A strain of Limosilactobacillus mucosae for regulating liver lipid metabolism and its application

Abstract

The present invention discloses a Lactobacillus mucosae that regulates liver lipid metabolism and its application, belonging to the field of microbial technology. The present invention has screened out a strain of Lactobacillus mucosae CCFM1277, with the preservation number of GDMCC No: 62777. The Lactobacillus mucosae CCFM1277 provided by the present invention has the effect of alleviating non-alcoholic fatty liver, specifically manifested in: effectively reducing lipid deposition in the liver of individuals with non-alcoholic fatty liver caused by a high-fat diet, reducing the size of adipocytes, and upregulating the enzymes related to fat decomposition in the liver, including lipoprotein lipase LPL, adipose triglyceride lipase ATGL, hormone-sensitive lipase HSL, serine hydrolase MGL, etc., as well as the mRNA expression of the key enzyme CPT-1 of fatty acid β-oxidation.

Description

Technical Field

[0001] The present invention relates to a mucosal Lactobacillus mucosae for regulating liver lipid metabolism and its application, belonging to the field of microbial technology. Background Art

[0002] With the change of diet structure towards high-fat and high-protein, the incidence of non-alcoholic fatty liver disease (NAFLD) induced thereby has been increasing year by year and shows a trend of getting younger. NAFLD has become one of the most common chronic liver diseases worldwide. For example, the prevalence of NAFLD in the United States has increased significantly in recent years and now accounts for one-fourth of the total population. In recent years, the incidence of NAFLD in China has also been rising year by year and has become the second most common chronic liver disease, second only to viral hepatitis. Among obese people, the prevalence of NAFLD is even higher. It has been found that the mortality rate of NAFLD patients is significantly higher than that of the normal population. NAFLD is the main inducing factor of various chronic liver diseases, such as liver cirrhosis and liver cancer. Due to the relatively complex pathogenesis of NAFLD, there is currently no ideal treatment drug or regimen.

[0003] Intestinal microbiota includes a variety of microorganisms (mainly bacteria), which contribute to digestion, energy extraction, and antagonizing the colonization of pathogenic bacteria. It can also stimulate the immune system of the gastrointestinal tract by competitively taking up nutrients and space, thereby reducing pathogens. Intestinal microbiota dysbiosis refers to the disruption of the normal intestinal flora, which can be caused by a series of environmental, immune, or host factors, as well as changes in bile flow, gastric pH, or intestinal motility disorders. A large amount of evidence shows that intestinal microbiota dysregulation is associated with the pathogenesis of human liver diseases and plays an important role in NAFLD and its related metabolic disorders. Therefore, the use of probiotics to intervene in non-alcoholic fatty liver has become a recent research hotspot. A large amount of evidence has shown that probiotics can relieve NAFLD by regulating the liver's fat decomposition ability and reducing liver lipid deposition.

[0004] Chinese Patent Application CN 113265361A discloses a compound probiotic preparation for relieving non-alcoholic fatty liver, its preparation method and application, but does not study the relieving effect of single bacteria on non-alcoholic fatty liver. This study focuses on the relieving effect of a single probiotic on NAFLD and obtains a strain of Lactobacillus mucosae with a good relieving effect. Summary of the Invention

[0005] The object of the present invention is to overcome the defects of the prior art. Through in-vivo research on animal experiments, a Limosilactobacillus mucosae CCFM1277 with the ability to prevent non-alcoholic fatty liver is obtained. It can reduce lipid deposition in the liver, decrease the size of adipocytes, up-regulate the expression of enzymes related to fat decomposition in the liver, such as lipoprotein lipase (LPL), adipose triglyceride lipase (ATGL), hormone-sensitive lipase (HSL), and serine hydrolase (MGL), etc., and up-regulate the expression of the key enzyme of fatty acid β-oxidation, carnitine palmitoyltransferase-1 (CPT-1), etc.

[0006] To achieve the above object, the present invention provides a Limosilactobacillus mucosae CCFM1277, which was deposited in the Guangdong Provincial Culture Collection Center of Microorganisms on September 26, 2022, with the deposit number GDMCC No: 62777, and the deposit address is the 5th floor of Building 59, No. 100 Compound, Xianlie Middle Road, Guangzhou.

[0007] In one embodiment, the Limosilactobacillus mucosae CCFM1277 has the following biological characteristics:

[0008] Cell characteristics: It is milky yellow;

[0009] Colony characteristics: On the MRS solid plate, the colonies are milky yellow, raised, with irregular edges, and are Gram-positive bacteria;

[0010] Growth characteristics: Under the condition of constant temperature and aerobic at 37°C, when cultured in MRS medium for about 16 h, it reaches the late logarithmic phase.

[0011] The present invention also provides the application of the Limosilactobacillus mucosae CCFM1277 in the preparation of drugs for preventing and / or treating non-alcoholic fatty liver.

[0012] In one embodiment, in the drug, the viable count of Limosilactobacillus mucosae CCFM1277 is not less than 1×10 10 CFU / g.

[0013] In one embodiment, the drug contains the Limosilactobacillus mucosae CCFM1277, a drug carrier, and / or pharmaceutical excipients.

[0014] The present invention also provides the application of the Limosilactobacillus mucosae CCFM1277 in the preparation of health products that help control body fat and / or help maintain a healthy blood lipid level.

[0015] The present invention also provides a product containing the Limosilactobacillus mucosae CCFM1277.

[0016] In one embodiment, the product is a fermenting agent containing Limosilactobacillus mucosae CCFM1277.

[0017] In one embodiment, the starter culture is obtained by culturing Lactobacillus mucosae CCFM1277 in a culture medium for a period of time, collecting the bacterial cells in the cell culture broth, and directly using it as a starter culture or obtaining a starter culture after treating the bacterial cells.

[0018] In one embodiment, the treatment includes, but is not limited to, one or more of the following treatment methods: washing, adding a protective agent, drying, etc.

[0019] In one embodiment, the method for preparing the starter culture is as follows: inoculate Lactobacillus mucosae CCFM1277 into a culture medium at an inoculum amount of 1-5% of the total mass of the culture medium, culture at 37 °C for 18 h to obtain a culture broth; centrifuge the culture broth to obtain bacterial cells; resuspend the bacterial cells with physiological saline to obtain a starter culture.

[0020] In one embodiment, the culture medium is MRS culture medium.

[0021] In one embodiment, in the product, the viable count of Lactobacillus mucosae CCFM1277 is not less than 1×10 10 CFU / g.

[0022] In one embodiment, the product includes food, health products or drugs.

[0023] In one embodiment, the drug contains Lactobacillus mucosae CCFM1277, a drug carrier and / or a pharmaceutical excipient.

[0024] In one embodiment, the food includes health foods containing Lactobacillus mucosae CCFM1277.

[0025] In one embodiment, the food includes dairy products, soy products, meat products or fruit and vegetable products produced using the starter culture of Lactobacillus mucosae CCFM1277.

[0026] In addition, the present invention also provides the application of Lactobacillus mucosae CCFM1277 in food additives, and the application includes, but is not limited to, being used as a food starter culture.

[0027] Beneficial Effects: This study demonstrates that Lactobacillus mucosae CCFM1277 reduces lipid deposition in the liver, decreases adipocyte size, upregulates the expression of enzymes involved in fat breakdown in the liver, such as lipoprotein lipase (LPL), adipose triglyceride lipase (ATGL), hormone-sensitive lipase (HSL), and serine hydrolase (MGL), and upregulates CPT-1, a key enzyme in fatty acid β-oxidation. This strain exhibits enhanced therapeutic or preventive effects on non-alcoholic fatty liver disease (NAFLD). Therefore, probiotic foods, health products, and pharmaceuticals for the prevention and treatment of NAFLD can be prepared, demonstrating broad potential for application.

[0028] Biomaterial Deposit

[0029] Mucosal mucus lactobacillus CCFM1277, classified as Limosilactobacillus mucosae, was deposited in the Guangdong Provincial Microbial Culture Collection on September 26, 2022. The deposit address is 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, and the deposit number is GDMCC No: 62777. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 The oil red staining of liver tissues in each group shows that Lactobacillus mucosa CCFM1277 reduces fat deposition in the liver; in the figure: different letters above the bar graph indicate significant differences (P<0.05).

[0031] Figure 2 The HE staining of perirenal adipose tissue in each group showed that Lactobacillus mucosa CCFM1277 reduced the size of adipocytes; in the figure: different letters above the bar graph indicate significant differences (P<0.05).

[0032] Figure 3 The figure shows the effect of Lactobacillus mucosa CCFM1277 on the relative expression of lipoprotein lipase LPL mRNA in each group of mice, indicating that Lactobacillus mucosa CCFM1277 can significantly increase the relative expression of LPL mRNA in mice; in the figure: in the figure: different letters above the bar graph indicate significant differences (P<0.05).

[0033] Figure 4 The relative expression levels of adipose triglyceride lipase ATGL mRNA in each group indicate that Lactobacillus mucosae CCFM1277 can increase the relative expression of ATGL mRNA in mice; In the figure: In the figure: Different letters above the bar graph indicate significant differences (P<0.05).

[0034] Figure 5Indicates the effect of Lactobacillus mucosae CCFM1277 on the change of hormone-sensitive lipase (HSL) in mice, showing that Lactobacillus mucosae CCFM1277 can significantly increase the relative expression of HSL mRNA in mice; In the figure: Different letters above the bar graphs indicate significant differences (P < 0.05).

[0035] Figure 6 Indicates the effect of Lactobacillus mucosae CCFM1277 on murine serine hydrolase MGL, showing that Lactobacillus mucosae CCFM1277 can significantly increase the relative expression of MGL mRNA in mice; In the figure: Different letters above the bar graphs indicate significant differences (P < 0.05).

[0036] Figure 7 Indicates the effect of Lactobacillus mucosae CCFM1277 on carnitine palmitoyl transferase CPT-1 in mice, showing that Lactobacillus mucosae CCFM1277 can significantly increase the relative expression of CPT-1 mRNA in mice; In the figure: Different letters above the bar graphs indicate significant differences (P < 0.05). Detailed implementation mode

[0037] The present invention can be better understood through the following embodiments.

[0038] In the present invention, unless otherwise specified, "%" or percentage used to indicate concentration or proportion is weight percentage.

[0039] The present invention relates to the following culture media:

[0040] MRS liquid medium: 10 g of tryptone, 10 g of beef extract, 5 g of yeast powder, 20 g of glucose, 2 g of diammonium hydrogen citrate, 5 g of sodium acetate, 2 g of dipotassium hydrogen phosphate, 0.5 g of magnesium sulfate heptahydrate, 0.25 g of manganese sulfate monohydrate, 1 mL of Tween 80, and add water to 1000 mL.

[0041] MRS solid medium is obtained by adding 1.5% agar based on the total weight of the liquid medium on the above basis.

[0042] Example 1: Collection, isolation and identification of Lactobacillus mucosae CCFM1277

[0043] Take 1 g of fecal sample from a 5-year-old male child in Lianzhou, Guangdong. After gradient dilution, it was spread on MRS solid medium and cultured at 37 °C in an aerobic environment for 72 h. The colony morphology was observed and recorded. The colonies were picked and streaked for purification, and then cultured in MRS liquid medium at 37 °C for 48 h. The obtained colonies were subjected to Gram staining and the strain morphology was recorded. Gram-negative strains and Gram-positive cocci in the colonies were discarded, and Gram-positive bacilli were selected. After catalase analysis, catalase-positive strains were discarded, and catalase-negative strains were retained. Fructose-6-phosphate kinase detection was used to discard negative strains. All the obtained strains were identified as Lactobacillus mucosae by 16S rDNA sequencing and named CCFM1277. The obtained Lactobacillus mucosae was subcultured, and the cells were collected and centrifuged at 3000 rpm for 10 min in a centrifuge tube and washed 3 times. The obtained cells were added to the matrix protectant for cryopreservation. Another strain of Lactobacillus mucosae screened in the same batch was named FGSYC17L3.

[0044] 16S rDNA amplification conditions: 95 °C for 5 min; 35 cycles (95 °C for 30 s, 55 °C for 30 s, 72 °C for 2 min); 72 °C for 10 min

[0045] Amplification primers: 27F: (5’-AGAGTTTGATCCTGGCTCAG-3’), 1492R: (5’-TACGGCTACCTTGTTACGACTT-3’). The purification and sequence alignment process of the amplification products were carried out according to the method described in the literature (Turroni Fetal. Exploring the Diversity of the Bifidobacterial Population in the Human Intestinal Tract[J]. Appl Environ Microb. 2009;75(6):1534–45).

[0046] Example 2: Preparation of Lactobacillus mucosae CCFM1277 bacterial suspension

[0047] (1) Activation culture:

[0048] Use MRS liquid medium and statically culture it in a 37 °C ordinary incubator.

[0049] Culture objective: Take the cryopreserved cells, pick a single colony into liquid MRS liquid medium, and statically culture it in a 37 °C ordinary incubator for about 24 h to activate Lactobacillus mucosae CCFM1277.

[0050] (2) Primary culture:

[0051] Use MRS liquid medium and statically culture it in a 37°C ordinary incubator.

[0052] Cultivation objective: Transfer the activated Lactobacillus mucosae CCFM1277 to MRS liquid medium at an inoculation amount of 1% (by volume of the medium) and passage it for two generations.

[0053] (3) Secondary culture:

[0054] Transfer the Lactobacillus mucosae CCFM1277 that has undergone primary culture to 1L of MRS liquid medium at an inoculation amount of 1% (by volume of the medium), statically culture it in a 37°C ordinary incubator for about 24 h, and collect the bacterial cells. Wash them twice with PBS at pH 7.4, then resuspend them in a 30% glycerol solution. The viable cell count is 2.2×10 10 CFU / mL. When used in subsequent experiments, centrifuge to discard the glycerol, wash once with physiological saline, and then resuspend in physiological saline.

[0055] Example 3: Determination of the gastrointestinal fluid tolerance of Lactobacillus mucosae CCFM1277

[0056] Method for determining the survival rate in simulated gastrointestinal fluid: The artificial simulated gastrointestinal fluid needs to be freshly prepared. Dissolve pepsin (Sinopharm Shanghai Test, 64008860) in PBS at pH 3.0 to a final concentration of 3 g / L, and prepare simulated gastric fluid after filtering through a 0.22 μm filter membrane. Dissolve trypsin (Shanghai Sangon Biotech Co., Ltd., A003319 - 0001) in PBS at pH 8.0 to a final concentration of 1 g / L, and prepare simulated intestinal fluid after filtering through a 0.22 μm filter membrane. Centrifuge the cultured Lactobacillus mucosae at 6000 rpm for 10 min at 4°C, collect the bacterial sludge, resuspend it with 0.85% physiological saline, and adjust the bacterial liquid density to 1×10 9 CFU / mL in simulated gastric fluid (pH 3.0). After mixing, place it in an incubator at 37°C for 2 h and then count the viable cell number. Take 1 mL of the bacterial liquid treated with simulated gastric fluid and add it to 9 mL of simulated intestinal fluid (pH 8.0), mix well, and culture at 37°C. After 4 h, detect the viable cell number. The percentage of the ratio of the viable cell number after treatment to the initial viable cell number is the survival rate.

[0057] The results show that the survival rate of CCFM1277 in simulated gastric fluid is 87.66%, and the survival rate in simulated intestinal fluid is 78.81%

[0058] Example 4: Improvement of liver fat accumulation in non - alcoholic fatty liver mice by Lactobacillus mucosae CCFM1277

[0059] 1. Experimental animals

[0060] Male SPF-grade C57BL / 6J mice (4 weeks old, weighing 20 - 23 g) were purchased from Vital River Laboratory Animal Technology Co., Ltd., Zhejiang, China. The mice were housed in polypropylene cages with 8 mice per group. The cages were equipped with food and water. The temperature was controlled at (22 °C), and the relative humidity was (50 ± 10%). The mice had free access to water. Control mice were fed a standard diet, while the model group and the CCFM1277 group were fed a 60% high-fat diet.

[0061] 2. Experimental methods

[0062] (1) Establishment of non-alcoholic fatty liver mouse model

[0063] Control mice were fed a standard diet, while the model group and the CCFM1277 group were fed a 60% high-fat diet. The mice had free access to water for 12 weeks. The body weight of the mice was measured weekly. After 12 weeks, if the body weight of the model group was 25% higher than that of the control group, the model was considered successfully established.

[0064] (2) Experimental grouping and administration

[0065] Thirty-two C57BL / 6J mice were randomly divided into 4 groups: control group, model group, CCFM1277 treatment group (Lactobacillus mucosae CCFM1277) and FGSYC17L3 treatment group (Lactobacillus mucosae FGSYC17L3), with 8 mice in each group. The administration method for the Lactobacillus mucosae treatment groups was intragastric administration, and the intragastric dose was 4×10 10 CFU / 100 μL / mouse / day. During the modeling period, the mice in the Lactobacillus mucosae treatment groups were intragastrically administered Lactobacillus mucosae every day, while the normal group and the model group were intragastrically administered normal saline as a control until the end of the experiment.

[0066] During the modeling period, the body weight of the mice was measured weekly.

[0067] After 12 weeks of modeling, since the body weight of the model group was 25.81% higher than that of the control group, the model was considered successfully established. After sacrificing the mice, some liver tissues were dissected, washed with normal saline to remove the blood, fixed with 4% paraformaldehyde for 24 h, dehydrated, embedded in paraffin, and frozen-sectioned. Next, the ultra-thin sections (4 μm) were rinsed with isopropanol and then stained with Oil Red O staining solution. The stained sections were covered with a cover slip using neutral gum as an adhesive. A pathological section scanner was used to record microscopic photos.

[0068] In the livers of non-alcoholic fatty liver mice, the accumulation of fat increased. In the Oil Red sections, an increase in the number and area of lipid droplets could be observed, which could reflect the severity of non-alcoholic fatty liver. The software Image Pro Plus was used to analyze the fat proportion in the sections, as Figure 1As shown, compared with the blank group, the fat percentage in the model group increased significantly from 2.74% to 22.61% (p < 0.05). The fat area in the mucosa mucilaginous lactobacillus CCFM1277 group was 10.84%, which could significantly alleviate the fat accumulation caused by a high-fat diet (p < 0.05), while FGSYC17L3 had no such effect.

[0069] Example 5: Mucosa mucilaginous lactobacillus CCFM1277 improves the perirenal fat accumulation in non-alcoholic fatty liver mice

[0070] The implementation methods of modeling and grouping treatment were the same as those in Example 4.

[0071] Dissect a part of the perirenal fat tissue, rinse the blood clean with physiological saline, fix it with 4% paraformaldehyde for 24 hours, dehydrate, embed it in paraffin, and section it. Next, take ultra-thin sections (4 μm), and stain them with hematoxylin and eosin (HE). The stained sections were covered with a cover glass using neutral gum as an adhesive. A pathological section scanner was used to record microscopic photos.

[0072] The fat cells of non-alcoholic fatty liver mice were larger and arranged loosely. The software FIJI was used to analyze the size of fat cells in the sections. As Figure 2 shown, compared with the blank group, the fat cell area in the model group increased significantly from 1020.09 μm 2 to 2928.23 μm 2 (p < 0.05), while that in the mucosa mucilaginous lactobacillus CCFM1277 group was 2380.25 μm 2 , which could significantly alleviate this situation (p < 0.05), while FGSYC17L3 had no such effect.

[0073] Example 6: Mucosa mucilaginous lactobacillus CCFM1277 regulates the relative expression level of lipoprotein lipase mRNA in non-alcoholic fatty liver mice

[0074] The implementation methods of modeling and grouping treatment were the same as those in Example 4.

[0075] Twenty-four hours after the last gavage, take some liver tissues and store them in a -80°C refrigerator for subsequent experiments. When conducting the experiment, take out the liver tissues, use the Trizol method to extract the total RNA of the fat tissues, reverse transcribe it into cDNA, and then use quantitative PCR (qPCR) to detect the relative expression level of lipoprotein lipase (LPL) mRNA in the mouse liver tissues.

[0076] Lipoprotein lipase can catalyze the hydrolysis of triglycerides in lipoproteins and participate in the metabolism of chylomicrons and very low-density lipoproteins. The expression of LPL in the liver can promote the decomposition of fat in the liver and avoid liver lipid deposition. As Figure 3As shown, compared with the control group, the relative mRNA expression level of LPL in the model group mice decreased to 0.64, while Lactobacillus mucosae CCFM1277 significantly increased the transcriptional level of LPL in mice, which could be increased to 2.78 (p<0.05).

[0077] Example 7: Regulation of relative mRNA expression level of adipose triglyceride lipase in non-alcoholic fatty liver mice by Lactobacillus mucosae CCFM1277

[0078] The implementation methods of modeling, grouping and treatment were the same as those in Example 4.

[0079] At 24 h after the last gavage, part of the liver tissue was taken and stored in a -80 °C refrigerator for subsequent experiments. When conducting the experiment, the liver tissue was taken out, and the total RNA of adipose tissue was extracted by the Trizol method. After reverse transcription into cDNA, the relative mRNA expression level of adipose triglyceride lipase (ATGL) in mouse liver tissue was detected by quantitative PCR (qPCR).

[0080] Adipose triglyceride lipase can specifically hydrolyze the first ester bond of triglyceride, promote the decomposition of triglyceride in the liver, and reduce liver lipid deposition. As Figure 4 shown, compared with the control group, the relative mRNA expression level of ATGL in the model group mice decreased to 0.33, while Lactobacillus mucosae CCFM1277 significantly increased the transcriptional level of ATGL in mice, which was increased to 1.18 (p<0.05), while FGSYC17L3 had no such effect.

[0081] Example 8: Regulation of relative mRNA expression level of hormone-sensitive lipase in non-alcoholic fatty liver mice by Lactobacillus mucosae CCFM1277

[0082] The implementation methods of modeling, grouping and treatment were the same as those in Example 4.

[0083] At 24 h after the last gavage, part of the liver tissue was taken and stored in a -80 °C refrigerator for subsequent experiments. When conducting the experiment, the liver tissue was taken out, and the total RNA of adipose tissue was extracted by the Trizol method. After reverse transcription into cDNA, the relative mRNA expression level of hormone-sensitive lipase (HSL) in mouse liver tissue was detected by quantitative PCR (qPCR).

[0084] Hormone-sensitive lipase can directly act on fat, hydrolyze triglyceride into diglyceride, promote the decomposition of triglyceride in the liver, and reduce liver lipid deposition. As Figure 5 shown, compared with the control group, the relative mRNA expression level of HSL in the model group mice decreased to 0.43, while in the Lactobacillus mucosae CCFM1277 group it increased to 3.87, which significantly increased the transcriptional level of HSL in mice (p<0.05).

[0085] Example 9: Regulation of relative mRNA expression levels of serine hydrolase in mice with non-alcoholic fatty liver by Lactobacillus mucosae CCFM1277

[0086] The implementation methods of modeling, grouping and treatment were the same as those in Example 4.

[0087] At 24 h after the last gavage, a part of liver tissue was taken and stored in a -80 °C refrigerator for subsequent experiments. When conducting the experiment, the liver tissue was taken out, and total RNA of adipose tissue was extracted using the Trizol method. After reverse transcription into cDNA, the relative mRNA expression levels of serine hydrolase (MGL) in mouse liver tissue were detected by quantitative PCR (qPCR).

[0088] Serine hydrolase can decompose triglycerides into free fatty acids and glycerol, promote the decomposition of triglycerides in the liver, and reduce liver lipid deposition. As Figure 6 shown, compared with the control group, the relative mRNA expression level of MGL in the model group mice was significantly decreased to 0.28 (p < 0.05), while that in the Lactobacillus mucosae CCFM1277 group was significantly increased to 2.07, indicating that Lactobacillus mucosae CCFM1277 can well increase the transcription level of HSL, while FGSYC17L3 has no such effect.

[0089] Example 10: Regulation of relative mRNA expression levels of carnitine palmitoyltransferase in mice with non-alcoholic fatty liver by Lactobacillus mucosae CCFM1277

[0090] The implementation methods of modeling, grouping and treatment were the same as those in Example 2.

[0091] At 24 h after the last gavage, a part of liver tissue was taken and stored in a -80 °C refrigerator for subsequent experiments. When conducting the experiment, the liver tissue was taken out, and total RNA of adipose tissue was extracted using the Trizol method. After reverse transcription into cDNA, the relative mRNA expression levels of carnitine palmitoyltransferase (CPT-1)

[0092] in mouse liver tissue were detected by quantitative PCR (qPCR).

[0093] Carnitine palmitoyltransferase can transfer long-chain acyl coenzyme A outside the mitochondria into the mitochondrial matrix for β-oxidation. It is the rate-limiting enzyme for fatty acid oxidation. By catalyzing the entry of fatty acids into the mitochondria for oxidation, it maintains the balance of blood glucose and energy supply. As Figure 7As shown, the relative mRNA expression level of CPT-1 in the model group mice decreased to 0.75, while Lactobacillus mucosae CCFM1277 had a significant effect of increasing the transcriptional level of CPT-1 in mice. The transcriptional level of CPT-1 in the Lactobacillus mucosae CCFM1277 group increased significantly to 3.08 (p<0.05), while FGSYC17L3 had no such effect.

[0094] The above results indicate that Lactobacillus mucosae CCFM1277 can effectively reduce lipid deposition in the liver, reduce the size of adipocytes, up-regulate the enzymes related to fat decomposition in the liver: lipoprotein lipase LPL, adipose triglyceride lipase ATGL, hormone-sensitive lipase HSL, and serine hydrolase MGL, as well as up-regulate the mRNA expression of key enzymes such as CPT-1 in fatty acid β-oxidation. It can be used to prepare drugs or health products for the prevention and treatment of non-alcoholic fatty liver, or produce foods beneficial for weight loss, such as food additives for probiotic beverages, acid soymilk, fermented jelly, fermented tea beverages, or dairy products (such as yogurt, cheese products, lactic acid bacteria, milk powder).

[0095] Example 11: Preparation of tablets containing Lactobacillus mucosae CCFM1277

[0096] The basic steps of the specific production process are as follows: strain activation → subculture → cell collection → preparation of cell suspension → freeze-drying → total mixing → tabletting

[0097] 1. Strain activation: Lactobacillus mucosae CCFM1277 was statically cultured in MRS liquid medium at an inoculation amount of 1% (by volume of the medium) in a 37°C ordinary incubator, and continuously activated for two generations.

[0098] 2. Subculture: The activated Lactobacillus mucosae CCFM1277 was transferred to 1 L of MRS liquid medium at an inoculation amount of 1% (by volume of the medium) for subculture, and statically cultured in a 37°C ordinary incubator for 24 h.

[0099] 3. Cell collection and preparation of cell suspension: After the subculture was completed, the cells were collected by low-temperature centrifugation at 4°C, washed twice with PBS (pH 7.4), and then made into a cell suspension of 1×10 10 CFU / mL with a 13% defatted milk aqueous solution by weight.

[0100] 4. Freeze-drying: The cell suspension was made into cell powder according to the conventional freeze-drying process.

[0101] 5. Total mixing: 2% stearic acid by weight of the cell powder was added as a lubricant, and 3% CMC-Na was added as a binder, and mixed evenly.

[0102] 6. Tabletting: Tablets were pressed by a tabletting machine according to the conventional tabletting process.

[0103] Example 12: Preparation of a powder containing the bacterial agent of Lactobacillus mucosae CCFM1277

[0104] The basic steps of the specific production process are as follows: strain activation → subculture → cell collection → preparation of cell suspension → freeze-drying. The steps of strain activation, subculture, cell collection, and preparation of cell suspension are as described above.

[0105] Freeze-drying: The cell suspension is made into a freeze-dried cell powder according to the conventional freeze-drying process.

[0106] Although the present invention has been disclosed above with preferred embodiments, it is not intended to limit the present invention. Any person familiar with this technology can make various modifications and alterations without departing from the spirit and scope of the present invention. Therefore, the protection scope of the present invention should be defined by the claims.

Claims

1. Lactobacillus mucosa ( Limosilactobacillus mucosae ) Use of CCFM1277 in the preparation of a drug for preventing and / or treating non-alcoholic fatty liver disease, characterized in that: The Limosilactobacillus mucosae CCFM1277 was deposited at the Guangdong Microbial Culture Collection Center on September 26, 2022, with the deposit number GDMCC No: 62777.

2. The application according to claim 1, wherein The viable count of mucosal Lactobacillus CCFM1277 in the drug is not less than 1×10 10 CFU / g.

3. The application according to claim 1, characterized in that, The drug contains the Limosilactobacillus mucosae CCFM1277 and a drug carrier.

4. The application according to claim 1, wherein The drug contains the Limosilactobacillus mucosae CCFM1277 and pharmaceutical excipients.

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