Lactobacillus rhamnosus and application thereof in anti-oxidative inflammation drugs

Abstract

The invention belongs to the field of biological medicine, and particularly discloses lactobacillus rhamnosus and application of the lactobacillus rhamnosus in anti-oxidative inflammation drugs, the lactobacillus rhamnosus strain (lactobacillus rhamnosus CQFP202412) is preserved in the general microbiological center of the China Committee for Culture Collection of Microorganisms, and the preservation number is CGMCC No.29611. The lactobacillus rhamnosus CQFP202412 has the advantages that the lactobacillus rhamnosus CQFP202412 can be used for preparing anti-oxidative inflammation drugs, and the lactobacillus rhamnosus CQFP202412 can be used for preparing anti-oxidative inflammation drugs. The lactobacillus rhamnosus CQFP202412 disclosed by the invention is found to be a probiotic with a good effect by investigating the protective effect of the lactobacillus rhamnosus CQFP202412 intervention on mouse motor function, intestinal permeability and brain tissue oxidative inflammation by inducing mouse injury and micro-ecological disorder through antibiotics and finding out that the lactobacillus rhamnosus CQFP202412 disclosed by the invention is a probiotic with a good effect; the reduction of mouse motor function, intestinal barrier injury and oxidative inflammation of brain nervous centralis caused by antibiotics are improved; good pharmaceutical prospects are realized.

Description

Technical Field

[0001] The invention belongs to the field of biomedicine, and specifically discloses a Lactobacillus rhamnosus and application thereof in anti-oxidation and inflammation drugs. Background Art

[0002] Antibiotics are common bactericidal drugs in clinical practice and are widely used in the treatment of human infectious diseases. However, in recent years, it has been found that the abuse of antibiotics can also endanger human health, causing allergies, renal toxicity, bacterial resistance, obesity, intestinal flora imbalance, depression and other brain neurological diseases. Among them, the imbalance of intestinal microbiota caused by the abuse of antibiotics will not only lead to various intestinal diseases, but also dysfunction of the central nervous system. The increased permeability of the intestinal and blood-brain barriers caused by microbial dysbiosis will produce various bacteria and various metabolites that reach the brain through the blood, leading to central nervous system lesions. The existence of intestinal flora is crucial to the body's digestion, immunity, metabolism and other physiological functions. It has been found that it is closely related to the occurrence and development of many diseases such as Alzheimer's disease, diabetes, anxiety, fatty liver, etc. The concept of "Microbiota-gut-brain axis" (MGBA) has attracted the attention of more scholars to use intestinal flora as a therapeutic target. Summary of the invention

[0003] In view of the above problems, the present invention discloses a Lactobacillus rhamnosus and application thereof in anti-oxidation and inflammation drugs.

[0004] The purpose of the present invention is achieved through the following technical solutions.

[0005] The invention discloses a lactobacillus rhamnosus strain. The lactobacillus rhamnosus strain (Lactobacillus rhamnosus) CQFP202412 is deposited in the General Microbiological Center of China Microbiological Culture Collection Administration, and the deposit number is CGMCC No.29611.

[0006] Furthermore, the present invention also discloses a live Lactobacillus rhamnosus product, which contains the live Lactobacillus rhamnosus mentioned above.

[0007] Furthermore, the present invention also discloses the use of the live Lactobacillus rhamnosus product in the preparation of a medicine for improving brain function.

[0008] Furthermore, the present invention also discloses the use of the live Lactobacillus rhamnosus product in the preparation of a medicine for improving intestinal function.

[0009] Furthermore, the present invention also discloses the use of the live Lactobacillus rhamnosus product in the preparation of a medicine for improving motor function.

[0010] Furthermore, in the above-mentioned pharmaceutical use, the drug is a drug for improving symptoms caused by abuse of antibiotics.

[0011] Furthermore, the present invention discloses a health product containing the above-mentioned live Lactobacillus rhamnosus product.

[0012] Furthermore, the present invention discloses a feed containing the above-mentioned live Lactobacillus rhamnosus product.

[0013] Furthermore, the present invention discloses a medicine for improving symptoms caused by abuse of antibiotics, comprising the above-mentioned live Lactobacillus rhamnosus product.

[0014] Furthermore, among the above symptoms, the symptom includes any one of the following: decreased motor function, intestinal barrier damage or oxidative inflammation of the central nervous system of the brain.

[0015] Compared with the prior art, the present invention has the following beneficial effects:

[0016] The present invention uses antibiotics to induce damage and microecological disorder in mice, and investigates the protective effect of Lactobacillus rhamnosus CQFP202412 intervention on the motor function, intestinal permeability and oxidative inflammation of brain tissue of mice. It is found that the Lactobacillus rhamnosus CQFP202412 disclosed in the present invention is a probiotic with good effect, which improves the decreased motor function, intestinal barrier damage and oxidative inflammation of the central nervous system of mice caused by antibiotics; and has good pharmaceutical and feed prospects.

[0017] The strain deposit information is as follows:

[0018] Name of depository: General Microbiology Center of China Microbiological Culture Collection Administration;

[0019] Address of the depository: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing;

[0020] Deposit date: January 12, 2024

[0021] Deposit number: CGMCC No.29611,

[0022] Taxonomic name: Lacticaseibacillus rhamnosus. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 :Changes in weight-bearing swimming time of mice. Values ​​shown in ab are mean ± standard deviation. According to Tukey's test, the mean values ​​with different letters in the same figure are significantly different (p < 0.05), the same below;

[0024] Figure 2 : Changes in running time of mice;

[0025] Figure 3 :Pathological changes in brain tissue of mice;

[0026] Figure 4 :The gene expression of AKT, BDNF, CREB and ERK1 in mouse brain tissue;

[0027] Figure 5 :ZO-1, occludin-1 and claudin-1 gene expression in mouse cecal tissue. DETAILED DESCRIPTION

[0028] In order to make the purpose, technical scheme and advantages of the present invention clearer, the present invention is further described in detail below through embodiments. However, it should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the scope of the present invention. In addition, in the following description, the description of known structures and technologies is omitted to avoid unnecessary confusion of the concept of the present invention.

[0029] It should be noted that, in the absence of conflict, the embodiments and features in the embodiments of the present application can be combined with each other. The present invention will be described in detail below in conjunction with the embodiments.

[0030] Example 1

[0031] 1 Materials and methods

[0032] 1.1 Materials and reagents

[0033] Thirty C57BL / 6J mice, male, 6-8 weeks old, were purchased from Chongqing Enswell Biotechnology Co., Ltd.

[0034] Neomycin, amphotericin B solution, ampicillin, metronidazole, and caffeic acid were purchased from Aladdin Reagent (Shanghai) Co., Ltd.; vancomycin was purchased from Shanghai MacLean Biochemical Technology Co., Ltd.; lipopolysaccharide, Trizol, and DEPC water were purchased from Beijing Solebold Technology Co., Ltd.; cDNA reverse transcription kit and SYBR premix were purchased from Thermo Fisher Biotechnology Co., Ltd., USA; malondialdehyde, superoxide dismutase, and glutathione kits were purchased from Nanjing Jiancheng Bioengineering Institute; chloroform was purchased from Chengdu Cologne Chemical Co., Ltd.; isopropanol and anhydrous ethanol were purchased from Chongqing Wansheng Chuandong Chemical Co., Ltd.; interleukin-6, interleukin-10, and tumor necrosis factor-α kits were purchased from Shanghai ELISA Biotechnology Co., Ltd.

[0035] 1.2 Instruments and Equipment

[0036] Varioskan LUX multifunctional microplate reader, SteponePlus automatic PCR amplification instrument, Thermo Fisher Biotechnology Co., Ltd., USA; UPH-II-20T UPH series ultrapure water purifier, Sichuan UPH Ultrapure Technology Co., Ltd.; Centrifuge 5 418R refrigerated centrifuge, Eppendorf, Germany; BSA124S electronic balance, Sartorius Scientific Instruments (Beijing) Co., Ltd.; ZH-PT mouse experimental treadmill, Anhui Zhenghua Biological Instrument Equipment Co., Ltd.; BX43 electron microscope, Olympus Corporation, Japan.

[0037] 1.3 Experimental methods

[0038] 1.3.1 Cultivation of experimental strains

[0039] A strain of Lactobacillus rhamnosus CQFP202412 screened from Jiangdong pickle juice in Kaixian County, Chongqing was registered and stored in the China General Microbiological Culture Collection Center (Accession No.: CGMCC No.29611). The experimental strain cryopreservation tube was taken out from -80℃, thawed and inoculated (200μL) into 8mL of MRS liquid medium, revived and cultured at 37℃ for 24h, and then mixed and 3% of the inoculum was transferred to MRS liquid medium for further culture for 18h. The Lactobacillus rhamnosus CQFP202412 at the bottom was collected by centrifugation (4℃, 4000rpm, 10min), and resuspended with physiological saline to an adjusted cell concentration of 1.0×10 9 CFU / kg·bw order of magnitude.

[0040] 1.3.2 Experimental animal models

[0041] Mice were housed in standard cages in the animal room at a room temperature of 23±2℃ and a relative humidity of 55±5%. Lights were turned on every 12 hours, and the mice were allowed to eat and drink freely. The mice were adaptively housed for one week. The experimental mice were divided into 5 groups, namely normal group, model group, caffeic acid group, Lactobacillus rhamnosus group, and high-temperature inactivated Lactobacillus rhamnosus group, with 6 mice in each group. The specific design was as follows: (1) Normal group (Normal), no treatment; (2) Model group (ABXs), intraperitoneal injection of mixed antibiotic preparations, gavage of normal saline; (3) Caffeic acid group (ABXs+CA), intraperitoneal injection of mixed antibiotic preparations, gavage of caffeic acid (10 mg / kg); (4) Lactobacillus rhamnosus group (ABXs+LR), intraperitoneal injection of mixed antibiotic preparations, gavage of Lactobacillus rhamnosus (10 9 CFU / kg·bw); (5) high temperature inactivated Lactobacillus rhamnosus group (ABXs+D-LR), intraperitoneal injection of mixed antibiotic preparation, oral gavage of high temperature inactivated (100℃, 15min) Lactobacillus rhamnosus (10 9CFU / kg·bw). Injection / oral gavage every other day, the dose for oral gavage and intraperitoneal injection was 0.1mL / 10g·bw. The experiment lasted for two weeks. Mixed antibiotic preparation: neomycin 5mg / mL, vancomycin 25mg / mL, amphotericin B 0.1mg / mL, ampicillin 10mg / mL, metronidazole 5mg / mL, lipopolysaccharide 1.5μg / mL.

[0042] 1.3.3 Weighted swimming test

[0043] A Φ60cm×5cm water tank was used, with a water depth of about 35cm and a water temperature of 20±2℃. 30 minutes after administration, a lead wire weighing 5% of body weight was tied to the tails of mice in the normal group, model group, caffeic acid group, Lactobacillus rhamnosus group, and high-temperature inactivated Lactobacillus rhamnosus group, and the mice were marked and put into the water in turn for a weighted swimming experiment. The timer was used to record that when the head of the mouse was immersed in the water for more than 8s and could not float to the surface, it was the exhaustion time of the mouse, and it was wiped dry with a dry towel after exercise.

[0044] 1.3.4 Treadmill test

[0045] The ZH-PT mouse experimental treadmill was used, with dimensions of 565×630×310mm in length×width×height, 55mm in width of a single runway, and 8 treadmills. Parameter settings: 5° inclination of the treadmill; 0.6mA of stimulation current; 2m / s^2 of acceleration; 20m / min of speed; 30min of exercise time; and 1min of rest time. Adaptive training began 4 days before the end of the experiment, once a day, 5min each time. The formal test was conducted on the last day of the experiment, and the final results were recorded.

[0046] 1.3.5 Tissue sample collection

[0047] After the last exercise test, the mice were weighed and blood was collected from the eyeballs after ether anesthesia. The brain, liver, and cecum tissues were dissected and collected after cervical dissection. The liver was separated and weighed. Part of the brain tissue was soaked in tissue fixative for slice preparation. The remaining tissues were stored at -80°C for subsequent experiments.

[0048] 1.3.6 Pathological observation

[0049] Brain tissue (0.5 cm 2 ) were fixed in tissue fixative for more than 48 h, then embedded in paraffin and sectioned (4 μm). Hematoxylin and eosin staining was used to observe pathological changes under an electron microscope.

[0050] 1.3.7 Determination of serum and brain tissue biochemical indicators

[0051] Serum preparation: Mouse blood was allowed to stand at 4°C for 2 h, centrifuged at 3000 rpm and 4°C for 15 min, and the upper serum was obtained and frozen at -80°C. Preparation of brain tissue homogenate: After adding 0.1 g of mouse brain tissue to 0.9 mL of normal saline, the tissue was homogenized three times at a rate of 6 m / s for 30 s. Index detection: The levels of malondialdehyde (MDA), glutathione (GSH), and superoxide dismutase (SOD) in mouse serum and brain homogenate were determined by kits; the levels of cytokines tumor necrosis factor-α (TNF-α), interleukin (IL)-6 and -10 in serum and brain tissue were determined by enzyme-linked immunosorbent assay kits.

[0052] 1.3.8 Real-time fluorescence quantitative PCR analysis

[0053] After homogenization of brain and cecal tissues, total RNA from brain and cecal tissues was extracted using TRIzol reagent and diluted to a concentration of 1.0 μg / μL. Total RNA was reverse transcribed into cDNA. Then, 1.0 μL cDNA, 2.0 μL forward and reverse primers (10 μm), and 10 μL premix were mixed and reacted for 40 cycles under the following conditions: 3 min at 95 °C, 30 s at 60 °C, and 1 min at 95 °C. β-actin was an internal reference gene, and the data were obtained by 2 -ΔΔCt Method Calculation: The primer sequences (SEQ ID No. 1-16) are shown in Table 1.

[0054] Table 1 Primer sequences

[0055]

[0056]

[0057] 1.3.9 Statistical analysis

[0058] All experimental data were statistically analyzed using SPSS 20.0 software (SPSS Inc, Chicago, IL, USA), combined with one-way analysis of variance and Tukey's multiple comparisons; p < 0.05 was considered statistically significant. GraphPad Prism 9.3 was used for drawing. Data are expressed as mean ± standard deviation (SD), repeated 3 times.

[0059] Example 2

[0060] 2 Results and Analysis

[0061] 2.1 Results of the weight-bearing swimming test in mice

[0062] Abuse of antibiotics can lead to decreased endurance in mice, while intervention with probiotics can improve endurance. Figure 1As shown in the figure, compared with the normal group, the weighted swimming time of mice in the model group decreased significantly (p < 0.05). After the intervention of Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) by oral gavage in advance, the weighted swimming time of mice increased significantly, approaching that of the normal group and the caffeic acid group. The Lactobacillus rhamnosus group was slightly better than the heat-killed Lactobacillus rhamnosus group.

[0063] 2.2 Results of mouse running experiment

[0064] We also conducted a running experiment to further investigate the endurance of mice. Figure 2 As shown in the figure, compared with the normal group, the running time of mice in the model group decreased significantly (p < 0.05). After the intervention of Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) and caffeic acid in advance, the weight-bearing running time of mice also increased significantly, and the effect of the Lactobacillus rhamnosus group and the heat-killed Lactobacillus rhamnosus group was similar. The difference is that the running time of mice in the Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) group was significantly higher than that of the model group, and lower than that of the caffeic acid group. In addition, the running time of mice in the caffeic acid group was lower than that of the normal group.

[0065] 2.3 Changes in mouse liver index

[0066] Antibiotic preparations affect the metabolism of mice and thus affect liver changes. As shown in Table 2, compared with the normal group, the liver index of mice in the model group increased and the difference was significant (p<0.05). It is worth noting that the liver index decreased after the early intervention of Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus), and the downward trend caused by the Lactobacillus rhamnosus group was more significant, close to the effect of the caffeic acid group.

[0067] Table 2 Mouse body weight, liver weight, liver index

[0068]

[0069] a-b The values ​​shown are mean ± standard deviation. According to Tukey's test, the means with different letters in the same row are significantly different (p<0.05). Organ index (%) = organ weight (g) / mouse body weight (g) × 100. The same below.

[0070] 2.4 Pathological changes in mouse brain tissue

[0071] HE staining results of mouse brain tissue showed that ( Figure 3), the morphology of the cerebral cortex neurons of the normal group mice was normal, the cell morphology was uniform and neatly arranged, and the nuclei were large and obvious. The number of brain tissue cells of the model group mice decreased, the color of the nuclei deepened, obvious pyknosis occurred, some showed point-like necrosis, the arrangement was disordered, the cortical neurons atrophied and became smaller, and the cell damage was serious. After pretreatment with Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus), the phenomenon of nuclear pyknosis in the brain tissue of mice was alleviated and the number of cells also increased. Some neurons tended to be normal, and a few showed reduction and atrophy. Among them, Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) and caffeic acid had similar effects.

[0072] 2.5 Mouse serum oxidation and inflammatory indicators

[0073] MDA, GSH, and SOD can reflect the body's oxidative damage and antioxidant capacity. As shown in Table 3, compared with the normal group, the MDA level in the serum of the model group increased, and the levels of GSH and CuZn-SOD decreased, and the difference was statistically significant (p<0.05). Compared with the model group, after pretreatment with caffeic acid and Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus), the MDA level in the serum decreased, and the levels of GSH and CuZn-SOD increased, and the difference was statistically significant (p<0.05). Among them, there was no significant difference between the effects of Lactobacillus rhamnosus and heat-killed Lactobacillus rhamnosus. IL-6, IL-10, and TNF-α reflect the degree of inflammation in the body. As shown in Table 3, compared with the normal group, the levels of pro-inflammatory factors TNF-α and IL-6 in the model group increased, while the level of anti-inflammatory factor IL-10 decreased (p<0.05). Compared with the model group, pretreatment with caffeic acid and Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) downregulated TNF-α and IL-6, but upregulated IL-10 (p<0.05), and the effects of the three were similar.

[0074] Table 3 Oxidation and inflammation indexes in mouse serum

[0075]

[0076] 2.6 Oxidative and inflammatory indicators in mouse brain tissue

[0077] We further tested the oxidative and inflammatory indicators of brain tissue. As shown in Table 4, compared with the normal group, the MDA level in the model brain tissue increased, and the GSH and CuZn-SOD levels decreased, and the difference was statistically significant (p<0.05). Compared with the model group, after intervention with Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus), the levels of GSH and CuZn-SOD increased, but Lactobacillus rhamnosus was more effective, close to caffeic acid. The MDA level decreased, and the effects of caffeic acid, Lactobacillus rhamnosus, and heat-killed Lactobacillus rhamnosus were similar. The results of brain tissue inflammation are shown in Table 4. Compared with the normal group, the levels of TNF-α and IL-6 in the model group increased, while the level of IL-10 decreased (p<0.05). Compared with the model group, pretreatment with caffeic acid and Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) had a down-regulatory effect on TNF-α and IL-6, with the order of heat-killed Lactobacillus rhamnosus > Lactobacillus rhamnosus > caffeic acid, but had an up-regulatory effect on IL-10, and the three effects showed opposite trends.

[0078] Table 4 Oxidation and inflammation indexes of mouse brain tissue

[0079]

[0080] 2.7 Expression of AKT, BDNF, CREB and ERK1 genes in mouse brain tissue

[0081] AKT, CREB, BDNF, ERK1, and IL-6 play an important role in cranial nerve injury. Figure 4 As shown in the figure, compared with the normal group, the gene expressions of AKT, CREB, BDNF, and ERK1 in the model group showed a significant decrease (p<0.05), and the expression increased after intervention with caffeic acid and Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus). Among them, the expression of CREB, BDNF, and ERK1 of heat-killed Lactobacillus rhamnosus was stronger than that of Lactobacillus rhamnosus, while the expression of AKT was the opposite. The expression of CREB in Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) was significantly higher than that in the caffeic acid and normal groups (p<0.05), and the expression of BDNF was similar to that of caffeic acid, but both were significantly higher than those in the normal group (p<0.05).

[0082] 2.8 ZO-1, occludin-1 and claudin-1 gene expression in mouse cecum

[0083] As classic tight junction proteins, ZO-1, occludin-1, and claudin-1 can reflect the mechanical barrier function of intestinal epithelial cells and the degree of damage to the tight junction structure. Figure 5As shown in the figure, compared with the normal group, the gene expressions of ZO-1, occludin-1 and claudin-1 in the model group all showed a significant decrease (p<0.05), and the expressions increased after intervention with caffeic acid and Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus). The expressions of occludin-1 and claudin-1 were similar and close to those in the normal group. In the expression of ZO-1, the effects of Lactobacillus rhamnosus and caffeic acid were similar, lower than those of heat-killed Lactobacillus rhamnosus, and all three were significantly lower than those in the normal group (p<0.05).

[0084] 3 Conclusion and discussion

[0085] Probiotics can reduce the body's oxidation degree, improve exercise capacity, provide necessary metabolites for skeletal muscle mitochondria, and regulate the key transcription factors and enzymes (such as PGC-1α, AMPK) involved in the mitochondrial biological metabolic process by playing an intestinal regulatory role. After the intervention of Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) in the present invention, mice showed longer weight-bearing swimming and running time, and endurance was significantly improved, but there was no significant difference between Lactobacillus rhamnosus and heat-killed Lactobacillus rhamnosus. Studies have shown that supplementing probiotics can improve lipid metabolism, reduce fat and lose weight. The weight of mice after the intervention of Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) in the present invention decreased, but it was not significant, which may be related to the fact that the supplementation time of probiotics is not long enough, but the liver index decreased significantly.

[0086] Adverse reactions caused by the abuse of antibiotics are often accompanied by a variety of diseases, involving oxidative changes in key physiological molecules, including proteins, lipids, carbohydrates, nucleic acids, and regulation of gene expression and inflammatory responses. Oxidative damage is one of the side effects of antibiotic abuse. Malondialdehyde (MDA) is the final metabolite of membrane lipid peroxidation in the body. MDA released from the cell membrane can react with proteins and nucleic acids to cause cross-linking polymerization, which damages the membrane structure and function and changes its permeability, thereby affecting the normal biochemical reactions of the body. Endogenous antioxidant enzymes SOD and non-enzyme GSH can effectively improve oxidative stress damage and enhance antioxidant capacity. Our studies have shown that the intake of Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) can reduce the level of MDA in serum and brain tissue, increase the levels of SOD and GSH, and inhibit the degree of peroxidation.

[0087] Inflammation plays an important role in the process of brain damage caused by antibiotics. Antibiotics and their secondary metabolites can destroy the dynamic balance between pro-inflammatory and anti-inflammatory reactions, and the level of inflammatory markers increases. Brain tissue and cells after damage will undergo pathological changes, and it can be observed by HE staining that neurons in the cortical area of ​​mice appear condensed, the nuclear color deepens, and the arrangement is disordered, and the release of pro-inflammatory cytokines such as IL-6 and TNF-α will increase, thereby activating the body's immune response, releasing more inflammatory cytokines, aggravating brain inflammation and neurotoxicity, and further inducing the excessive production of free radicals and lipid peroxidation. In the present invention, Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) reduces the levels of IL-6 and TNF-α pro-inflammatory factors in serum and brain tissue, increases the level of IL-10 anti-inflammatory factors, and suppresses the degree of inflammation.

[0088] As the main component of the intestinal barrier system, intestinal epithelial cells prevent harmful substances and potential pathogens from entering the body while allowing ions, water and nutrients to enter the body to maintain the body's normal metabolism. Studies have found that antibiotic exposure can cause damage to the mechanical barrier function and tight junction (TJ) structure of intestinal epithelial cells. An important manifestation of the polarity of epithelial cells and the adhesion between cells is the tight junctions formed between adjacent cells. The high concentration of tight junction protein expression is the basis of the above functions. Claudin-1, occludin and zonula occludens-1 (ZO-1) as tight junction proteins are core proteins of intestinal epithelial cells, which can improve the intestinal barrier, reduce the translocation of intestinal flora and intestinal toxins to the intestinal mucosa and intestinal inflammation. Occludin protein can increase transmembrane resistance, maintain the permeability between cells within the normal range, and maintain intestinal epithelial cells and tight junction structure. Claudin protein regulates the transport of substances in intestinal epithelial cells and plays a supporting role in the TJ structure. Its high expression makes the epithelial cells tighter. ZO-1 forms a link with the actin cytoskeleton between different cells and plays a role in mediating the opening and closing of tight junctions

[21] The present invention shows that intervention with Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) can upregulate the gene expression of Claudin-1, Occludin-1 and ZO-1 in the intestine, thereby improving the damage to the intestinal barrier.

[0089] The protein kinase B / CAMP response element binding protein / brain-derived neurotrophic factor (AKT / CREB / BDNF) signaling pathway plays an important role in brain neurological diseases. AKT is an important effector of the AKT / CREB / BDNF signaling pathway, which is involved in regulating cell growth, proliferation, apoptosis, intracellular metabolism, and cell migration and movement. CREB is a substrate of AKT, which enhances the transcription of CRE sequences and regulates the transcription of target genes related to neuronal growth, injury, regeneration, synaptic plasticity, and learning and memory abilities. BDNF is one of the targets of CREB downstream transcriptional regulation. Its promoter region contains CRE sequences, so it is regulated by p-CREB. The binding of BDNF to its receptor tyrosine kinase B (TrKB) leads to the activation of the downstream Ras-Raf-MEK-ERK pathway, and the activated ERK1 / 2 can phosphorylate kinases such as Raf and MEK, thereby activating substrates such as CREB, initiating transcription and translation, and forming new synapses. The results of the present invention show that after intervention with Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus), the gene expression of AKT, CREB, BDNF, and ERK1 in brain tissue is upregulated, and the pro-inflammatory factor IL-6 is downregulated, indicating that phosphorylated AKT can activate CREB, upregulate BDNF and ERK1 expression, and inhibit brain cell damage.

[0090] In summary, the present invention uses a mixed antibiotic preparation (neomycin 5 mg / mL, vancomycin 25 mg / mL, amphotericin B 0.1 mg / mL, ampicillin 10 mg / mL, metronidazole 5 mg / mL, lipopolysaccharide 1.5 μg / mL) to induce decreased motor function, intestinal barrier damage, and brain tissue oxidation and inflammation in mice by intraperitoneal injection. Intervention with Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) CQFP202412 can improve the endurance of mice, and the exhaustion time of weighted swimming and running is improved. Intervention with Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) CQFP202412 can also alleviate the oxidation and inflammatory response of antibiotic-induced mice, reduce the levels of MDA, IL-6, TNF-α oxidation and inflammatory indicators in serum and brain, and increase the levels of SOD, GSH and IL-10 antioxidant and anti-inflammatory indicators. HE staining showed that Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) CQFP202412 intervention improved the cell morphology of mouse brain tissue, reduced shrinkage and increased the number. Real-time PCR analysis showed that Lactobacillus rhamnosus (or heat-killed Lactobacillus rhamnosus) CQFP202412 activated and enhanced the gene expression of AKT / CREB / BDNF / ERK1 pathway in the brain, reduced the gene expression of proinflammatory factor IL-6, and played a role in the protection of the central nervous system of the brain; it also enhanced the gene expression of ZO-1, occludin-1 and claudin-1 in cecal tissue, and improved the intestinal epithelial cell barrier damage caused by antibiotics.

[0091] Therefore, the Lactobacillus rhamnosus CQFP202412 in the present invention is a probiotic with good effects, which improves the decreased motor function, intestinal barrier damage and oxidative inflammation of the central nervous system of mice caused by antibiotics.

[0092] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention rather than to limit the protection scope of the present invention. Therefore, based on the innovative concept of the present invention, changes and modifications to the embodiments described herein, or equivalent structural or equivalent process transformations made using the contents of the specification and drawings of the present invention, directly or indirectly applying the above technical solutions to other related technical fields are all included in the protection scope of the patent of the present invention.

Claims

1. A strain of Lactobacillus rhamnosus, It is characterized in that The Lactobacillus rhamnosus strain CQFP202412 is deposited in the General Microbiology Center of China National Microbiological Culture Collection Administration, with a deposit number of CGMCC No.29611.

2. A live Lactobacillus rhamnosus product, It is characterized in that Contains the live Lactobacillus rhamnosus bacteria as claimed in claim 1.

3. Use of the live Lactobacillus rhamnosus product as claimed in claim 2 in the preparation of a medicine for improving brain function.

4. Use of the live Lactobacillus rhamnosus product according to claim 2 in the preparation of a medicament for improving intestinal function.

5. Use of the live Lactobacillus rhamnosus product according to claim 2 in the preparation of a medicament for improving motor function.

6. The use according to any one of claims 3 to 5, It is characterized in that The medicine is a medicine for improving symptoms caused by abuse of antibiotics.

7. A health product, It is characterized in that A product containing the live Lactobacillus rhamnosus bacteria as claimed in claim 2.

8. A feed, It is characterized in that A product containing the live Lactobacillus rhamnosus bacteria as claimed in claim 2.

9. A drug that improves symptoms caused by the misuse of antibiotics. It is characterized in that A product containing the live Lactobacillus rhamnosus bacteria as claimed in claim 2.

10. The drug for improving symptoms caused by abuse of antibiotics according to claim 9, It is characterized in that The symptoms include any of the following: decreased motor function, intestinal barrier damage, or oxidative inflammation of the central nervous system of the brain.

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