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Application of m6A modified protein in preparation of toxoplasma gondii infection resisting medicine

A technology of Toxoplasma gondii and protein, applied in the field of molecular biology, can solve problems affecting the level of modification

Pending Publication Date: 2021-12-31
NANJING MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But in the macrophage immune response induced by parasites, especially the parasitic protozoa-toxoplasma gondii, does the parasite affect the host m 6 A modifies protein expression, thereby affecting the m of macrophage RNA 6 The level of A modification and its function have not been reported yet

Method used

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  • Application of m6A modified protein in preparation of toxoplasma gondii infection resisting medicine
  • Application of m6A modified protein in preparation of toxoplasma gondii infection resisting medicine
  • Application of m6A modified protein in preparation of toxoplasma gondii infection resisting medicine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Embodiment 1 Materials and methods

[0055] 1 Materials and Reagents

[0056] The HFF human foreskin fibroblast cell line was donated to the laboratory of Teacher Yu Li, Department of Pathogen Biology, Anhui Medical University; the THP-1 human peripheral blood mononuclear cell line was donated to the laboratory of Teacher Zhang Mingshun, Department of Immunology.

[0057] Toxoplasma gondii RH strain tachyzoites and RH strain tachyzoites expressing green fluorescent protein (GFP) were donated to the laboratory of Teacher Yu Li, Department of Pathogen Biology, Anhui Medical University.

[0058] 2 methods

[0059] 2.1 Culture of HFF human foreskin fibroblast cell line

[0060] The HFF human foreskin fibroblasts in the cryopreservation tube were revived and subcultured, and when the cell density was about 90%, they were frozen and stored for later use.

[0061] 2.2 Culture of THP-1 human peripheral blood mononuclear cell line

[0062] 2.2.1 Resuscitate and subculture th...

Embodiment 2

[0067] Example 2 Characteristics of the immune response of macrophages infected with Toxoplasma gondii

[0068] In order to understand the effect of Toxoplasma gondii infection on the function of human macrophages, the tachyzoites of Toxoplasma gondii RH strain were used to infect THP-1-derived macrophages, and the expression of inflammation-related genes and secretion of inflammation-related cytokines in THP-1 macrophages were observed .

[0069] 1 Expression of inflammation-related genes

[0070] First, THP-1 macrophages were infected with tachyzoites of Toxoplasma gondii RH strain for 6h, 12h, 18h and 24h, the ratio of cells to tachyzoites was 1:1, and qRT-PCR was used to detect inflammation-related genes TNF-α, iNOS , Arg-1, IL-1β, IL-10, etc. in the expression of 6h, 12h, 18h and 24h. The result is as figure 1 As shown, the mRNA levels of TNF-α, IL-1β and IL-6 were significantly up-regulated at 6h, 12h, 18h and 24h after infection, and the up-regulation range was IL-6>...

Embodiment 3

[0073] Example 3 Toxoplasma gondii infection to m 6 Effect of A modified protein

[0074] In the process of pathogens regulating host immune response, the regulation of transcription level is only one aspect of gene expression regulation. Moreover, compared with transcriptional regulation, post-transcriptional mRNA regulation can help cells respond to external stimuli more quickly. RNA m 6 As an important way of post-transcriptional regulation, A modification is involved in the regulation of host immune response in various pathogen infection models. Therefore, the present invention firstly observed the effect of toxoplasma infection on m 6 A Modified protein effect.

[0075] 1 Toxoplasma infection on m 6 Effect of A modified protein mRNA level expression

[0076] Toxoplasma gondii RH strain tachyzoites were used to infect THP-1 macrophages for 6h, 12h, 18h and 24h. The ratio of cells to tachyzoites was 1:1, and m 6 A The mRNA levels of methyltransferases, demethylases a...

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Abstract

The invention discloses an application of m6A modified protein in preparation of a toxoplasma gondii infection resisting medicine, and belongs to the field of molecular biology. The application achieves the purpose of resisting toxoplasma gondii infection by regulating the expression level of the m6A modified protein and regulating the expression of an inflammatory factor TNF-alpha. The invention finds that m6A modification sites exist on human TNF-alpha mRNA, after toxoplasma gondii infection, the modification degree of the modification sites on the TNF-alpha mRNA can be reduced by up-regulating expression of demethylase FTO, and degradation of the TNF-alpha mRNA is reduced and TNF-alpha expression is induced under the help of recognition protein YTHDF2. From the perspective of m6A methylation modification for the first time, the invention discloses a molecular mechanism of TNF-alpha expression induced by toxoplasma gondii infection, and changes of m6A modification sites and modification levels on TNF-alpha after human macrophages are infected by toxoplasma gondii, and clarifies a mechanism of regulating TNF-alpha expression by m6A modification.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a m 6 Application of A modified protein in preparation of anti-toxoplasma infection medicine. Background technique [0002] Toxoplasma gondii (T.gondii) is a common obligate intracellular parasitic protozoa. Toxoplasma gondii can parasitize most of the nucleated cells when it invades the host, and can infect almost all warm-blooded animals including humans. Toxoplasma gondii is an important opportunistic pathogenic protozoa. After people with normal immune function are infected with Toxoplasma gondii, the parasites usually live in the form of cysts in the brain, eyes and skeletal muscles, and usually do not show obvious clinical symptoms ; but in immunocompromised populations such as the elderly and AIDS patients, Toxoplasma infection can develop into acute toxoplasmosis, causing serious diseases such as encephalitis, eye disease or pneumonia, and even death; in addition, Toxop...

Claims

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Application Information

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IPC IPC(8): A61K38/45A61K38/44A61P33/02
CPCA61K38/45A61K38/44A61P33/02
Inventor 邱竞帆王勇秦敏邵天业缪婷婷张戎刘新建
Owner NANJING MEDICAL UNIV
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