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Application of IncRNA in regulating polarization of macrophages in viral myocarditis

A viral myocarditis and macrophage technology, applied in the field of bioengineering, can solve the problem that the mechanism of macrophage polarization remains to be explored.

Active Publication Date: 2019-11-22
THE FIRST AFFILIATED HOSPITAL OF WANNAN MEDICAL COLLEGE YIJISHAN HOSPITAL OF WANNAN MEDICAL COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, despite the importance of this process for VM, the mechanism of macrophage polarization remains to be explored

Method used

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  • Application of IncRNA in regulating polarization of macrophages in viral myocarditis
  • Application of IncRNA in regulating polarization of macrophages in viral myocarditis
  • Application of IncRNA in regulating polarization of macrophages in viral myocarditis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0095] The applicants performed microarray analysis on M1 and M2 macrophages polarized in vitro to identify lncRNAs involved in macrophage polarization, and the results were as follows figure 1 As shown in A. Depend on figure 1 A shows that: when the threshold of differential expression is set to fold change ≥ 2, and P<0.05, there are 627 highly expressed lncRNAs in M1 macrophages, and 624 highly expressed lncRNAs in M2 macrophages.

[0096] According to lncRNA reverse target gene prediction combined with the adjacent gene function of differentially expressed lncRNAs, the applicant selected several lncRNAs, and used RT-qPCR to verify the chip results, and the results are shown in Figure 1B. Depend on figure 1 B: After RT-qPCR analysis, the results of 4 lncRNAs (AK048798, AK085865, AK083884, AK153212) with significant expression differences in M1 / M2 were consistent with the chip.

[0097] Previous studies have found that there are differences in the phenotypes of myocardial ...

Embodiment 2

[0099] Since many lncRNAs have been proven to have positive or negative regulatory effects on their neighboring genes, the genomic location of their neighboring genes needs to be further analyzed, such as figure 2 As shown in A. AK085865 is located on chromosome 6 of the mouse and is transcribed in the second intron of the protein-coding gene PPARγ.

[0100] like figure 2 As shown in B, the applicant used Rapid Amplification of cDNA Ends (RACE) to determine the length of AK085865 to be 1266 bp.

[0101] After nucleoplasm separation, RNA was extracted for RT-qPCR detection, and the results were as follows: figure 2 C shown. Depend on figure 2 C It can be seen that about 70% of AK085865 transcripts are located in the nucleus.

[0102] like figure 2 As shown in D, fluorescence in situ hybridization (FISH) also shows that AK085865 is mainly located in the nucleus, suggesting that AK085865 may exert its biological function in the nucleus. Consistent with the definition ...

Embodiment 3

[0105] The applicant used RT-qPCR to detect the expression level of AK085865 during the polarization process of macrophages (M1 and M2), and GAPDH was used as an internal reference. The results are as follows image 3 As shown in A. Depend on image 3 A shows that the AK085865 level of M2 macrophages is significantly higher than that of M1 macrophages.

[0106] The applicant used LPS and IFN-γ to stimulate M2 macrophages or IL-4 to stimulate M1 macrophages to reverse the phenotype of macrophages to determine whether AK085865 contributes to the plasticity of macrophage polarization . M1 macrophages were cultured in fresh medium containing IL-4 for another 2 days to induce the transition from M1 to M2; M2 macrophages were cultured in fresh medium containing LPS and IFN-γ for 2 days, To induce the transformation of M2 to M1; RT-qPCR detects the level of AK085865, and the results are as follows image 3 B. image 3 C shown. Depend on image 3 B and image 3 C shows that th...

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Abstract

The invention discloses application of long-chain non-coding RNA in regulating polarization of macrophages in viral myocarditis. The applicant found that the differential expression of lncRNA AK085865in M1 / M2 macrophages is the most significant, and that the expression level of AK085865 in M2 macrophages is higher than the expression level of AK085865 in M2 macrophages. The down-regulation of AK085865 gene reduces the phenotypic expression of M2 and promotes the polarization of M1 phenotype. Meanwhile, the susceptibility of AK085865- / - gene knockout mice to CVB3-induced VM increases. In the AK085865- / - gene knockout VM model mice, M1 macrophages increase significantly while the number of M2 cells decreases. The AK085865 specifically interacts with an interleukin enhancer binding factor 2,and plays a negative regulation role in the ILF2 / ILF3 complex-mediated microRNA processing pathway and promotes the polarization of M2 macrophages.

Description

technical field [0001] The invention relates to the technical field of bioengineering, in particular to the application of a long-chain non-coding RNA regulating macrophage polarization in viral myocarditis. Background technique [0002] Long non-coding RNAs (Long non-coding RNAs, lncRNAs) are a class of transcripts with a length of more than 200 bases that do not have protein-coding potential. These RNA molecules can be intergenic (between protein-coding genes; long intergenic non-coding RNA [lincRNA]), intronic, natural antisense transcripts (NATs), or transcribed from different enhancers and promoters form. LncRNAs regulate gene transcription by binding to chromatin modifiers, nuclear heterogeneous nuclear proteins (hnRNPs) or transcription factors. In addition, lncRNAs target and regulate the splicing, modification or translation of host mRNAs through post-transcriptional mechanisms. Although lncRNAs have been identified in almost all immune cells, their functions in ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6883A61K31/7105A61P9/00A61P31/14
CPCA61K31/7105A61P9/00A61P31/14C12Q1/6883C12Q2600/158C12Q2600/178
Inventor 吕坤张莺莺李雪琴
Owner THE FIRST AFFILIATED HOSPITAL OF WANNAN MEDICAL COLLEGE YIJISHAN HOSPITAL OF WANNAN MEDICAL COLLEGE
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