| Hyperlipidemia is a chronic disease caused by abnormal lipid metabolism in the body,the number of people who die from related diseases directly or indirectly caused by hyperlipidemia rises year by year.Clinical evidence shows that the intake of probiotics helps reduce blood lipids level,and maintain the body's blood lipid index balance.Taking reasonable doses of probiotics without side effects make probiotics a hot research topic for lipid-lowering drug alternatives.Therefore,based on the human intestinal micro-ecology,screen probiotics with cholesterol-lowering effects,comparing the effects of different intervention frequencies and cycles of probiotics on relevant physiological and biochemical indicators of hyperlipidemia rats,and determining the optimal intervention frequency and the relationship between the intervention effect and the intervention cycle,have certain research value,and it is also the key to research hypolipidemic products using probioticsThis paper screened probiotic strains isolated from the intestinal tract of the longevity population in Bama,Guangxi,and selected strains with a good cholesterol-lowering effect in vitro,and used them in a hyperlipidemia rat model to study the frequency and cycle of probiotic intervention on hyperlipidemia Liver physiology and biochemistry,antioxidant function,hypolipidemic effect,fecal short-chain fatty acid content,and intestinal flora in order to improve the utilization of probiotics,and also provide theoretical basis for clinical quantification of probiotics intervention therapy.1.Screening of strains with cholesterol-lowering effect in vitro.Lactic acid bacterias with cholesterol-lowering effect were selected as target strains for subsequent animal experiments from 40 strains of human-produced lactic acid bacterias which were deposited in the laboratory.The results showed that among the 40 strains of lactic acid bacteria,the highest comprehensive score of principal component analysis(PCA)was Lactobacillus.plantarum 67 and Lactobacillus.fermentum 83,the survival rates of these two strains in artificial gastric juice were 233.33%and 50.88%,and the survival rate in artificial intestinal juice was 345.00%and 76.00%,the survival rates in the 0.3%bile salt medium are 26.60%and 27.08%,the TC degradation rates are 3.67%and 0.88%,the TG degradation rates are 12.5%and 3.24%,and the hydroxyl radical scavenging rate are 85.01%and 84.40%,DPPH free radical scavenging rates are 93.66%and 94.29%,reducing forces are 2.04 and 2.19,surface hydrophobicities are 89.94%and 36.35%,self-polymerizing abilities are 85.27%and 37.48%,and comprehensive scores are 2.29 and 0.82,and they both have bile salt hydrolase.2.Construction of hyperlipidemia rat models and animal experiment.126 male wistar rats were purchased and divided into a blank control group(NC)and a model group.After being fed with high-fat diet for 4 weeks,the rats in the model group had significantly higher TC,TG,and LDL-C levels in serum than the NC group(p<0.05),the HDL-C level decreased significantly(p<0.05),indicating that the hyperlipidemia rat model was successfully constructedAfter successful modeling,the model group was divided into high-fat model group(MC),6L group which has continuous intervention 7 times a week with L.plantarum 67,65 group which has intervention 5 times a week,6G group which has intervention every other day in a week,HL group which has continuous intervention 7 times a week with mixed strain of L.plantarum 67 and L.fermentum 83,which mixing ratio is 1:1,H5 group which has intervention 5 times a week,HG group which has intervention every other day in a week.The blank group was fed with normal feed every day and was gavaged with normal saline(1mL/100g)according to body weight,the high-fat model group was fed with high-fat feed each day with normal saline(1mL/100g)and the probiotic intervention groups were fed high fat feed,gavaged different probiotic suspensions(108 CFU/mL,1mL/100g)were administered to different groups according to body weight.After gavaging for 4w,6w and 8w,the rats were banned from feeding and drinking for 12 hours,weighed the next morning and killed by dislocation,and took relevant samples for tissue measurement.3.Effects of intermittent probiotics intervention on liver physiology and biochemistry in hyperlipidemia rats.Measured three indexes of liver function(ALT,ALB and TP)in serum,calculated liver index of rats,and observed HE stained sections of liver.The results showed that after probiotics intervention,compared with the MC group,the rats' weight gain was controlled,the liver index decreased significantly(p<0.05),and the ALT index decreased significantly(p<0.05).During the 6w-8w of the intervention,the ALB and TP indicators increased significantly(p<0.05).Liver sections showed intact hepatocytes after intervening with probiotics,and there were fewer intercellular lipid droplets,but a few large lipid droplets were still visible.Among the intervention groups,the 65 group had the best effect.Therefore,the strain L.plantarum 67 screened in this paper with good cholesterol-lowering effect in vitro can effectively control the body weight of hyperlipidemia rats,improve liver index and serum liver function by intervention 5 times a week,and can effectively restored liver cells.And the effects of intervention on body weight,liver index and ALT are directly proportional to the intervention time,the effects of intervention on ALB and TP indicators are not directly proportional to the intervention time.Validation experiments using mixed strains yielded the same conclusion4.Effects of intermittent probiotics intervention on lipid peroxidation in serum in hyperlipidemia rats.Measured five antioxidant indexes(GSH-Px,T-SOD,CAT,MDA,T-AOC)and four lipid indexes(TC,TG,HDL-C,LDL-C)in serum in rats and calculated the atherosclerosis index.The results showed that after probiotics intervention,compared with the MC group,the levels of GSH-Px,T-SOD,CAT,and MDA in the serum of rats were significantly decreased(p<0.05),and the T-AOC capacity was significantly increased(p<0.05),the TC,TG,LDL-C and AI values of each intervention group decreased significantly(p<0.05),HDL-C level increased significantly(p<0.05),65 group had best effect on the four anti-oxidation indicators,serum lipid levels and AI values.Therefore,intervened 5 times a week with L.plantarum 67 can effectively improved the anti-oxidation ability and effectively reduced the blood lipid index of hyperlipidemia rats,and the effect of intervention on TC and LDL-C indicators is directly proportional to the intervention time,and the improvement effect of TG,LDL-C and AI indicators is not directly proportional to the intervention time.According to the mixed strain intervention verification experiment,it is known that the conclusion which intervented 5 times a week can indeed increase the antioxidant capacity of rats,reduce blood lipid indexes,and there is a time dependence on the intervention of MDA,T-AOC,TC and LDL-C indexes,the improvement effect of TG,LDL-C and AI indicators is not directly proportional to the intervention time are consistent with that of single bacteria.The conclusion that the effect of intervention on GSH-Px,T-SOD,and CAT levels is not directly proportional to the intervention time is different from that of the single bacteria group,but the three indicators of the single bacteria group are significantly higher than the mixed bacteria group(p<0.05).5.Effects of intermittent probiotics intervention on short-chain fatty acids and intestinal flora in feces of hyperlipidemia rats.Measured the SCFAs in the feces and the intestinal flora of the rats,which were stored in the-80? refrigerator,and analyzed the structural changes at the phylum and genus classification levels and the species abundance of the intestinal flora.The results can be obtained that the acetic acid content decreased significantly(p<0.05),and the isobutyric acid content decreased to varying degrees during the whole process of probiotic intervention.During the 6w-8w of the intervention,the content of propionic acid and butyric acid increased significantly(p<0.05),and the content of isovaleric acid decreased significantly(p<0.05),valproic acid content increased significantly after probiotics intervention for 8w(p<0.05),the 6L group had the best effect on the content of SCFAs in rat feces.Therefore,L.plantarum 67 continuous intervention 7 times a week can effectively improve the levels of six SCFAs in the feces of hyperlipidemia rats,and the intervention effect on butyric acid,isobutyric acid,valeric acid and isovaleric acid content is directly proportional to the intervention time The effect of intervention on acetic acid and propionic acid content was not directly proportional to the intervention time.From the mixed bacteria verification experiment,it can be seen that 7 times a week intervention can indeed improve the content of SCFAs in feces of rats.The conclusion that the effect of acetic acid and propionic acid is not directly proportional to the time of intervention is consistent with that of single bacteria.The conclusion that the intervention time is not proportional is inconsistent with that of the single bacterium,but there is no significant difference with the indicator level of the single bacterium group(p<0.05).Compared with the MC group,the relative abundance of the Firmicute,Bacteroidetes,Proteobacteria and Actinobacteria were decreased significantly(p<0.05),and the relative abundance of the Spirochaetes increased significantly(p<0.05)among the probiotics intervention groups after interventing for 8 weeks.There were significant differences in the relative abundance of the three intervention groups(p<0.05)at every phylum level.The relative abundances of Turicibacter.spp decreased significantly(p<0.05),and the relative abundances of Lactobacillus.spp,Roseburia.spp and Treponema.spp increased significantly(p<0.05).There were significant differences in the relative abundance of the three intervention groups(p<0.05)at every genus level.Among the three intervention groups,the 6L group had the largest abundance of OUT and the highest Shannon index,indicating that probiotic intervention can improve the species diversity and richness of the intestinal flora of hyperlipidemia rats and the L.plantarum 67 continuous intervention 7 times a week had best effect. |
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