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Long non-coding RNA (Ribonucleic Acid) and application thereof in diagnosis/treatment of preeclampsia

A preeclampsia, non-coding technology, applied in the field of genetic engineering, can solve the problems of incomplete functional research of lncRNAs and achieve high transfection efficiency

Active Publication Date: 2017-05-31
JIANGSU PROVINCE HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Recently, the role of miRNAs in various aspects of cellular processes has been confirmed, however, the function of lncRNAs is not very thorough

Method used

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  • Long non-coding RNA (Ribonucleic Acid) and application thereof in diagnosis/treatment of preeclampsia
  • Long non-coding RNA (Ribonucleic Acid) and application thereof in diagnosis/treatment of preeclampsia
  • Long non-coding RNA (Ribonucleic Acid) and application thereof in diagnosis/treatment of preeclampsia

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0092] Detect the expression of PVT1 in tissues and cells

[0093] Take 0.1g tissue, grind it sufficiently with liquid nitrogen (into a powder form) or discard the culture medium for 1-5×107 cells, and rinse twice with pre-cooled PBS. Add 1ml of Trizol lysate, pipette and mix with an enzyme-free pipette tip, let stand for 5min, and transfer the lysate into a pre-labeled 1.5ml enzyme-free centrifuge tube. Centrifuge at 7500g at 4°C for 5 minutes, take the supernatant and add 1 / 5 volume of chloroform, invert and mix for 30 seconds, and let stand for 2 minutes. Centrifuge at 12000g for 15min at 4°C. The solution is divided into three layers (aqueous phase-white precipitate-red organic matter), transfer the aqueous phase layer to a new 1.5ml centrifuge tube, try not to absorb the white precipitate. Add an equal volume of isopropanol, mix gently by inversion, and let stand for 5-10min. Centrifuge at 12000g for 10min at 4°C. Discard the supernatant, add 1ml of 75% ethanol (prepa...

Embodiment 2

[0120] To study the effect of PVT1 on the phenotype of normal trophoblast HTR-8 / SVneo cells.

[0121] First, the normal trophoblast HTR-8 / SVneo cell line was selected as the research object of this experiment, and the PVT1 interference sequence si-PVT 1#GCUUGGAGGCUGAGGAGUUTT (SEQ ID NO: 2) and si-PVT2# AACUCCUCAGCCUCCAAGCTT ( SEQ ID NO: 3) The pathogenesis of preeclampsia was simulated by knocking down the expression of PVT1. The MTT proliferation assay found that HTR-8 / SVneo cells were transfected with the interfering sequence of PVT1, and the cell growth was inhibited after knocking down the expression of PVT1. In contrast, the PVT1 plasmid was transfected in normal trophoblast cells to further verify the functional role of PVT1, and found that the overexpression of PVT1 promoted the proliferation of HTR-8 / SVneo cells ( figure 2 A-D). In addition, the results of colony formation assays showed that downregulation of PVT1 weakened the colony formation ability of HTR-8 / SVneo ...

Embodiment 3

[0123] Effects of PVT1 on cycle and apoptosis of placental trophoblast cell HTR-8 / SVneo

[0124] In order to study whether PVT1 affects the cell cycle transition on the proliferation of HTR-8 / SVneo cells, the normal trophoblast HTR-8 / SVneo cell line was used as the research object, and lip2000 was used as the carrier to transfect the PVT1 interference sequence si-PVT1#GCUUGGAGGCUGAGGAGUUTT( SEQ ID NO: 2) and si-PVT 2#AACUCCUCAGCCUCCAAGCTT (SEQ ID NO: 3) were used to knock down the expression of PVT1 to simulate the pathogenesis of preeclampsia. Cell cycle analysis by flow cytometry. The results showed that after HTR-8 / SVneo cells were transfected with PVT1 interference sequence, it was found that the cell cycle was arrested in the G1 / G0 phase after knocking down the expression of PVT1. On the contrary, the PVT1 plasmid was transfected in normal trophoblast cells to further verify the function of PVT1 , it was found that the proportion of cell cycle arrest in G1 / G0 phase decre...

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Abstract

The invention belongs to the field of genetic engineering, and in particular relates to application of PVT1 in preparation and diagnosis of preeclampsia and target drug therapy. The down-regulation of PVT1 in a placenta tissue of a preeclampsia pregnant woman is related to the generation and development of preeclampsia, and the low-expression-level PVT1 is closely related to the pathogenesis of preeclampsia. The proliferation, apoptosis, invasion, migration and the like of trophoblast cells of the preeclampsia pregnant woman are affected by changing the expression of PVT1, and the proliferation, invasion and migration of trophoblast cells can be promoted by enhancing the expression of PVT1.

Description

technical field [0001] The invention belongs to the field of genetic engineering, and particularly relates to the application of long non-coding RNA-PVT1 in diagnosis and preparation of drugs for treating preeclampsia. Background technique [0002] Preeclampsia is one of the most common pregnancy complications in the world, and it is the main cause of death related to pregnancy complications. According to the statistics of the World Health Organization, preeclampsia or eclampsia directly leads to about 14% (about 8500) of the world's annual Maternal death, which is one of the most common causes of preterm birth and fetal growth restriction and maternal death. Although the current medical treatment has been greatly developed, but the overall incidence of patients is still high. With the rapid development of sequencing technology and molecular biology, gene diagnosis and molecular targeted therapy have become a hot issue in the treatment of preeclampsia. Therefore, the study...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12Q1/68C12N15/11A61K48/00A61K31/7105A61P15/00A61P9/12
CPCA61K31/7105C12N15/113C12N2310/10C12N2310/141C12Q1/6883C12Q2600/136C12Q2600/158C12Q2600/178
Inventor 孙丽洲许叶涛左青黄诗韵葛志平吴丹
Owner JIANGSU PROVINCE HOSPITAL
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