Construction of Recombinant Lactococcus lactis Expressing Porcine Epidermal Growth Factor and Its Repair Effects on Intestinal Damage of Colitis Model in Mice

  • LIU Shujie ,
  • TAO Xin ,
  • DENG Bo ,
  • MEN Xiaoming ,
  • XU Ziwei
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  • Institute of Animal Husbandry and Veterinary Science, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China

Received date: 2020-10-28

  Online published: 2021-04-15

Supported by

 

Abstract

This study aimed to construct recombinant Lactococcus lactis (L. lactis) expressing porcine epidermal growth factor (pEGF), evaluate its repair effects on intestinal damage of colitis model induced by dextra sulfate sodium (DSS) in mice, and further provide an important foundation in order that recombinant L. Lactis could be applied to intestinal protection in weaned piglets. Three copies of mature pEGF with the same nucleotide sequence were designed by tandem repeat strategy. The gene sequence of designed 3pEGF was further optimized based on codon bias of L. lactis. The resulting fragment was synthesized, and then was cloned into pNZ8148 vector. Recombinant L. lactis expressing pEGF was constructed. Thirty-two BALB/C mice were randomly allotted in four groups with eight mice per group. Colitis model was induced by 4% DDS in mice. The mice in the four groups were treated as follow:normal control group received normal drinking water on the 1st to 12th day; DSS model control group received 4% DSS solution in drinking water on the 1st to 7th days and followed with normal drinking water on the 8th to 12th days; L. lactis group received the same treatment of DSS as DSS model control group, meanwhile, these mice were orally administered with 1×1012 CFU/d of L. lactis for consecutive 12 days; recombinant L. lactis group received the same treatment of DSS as DSS model control group, meanwhile, these mice were orally administered with 1×1012 CFU/d of recombinant L. lactis for consecutive 12 days. The results showed as follows:1) by sequence identification, the 3pEGF had been successfully transformed into L. lactis. Western blot analysis results revealed that pEGF could react with the specific antibody, which primarily indicated that pEGF had biological activity. 2) DSS treatment significantly shortened colon length compared with normal control group (P<0.05), and oral administration of recombinant L. lactis significantly increased colon length compared with DSS model control group (P<0.05). 3) DSS treatment significantly decreased colonic occludin concentration (P<0.05), significantly increased colonic D-lactate (D-LAC) concentration (P<0.05) and diamine oxidase (DAO) activity (P<0.01) compared with normal control group. Oral administration of recombinant L. lactis significantly increased colonic occludin concentration (P<0.05), and significantly decreased colonic D-LAC concentration (P<0.05) and DAO activity (P<0.01) compared with DSS model control group. 4) DSS treatment significantly decreased colonic interleukin-10 (IL-10) (P<0.01) and interleukin-4 (IL-4) concentrations (P<0.05), and significantly increased colonic tumor necrosis factor-α (TNF-α) concentration (P<0.01) compared with normal control group. Oral administration of recombinant L. lactis significantly enhanced colonic IL-10 (P<0.01) and IL-4 concentrations (P<0.05), and significantly decreased colonic TNF-α concentration compared with DSS model control group. In conclusion, recombinant L. lactis expressing pEGF is successfully constructed, and recombinant pEGF possesses good biological activity. Oral administration of recombinant L. lactis in mice can prevente colon shortening, improve intestinal permeability, and regulate the concentrations of intestinal cytokines to normal levels, these show that recombinant L. lactis expressing pEGF can maintain the integrity of intestinal mucosal barrier, and has a certain effect for repairing the damaged intestinal tissue.

Cite this article

LIU Shujie , TAO Xin , DENG Bo , MEN Xiaoming , XU Ziwei . Construction of Recombinant Lactococcus lactis Expressing Porcine Epidermal Growth Factor and Its Repair Effects on Intestinal Damage of Colitis Model in Mice[J]. Chinese Journal of Animal Nutrition, 2021 , 33(4) : 2253 -2262 . DOI: 10.3969/j.issn.1006-267x.2021.04.044

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