Recombinant Lactobacillus paracasei Expressing Porcine β-Defensin 2 Attenuates DSS-Induced Colitis by Restoring Intestinal Barrier Integrity and Modulating Immune Homeostasis

Description

Figure 1. DSS-induced colitis model. Figure 2. The recombinant L. paracasei 27-2 strains expressing pBD2 were identified by PCR, Western blot and IFA. (a). Double enzyme digestion results of pPG-PPT plasmid. (b) PCR amplification of the N1-pBD2 fragment. (c) PCR identification of the pPG-N1-pBD2 plasmid. (d) PCR Identification Results of recombinant L. paracasei 27-2. (e) Western Blot identification of the supernatant and precipitation from pPG-N1-pBD2/27-2 culture. (f) Identification of proteins expressed in the pPG-N1-pBD2/27-2. Figure 3. The growth characteristics and stability of recombinant L. paracasei 27-2 were analyzed. (a) The determination of growth curve of pPG-N1-pBD2/27-2. (b-c) The genetic and protein expression stability of pPG-N1-pBD2/27-2 underwent continuous subculturing for 20 generations. Figure 4. Analysis of the activity of pBD2 in vitro. The toxicity assessment (a), proliferation assay (b) and migration effect (c-d) of pBD2 protein expressed by recombinant L. paracasei 27-2 on IPEC-J2 were analyzed. Figure 5. Protective effects of orally administered recombinant L. paracasei 27-2 on intestinal injury in a mice Model of Colitis. (a) The mice were scored according to the DAI scoring table (b) After the completion of the modeling, a section of the colon (approximately 1cm in length) near the rectum was excised from the mice of each group for H&E staining. Note: Green arrows indicate epithelial cell shedding and hemorrhage; dark blue arrows indicate inflammatory cell infiltration; yellow arrows indicate near-complete loss of crypt architecture; red arrows indicate reduced and atrophic glands; light blue arrows indicate decreased and deformed goblet cells. (c) The length of the colon of each group of mice was measured. Figure 6. Measurement results of colonic tight junction protein content in each group of mice (a) Western blot analysis of tight junction protein expression in mouse colon. (b) qPCR analysis of tight junction protein expression in mouse colon. Figure 7. Effects of recombinant L. paracasei 27-2 on colitis-related enzyme levels in the colon and serum cytokine levels in mice. (a) The activity of MPO, EPO, and NAG in mouse colons across treatment groups. (b) Measurement results of iNOS and COX-2 related enzyme concentrations in each group of mice. (c) Detection results of inflammatory cytokine secretion levels in mice of each group. Figure 8. Proportions of immune cell populations in the spleens of mice from different groups. Proportions of B lymphocytes (a), T lymphocytes (b), macrophages (c) and NK cells (d),in the spleens of mice from different treatment groups.

Institutions

• Northeast Agricultural University

  Harbin, Harbin

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