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Polypeptides for promoting pig body to generate broad-spectrum immune response and application of polypeptides

An immune response and pig body technology, applied in the field of biomedicine, can solve problems such as complex components, lack of immune enhancers, and no data to prove broad-spectrum effects, so as to enhance immune response, promote broad-spectrum immune response, and promote antigen specificity The effect of sexual immune response

Active Publication Date: 2021-01-01
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

It has been reported that CVC1320 is used as an immune enhancer for foot-and-mouth disease vaccines (Studies on how immune enhancers improve the immune efficacy of porcine foot-and-mouth disease inactivated vaccines, Yu Xiaoming, 2019), but the immune enhancer is a compound preparation with complex components and large dosages. relatively high production costs
Moreover, the preparation is only verified to be applicable to FMDV vaccine, and there is no data to prove that it has a broad-spectrum effect
Tang Bo et al. (Studies on Immunopotentiators Improving the Immune Efficacy of Inactivated Vaccines for Pigs, 2016) used VA5-containing immunopotentiators as companions of porcine parvovirus and JE dual inactivated vaccines, which could increase the antibody titer, but there was no data to prove its efficacy. have a broad-spectrum effect
Currently, there is an urgent need for an immune enhancer that induces a broad-spectrum immune response in pigs

Method used

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  • Polypeptides for promoting pig body to generate broad-spectrum immune response and application of polypeptides
  • Polypeptides for promoting pig body to generate broad-spectrum immune response and application of polypeptides
  • Polypeptides for promoting pig body to generate broad-spectrum immune response and application of polypeptides

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062]Example 1 Solid-phase synthesis and purity detection of polypeptide

[0063]The polypeptides of the present invention are as follows:

[0064]SEQ ID NO.1: MMIFLTYDL;

[0065]SEQ ID NO. 2: LSIGYTLLF;

[0066]SEQ ID NO. 3: HLNGFVSTL;

[0067]SEQ ID NO. 4: ASGGGPAP;

[0068]SEQ ID NO.5: GIALDLSTL;

[0069]The above-mentioned peptides were synthesized by Shanghai Shenggong Biological Engineering Co., Ltd.

[0070]The detection wavelength is 214nm. The purity of the final polypeptide purification product>98%, and the structure was identified by ESI-MS, see the identification resultsFigure 1~Figure 10 ,among themfigure 1 Is the identification chromatogram of AP1,figure 2 It is the mass spectrum of AP1, the identified molecular weight is 1146g / mol, which is consistent with the theoretical value;image 3 Is the identification chromatogram of AP2,Figure 4 It is the mass spectrum of AP2, the identified molecular weight is 1026.6g / mol, which is consistent with the theoretical value;Figure 5 Is the identification...

Embodiment 2

[0071]Example 2 Porcine lymphocyte proliferation experiment

[0072]1. Immunize pigs with ASFV inactivated virus.

[0073]Five 90-day-old male Landrace pigs were immunized with the ASFV epidemic strain inactivated virus (from the African Swine Fever Regional Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences) (10HID50), a booster immunization after one month, and implementation on pigs 7 days later Euthanize and remove the spleen after dissection. Use unimmunized healthy pigs as a negative control group.

[0074]2. Preparation, culture and proliferation of splenocytes.

[0075]1) Aseptically treat the collected pig spleen with 75% alcohol. After washing three times with PBS, cut the spleen into small pieces, place them in folded sterile gauze (2 layers), and add 5mL serum-containing 1640 medium Grind the spleen in a plate.

[0076]2) Then suck the liquid in a 15ml centrifuge tube, centrifuge at 1000rpm for 5min.

[0077]3) Discard the supernatant, knock the pe...

Embodiment 3

[0085]Example 3 Detection of IFN-γ secretion by different subtypes of splenic lymphocytes.

[0086]The porcine immunization procedure and the separation and culture of spleen cells are the same as in Example 2. After obtaining the dispersed spleen cells, prepare a single cell suspension with RPMI1640 complete medium at a concentration of 1×106 / ml. Inoculate into a 24-well plate, add 0.2 μg of the polypeptide provided by the present invention to each well, and place it in a CO2 incubator for 60 hours. Collect the cells in each well, respectively label the cells with pig CD3, CD4, CD8 and IFN-γ specific antibodies, then wash them twice with 2% serum-containing PBS buffer, and finally use the washing solution to disperse into a cell suspension. Cytometer to detect and determine CD4 separately+T lymphocytes and CD8+The level of IFN-γ secreted by T lymphocytes.

[0087]See the test resultsFigure 13 ~ Figure 17 . among themFigure 13 To promote AP1 to secrete IFN-γ levels of immune cells of diff...

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Abstract

The invention provides a class of polypeptides for promoting a pig body to generate broad-spectrum immune response and application of the polypeptides, and belongs to the technical field of biomedicine. The polypeptides include one or more of a first polypeptide, a second polypeptide, a third polypeptide, a fourth polypeptide and a fifth polypeptide; and the amino acid sequences of the first polypeptide, the second polypeptide, the third polypeptide, the fourth polypeptide and the fifth polypeptide are respectively shown as SEQ ID NO.1 to SEQ ID NO.5. The polypeptides disclosed by the invention can obviously promote the activation of an IFN-beta signal path in the pig body, promote the secretion of IFN-gamma by mononuclear macrophages, T cells and B cells and promote the proliferation of spleen lymphocytes and mononuclear macrophages, so that the pig body is promoted to generate broad-spectrum immune response; and moreover, the polypeptides disclosed by the invention can promote the ASFV antigen specific immune response in the pig body, so that the polypeptides can be used as an effective component of an ASFV subunit vaccine to enhance the immune response of the pig.

Description

Technical field[0001]The present invention relates to the field of biomedicine technology, in particular to a class of polypeptides that promote pigs to produce a broad-spectrum immune response and their applications.Background technique[0002]Immune enhancer is a type of use alone or in combination with antigens. It enhances the animal by enhancing the activity of macrophages, enhancing the immunogenicity and stability of antigenic substances, and promoting the synthesis and secretion of various immune factors and specific antibodies. The body's immune response substance.[0003]Immune enhancers can stimulate the body to produce humoral-mediated and cell-mediated immune responses, so as to eliminate invading pathogens and protect animals from the harm of pathogens.[0004]There are many types of immune enhancers, which are roughly divided into three categories according to their composition characteristics: Chinese herbal medicine extracts, chemical compounds and cytokines. There are fe...

Claims

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Application Information

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IPC IPC(8): C07K7/06A61K39/39A61K39/12A61P37/04A61P31/20
CPCC07K7/06A61K39/39A61K39/12A61P37/04A61P31/20A61K2039/55516C12N2710/12034Y02A50/30
Inventor 郑海学毛箬青周晓丽孙德惠朱昱茜张克山张忠献杨帆刘华南王尧河刘湘涛
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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