CN114410533A - Application of lactobacillus reuteri CCFM1040 in relieving and preventing atherosclerosis - Google Patents
Application of lactobacillus reuteri CCFM1040 in relieving and preventing atherosclerosis Download PDFInfo
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Abstract
The invention discloses an application of lactobacillus reuteri CCFM1040 in relieving and preventing atherosclerosis, and belongs to the technical field of microorganisms. The lactobacillus reuteri CCFM1040 can reduce the level of plasma TMAO; can improve the structure of intestinal flora, recover intestinal flora disorder caused by high choline, improve abundance index of observed species, improve abundance of beneficial bacteria, and reduce risk of constipation, enteritis, lactose intolerance, infection, inflammation, allergy, glycolipid metabolism disorder, obesity, non-alcoholic fatty liver, type II diabetes, and cardiovascular disease. Therefore, the lactobacillus reuteri CCFM1040 has wide application value in preparing products for relieving and preventing atherosclerosis.
Description
Technical Field
The invention relates to application of lactobacillus reuteri CCFM1040 in relieving and preventing atherosclerosis, and belongs to the technical field of microorganisms.
Background
Cardiovascular disease is the leading cause of morbidity and mortality worldwide, and Atherosclerosis (AS) is the pathological basis of cardiovascular disease and the most prominent factor in the induction of cardiovascular disease. Currently, the accepted initiating factors of AS are arterial wall endothelial damage and lipid deposition; the risk factors include hypertension, blood lipid increase, inflammation, oxidized choline, obesity, smoking, etc.
The genome of the intestinal microorganisms and the genome of the human body affect a plurality of important physiological functions of the host such as food digestion and metabolism, immune response and inflammation, nervous activity and the like through interaction with environmental factors. Intestinal microorganisms are considered the second genome of the human body. The interaction between the intestinal flora and its metabolites with the host is essential to maintain the health of the host. Disorders of intestinal microecology are associated with a number of diseases, such as diabetes, obesity, inflammatory bowel disease, neurodegenerative diseases and tumours.
Research shows that intestinal microorganisms mainly affect atherosclerosis through three ways of bacterial infection, cholesterol and lipid metabolism regulation and food and microbial metabolites. TMAO generated by diet-intestinal microorganism-liver-trimethylamine oxide (TMAO) can promote cardiovascular diseases. The microorganism-dependent Trimethylamine (TMA)/TMAO pathway has been shown to be involved in the pathogenesis of cardiovascular disease and is an important diagnostic and therapeutic target for cardiovascular disease. Intervening in the metabolism of intestinal flora may become one of the methods for preventing and treating cardiovascular diseases.
Probiotics have found wide acceptance by consumers as dietary supplements. The supplementary probiotics can directly inject a large amount of beneficial flora into the intestinal tract of a human body, and help to improve the metabolic function of the flora.
Disclosure of Invention
The invention aims to provide application of Lactobacillus reuteri (Lactobacillus reuteri) CCFM1040 in preparing a product for relieving and preventing atherosclerosis.
In one embodiment, the Lactobacillus reuteri CCFM1040 is deposited at the Guangdong province microbial culture collection center 12/14 in 2018, and is deposited at Guangdong province microbial research institute of No. 59 building, No. 5 building, Guangzhou province, Michelia furiosa No. 100, Michelia.
In one embodiment, the amount of Lactobacillus reuteri CCFM1040 in the product is more than or equal to 1 × 106CFU/mL or more than or equal to 1X 106CFU/g。
In one embodiment, the product has at least one of the following effects:
(1) reducing plasma TMAO levels in high choline diet mammals;
(2) increasing the abundance of Bifidobacterium genus in a high choline diet mammal.
In one embodiment, the mammal includes, but is not limited to, a human.
In one embodiment, the product comprises a food or pharmaceutical product.
In one embodiment, the medicament comprises lactobacillus reuteri CCFM1040, a pharmaceutical carrier and/or a pharmaceutical excipient.
In one embodiment, the drug carrier comprises a microcapsule, microsphere, nanoparticle, and/or liposome.
In one embodiment, the pharmaceutical excipient comprises a filler, a binder, a wetting agent, a disintegrant, a lubricant, and/or a flavoring agent.
In one embodiment, the pharmaceutical product is in the form of a powder, granule, capsule, tablet, pill, or oral liquid.
In one embodiment, the food product comprises a solid food product, a liquid food product, a semi-solid food product.
In one embodiment, the food product comprises dairy products, soy products, fruit and vegetable products.
In one embodiment, the dairy product comprises milk, sour cream, cheese.
In one embodiment, the fruit and vegetable product comprises one or more of cucumber, carrot, beet, celery, cabbage and other edible fruits and vegetables.
Has the advantages that:
1. in a high-choline model apoE-/-mouse experiment, the plasma TMAO level can be obviously reduced by taking the lactobacillus reuteri CCFM1040, and the plasma TMAO level of the mice in the lactobacillus reuteri CCFM1040 group is reduced by 30.33 percent compared with that of the mice in the model group;
2. in a high-choline model apoE-/-mouse experiment, the abundance of beneficial bacteria, namely the genus Bifidobacterium is improved by taking the lactobacillus reuteri CCFM1040, the abundance of the beneficial bacteria, namely the genus Bifidobacterium is improved by 4.7 times compared with that of the model group, the abundance of the beneficial bacteria, namely the genus Bifidobacterium is improved, and the risks of constipation, enteritis, lactose intolerance, infection, inflammation, allergy, glycolipid metabolic disorder, obesity, nonalcoholic fatty liver, type II diabetes and cardiovascular diseases are further reduced.
3. The lactobacillus reuteri CCFM1040 has the function of regulating intestinal flora, can effectively reduce the level of plasma TMAO, expands the application range of the lactobacillus reuteri as probiotics, and has very important significance for deeply exploring the functions of the probiotics and developing the probiotics with higher health care value.
Biological material preservation
Lactobacillus reuteri (Lactobacillus reuteri) CCFM1040, which is classified and named as Lactobacillus reuteri, has been preserved in Guangdong province microorganism strain preservation center in 12 and 14 months in 2018, has the preservation address of No. 59 building, No. 5 building, Guangdong province microorganism research institute of Michelia Tokoro No. 100 college in Guangzhou city, and has the preservation number of GDMCC No. 60515.
Drawings
FIG. 1 shows the colony morphology of Lactobacillus reuteri CCFM 1040.
FIG. 2 is a graph of the effect of Lactobacillus reuteri CCFM1040 on plasma TMAO in choline-fed apoE-/-mice; wherein P <0.01, P < 0.0001.
FIG. 3 is a graph of the effect of Lactobacillus reuteri CCFM1040 on the cecum Bifidobacterium genus of choline-fed apoE-/-mice; wherein P < 0.05.
FIG. 4 is a graph of the effect of different Lactobacillus reuteri on plasma TMAO in choline-fed apoE-/-mice; wherein P <0.01, P < 0.0001.
Detailed Description
Example 1: effect of Lactobacillus reuteri CCFM1040 on plasma TMAO
The colony morphology of Lactobacillus reuteri CCFM1040 is shown in FIG. 1.
The influence and action pathway of lactobacillus reuteri on hyperuricemia are researched by lactobacillus reuteri CCFM1040 and lactobacillus reuteri FHNXY12L1 (lactobacillus reuteri FHNXY12L1 reported in Nissan. [ D]Jiangnan university, 2021) was cultured in MRS medium at 37 deg.C, and the cells were collected and resuspended in physiological saline to a concentration of 1X 109CFU/mL of bacterial suspension.
24 healthy female apolipoprotein E gene knockout mice (apoE-/-mice) weighing 18-20g were taken, acclimated for 1 week, and randomly divided into 4 groups of 6 apoE-/-mice per group: control group (Control), model group (Choline ), Lactobacillus reuteri CCFM1040 group (CCFM1040), Lactobacillus reuteri FHNXY12L1 Control group (FHNXY12L 1). The grouping and treatment method of experimental animals is shown in table 1:
TABLE 1 grouping and treatment method of experimental animals
| Group of | Number/group only | Time of treatment | Feed stuff | Processing method |
| Control group | 6 | 2-24 weeks | Common feed | Gavage 0.2mL physiological saline every day |
| Model set | 6 | 2-24 weeks | Choline feed | Gavage 0.2mL physiological saline every day |
| CCFM1040 | 6 | 2-24 weeks | Choline feed | Gavage 0.2mL per day of Lactobacillus reuteri CCFM1040 bacterial suspension |
| FHNXY12L1 | 6 | 2-24 weeks | Choline feed | Gavage 0.2mL per day of Lactobacillus reuteri FHNXY12L1 bacterial suspension |
Week 2-24: control groups of apoE-/-mice were fed with normal feed, and the remaining three groups of apoE-/-mice were fed with Choline feed. Common feed (LAD 3001M, choline content 0.1%) and choline feed (LAD 3001M, choline content 1.0%) were purchased from Nantong Telofil feed science and technology Co.
Before the end of the experiment, 24 apoE-/-mice were fasted for 4h and bled via periorbital capillaries. The blood sample is centrifuged for 15min at 4000 Xg and 4 ℃, the supernatant is taken and frozen in a refrigerator at-80 ℃,20 mu L of the plasma sample is taken and added with 80 mu L (V: V, 1: 4) of acetonitrile to precipitate the protein in the plasma sample, and simultaneously, the deuterated trimethylamine oxide (d9-TMAO) with the final concentration of 2.0 mu M is added into the plasma sample as an internal standard. Mixing, standing at-80 deg.C for 2h, at 4 deg.C for 12000 Xg for 15min, sucking supernatant into a sample bottle, storing in-80 deg.C refrigerator, and measuring plasma TMAO level by HPLC-MS/MS.
As shown in FIG. 2, the plasma TMAO of the apoE-/-mice in the model group was significantly higher than that of the control group, and the plasma TMAO of the apoE-/-mice in the L.gastri CCFM1040 was significantly reduced by 30.33% compared with that of the apoE-/-mice in the model group. Lactobacillus gasseri FHNXY12L1 had no significant effect on the level of TMAO in plasma.
Example 2: effect of Lactobacillus reuteri CCFM1040 on the genus Bifidobacterium in the intestinal tract of mice
Grouping, modeling and treatment of apoE-/-mice were performed as in example 3.
After the experiment is finished, 24 apoE-/-mice are fasted and water-deprived for 12h, the cecum is taken after being anesthetized by injecting 10% chloral hydrate solution into the abdominal cavity, cecum DNA is extracted according to the method of the fecal DNA kit, and 16S rDNA flora analysis is carried out on a V3-V4 region of the cecum by using a second-generation sequencer.
The results of the experiment are shown in FIG. 3. The relative abundance of the caecum Bifidobacterium of the model group apoE-/-mice is obviously lower than that of the control group, compared with the model group apoE-/-mice, the relative abundance of the caecum Bifidobacterium of the lactobacillus gasseri CCFM1040 is obviously improved by 4.7 times, compared with the model group apoE-/-mice, the relative abundance of the caecum Bifidobacterium of the lactobacillus gasseri CCFM1040 is further improved by 4.7 times, and the risks of constipation, enteritis, lactose intolerance, infection, inflammation, allergy, metabolic glycolipid disorder, obesity, nonalcoholic fatty liver, type II diabetes and cardiovascular disease are further reduced.
Example 3: comparison of the Effect of different Lactobacillus reuteri on apoE-/-mouse plasma TMAO
As in example 1, 16 different strains of Lactobacillus reuteri were respectively used for gavage of the apoE-/-mice to detect plasma TMAO, as shown in FIG. 4, only CCFM1040 had the ability to significantly reduce the plasma TMAO of the apoE-/-mice by 30.33%, and the other Lactobacillus reuteri could not significantly reduce the plasma TMAO of the apoE-/-mice.
Example 4: fermented food prepared from Lactobacillus reuteri CCFM1040
Selecting fresh vegetables, washing, juicing, carrying out high-temperature instant sterilization, carrying out high-temperature heat sterilization at 140 ℃ for 2 seconds, immediately cooling to 37 ℃, and inoculating the lactobacillus reuteri CCFM1040 microbial inoculum starter prepared by the invention to ensure that the concentration of the starter reaches 108More than CFU/mL, and storing at 4 ℃ in a refrigerated way, thus obtaining the fruit and vegetable beverage containing the viable bacteria of the lactobacillus reuteri CCFM 1040.
The lactobacillus reuteri CCFM1040 can be used for fermentation production and preparation of other fermented foods, wherein the fermented foods comprise solid foods, liquid foods and semi-solid foods. The fermented food comprises dairy products, bean products and fruit and vegetable products, wherein the dairy products comprise milk, sour cream and cheese; the fruit and vegetable products comprise cucumber, carrot, beet, celery and cabbage products.
When the fermented food prepared by the method is fed to a high-choline model mouse, the level of the blood plasma TMAO can be reduced; can improve the structure of intestinal flora, increase the abundance of beneficial bacteria, and reduce the risk of constipation, enteritis, lactose intolerance, infection, inflammation, allergy, glycolipid metabolic disorder, obesity, non-alcoholic fatty liver, type II diabetes, and cardiovascular diseases.
Although the present invention has been described with reference to the preferred embodiments, it should be understood that various changes and modifications can be made therein by those skilled in the art without departing from the spirit and scope of the invention as defined in the appended claims.
Claims (10)
1. The application of Lactobacillus reuteri (Lactobacillus reuteri) CCFM1040 in preparing products for relieving and preventing atherosclerosis; the lactobacillus reuteri CCFM1040 is preserved in Guangdong province microbial strain preservation center in 2018, 12 and 14 months, the preservation address is Guangzhou city, Jielizhou 100 college No. 59 building, 5 building, Guangdong province microbial research institute, and the preservation number is GDMCC No. 60515.
2. The use according to claim 1, wherein the amount of Lactobacillus reuteri CCFM1040 is 1 x 10 or more6CFU/mL or more than or equal to 1X 106CFU/g。
3. Use according to claim 1, wherein the product has at least one of the following effects:
(1) reducing plasma trimethylamine oxide levels in a high choline diet mammal;
(2) increasing the abundance of Bifidobacterium genus in a high choline diet mammal.
4. The use according to claim 3, wherein said mammal includes but is not limited to a human.
5. Use according to claim 1, wherein the product comprises a food or a pharmaceutical product.
6. The use according to claim 5, wherein the medicament comprises Lactobacillus reuteri CCFM1040, a pharmaceutical carrier and/or a pharmaceutical excipient.
7. Use according to claim 6, wherein the drug carrier comprises microcapsules, microspheres, nanoparticles and/or liposomes.
8. The use of claim 6, wherein the medicament is in the form of a powder, granules, capsules, tablets, pills or oral liquid.
9. Use according to claim 5, wherein the food product comprises a solid food product, a liquid food product, a semi-solid food product.
10. Use according to claim 9, wherein the food product comprises dairy products, soy products, fruit and vegetable products.
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