CAJ | 학술논문

目的在果蝇S2细胞中表达人趋化素样因子1(chemokine-like factor 1,CKLF1),并对其分泌形式进行功能研究.方法构建pMT/V5-His-CKLF1表达质粒,转染S2细胞,筛选并鉴定阳性细胞克隆,用兔抗人CKLF1多肽抗体对其培养上清进行Western blot检测,并分析其趋化活性和促C2C12细胞增殖活性.结果 RT-PCR证明CKLF1在S2细胞中高效转录;通过Western blot在细胞培养上清中可检测到表达的重组CKLF1蛋白;其对人外周血中性粒细胞和淋巴细胞有较弱的趋化活性,对C2C12细胞具有增殖促进作用.结论在果蝇S2细胞中成功表达了人重组CKLF1,并证实其存在分泌形式且具有趋化活性和促C2C12细胞增殖活性.
목적 재과승S2세포중표체인추화소양인자1(chemokine-like factor 1,CKLF1),병대기분비형식진행공능연구.방법 구건pMT/V5-His-CKLF1표체질립,전염S2세포,사선병감정양성세포극륭,용토항인CKLF1다태항체대기배양상청진행Western blot검측,병분석기추화활성화촉C2C12세포증식활성.결과 RT-PCR증명CKLF1재S2세포중고효전록;통과Western blot재세포배양상청중가검측도표체적중조CKLF1단백;기대인외주혈중성립세포화림파세포유교약적추화활성,대C2C12세포구유증식촉진작용.결론 재과승S2세포중성공표체료인중조CKLF1,병증실기존재분비형식차구유추화활성화촉C2C12세포증식활성.
Objective To express human chemokine-like factor 1 (CKLF1) in Drosophila S2 cells and study its function. Methods The pMT/V5-His-CKLF1 expression plasmid was constructed and transfected into Drosophila S2 cells. The positive clones were selected through PCR and RT-PCR. The culture medium was analyzed by Western blot with anti-CKLF1 polyclonal antibody. Chemotaxis and MTT assays on human peripheral blood and C2C12 cells, respectively, were then carried out with the medium. Results CKLF1 was transcribed efficiently in S2 cells. The expressed CKLF1 protein could be detected in the culture supernatant by Western blot, which showed weak chemotactic activity on both human peripheral blood neutrophils and lymphocytes as well as enhancing effect on the proliferation of C2C12 cells. Conclusion CKLF1 was expressed successfully in Drosophila S2 cells and secreted into the culture medium. The recombinant CKLF1 expressed in Drosophila cells can chemoattract leucocytes and promote the proliferation of C2C12 cells.