核仁素通过调控胸苷激酶1影响淋巴瘤增殖的研究  

Effect of Nucleolin on Lymphoma Proliferation by Regulating Thymidine Kinase 1

作　　者：梅序桥 胡建达 杨婷 吴阿阳 许煜煌 林子杭 林聪猛 MEI Xu-Qiao;HU Jian-Da;YANG Ting;WUA-Yang;XU Yu-Huang;LIN Zi-Hang;LIN Cong-Meng(Department of Clinical Laboratory,Zhangzhou Afiliated Hospital of Fujian Medical University,Zhangzhou 363000,Fujian Province China;Fujian Provincial Key Laboratory of Hematology,Fujian Institute of Hematology,Fujian Medical University Union Hospital,Fuzhou 350001,Fujian Province,China;Department of Hematology,Zhangzhou Affiliated Hospital of Fujian Medical University,Zhangzhou 363000,Fujian Province,China)

机构地区：[1]福建医科大学附属漳州市医院检验科,福建漳州363000 [2]福建医科大学附属协和医院福建省血液病研究所,福建福州350001 [3]福建医科大学附属漳州市医院血液科,福建漳州363000

出　　处：《中国实验血液学杂志》2023年第3期699-706,共8页Journal of Experimental Hematology

基　　金：福建省中医药科技项目(2017FJZYZY209);福建省自然科学基金项目(2020J011302,2020J011299);漳州市自然科学基金项目(ZZ2020J05)。

摘　　要：目的:探讨核仁素通过胸苷激酶1(TK1)参与淋巴瘤增殖调控的机制。方法:选取弥漫性大B细胞淋巴瘤患者23例,纳入初治组14例,复发/难治组9例,检测患者血清TK1以及外周血单个核细胞中C23蛋白。以Burkitt淋巴瘤细胞株CA46为研究对象,通过慢病毒介导的转染方法,获得RNAi核仁素基因表达下调的细胞模型CA46-NCL-KD及阴性对照组细胞模型CA46-NCL-KNC;采用细胞增殖法(MTS法)分别检测CA46-NCL-KD、CA46-NCL-KNC和CA46对阿霉素的增殖半数抑制率(IC_(50))。通过Q-PCR和Western blot分别检测CA46-NCL-KD、CA46-NCL-KNC细胞NCL mRNA和蛋白表达水平,用流式细胞术对CA46-NCL-KD、CA46-NCL-KNC和CA46细胞进行细胞周期检测,用酶免疫点印记化学发光法分析CA46-NCL-KD、CA46-NCL-KNC细胞TK1蛋白的表达情况。结果:初治组患者血清TK1水平为0.43(0.30-1.01)pmol/L,低于复发/难治组患者的10.56(2.19-14.99)pmol/L(P<0.01),且初治组患者外周血单个核细胞NCL蛋白相对表达量明显低于复发/难治组。CA46-NCL-KD对阿霉素的IC_(50)为(0.147±0.02)μg/ml,明显低于CA46-NCL-KNC的(0.301±0.04)μg/ml以及CA46的(0.338±0.05)μg/ml(P<0.05);与CA46-NCL-KNC相比,CA46-NCL-KD中NCL mRNA和蛋白水平表达下降,细胞周期中的G0/G1期比例升高,S期及G2/M期比例下降,TK1蛋白表达下降。结论:在弥漫大B细胞淋巴瘤细胞以及淋巴瘤细胞株CA46细胞中NCL表达升高,提示细胞对药物敏感度降低,NCL可能通过影响TK1表达参与淋巴瘤细胞增殖的调控,从而影响淋巴瘤细胞对药物的敏感性。Objective:To investigate the mechanism of nucleolin(NCL) involved in lymphoma proliferation by regulating thymidine kinase 1(TK1).Methods:Twenty-three patients with diffuse large B-cell lymphoma(DLBCL)were selected and divided into initial treatment group(14 cases) and relapsed/refractory group(9 cases).Serum TK1and C23 protein in peripheral blood mononuclear cells were detected.Cell models of CA46-NCL-KD(CA46-NCLknockdown) and CA46-NCL-KNC(CA46-NCL-knockdown negative control) were established by lentivirus vector mediated transfection in Burkitt lymphoma cell line CA46.The half maximal inhibitory concentration(IC_(50)) of CA46-NCL-KD,CA46-NCL-KNC,and CA46 to adriamycin were detected by cell proliferation assay(MTS).The expression of NCL mRNA and protein in CA46-NCL-KD and CA46-NCL-KNC cells were dectected by Q-PCR and Western blot,respectively.The cell cycle of CA46-NCL-KD,CA46-NCL-KNC,and CA46 cells were detected by flow cytometry.The expression of TK1 protein in CA46-NCL-KD and CA46-NCL-KNC cells was detected by an enhanced chemiluminescence(ECL) dot blot assay.Results:The level of serum TK1 in the initial treatment group was 0.43(0.30-1.01) pmol/L,which was lower than 10.56(2.19-14.99) pmol/L in the relapsed/refractory group(P<0.01),and the relative expression level of NCL protein in peripheral blood was also significantly lower.The IC_(50) of CA46-C23-KD cells to adriamycin was(0.147±0.02) μg/ml,which was significantly lower than(0.301±0.04)μg/ml of CA46-C23-KNC cells and(0.338±0.05) μg/ml of CA46 cells(P<0.05).Compared with CA46-NCLKNC cells,the expression of NCL mRNA and protein,TK1 protein decreased in CA46-NCL-KD cells,and the proportion of S phase and G_(2)/M phase also decreased,while G_(0)/G_1 phase increased in cell cycle.Conclusion:The increased expression of NCL in DLBCL and CA46 cells indicates low sensitivity to drug.NCL may participate in regulation of lymphoma proliferation by affecting TK1 expression,thereby affecting the drug sensitivity.

关 键 词：CA46细胞 淋巴瘤 核仁素 胸苷激酶1 细胞周期 

分 类 号：R733.1[医药卫生—肿瘤]