[Expression of human insulin in lactic acid bacteria and its oral administration in non-obese diabetic mice].
Type 1 diabetes mellitus (T1DM) is an auto-immune disease while oral administrating its autoantigens could be a treatment of T1DM. To express human insulin gene (ins) in lactic acid bacteria (LAB) for oral vaccine, ins gene was replaced by LAB bias codon and an 8-amino-acid-residue linker peptide forming a beta-turn was designed to link insulin chain A and B. After synthesized by primer annealing method, the whole ins gene was fused with signal peptide sequence SP(Usp45), subcloned into a LAB secretory expressive vector pSW501 and then introduced to Lactococcus lactis (L. lactis) MG1363 and Lactobacillus casei (Lb. casei ) ATCC27092 respectively. Western blot showed that the expression product (SP(Usp45)-INS protein) targeted mainly at the cell wall while little was found in cytoplasm or supernatant. The highest expression level emerged in exponential phase when the optical density at 600nm of the culture was 0.4. The culture of the recombinant strain Lb. casei/pSW501 was administered to non-obese diabetic (NOD) mice orally. ELISA and Western blot results showed that the recombinant strain could induce SP(Usp45)-INS-specific antibodies and raise IL-4 level (38.583 +/- 2.083 pg/mL, P < 0.05) in the mice' s sera. Expression of insulin in the food-grade vehicle LAB could induce oral immune tolerance in NOD mice and protect it from pancreas injury, suggesting it might be a new way to the treatment of T1DM.
Chen
Si-Wei
SW
State Key Laboratory of Microbial Resources, Chinese Academy of Sciences, Beijing 100101, China. chensiwei@mails.gucas.ac.cn
Zhong
Jin
J
Huan
Lian-Dong
LD
D000284:Administration, Oral; D000818:Animals; D001483:Base Sequence; D003922:Diabetes Mellitus, Type 1 / Q000188:drug therapy*; D006801:Humans; D007328:Insulin / Q000235:genetics* / Q000627:therapeutic use; D007780:Lacticaseibacillus casei / Q000235:genetics*; D013294:Lactococcus lactis / Q000235:genetics*; D051379:Mice; D016688:Mice, Inbred NOD; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000627:therapeutic use
D007328:Insulin; D011994:Recombinant Proteins