College of Biological and Food Engineering, Huaihua University, Huaihua, Hunan 418000, China.
Center for Reproduction and Health Development, Shenzhen Institutes of Advanced Technology, Chinese Academy of Science, Shenzhen, Guangdong 518055, China.
Int J Med Sci. 2021 Jan 1;18(1):207-215. doi: 10.7150/ijms.51900. eCollection 2021.
Placental-like chondroitin sulfate A (pl-CSA) is exclusively expressed in cancerous and placental tissues and is highly correlated with the degree of malignancy. However, the mechanism through which pl-CSA regulates tumorigenesis and metastasis in choriocarcinoma remains unclear. Stable transfectants of the JEG3 choriocarcinoma cell line, including a negative control (NC) line and a cell line with knockout of the biosynthetic enzyme CS synthase-2 (ChSy-2) (ChSy-2), were obtained using CRISPR/Cas9 systems and identified by immunofluorescence, flow cytometry, western blots and enzyme-linked immunosorbent assays (ELISAs). The proliferation, migration, invasion and colony formation of the cells were determined by a cell counting kit, scratch-wound assays, transwell assays and soft agar colony formation assays , respectively. The tumorigenesis and metastasis of choriocarcinoma were also investigated through two xenograft models . The ChSy-2 protein in the ChSy-2group was below the detection threshold, which was accompanied a significant reduction in the pl-CSA level. Reducing pl-CSA through ChSy-2 knockout significantly inhibited cell proliferation, migration, invasion and colony formation and tumorigenesis and metastasis of choriocarcinoma, with deceases in tumor volume and metastatic foci and a high percent survival compared to the NC . pl-CSA, as a necessary component of JEG-3 cells, was efficiently reduced through ChSy-2 knockout, which significantly inhibited the tumorigenesis and metastasis of choriocarcinoma. ChSy-2/pl-CSA could be alternative targets for tumor therapy.
胎盘样硫酸软骨素 A（pl-CSA）仅在癌组织和胎盘组织中表达，与恶性程度高度相关。然而，pl-CSA 调节绒癌发生和转移的机制尚不清楚。
利用 CRISPR/Cas9 系统获得了 JEG3 绒癌细胞系的稳定转染子，包括阴性对照（NC）系和生物合成酶 CS 合酶-2（ChSy-2）敲除的细胞系（ChSy-2），并通过免疫荧光、流式细胞术、western blot 和酶联免疫吸附测定（ELISA）进行了鉴定。通过细胞计数试剂盒、划痕试验、Transwell 试验和软琼脂集落形成试验分别测定了细胞的增殖、迁移、侵袭和集落形成能力。还通过两种异种移植模型研究了绒癌的致瘤性和转移。ChSy-2 组中的 ChSy-2 蛋白低于检测阈值，同时 pl-CSA 水平显著降低。通过 ChSy-2 敲除降低 pl-CSA 显著抑制绒癌细胞的增殖、迁移、侵袭和集落形成以及绒癌的致瘤性和转移，与 NC 相比，肿瘤体积和转移灶减少，存活率高。
pl-CSA 作为 JEG-3 细胞的必需成分，通过 ChSy-2 敲除得到有效降低，显著抑制绒癌的发生和转移。ChSy-2/pl-CSA 可能是肿瘤治疗的替代靶点。
