Therapeutic effect of daphnetin on the autoimmune arthritis through demethylation of proapoptotic genes in synovial cells.
Therapeutic effect of daphnetin on the autoimmune arthritis through demethylation of proapoptotic genes in synovial cells.
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瑞香素通过滑膜细胞促凋亡基因去甲基化对自身免疫性关节炎的治疗作用
DOI：
10.1186/s12967-014-0287-x
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发表时间：
2014-10-14
影响因子：
7.4
通讯作者：
Min W
中科院分区：
文献类型：
--
作者：
Shu K;Kuang N;Zhang Z;Hu Z;Zhang Y;Fu Y;Min W
背景我们以前曾报道过，在胶原诱导的关节炎（CIA）大鼠模型中，去啡肽对类风湿性关节炎（RA）有治疗作用。然而，瑞香素对滑膜细胞中促凋亡基因去甲基化的分子机制和作用仍有待进一步阐明。方法（1）采用MTT法（3-（4，5）-dimethylthiahiazo（-z-y1）-3，5-di-phenylterazoliumromide）检测瑞香素对CIA大鼠滑膜细胞增殖的抑制作用;（2）甲基化特异性PCR（MSP）检测促凋亡基因DR 3的甲基化（死亡受体3），PDCD 5（3）采用真实的时间PCR检测DR 3、PDCD 5、FasL、p53和DNA甲基转移酶（DNMT）DNMT 1、DNMT 3a和DNMT 3b的mRNA表达;（4）流式细胞术检测DR 3、PDCD 5、FasL和p53蛋白表达：（5）Annexin V和PI法检测滑膜细胞凋亡率;（6）采用扫描电镜（SEM）和透射电镜（TEM）观察CIA大鼠滑膜细胞结构的变化结果（1）在1.25 μg/mL ~ 40 μg/mL范围内，瑞香素和5-aza-dc对CIA大鼠滑膜细胞的抑制作用呈剂量依赖性和时间依赖性。(2)瑞香素和5-aza-dc对CIA大鼠滑膜细胞的促凋亡基因DR 3、PDCD 5、FasL和p53具有去甲基化作用。(3)瑞香素和5-aza-dc降低甲基转移酶DNMT 1、DNMT 3a和DNMT 3b的基因表达，增加促凋亡基因DR 3、PDCD 5、FasL和p53的表达，这转化为DR 3、PDCD 5、FasL和p53的蛋白表达增加。(4)结论瑞香素可降低DNMT 1、DNMT 3a和DNMT 3b的表达，并可能导致促凋亡基因DR 3、PDCD 5、FasL和p53的去甲基化。因此，瑞香素可以增加促凋亡基因和蛋白的表达，导致结构凋亡的变化和增加早期和晚期CIA大鼠滑膜细胞凋亡。
BackgroundWe have previously reported that dephnetin is therapeutically effective in the treatment of rheumatoid arthritis (RA) in collagen-induced arthritis (CIA) rat model. However, the molecular mechanism and the effect of daphnetin on demethylating proapoptotic genes in the synovial cells remains further clarified. This study may provide a deeper insight into the medicinal application of daphnetin as a treatment for RA.Methods(1) The proliferation inhibition of CIA rat synovial cells was determined by an MTT (3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenyterazoliumromide) assay; (2) Methylation specific PCR (MSP) was used to measure the methylation of the proapoptotic genes DR3 (death receptor 3), PDCD5 (programmed cell death 5), FasL and p53; (3) Real time-PCR was performed to determine the mRNA expression of DR3, PDCD5, FasL, p53 and DNA methyltransferases (DNMTs) DNMT1, DNMT3a and DNMT3b; (4) Flow cytometry was applied to detect the protein expression of the DR3, PDCD5, FasL and p53; (5) The apoptotic rate of synovial cells was assessed by flow cytometry with Annexin V and propidium iodide (PI); (6) Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were used to observe the changes of CIA rat synovial cell structure.Results(1) In the range of 1.25 μg/mL to 40 μg/mL, daphnetin and 5-aza-dc had a dose-dependent and time-dependent degree of inhibition to the CIA rat synovial cells. (2) Daphnetin and 5-aza-dc had a demethylating role on the proapoptotic genes DR3, PDCD5, FasL and p53 of CIA rat synovial cells. (3) Daphnetin and 5-aza-dc decreased the gene expression of methyltransferases DNMT1, DNMT3a and DNMT3b, and increased expression of proapoptotic genes DR3, PDCD5, FasL and p53, which translated into an increased protein expression of DR3, PDCD5, FasL and p53. (4) Daphnetin and 5-aza-dc changed the structure of CIA rat synovial cells to show apoptotic changes and increased the rate of apoptosis.ConclusionsDaphnetin can reduce the expression of DNMT1, DNMT3a and DNMT3b, which could result in the proapoptotic genes DR3, PDCD5, FasL and p53 being demethylated. Therefore, daphnetin can increase proapoptotic gene and protein expression resulting in structural apoptotic changes and an increase in early and late CIA rat synovial cell apoptosis.
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影响因子：
8
作者：
Kim, J. K.;Samaranayake, M.;Pradhan, S.
通讯作者：
Pradhan, S.
影响因子：
30.8
作者：
通讯作者：
--
影响因子：
3.2
作者：
Xiaofang L;Kun T;Shaoping Y;Zaiqiu W;Hailong S
通讯作者：
Hailong S
影响因子：
2.7
作者：
Buchi, Francesca;Spinelli, Elena;Santini, Valeria
通讯作者：
Santini, Valeria
成纤维细胞样滑膜细胞中的 p53 可以调节活动性类风湿关节炎患者的 T 辅助细胞功能。
影响因子：
6.1
作者：
Tang Bi-xia;You Xin;He Wei
通讯作者：
He Wei