Application of bovine beta defensin 5 as novel mucosal immunologic adjuvant

A technology of immune adjuvant and beta defensin, applied in the field of tuberculosis prevention and treatment

Active Publication Date: 2021-06-04
CHINA AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Bacillus Calmette-Guerin (BCG) is currently a vaccine widely used to prevent tuberculosis. It can prevent tuberculo

Method used

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  • Application of bovine beta defensin 5 as novel mucosal immunologic adjuvant
  • Application of bovine beta defensin 5 as novel mucosal immunologic adjuvant

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] The preparation method of embodiment 1 recombinant protein B5

[0022] 1.1 Induced expression of B5

[0023] Refer to Kang Jingjing (Kang, J.; Zhao, D.; Lyu, Y.; et al. Antimycobacterial activity of Pichia pastoris-derived mature bovine neutrophilβ-defensins 5. Eur. J. Clin. Microbiol. Infect. Dis. 2014 , 33, 1823–1834) and other previous studies to prepare bovine β-defensin 5 methods. Use 50 mL of YPD medium to cultivate recombinant Pichia pastoris GS115 expressing bovine β-defensin 5 (the expression vector is pPIC9K), shake at 28°C, culture at 180rpm for 24h, centrifuge at 4000rpm for 5min, discard the YPD medium, and transfer the cells to a 200mL BMGY medium, shaker at 28°C, culture at 180rpm for 24h, centrifuge at 4000rpm for 5min, discard BMGY medium, transfer the bacteria to 1L of BMMY medium, shaker at 28°C, culture at 180rpm for 72h, add methanol every 24h The final concentration is 1%. Centrifuge at 8000 rpm for 3 min at 4°C, and collect the culture supernat...

Embodiment 2

[0026] Example 2 Evaluation of the immune effect of B5-promoting subunit vaccine AH

[0027] The subunit vaccine AH contains the dominant antigens Ag85A and HspX of Mycobacterium tuberculosis, with a flexible Linker (GGGGS) 3 Connect Ag85A and HspX, insert the prokaryotic expression vector pET-30a(+) through KpnI and BamH I double digestion, and transform into Escherichia coli competent cells BL21(DE3), add IPTG to induce expression, and the expressed inclusion body protein is purified, The fusion protein AH was obtained by endotoxin removal and filter sterilization.

[0028] Sixty 6-8-week-old female C57BL / 6 mice were randomly divided into 6 groups, 10 in each group, divided into normal control group (Control), BCG control group (BCG), polymyocyte and AH immune group (AH- P), B5 combined with polymyocytes and AH immunization group (AH-PB), polymyocytes combined with B5 adjuvant control group (PB), challenge model group (M.bovis). The immunization doses of polymyocytes, B5 a...

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Abstract

The invention provides application of bovine beta defensin 5 as a mucosal immunologic adjuvant in prevention of tuberculosis. A C57BL/6 mouse is immunized by expressing and purifying recombinant protein bovine beta defensin 5, combining polyinosinic-polycytidylic acid (PolyI: C) and a mycobacterium tuberculosis protein subunit vaccine AH in a nasal drop mode, and the effect of promoting the immune effect of the protein subunit vaccine AH is proved. Compared with polyinosinic-polycytidylic acid and AH immunization, the bovine beta defensin 5 can promote antigen AH specific cellular immunization and humoral immunization levels and promote the protective effect of subunit vaccine AH on resisting bovine mycobacterium infection.

Description

technical field [0001] The invention belongs to the field of tuberculosis prevention and control and the field of vaccines. Specifically, the application provides a mucosal immune adjuvant containing bovine beta defensin 5, which is especially suitable for tuberculosis subunit vaccines. Background technique [0002] Tuberculosis is a chronic infectious disease caused by Mycobacterium tuberculosis complex. Mycobacterium tuberculosis and Mycobacterium bovis (M.bovis for short) are the main pathogenic bacteria. Bovine tuberculosis caused by Mycobacterium bovis is a It is an important zoonotic disease that endangers the development of breeding industry and public health security. Mycobacterium bovis can break through the interspecies barrier and spread among other mammals and humans, with a wide range of hosts. The current investigation and research shows that bovine tuberculosis in my country has the characteristics of easy recurrence and national distribution, and it is trans...

Claims

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Application Information

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IPC IPC(8): A61K39/39A61K39/04A61P31/06
CPCA61K39/39A61K39/04A61P31/06A61K2039/55516A61K2039/55511A61K2039/543Y02A50/30
Inventor 周向梅梁正敏刘一朵王元智葛昕
Owner CHINA AGRI UNIV
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