长效促胰岛素降糖酵母的构建及其对糖尿病模型小鼠的治疗效果

益生菌生物药物是指通过口服表达药用多肽(蛋白)的重组益生菌活细胞达到治疗疾病的新型口服给药系统。为了构建一种能有效防治2型糖尿病的酵母生物药物,文章首先构建了酿酒酵母(S.cerevisiae)整合型表达载体pNK1-PGK,并且通过绿色荧光蛋白(GFP)证明其表达功能正常,利用该载体将10×GLP-1 (Glucagon-like peptide-1)基因转化到酿酒酵母INVSc1中,通过营养缺陷型和Western blotting成功筛选出表达10×GLP-1的长效促胰岛素降糖酵母(Long-acting GLP-1 hypoglycemic yeast, LHY)。该酵母生长迅速,外源基因10×GLP-1表达稳定,表达量达到1.56 mg/g细胞湿重。通过链脲佐菌素和高脂高糖饮食联合诱导的方法构建了2型糖尿病小鼠模型,用LHY对其进行口服灌胃治疗,证明LHY具有较好疗效,明显降低血糖水平。

Construction of yeast strains expressing long-acting glucagon-like peptide-1 (GLP-1) and their therapeutic effects on type 2 diabetes mellitus mouse model

Probiotics, i.e., bacteria expressing therapeutic peptides (protein), are used as a new type of orally administrated biologic drugs to treat diseases. To develop yeast strains which could effectively prevent and treat type 2 diabetes mellitus, we firstly constructed the yeast integrating plasmid pNK1-PGK which could successfully express green fluorescent protein (GFP) in Saccharomyces cerevisiae. The gene encoding ten tandem repeats of glucagon-like peptide-1(10×GLP-1) was cloned into the vector pNK1-PGK and the resulting plasmids were then transformed into the S. cerevisiae INVSc1. The long-acting GLP-1 hypoglycemic yeast (LHY) which grows rapidly and expresses 10×GLP-1 stably was selected by nutrition screening and Western blotting. The amount of 10×GLP-1 produced by LHY reached 1.56 mg per gram of wet cells. Moreover, the oral administration of LHY significantly reduced blood glucose level in type 2 diabetic mice induced by streptozotocin plus high fat and high sugar diet.