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lncRNA marker lncAIS of adolescent idiopathic scoliosis (AIS) and application of lncAIS in preparation of AIS detection, prognosis, prevention or treatment products

A scoliosis, idiopathic technology, applied in the field of biomedicine, can solve the problem of unclear biological role of lncRNA

Active Publication Date: 2019-03-01
PEKING UNION MEDICAL COLLEGE HOSPITAL CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the biological roles of lncRNAs in the pathogenesis of AIS are still unclear

Method used

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  • lncRNA marker lncAIS of adolescent idiopathic scoliosis (AIS) and application of lncAIS in preparation of AIS detection, prognosis, prevention or treatment products
  • lncRNA marker lncAIS of adolescent idiopathic scoliosis (AIS) and application of lncAIS in preparation of AIS detection, prognosis, prevention or treatment products
  • lncRNA marker lncAIS of adolescent idiopathic scoliosis (AIS) and application of lncAIS in preparation of AIS detection, prognosis, prevention or treatment products

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0095] Example 1: Identification of lncAIS as a Significantly Differentially Expressed lncRNA Downregulated in AIS Patient Bone Marrow Mesenchymal Stem Cells (BM-MSCs)

[0096] To identify key lncRNAs involved in adolescent idiopathic scoliosis (AIS), we performed microarray analysis of BM-MSCs from 5 healthy donors and 12 AIS patients. 1483 lncRNAs showed differential expression between normal BM-MSCs and BM-MSCs of patients with AIS, of which 718 were up-regulated and 765 were down-regulated, such as figure 1 as shown in a. Among the most downregulated lncRNAs in BM-MSCs of AIS patients, we focused on an uncharacterized lncRNA we termed lncAIS (gene symbol: ENST00000453347). lncAIS is located on human chromosome 1, contains 4 exons, and is a full-length 476nt transcript. lncAIS was identified as a conserved locus, as figure 1 as shown in b.

[0097] Perform real-time qPCR according to routine operations, analyze lncAIS transcripts in BM-MSCs of normal healthy donors and ...

Embodiment 2

[0103] Example 2: lncAIS knockdown inhibits osteogenic differentiation and bone formation

[0104] To explore the physiological role of lncAIS in the pathogenesis of AIS, we knocked down lncAIS in healthy normal BM-MSCs using lentivirus-mediated short hairpin RNA (shRNA) (the shLncAIS gene sequence for lncAIS knockdown is shown in Table 2 above shown in SEQ ID NO:4-6), and confirmed the knockdown efficiency by real-time quantitative PCR, as shown in figure 2 as shown in a. Normal BM-MSCs were infected with shlncAIS-expressing lentivirus and cultured in MSC medium for 3 days, then the mRNA level of lncAIS was detected by real-time qPCR. The primer sequences used for cDNA amplification are shown in Table 3 above. Relative gene expression fold changes were normalized to endogenous β-actin and counted as mean ± S.D. **, P<0.01. Data are from three independent experiments using BM-MSCs derived from 3 healthy donors.

[0105] Cell proliferation of shlncAIS BM-MSCs and shCtrl (...

Embodiment 3

[0113] Example 3: Overexpression of lncAIS promotes osteogenic differentiation and bone formation

[0114] We overexpressed lncAIS in normal BM-MSCs by lentivirus, first constructed the target sequence into the pSicoR plasmid, and used pMD2.G and psPAX2 vectors to package the virus, and then infected the target BM-MSCs. lncAIS overexpression (oeLncAIS) significantly increased cell proliferation of normal BM-MSCs ( image 3 a). Consistently, lncAIS overexpression enhanced the expression levels of self-renewal-related genes (NANOG, POU5F1, and SOX2) and osteogenic differentiation genes (ALPL, BSP, RUNX2, LPL, and PPAR) ( image 3 b, 3c). Furthermore, lncAIS overexpression also increased osteogenic marker genes such as osteopontin (OPN), COL1A1 and IBSP ( image 3 d) Protein levels. Therefore, by ALP staining ( image 3 e) and Von Kossa staining ( image 3 f) Overexpression of lncAIS enhanced osteogenic differentiation of BM-MSCs. Finally, lncAIS-overexpressed BM-MSCs sign...

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Abstract

The invention discloses an lncRNA marker lncAIS of adolescent idiopathic scoliosis (AIS) and application of the lncAIS in preparation of AIS detection, prognosis, prevention or treatment products. Theinvention provides a kit for AIS detection and prognosis. The invention further provides a pharmaceutical composition for preventing and / or treating the AIS. The pharmaceutical composition comprisesthe lncAIS or an agonist or an analogue of the lncAIS, wherein the agonist or the analogue of the lncAIS is used as an active ingredient. The lncAIS can be used for early detection of AIS risk or disease status, and can be used for clinical application such as gene therapy and drug therapy.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to an lncRNA marker related to the detection and prognosis of adolescent idiopathic scoliosis (AIS) and its application. Background technique [0002] Adolescent idiopathic scoliosis (AIS) is a complex three-dimensional deformity of the spine that occurs mainly in girls aged 10 to 16 years during the pubertal growth period. The progression of scoliosis may lead to appearance deformity, back pain, and functional impairment. In severe cases, physiological function problems such as limited cardiopulmonary function will occur, which will affect social activities to varying degrees. Epidemiological survey results show that the global incidence of AIS among adolescents is about 2-4%, and about 10% of those diagnosed with AIS require treatment. The current treatment of AIS includes bracing therapy and surgical correction; full-time bracing therapy may cause back pain and psychological...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883A61K45/00A61P19/08
CPCA61K45/00A61P19/08C12Q1/6883C12Q2600/178C12Q2600/118C12Q2600/158
Inventor 庄乾宇仉建国惠尚懿范祖森邱贵兴吴志宏叶步青赵春华李静李娜王升儒张延斌杨阳林莞峰
Owner PEKING UNION MEDICAL COLLEGE HOSPITAL CHINESE ACAD OF MEDICAL SCI
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