【作者】 王珺；

【导师】 张家超；

【作者基本信息】 海南大学 ， 食品科学与工程， 2024， 硕士

【摘要】 高脂血症(hyper-lipidemia,HLP)是一种由于体内脂质代谢紊乱引起的疾病。主要表现为血清中总胆固醇(TC)、甘油三酯(TG)、低密度脂蛋白胆固醇(LDL-C)水平过高和高密度脂蛋白胆固醇(HDL-C)水平过低。近年来随着微生物组学和代谢组学的发展,研究者发现肠道菌群与宿主脂质代谢等疾病有着密切关系。肠道菌群可以通过代谢和免疫调节等途径作用于宿主,并最终参与宿主HLP等代谢性疾病的发生和发展,因而肠道菌群可以作为HLP的重要生物标志物。为了探究不同种乳杆菌对于高脂饮食引起的金黄地鼠高脂血症的预防和缓解作用及其差异。本研究选用了四株乳杆菌株(Limosilactobacillus fermentum CKCC1858(CKCC1)、Limosilactobacillus fermentum CKCC1369(CKCC2)、Lactiplantibacillus plantarum CKCC1312(CKCC3)、Lactobacillus gasseri CKCC1913(CKCC4))进行高脂血症的预防和治疗实验。首先,通过分析四株乳杆菌的全基因组信息,了解菌株遗传物质中与功能性基因、致病相关的已知毒力因子和抗性基因等存在情况。结果表明,四株乳杆菌基因组胞嘧啶和鸟嘌呤(GC)含量分别为52.88、52.91、47.24、35.38%,表明四株乳杆菌可能具有较强的稳定性、耐碱性和耐高温性;功能基因预测结果显示,四株乳杆菌的碳水化合物和脂质代谢功能丰富,可能对肠道菌群代谢过程产生影响。其次,对六周龄金黄地鼠进行六周的高脂饮食单独喂养以及乳杆菌干预实验,探究乳杆菌对高脂饮食引起的高脂血症的预防效果。研究结果表明,与模型组相比,CKCC1干预使高脂饮食金黄地鼠血清中TC、TG、LDL-C、肿瘤坏死因子α(TNF-α)和白细胞介素6(IL-6)水平分别降低了53.46、55.72、46.61、19.66和13.69%(P<0.05),减轻了高脂饮食金黄地鼠的炎症反应。另外,与模型组相比,经过CKCC1的干预,高脂饮食金黄地鼠血清中谷草转氨酶(AST)和谷丙转氨酶(ALT)的活性显著下降了8.74%和35.52%;CKCC1干预后的金黄地鼠肝脏中的炎症浸润和肝细胞中脂质含量也减少,进而缓解了肝脏的炎症和脂质堆积。通过分析肠道菌群发现,乳杆菌的干预改变了高脂饮食金黄地鼠肠道菌群多样性,更重要的是CKCC1的干预增加了产生短链脂肪酸(SCFAs)的细菌丰度(Bifidobacterium pseudolongum和Bacteroides vulgatus)和改变了肠道菌群的代谢功能(促进丙酮酸发酵至丙酸I和抑制脂质合成途径),从而减轻高脂饮食诱导的肠道菌群失调,这可能是CKCC1对预防高脂血症有显著效果的原因。与此同时,通过六周高脂饮食喂养构建金黄地鼠高脂血症模型,随后利用四株乳杆菌对高脂血症金黄地鼠进行了为期六周的治疗。与预防实验结果相似,与其他菌株相比,CKCC1在缓解高脂血症、肝损伤和炎症方面的能力尤为突出。结果显示,CKCC1和洛伐他汀的干预可以显著降低高脂饮食中金黄地鼠的血脂水平和肝脏脂肪堆积,更重要的是,与CKCC1相比,洛伐他汀(LOV)的摄入使AST的活性增加了21.51%,可能对肝脏造成损害。LOV干预显著降低了结肠中SCFAs的含量,并下调了产生丁酸的细菌(Lactobacillus reuteri)的丰度,但CKCC1的摄入增加了丁酸含量,从而可以降低高脂血症金黄地鼠的血脂水平并保护肝脏。本研究阐明了乳杆菌通过肠道菌群预防和治疗高脂血症的可能作用机制,并为乳杆菌改善脂质代谢疾病提供了数据支撑和理论依据。更多还原

【Abstract】 Hyperlipidemia(HLP)is a condition caused by abnormal disorders of lipid metabolism in the body.It is mainly characterized by various abnormalities of lipid composition in plasma,including high levels of total cholesterol(TC),triglycerides(TG),low-density lipoprotein cholesterol(LDL-C),and low levels of high-density lipoprotein cholesterol(HDL-C).In recent years,with the development of microbiomes and metabolomics,gut microbiota has been found to have a close relationship with lipid metabolism and other diseases of the host.Gut microbiota can act on the host through metabolic and immunomodulatory pathways,and ultimately participate in the occurrence and development of HLP and other metabolic diseases in the host,thus gut microbiota can be used as an important biomarker related to HLP.To investigate the preventive and treatment effects of different species of probiotics on hyperlipidemia caused by a high-fat diet.In this study,four Lactobacillus strains(Limosilactobacillus fermentum CKCC1858(CKCC1),Limosilactobacillus fermentum CKCC1369(CKCC2),Lactiplantibacillus plantarum CKCC1312(CKCC3),Lactobacillus gasseri CKCC1913(CKCC4))were selected for experiments on prevention and treatment of hyperlipidemia.First,whole genome sequencing(WGS)information of the four Lactobacillus strains was analyzed to understand the presence of functional genes,known virulence factors related to pathogenicity,and resistance genes in the genetic material of the strains.The results showed that the genomic Cytosine and Guanine(GC)contents of the four Lactobacillus strains were 52.88,52.91,47.24,and 35.38%,respectively,indicating that they have more stable,high alkaline,and high-temperature resistance.The results of functional gene prediction showed that the four strains were rich in carbohydrate and lipid metabolism,which might have a role in the metabolic process of the gut microbiota.Secondly,6-week-old golden hamsters were fed a high-fat diet alone for 6 weeks as well as Lactobacillus interventions to investigate the preventive effects of Lactobacillus on high-fat diet-induced hyperlipidemia.The results showed that CKCC1 reduced the serum levels of TC,TG,LDL-C,TNF-α,and IL-6 by 53.46,55.72,46.6,19.66 and 13.69%,respectively,and lowered the inflammatory response in golden hamsters on a high-fat diet compared with the model group(P<0.05).In addition,after the intervention of CKCC1,the activities of serum alanine aminotransferase(AST)and alanine transaminase(ALT)in high-fat diet golden hamsters were significantly decreased by 8.74 and 35.52%compared with the model group;the inflammatory infiltration in the liver and the lipid content of hepatocytes in the liver of golden hamsters after the intervention of CKCC1 were also reduced,which alleviated the inflammation and the accumulation of lipids in the liver.Analysis of the gut microbiota revealed that the Lactobacillus intervention altered the diversity of the gut microbiota in golden hamsters on a high-fat diet,and more importantly,the CKCC1 intervention increased the abundance of bacteria producing SCFAs(Bifidobacterium pseudolongum and Bacteroides vulgatus)and altered the metabolic functions of the gut microbiota(promoting pyruvate fermentation to propionate I and inhibition of lipid synthesis pathways)to attenuate high-fat diet-induced dysbiosis of the gut microbiota,which may account for the significant effect of CKCC1 in preventing hyperlipidemia.Meanwhile,a model of hyperlipidemia in golden hamsters was constructed by feeding a high-fat diet for six weeks,followed by a six-week treatment of hyperlipidemic golden hamsters using four strains of Lactobacillus.Similar to the results of the prophylaxis experiment,CKCC1 was particularly effective in treating hyperlipidemia and alleviating liver injury and inflammation compared to the other strains.The results showed that intervention with CKCC1 and lovastatin significantly reduced blood lipid levels and liver fat accumulation in golden hamsters on a high-fat diet,and more importantly,compared with CKCC1 group,lovastatin intake increased AST activity by21.51%,which may be detrimental to the liver.The LOV intervention significantly reduced short-chain fatty acids(SCFAs)in the colon and down-regulated butyric acid-producing bacteria(Lactobacillus reuteri)abundance,but CKCC1 intake increased butyric acid content,which could reduce lipid levels and protect the liver in hyperlipidemic golden hamsters.This study elucidated the possible mechanism of action of Lactobacillus in preventing and treating hyperlipidemia in golden hamsters through gut microbiota and provided data support and theoretical basis for Lactobacillus intervention to improve lipid metabolism.更多还原