实验海洋生物学重点实验室知识库

肠道微生物在包括2型糖尿病（T2DM）在内的代谢性疾病中起着关键作用。

3-溴-4,5-二（2,3-二溴-4,5-二羟基苄）-1,2-苯二酚（BDB）是从海洋红藻松节藻中分离得到的一种天然溴酚化合物。最新的研究表明，BDB可缓解2型糖尿病BKS db小鼠的高血糖病症。为了了解BDB是否在T2DM治疗期间调节肠道菌群的组成，我们将BKS db糖尿病小鼠随机分组，分别给予BDB、二甲双胍或安慰剂治疗7周。与模型组相比，BDB或二甲双胍治疗组糖尿病小鼠在第7周的空腹血糖水平显著降低。利用群落16S rRNA测序和宏基因组学分析，探究了BDB对肠道菌群结构以及代谢功能途径的影响。结果表明，在科水平上，BDB和二甲双胍治疗分别能使产短链脂肪酸（SCFAs）细菌疣微菌科（Verrucomicrobiaceae）和拟杆菌科（Bacteroidaceae）的相对丰度明显提升。在属水平上，阿克曼氏菌属（Akkermansia）的相对丰度在BDB治疗组中显著升高。宏基因组学结果表明，BDB可能通过促进丙酸代谢、抑制淀粉和蔗糖代谢、氨基糖和核苷酸糖代谢来缓解糖尿病小鼠的代谢紊乱。

通过系统发育分析，探究了BKS db小鼠肠道可培养细菌的多样性，从小鼠粪便样本中，共获得209株细菌，分属于4个门16个属25个种。其中含量最丰富的是乳酸杆菌属，占到了总分离菌株数的42.6%。

通过对25株分离菌株进行体外益生菌特性检测分析表明，Lactobacillus菌株具有较强的耐酸、耐碱和抗氧化活性。Lactobacillus johnsonii BM611在模拟胃液中3 h存活率能够达到81.90%，在模拟肠液中6 h存活率能够达到81.98%。菌株Lactobacillus johnsonii BM611细胞破碎液和无细胞分泌液均表现出较强的DPPH自由基清除能力，清除率能够达到44.85%和13.05%，并且具有较强的羟自由基清除能力，清除率能够达到17.16%和15.11%。Lactobacillus johnsonii BM611能显著抑制大肠杆菌等条件致病菌的生长，并促进C2C12细胞的糖消耗水平。

通过结合动物实验，探究了菌株Lactobacillus johnsonii BM611对BKS db小鼠的降糖作用以及对脂质代谢相关因子的影响。研究发现Lactobacillus johnsonii BM611能够显著降低BKS db小鼠的空腹血糖水平，改善2型糖尿病小鼠的葡萄糖耐量以及脂代谢异常，与BDB联合治疗的效果要显著优于单菌治疗；通过组织学病理分析并未发现Lactobacillus johnsonii BM611对组织产生不良影响，反而能够改善小鼠胰腺和肝组织的状态。

本研究不仅探究了海洋溴酚化合物BDB对糖尿病小鼠肠道菌群的调节作用，而且为BKS db模型小鼠肠道菌群多样性的研究提供了参考，为进一步开发益生降血糖菌株提供了资源。

Gut microbiota plays a key role in metabolic diseases including type 2 diabetes mellitus (T2DM).

3-bromo-4,5-bis (2,3-dibromo-4,5-dihydroxybenzyl) - 1,2-catechol (BDB) is a natural bromophenol compound isolated from marine red algae pineal algae. Rencently， the research showed that BDB can alleviate hyperglycemia in type 2 diabetic BKS db mice. In order to research whether BDB regulates the composition of gut microbiota during T2DM treatment, we randomly divided BKS db diabetic mice into groups and treated them with BDB, metformin, or placebo for 7 weeks. Compared with the model group, the fasting blood glucose level of diabetic mice in BDB or metformin treatment group was decreased significantly after 7 weeks. The effects of BDB on the structure and metabolic function of gut microbiota were studied by using 16S rRNA sequencing and metagenomics. At the family level, the results showed that, BDB and metformin treatment can significantly increase the relative abundance of short-chain fatty acid (SCFAs) producing bacteria Verrucomicrobiaceae and Bacteroidaceae, respectively. At the genus level, the relative abundance of Akkermansia increased significantly in BDB treatment group. The results of metagenomics showed that BDB could alleviate the metabolic disorder of diabetic mice by promoting propionic acid metabolism, inhibiting starch and sucrose metabolism, amino sugar, and nucleotide sugar metabolism.

Through phylogenetic analysis, the diversity of bacteria in the intestinal tract of BKS db mice was studied. 209 strains of bacteria were obtained from the fecal samples of mice, including 4 phyla, 16 genera, and 25 species. The most abundant genus was Lactobacillus, accounting for 42.6% of the total isolated bacteria.

The in vitro probiotic characteristics detection and analysis of 25 isolated strains showed that Lactobacillus strains all have high acid resistance, alkali resistance, and antioxidant activity. The survival rate of Lactobacillus johnsonii BM611 in simulated gastric juice can reach 81.90% for 3 h, and the survival rate for 6 h in simulated intestinal juice can reach 81.98%. Lactobacillus johnsonii BM611 showed strong DPPH free radical scavenging ability in cell disruption solution and cell-free excretory supernatants, with scavenging rates of 44.85% and 13.05%, and strong hydroxyl free radical scavenging ability, with scavenging rates of 17.16% and 15.11%. Lactobacillus johnsonii BM611 can significantly inhibit the growth of opportunistic bacteria such as Escherichia coli, and promote the glucose consumption of C2C12 cells.

The hypoglycemic effect of Lactobacillus johnsonii BM611 on BKS db mice and its effect on lipid metabolism related factors were studied by animal experiments. The results showed that Lactobacillus johnsonii BM611 could significantly reduce the fasting blood glucose level of BKS db mice, improve the glucose tolerance and lipid metabolism of type 2 diabetic mice, and the effect of combined treatment with BDB was significantly better than that of single bacterial treatment; Histological and pathological analysis showed that Lactobacillus johnsonii BM611 had no adverse effect on the tissue, but could improve the status of the pancreas and liver tissue in mice.

This study not only explored the regulatory effect of marine bromophenol compound BDB on gut microbiota of diabetic mice, but also provided reference for the development of gut microbiota diversity of BKS db model mice, and provided resources for the further development of probiotic hypoglycemic strains.