Abstract:

Objective To investigate the characteristics of cellular senescence， neoplastic stem cell and cell proliferation of polyploid giant cancer cells induced by Docetaxel （Doc） in non-small cell lung cancer， so as to analyze the potential role of polyploid giant cancer cells in neoplasm recurrence. Methods Non-small cell lung cancer NCI-H1299 cells were treated with dimethyl sulfoxide （DMSO） or 100 nmol/L Doc for 24 h， and then cultured for another 3 d after replacing with fresh complete medium. The cells were recorded as control group and Doc group， respectively. The cell growth was observed under a microscopy. The nuclear morphology was detected by immunofluorescence staining. The DNA content was detected by flow cytometry. The cell density was counted by trypan blue staining. A senescence-associated β-galactosidase staining kit was used to detect the β-galactosidase activity. The mRNA expression levels of the transcription factors Krüppel-like factor 4 （KLF4） and NANOG were detected by real-time quantitative polymerase chain reaction （PCR）. The cells in the Doc group were cultured for a long time to observe the growth of polyploid giant cancer cells. Results After Doc treatment， the cell and nuclear volume were increased in NCI-H1299 cells. The percentage of polyploidy in the Doc group was 45.80%±1.73%， which was significantly higher than that in the control group （4.83%±0.72%）， with a statistically significant difference （t=-29.01， P<0.001）. The cell density in the Doc group was （8.18±0.54）×10⁴/cm²， which was significantly higher than that in the control group [（0.75±0.08）×10⁴/cm²]， with a statistically significant difference （t=23.55， P<0.001）. The senescence-associated β-galactosidase activity was significantly increased and the percentage of positive cells （blue coloring） was 56.21%±3.38% in the Doc group， which was significantly higher than that in the control group （4.20%±0.79%）， with a statistically significant difference （t=-47.37， P<0.001）. Compared with the control group （1.00±0.16）， the mRNA expression level of KLF4 was significantly higher in the Doc group （2.47±0.18）， with a statistically significant difference （t=-10.74， P<0.001）. Compared with the control group （0.99±0.07）， the mRNA expression level of NANOG was significantly higher in the Doc group （5.58±0.26）， with a statistically significant difference （t=-30.09， P<0.001）. After being cultured for another 5 d， the cells in the Doc group maintained a large cellular state and there were less cells in the visual field. After being cultured for 7 d， a few small daughter cells appeared around the large cells and there were still few cells in the visual field. After being cultured for 13 d， the number of daughter cells was significantly increased. And the daughter cells finally occupied the entire field of view by 15 d. Conclusion Doc induces non-small cell lung cancer NCI-H1299 cells to generate polyploid giant cancer cells. These cells exhibit characteristics of the senescence， stemness and reproliferation， which may contribute to neoplasm recurrence.

Key words: Lung neoplasms, Polyploid, Cellular senescence, Neoplastic stem cell, Cell proliferation, Neoplasm recurrence