论文部分内容阅读
目的探讨体外沉默白血病细胞核干细胞因子(NS)基因转染后细胞形态学和细胞化学特征改变与细胞生物学行为的关系。方法体外合成NS特异性短发夹状RNA(shRNA)转染HL-60白血病细胞,逆转录-聚合酶链反应(RT-PCR)和免疫印迹法评价NS基因沉默和NS蛋白合成抑制效果,倒置显微镜观察活体细胞形态,Wright-Giemsa染色观察细胞形态学改变,血液分析仪测定细胞大小、粒度和均一性变化,髓过氧化物酶(MPO)、α-乙酸萘酯酶(α-NAE)和过碘酸-雪夫反应(PAS)测定反映细胞化学特征变化。结果沉默NS基因导致HL-60白血病细胞发生了形态学变化,细胞核和细胞质出现了进一步分化和成熟特征,Wright-Giemsa染色观察到细胞核碎裂现象,血液分析仪发现细胞变得大小不均并有较多核碎裂和无核的细胞碎片存在,细胞内定MPO、α-NAE酶活性和PAS反应增强。结论沉默NS基因表达能促使HL-60细胞进一步分化、成熟,细胞形态学和细胞化学特征发生相应改变,这些形态学变化可以作为研究细胞生物学行为改变的依据之一。
Objective To investigate the relationship between cell morphological and cytochemical characteristics and cell biological behavior after transfection of nucleus stem cell factor (NS) gene in leukemic cells in vitro. Methods NS-specific short hairpin RNAs (shRNAs) were transfected into HL-60 leukemia cells in vitro. The inhibitory effect of NS gene silencing and NS protein synthesis was evaluated by reverse transcription-polymerase chain reaction (RT-PCR) and immunoblotting. The morphological changes of cells were observed by Wright-Giemsa staining. The changes of cell size, granularity and homogeneity were determined by hematology analyzer. Myeloperoxidase (MPO), α-NAE and α- Periodic acid-Schiff reaction (PAS) assay reflects changes in cytochemical characteristics. Results Silencing the NS gene resulted in the morphological changes of HL-60 leukemia cells, further differentiation and maturation of nuclei and cytoplasm, Wright-Giemsa staining of nuclear fragmentation, hematology analyzer found that cells became uneven and had More nuclear fragmentation and non-nuclear cell debris exist, intracellular MPO, α-NAE activity and PAS response increased. Conclusion Silencing the expression of NS gene can promote the further differentiation and maturation of HL-60 cells. The morphological and cytochemical characteristics of HL-60 cells change accordingly. These morphological changes can be used as a basis for studying the changes of cell biology behavior.