[Inducement of tumor cell autophagy and cell cycle analysis].
OBJECTIVE
Autophagy is the main phenomenon of type II programmed cell death which is also named as autophagic cell death, and autophagy has a close relationship with autophagic cell death. The relationship of apoptosis and cell cycle has been explored deeply, but little is known about the relationship of autophagic cell death and cell cycle. This study was to observe the correlation of autophagy induced by different methods to cell cycle.


METHODS
Exponentially growing HeLa and SW480 cells, and peripheral blood lymphocytes (PBLs) from healthy donors, with or without 48 h stimulation of phytohemagglutinin (PHA), were treated with Hanks' solution (to produce starvation) or vincristine. Confocal laser microscope and transmission electron microscope (TEM) were used to detect autophagy; flow cytometry (FCM) was innovatively used to detect the cell cycle of autophagic cells with dipl-parameters of microtubule-associated protein 1 light chain 3 (MAP1-LC3-II)/PI.


RESULTS
Autophagy of HeLa and SW480 cells induced by starvation or vincristine was observed in G1, S, and G2/M phases and increased along with the inducement time; no autophagy was observed in unstimulated PBLs. The positive rate of LC3-II, indicating the occurrence of autophagy, was lower than 2.62% when induced by starvation in Hanks' solution for 48 h, or 6.16% when induced by vincristine for 48 h. After PBLs were stimulated into cell cycle by PHA, autophagy was markedly detected 2 h after the indicated inducements.


CONCLUSIONS
MAP1-LC3-II/DNA dipl-parameter analysis by FCM is a convenient and reliable method for simultaneously analyzing autophagy and cell cycle. Autophagy could be induced when cells are in cell cycle, while the cells in G0 phase are insensitive to the inducers.
Han
Zhong-Bo
ZB
Cancer Research Institute/Department of General Surgery, Tongji Hospital, Huazhong University of Science and Technology, Wuhan, Hubei, 430030, P. R. China.
Zhang
Peng
P
Fu
Qiang
Q
Li
Xiao-Lan
XL
Ge
Jun-Na
JN
Tao
De-Ding
DD
Hu
Jun-Bo
JB
Gong
Jian-Ping
JP
D001343:Autophagy / Q000187:drug effects*; D002453:Cell Cycle / Q000187:drug effects*; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D003110:Colonic Neoplasms / Q000378:metabolism / Q000473:pathology*; D005434:Flow Cytometry; D006367:HeLa Cells; D006801:Humans; D008214:Lymphocytes / Q000166:cytology / Q000378:metabolism; D018613:Microscopy, Confocal; D046529:Microscopy, Electron, Transmission; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D014750:Vincristine / Q000494:pharmacology*
C502070:MAP1LC3B protein, human; D008869:Microtubule-Associated Proteins; D014750:Vincristine