphila. The present invention also screens anxiety-related metabolites based on fecal metabolome, and comprehensively screens out the key metabolite lactic acid.

[0013] Compared with the existing methods, the present invention optimizes the generation of 5-HT in a biological way, and has the characteristics of low side effects and high safety. The present invention uses 16s rRNA sequencing and metabolomics to screen and propose for the first time a scheme for increasing the body's 5-HT level by combining Akkermansia muciniphila with lactic acid, and verifies its ability to increase 5-HT levels through animal experiments, providing a new treatment strategy for 5-HT deficiency-related diseases.

[0014] The present invention uses Akkermansia muciniphila and lactic acid in combination to increase the body's 5-HT level, and has the characteristics of safety and high efficiency. The present invention promotes the production of intestinal 5-HT and increases serum 5-HT levels through flora regulation, providing a new treatment idea with low side effects for 5-HT deficiency-related diseases, and has important clinical application potential in the field of mental health.

[0015] Compared with the prior art, the technical effect of the present invention is positive and obvious. The present invention promotes the production of 5-HT in the body and restores 5-HT metabolic disorder by combining Akkermansia muciniphila with lactic acid, and has important practical application value in the field of mental and psychological diseases characterized by 5-HT depletion, such as depression and anxiety. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The diagram shows that the relative abundance of Akkermansia muciniphila in feces is significantly negatively correlated with the anxiety index GAD-7 score.

[0017] Figure 2 Schematic diagram showing that fecal lactic acid levels in the anxiety group were significantly reduced compared to the healthy group.

[0018] Figure 3 It is a schematic diagram of serum 5-HT levels of SPF mice after antibiotic detoxification in which a group of mice were orally administered Akkermansia muciniphila alone, a group of mice were orally administered lactic acid alone, and a group of mice were orally administered Akkermansia muciniphila and lactic acid together.

[0019] Figure 4It is a schematic diagram of serum 5-HT levels of germ-free mice of the present invention which orally administered Akkermansia muciniphila alone, lactic acid alone, and Akkermansia muciniphila and lactic acid together. DETAILED DESCRIPTION

[0020] In order to better understand the technical solution of the present invention, the embodiments of the present invention are described in detail below with reference to the accompanying drawings. Example 1

[0021] The invention discloses a medicine combining Akkermansia muciniphila and lactic acid for improving the level of 5-HT.

[0022] 1. 16s rRNA sequencing was performed using fecal samples of volunteers in an anxious state (specific screening criteria were GAD-7>4 points), and the obtained high-throughput data were subjected to bioinformatics analysis. The specific process included sequence quality control through USEARCH and VSEARCH to remove low-quality bases and sequence fragments, remove adapter primers and chimera sequences, merge double-end sequencing data and denoise, and obtain ASV based on sequence similarity alignment. ASV data was used for lefse screening of differential microorganisms (genus level) through GALAXY, and the specific standard was species with LDA>2.0. Prism9 was further used to perform spearman correlation analysis on the obtained differential species and the GAD-7 scores of the volunteers' psychological scale, with specific criteria of r<0, p<0.05. Finally, the microorganism Akkermansia muciniphila was screened, which was significantly reduced in the anxious samples and significantly negatively correlated with the anxiety index.

[0023] Figure 1 The results showed that the relative abundance of Akkermansia muciniphila in feces was significantly negatively correlated with the anxiety index GAD-7 score.

[0024] 2. The fecal samples of the volunteers were tested by liquid chromatography-mass spectrometry (the screening criteria for the anxiety group was GAD-7>4 points, and the screening criteria for the healthy group was GAD-7<4), followed by non-targeted metabolomics analysis (peak integration, calibration and quantification) by TMBQ. After filtering out low-quality features, iMAP was used to standardize the data of QC samples, and characteristic variables were obtained based on PLS-DA. Combined with the specific criteria for screening differential metabolites, p<0.05, VIP>1, the metabolite lactic acid with significantly changed relative content in the anxiety group compared with the healthy group was obtained.

[0025] Figure 2 Results showed that the fecal lactic acid content in the anxiety group was significantly lower than that in the healthy group.

[0026] 3. The dosage of Akkermansia muciniphila and lactic acid to increase the 5-HT level is: Akkermansia muciniphila 2x10^8 CFU / d, lactic acid 750 mg / kg / d. Example 2

[0027] SPF mice (purchased from Shanghai Jiesijie Experimental Animal Co., Ltd.) were used for the combined intervention experiment of Akkermansia mucicola and lactic acid.

[0028] Take 1.68x10^10 CFU of Akkermansia muciniphila (ATCC BAA-835), centrifuge at 12000g for 5 minutes at 4 degrees Celsius, take the precipitate and dissolve it in 8.4mL of normal saline (containing 25% glycerol by volume) for later use. The final Akkermansia muciniphila content of the solution is 2x10^8 CFU / 100µL.

[0029] 1.26 g of lactic acid (purchased from MCE) was dissolved in 8.4 mL of normal saline for later use. The final concentration of lactic acid in the solution was 150 mg / ml.

[0030] All SPF mice (n = 28) were treated with antibiotic-reduced drinking water containing mixed antibiotics (ampicillin 100 mg / kg, neomycin 100 mg / kg, metronidazole 100 mg / kg, and vancomycin 50 mg / kg) for 7 days.

[0031] The SPF mice required for the experiment were weighed. Akkermansia muciniphila (n=6) was gavaged at a dose of 2x10^8CFU / d, the lactic acid group (n=6) was gavaged at a dose of 750mg / kg / d, Akkermansia muciniphila and lactic acid (n=6) were gavaged at 2x10^8CFU / d and 750mg / kg / d, respectively, and the control group (n=6) was gavaged with the same amount of solvent. After 7 consecutive days of administration, the serum 5-HT level of mice was detected and recorded using the Elabscience 5-HT kit.

[0032] Figure 3 It showed that supplementation of Akkermansia muciniphila or lactic acid alone could not increase the serum 5-HT level of mice, while the combined use of Akkermansia muciniphila and lactic acid could significantly increase the serum 5-HT level of mice (p<0.05). Example 3

[0033] Germ-free mice (purchased from Shenzhen Jingtuo Biotechnology Co., Ltd.) Akkermansia muciniphila intervention experiment Take 1.12x10^10 CFU of Akkermansia muciniphila (ATCC BAA-835), centrifuge at 12000g for 5 minutes at 4 degrees Celsius, take the precipitate and dissolve it in 5.6mL of normal saline (containing 25% glycerol by volume) for later use. The final Akkermansia muciniphila content of the solution is 2x10^8 CFU / 100µL.

[0034] Take 0.84 g of lactic acid (purchased from MCE) and dissolve it in 5.6 mL of normal saline for later use. The final concentration of lactic acid in the solution is 150 mg / ml.

[0035] The body weight of germ-free mice required for the experiment was weighed, and Akkermansia muciniphila (n=4) was gavaged at a dose of 2x10^8CFU / d, the lactic acid group (n=4) was gavaged at a dose of 750mg / kg / d, Akkermansia muciniphila and lactic acid (n=4) were gavaged at 2x10^8CFU / d and 750mg / kg / d, respectively, and the control group (n=4) was gavaged with an equal amount of solvent. After 7 consecutive days of administration, the serum 5-HT level of mice was detected and recorded using the Elabscience 5-HT kit.

[0036] Figure 4 It showed that supplementation of Akkermansia muciniphila or lactic acid alone could not increase the serum 5-HT level of germ-free mice, while the combined use of Akkermansia muciniphila and lactic acid could significantly increase the serum 5-HT level of germ-free mice (p<0.05).

Claims

1. Use of Akkermansia muciniphila and lactic acid in combination for preparing a drug for increasing the level of 5-hydroxytryptamine in the body.

2. The use according to claim 1, characterized in that The amount of Akkermansia muciniphila is 2x10 8 CFU / d, the lactic acid dosage is 750mg / kg / d.

3. Use of Akkermansia muciniphila and lactic acid in combination for preparing a drug for treating or improving diseases associated with serotonin deficiency.

4. The use according to claim 3, characterized in that The amount of Akkermansia muciniphila is 2x10 8 CFU / d, the lactic acid dosage is 750mg / kg / d.

5. A pharmaceutical composition for increasing the level of serotonin in the body, characterized in that: Contains Akkermansia muciniphila and lactic acid.

6. A pharmaceutical composition for treating or improving diseases related to 5-hydroxytryptamine deficiency, characterized in that: Contains Akkermansia muciniphila and lactic acid.

7. The pharmaceutical composition according to claim 5 or 6, characterized in that It also contains carriers and / or excipients acceptable in pharmaceutical or food industry.

8. The pharmaceutical composition according to claim 5 or 6, characterized in that The dosage form is tablet, capsule, granule, pill, ointment, solution, suspension, emulsion, cream, spray, drop, patch or tube feeding preparation.

Images