摘要
目的揭示SOX2蛋白及趋化相关基因CCR1、CCR2和MCP-1影响骨髓瘤细胞生物学特性的机制。方法本研究以骨髓瘤细胞株H929为研究对象,采用随机数表法将细胞分为空白对照组、阴性对照组和实验组。空白对照组为不进行任何处理的骨髓瘤细胞株H929,阴性对照组为转染慢病毒r LV-ZPP的稳转株H929-ZPP,实验组为转染慢病毒r LV-hsox2-Zs Green-Puro的稳转株H929-hsox2。采用Western blot检测各组细胞中SOX2蛋白表达,采用实时荧光定量PCR反应(RT-PCR)测定各组细胞中CCR1、CCR2和MCP-1基因表达,相应基因的相对表达水平用2-ΔΔCT代表。采用酶标仪测定细胞在450 nm处的吸光度值,计算细胞生存率。采用流式细胞仪计算各组细胞的凋亡率。结果实验组的SOX2蛋白表达为(1.38±0.07),高于空白对照组的(1.04±0.06)和阴性对照组的(0.72±0.07),差异均有统计学意义(P<0.05);实验组的CCR1、CCR2和MCP-1基因表达水平分别为(4.37±0.33)、(4.45±0.38)、(10.72±1.14),高于空白对照组的(2.14±0.15)、(2.21±0.17)和(5.17±0.35)和阴性对照组的(1.22±0.10)、(1.16±0.09)和(1.59±0.09),差异均有统计学意义(P<0.05);实验组的细胞生存率为(40.53±2.21)%,低于空白对照组的(86.85±5.17)%和阴性对照组的(95.07±7.08)%,差异均有统计学意义(P<0.05);实验组的细胞凋亡率为(58.56±3.25)%,高于空白对照组的(12.15±1.14)%和阴性对照组的(4.93±2.12)%,差异均有统计学意义(P<0.05)。结论 SOX2基因在细胞周期中特定时期的高表达会导致骨髓瘤细胞的凋亡。CCR1、CCR2和MCP-1基因参与了骨髓瘤细胞恶性增殖,其在细胞中的异常表达为骨髓瘤细胞提供合适的环境,导致疾病复发和抗凋亡的发生。
Objective To reveal the molecular mechanism of SOX2 protein and chemotaxis related genes CCR1, CCR2 and MCP-1 on the biological characteristics of myeloma cells. Methods In this study, myeloma cell line H929 was taken as the research objects. Using random number method, the cells were divided into blank control group（myeloma cell strain H929 without any treatment）, negative control group（stable strain H929-ZPP transfected with lentivirus r LV-ZPP） and experimental group（stable transgenic strain H929-hsox2 transfected with lentivirus r LV-hsox2-ZsGreen-Puro）. Western blot was used to detect the expression of SOX2 protein in each group. The expression of CCR1,CCR2 and MCP-1 genes in the cells of each group were detected by real-time quantitative PCR reaction（RT-PCR）. The relative expression level of the corresponding genes was represented by 2-ΔΔCT. The absorbance value of cells at 450 nm was measured by enzyme labelling apparatus, and cell survival rate was calculated. The apoptotic rate of each group was calculated by flow cytometry. Results The expression of SOX2 protein in the experimental group was（1.38 ± 0.07）,which was significantly higher than（1.04±0.06） in the blank control group and（0.72 ± 0.07） in the negative control group（P〈0.05）. The expression levels of CCR1, CCR2 and MCP-1 were（4.37±0.33）,（4.45±0.38）,（10.72±1.14） in the experimental group, significantly higher than（2.14±0.15）,（2.21±0.17）,（5.17±0.35） in the blank control group and（1.22±0.10）,（1.16 ± 0.09）,（1.59 ± 0.09） in the negative control group（P〈0.05）. The cell survival rate of the experimental group was（40.53±2.21）%, significantly lower than（86.85±5.17）% in the blank control group and（95.07±7.08）% in the negative control group（P〈0.05）. The apoptosis rate of the experimental group was（58.56±3.25）%, significantly higher than（12.15±1.14）% in the blank control group and（4.93±2.12%） in the negative control group（P〈0.05）. Conclusion High expression of SOX2 gene in the cell cycle will lead to the apoptosis of myeloma cells. CCR1, CCR2 and MCP-1 genes are involved in the malignant proliferation of myeloma cells. The abnormal expression in the cells provides a suitable environment for myeloma cells, which leads to the recurrence of disease and the occurrence of anti-apoptosis.
作者
柯金勇
汪玉芳
陆亚岚
张欣
柯善栋
KE Jin-yong;WANG Yu-fang;LU Ya-lan;ZHANG Xin;KE Shan-dong(Department of Hematology,the Central Hospital of Huangshi City,Huangshi 435000,Hubei,CHINa)
出处
《海南医学》
CAS
2018年第17期2377-2380,共4页
Hainan Medical Journal
