目的:家族性地中海热(FMF)是一种遗传性自身炎症性疾病,由地中海热(MEFV)基因突变引起,并与促炎细胞因子的增加有关。如白细胞介素-1β(IL-1β)和白细胞介素-18(IL-18),导致过量的炎症。秋水仙碱是广泛用于治疗FMF发作的常见药物,但约5-15%的患者对常规秋水仙碱治疗表现出抗性。在这项研究中,我们使用了二甲基氨基小白菊内酯(DMAPT),作为核因子-κB(NF-κB)的小分子抑制剂,含NLR家族Pyrin结构域3(NLRP3),和FMF来源的外周血单核细胞(PBMC)上的半胱氨酸-天冬氨酸蛋白酶1(Caspase-1)。
方法:通过MTT和AnnexinV/PI测定评估DMAPT和秋水仙碱对FMF来源的PBMC代谢活性和凋亡的影响,分别。此外,NF-κB的表达水平,NLRP3,MEFV,使用TaqMan实时PCR研究了CASP1和IL-1βmRNA,和IL-1β的蛋白质水平,通过LPS/ATP刺激的PBMC中的酶联免疫吸附测定(ELISA)评估IL-18和IL-37。
结果:DMAPT降低NFκB的表达水平(0.38±0.096,P<0.0001),NLRP3(0.39±0.12,P<0.001),MEFV(0.384±0.145,P<0.001),CASP1(0.48±0.13,P=0.0023),和IL-1β(0.09±0.09,P<0.0001),并降低IL-1β的分泌水平(8.92±5.3vs.149.85±20.92,P<0.0001),IL-18(135±32.1vs.192±22.18,P=0.01),和IL-37(27.5±6.3vs.与未处理的细胞相比,78.19±14.3,P<0.0001)。
结论:鉴于获得的结果与以前的研究相比,DMAPT的未来临床发展可能导致FMF疾病的新抗炎治疗方法的扩展。
OBJECTIVE: Familial Mediterranean Fever (FMF) is a hereditary auto-inflammatory disorder that is caused by mutations in the Mediterranean fever (
MEFV) gene and is associated with an increase in pro-inflammatory cytokines, such as interleukin-1β (IL-1β) and interleukin-18 (IL-18), leading to excess inflammation. Colchicine is a common drug widely used for treatment of FMF attacks, but about 5-15% of the patients show resistance to the regular colchicine treatment. In this study, we used dimethylamino-parthenolide (DMAPT), as a small molecule inhibitor of Nuclear factor-κB (NF-κB), NLR family Pyrin domain containing 3 (NLRP3), and cysteine-aspartic acid protease 1(Caspase-1) on FMF-derived peripheral blood mononuclear cells (PBMCs).
METHODS: The effects of DMAPT and colchicine on metabolic activity and apoptosis of FMF-derived PBMCs were evaluated by MTT and Annexin V/PI assays, respectively. Also, the expression levels of NF-κB, NLRP3,
MEFV, CASP1, and IL-1β mRNA were investigated using a TaqMan real-time PCR, and the protein levels of IL-1β, IL-18, and IL-37 were assessed via an enzyme-linked immunosorbent assay (ELISA) in LPS/ ATP-stimulated PBMCs.
RESULTS: DMAPT decreased the expression levels of NFκB (0.38±0.096, P<0.0001), NLRP3 (0.39±0.12, P<0.001),
MEFV (0.384±0.145, P<0.001), CASP1 (0.48±0.13, P=0.0023), and IL-1β (0.09±0.09, P<0.0001) and reduced the secretion levels of IL-1β (8.92±5.3 vs. 149.85±20.92, P<0.0001), IL-18 (135±32.1 vs. 192±22.18, P=0.01), and IL-37 (27.5±6.3 vs. 78.19±14.3, P<0.0001) as compared to untreated cells.
CONCLUSIONS: Given the obtained results in comparison with previous research, the future clinical development of DMAPT could result in the expansion of new anti-inflammatory therapeutics for FMF disorder.