[EGR-1 mRNA expression during 12-0-tetradecanoylphorbol-13-acetate-induced K562 cell differentiation].
OBJECTIVE
12-0-tetradecanoylphorbol-13 acetate (TPA) plays an important role in precipitating cell differentiation for various tumor cells, especially leukemic cells. Changes of many genes may be involved in this process. The purpose of this study was to observe the relationship between the EGR1mRNA expression and cell differentiation during TPA-induced K562 cell differentiation.


METHODS
Incubation of human K562 cells in vitro was applied to cultivate K562 cells. The cells were treated in two different ways. K562 cells of experiment group were treated with TPA and those of control group were treated without TPA. Using morphology (Wright's staining and NSE staining) and flow cytometry (FCM), the investigators observed the differentiation characteristics of K562 cells, cell-cycle and the differentiation antigen expressions of CD33 and CD14 on cell membranes. RT-PCR was carried out to assay EGR1 mRNA expression.


RESULTS
After treated with TPA for 7 d, the morphology of K562 cells obviously tended to mature differentiation, like monocytes. The differentiation rate of induced K562 cells was up to 95% in experiment group and 4.5% in control group, respectively. Using SPSS software, the above result showed statistical significance (P < 0.01). Using NSE staining, K562 cells showed positive reaction. Some of them were densely stained. The positive rate was up to 86%. More than half of the positive cells could be inhibited by NaF. The inhibiting rate of NaF was up to 58.72%, showing statistical difference when compared with that of control group. FCM analysis showed that most of K562 cells stimulated by TPA underwent G1/S phase cell-cycle arrest. The composing rate of cell-cycle in TPA-treated group showed that (53.7 +/- 1.25)% of cells were at G0 + G1 phase and (44.3 +/- 1.32)% were at S phase (P < 0.05). The level of CD33 expression on cell membranes was mildly decreased from 0.997% to 0.893% (P > 0.05). However, the level of CD14 expression was significantly increased from 0.049% to 0.387% (P < 0.05).


CONCLUSIONS
K562 cells could express EGR1mRNA during TPA-induced differentiation, which suggested that EGR1mRNA might participate in the process of K562 cells differentiating into monocyte/macrophages, and might play an important role in precipitating and maintaining cell differentiation for leukemic cells.
Fang
Ding-zhu
DZ
Department of Pediatrics, The West China Second Hospital, Sichuan University, Chengdu 610041, China.
Liao
Qing-kui
QK
Gao
Jiu
J
Yang
Xian-jun
XJ
Yuan
Li-xing
LX
Jia
Guo-cun
GC
D015703:Antigens, CD / Q000378:metabolism; D015214:Antigens, Differentiation, Myelomonocytic / Q000378:metabolism; D002273:Carcinogens / Q000494:pharmacology; D002453:Cell Cycle / Q000187:drug effects / Q000235:genetics; D002454:Cell Differentiation / Q000187:drug effects* / Q000235:genetics; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002462:Cell Membrane / Q000737:chemistry / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000235:genetics*; D051766:Early Growth Response Protein 1; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics*; D020014:K562 Cells; D018950:Lipopolysaccharide Receptors / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D063268:Sialic Acid Binding Ig-like Lectin 3; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014157:Transcription Factors / Q000235:genetics*
D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; C571163:CD33 protein, human; D002273:Carcinogens; D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D017874:Immediate-Early Proteins; D018950:Lipopolysaccharide Receptors; D012333:RNA, Messenger; D063268:Sialic Acid Binding Ig-like Lectin 3; D014157:Transcription Factors; D013755:Tetradecanoylphorbol Acetate