Inhibitory Effect of Tetramerized Single-Chain Variable Fragment of Anti-Cyclic Citrullinated Peptide Antibodies on the Proliferation, Activation, and Secretion of Cytokines of Fibroblast-Like Synoviocytes in Rheumatoid Arthritis In Vitro Co-Culture Syste
Inhibitory Effect of Tetramerized Single-Chain Variable Fragment of Anti-Cyclic Citrullinated Peptide Antibodies on the Proliferation, Activation, and Secretion of Cytokines of Fibroblast-Like Synoviocytes in Rheumatoid Arthritis In Vitro Co-Culture Syste
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抗环瓜氨酸肽抗体四聚化单链可变片段对类风湿性关节炎成纤维样滑膜细胞增殖、活化和细胞因子分泌的抑制作用体外共培养体系
DOI：
10.1007/s10753-020-01292-z
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发表时间：
2020-08
期刊：
影响因子：
5.1
通讯作者：
Kang Xixiong
中科院分区：
文献类型：
--
作者：
Wang Jing;Tie Ning;Li Hongbin;Kang Xixiong
抗环瓜氨酸多肽四聚化单链可变区(Teab-CCP)是由抗环瓜氨酸肽(CCP)抗体四聚化而成，具有P53四聚体结构域，旨在研究其对外周单个核细胞(PBMC)和滑膜细胞体外培养体系中成纤维细胞样滑膜细胞(Fls)增殖、迁移、侵袭和炎性介质产生的影响。用P53四聚体结构域修饰抗CCP单价单链抗体，以提高其亲和力，构建Teab-CCP。从类风湿性关节炎(RA)患者和对照组分离培养Fls。采用外周血单个核细胞(PBMC)与外周血单核细胞(FLSS)共培养体系。用四甲基偶氮唑盐比色法、划痕试验和Transwell小室检测细胞的增殖、迁移和侵袭能力。用Luminex液相蛋白芯片和酶联免疫吸附试验检测培养上清液中细胞因子、趋化因子、金属蛋白酶和抗CCP抗体。TEAB-CCP以剂量依赖的方式抑制FLSS的增殖，在与PBMC和FLSS共同培养的第7天，浓度为100μg/ml时作用最强，但不能单独作用于FLSS。与对照组相比，Teab-CCP显著抑制Flss的迁移和侵袭能力。联合培养上清液中IL-6、IL-8、RANKL、精氨酸脱亚氨酶(PAD)-2、PAD4、基质金属蛋白酶(MMP1)、MMP3及抗CCP抗体水平显著降低。而转化生长因子-β(转化生长因子-β)和金属蛋白酶组织抑制因子-2(TIMP-2)在TEA-CCP组显著升高。IL-1a、IL-10、IL-17、肿瘤坏死因子α、血管内皮生长因子、成纤维细胞生长因子在两组间差异无统计学意义。作为一种封闭抗体，Teab-CCP能显著抑制RA患者PBMC产生促炎介质，进而抑制FL的增殖、激活、迁移和侵袭。因此，瓜氨酸修饰的自身表位可能成为类风湿关节炎治疗的新靶点。
Tetramerized single-chain variable fragment (ScFv) of anti-cyclic citrullinated peptide (TeAb-CCP) is a constructed tetramerized ScFv of anti-cyclic citrullinated peptide (CCP) antibodies with p53 tetrameric domain, aim to investigate its effect on fibroblast-like synoviocytes (FLSs) proliferation, migration, invasion, and production of inflammatory mediators in thein vitroco-culture system of peripheral mononuclear cells (PBMCs) and FLSs. TeAb-CCP was constructed by modifying a monovalent ScFv antibody to CCP with p53 tetrameric domain to improve its affinity. FLSs were isolated and cultured from rheumatoid arthritis (RA) patients and control subjects. A co-culture system of peripheral mononuclear cells (PBMCs) and FLSs was used. FLSs proliferation, migration, and invasion were measured by MTT, scratch test, and Transwell chamber. Supernatants were measured for cytokines, chemokines, metalloproteinases, and anti-CCP antibodies by Luminex liquid phase protein chip and ELISA. TeAb-CCP significantly inhibited FLSs proliferation in a dose-dependent mode, with maximal action at concentration of 100 μg/ml on the 7th day in the co-culture system with PBMCs and FLSs, but not the same with only FLSs. TeAb-CCP significantly suppressed FLSs migration and invasive ability compared with the controls. Significantly lower levels of interleukin (IL)-6, IL-8, RANKL, protein arginine deiminase (PAD)-2, PAD4, metalloproteinase (MMP)-1 and MMP-3 and anti-CCP antibodies were found in co-culture supernatant of TeAb-CCP group. In contrast, transforming growth factor-β (TGF-β) and tissue inhibitor of metalloproteinases-2 (TIMP-2) was significantly increased in the TeAb-CCP group. No significant difference of IL-1a, IL-10, IL-17, TNFα, VEGF, and FGF was found between two groups. As a blocking antibody, TeAb-CCP can significantly inhibit PBMCs of RA to produce pro-inflammatory mediators, and furthermore, inhibit the proliferation, activation, migration, and invasion of FLSsin vitro. In turn, it is suggested that citrullinated modified self-epitopes may be a new target for RA therapy.
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DOI：
10.2147/dddt.s41645
发表时间：
2013-12-27
期刊：
Drug design, development and therapy
影响因子：
--
作者：
Mok CC
通讯作者：
Mok CC
影响因子：
4.9
作者：
Bustamante MF;Garcia-Carbonell R;Whisenant KD;Guma M
通讯作者：
Guma M
影响因子：
--
作者：
Ahmad ZA;Yeap SK;Ali AM;Ho WY;Alitheen NB;Hamid M
通讯作者：
Hamid M
影响因子：
--
作者：
Hong-bin Li;Ning Tie;Yong-feng Jia;Lin Shi;Yan Su;Guizhi Zhang;Yong Wang;L. Bai;J. Zhao;Jing Wang;Zhen Xiao
通讯作者：
Hong-bin Li;Ning Tie;Yong-feng Jia;Lin Shi;Yan Su;Guizhi Zhang;Yong Wang;L. Bai;J. Zhao;Jing Wang;Zhen Xiao
影响因子：
27.4
作者：
Sun, Meng;Rethi, Bence;Catrina, Anca Irinel
通讯作者：
Catrina, Anca Irinel