Analysis of the effect of siRNA silencing α1 antitrypsin on rheumatoid arthritis fibroblasts
10.3760/cma.j.issn.1007-7480.2019.09.008
- VernacularTitle: 干扰小RNA沉默α1抗胰蛋白酶对类风湿关节炎成纤维细胞的作用探讨
- Author:
Yan ZHAO
1
,
2
;
Shengnan CAO
3
;
Guodong SUN
3
;
Jihong PAN
1
;
Xiaotian CHANG
4
;
Qingsong MENG
5
Author Information
1. Shandong Medicinal Biotechnology Center, Shandong First Medical University & Shandong Academy of Medical Sciences, Jinan 250062, China
2. Shandong Normal University, College of Life Sciences, Jinan 250014, China
3. Department of Rehabilitation Medicine, Affiliated Hospital of Shandong Academy of Medical Sciences, Jinan 250031, China
4. Laboratory Department, Shandong Provincial Qianfoshan Hospital, the First Hospital Affiliated with Shandong First Medical University, Jinan 250014, China
5. Medical Laboratory Center, Shandong Provincial Qianfoshan Hospital, the First Hospital Affiliated with Shandong First Medical University, Jinan 250014, China
- Publication Type:Journal Article
- Keywords:
Alpha-antitrypsin;
Arthritis, rheumatoid;
Fibroblasts;
RNA, small interferaing
- From:
Chinese Journal of Rheumatology
2019;23(9):617-622,插2
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effect of siRNA silencing α1 antitrypsin (α1-AT) gene on the biological behavior of rheumatoid arthritis fibroblast-like synoviocytes (RA-SFs).
Methods:Primary culture of knee synovial tissue from 5 patients with rheumatoid arthritis (RA) was performed. The artificially synthesized silencing α1-AT siRNA specifically inhibits the expression of α1-AT in RA-SFs. After 24 and 36 hours of transient transfection, the inhibition efficiency was detected by Quantitative reverse transcription polymerase chain reaction (RT-qPCR), and the expression of related genes after α1-AT gene silencing was detected.Furthermore, ethyl thiazolyl tetrazolium (MTT) assay, Trans-well chamber, cell scratch and enzyme-linked immunosorbent assay (ELISA) were used to detect the effects of interfering α1-AT expression on cell proliferation, invasion and migration, and secretion of interleukin (IL)-17, Tumor necrosis factor (TNF)-α, IL-1α, IL-1β and other related inflammatory factors. At the same time, when the pathway inhibitor (ERK inhibitor, signal transducer and activator of transcription 3 (STAT3) inhibitor, NF-κB inhibitor) stimulated cells, the effect on α1-AT was changed. One-way analysis of variance was used for comparison between the two groups; further pairwise comparison using LSD-t test; The count data was checked by χ2 test.
Results:Compared with the negative control group, the siRNA α1-AT group was transfected for 24 hours, the α1-AT mRNA level was significantly inhibited (P<0.05), and the proliferation rate of RA-SFs was not affected (P>0.05), and the invasion and migration ability of cells decreased significantly (P<0.05). The secretion of IL-17 and IL-1β was slightly decreased (P>0.05), while TNF-α and IL-1α were not affected (P>0.05). In addition, the expression of α1-AT downstream genes Matrix metallop roteinases (MMP)-2 169, Hu-Bax1, MMP-9, Hu Bax and Hu Bcl-xl were significantly decreased (P<0.05). Moreover, the signaling pathway inhibitors ERK inhibitor (PD98059) and NF-κB inhibitor (PDTC) had significant effects on α1-AT (P<0.05).
Conclusion:The α1-AT gene silencing has potential effect on the biological behavior of RA SFs. Further study of this mechanism is helpful to provide evidence for the treatment of RA.
