The effect of tymosin β4 and acsdkp on the growth of HL-60 cells and their mechanism

W. Q. Huang*, Q. R. Wang

*Corresponding author for this work

Research output: Contribution to journalArticle

Abstract

Thymosin β4 (Tβ4) consists of 43 peptides with a molecular weight of 4963. It contains at its N-terminal the complete sequence of the acetylated tetrapeptide AcSDKP. Both of them could inhibit the growth of normal hematopoietic cells. The present study was conducted to investigate their effect on the proliferation, differentiation and apoptosis of HL-60 cells. The results showed that Tβ4 (10-"~10-7mol/L) and AcSDKP (10-"-10-5mol/ L) had the dose-dependent inhibitory effect on the growth of HL-60 cells. The rate of 3HTdR incorporation and the number of HL-60 cells was diminished in 10"~10~7mol/L Tβ4 and 10-'~10Jmol/L AcSDKP culture system. Cell morphological examination and NET reduction test demonstrated that Tβ4 and AcSDKP could induce HL-60 cells to differentiate into mature granulocytes. Moreover, the cell morphological observation discovered that Tβ4 and AcSDKP induced apoptosis of HL-60 cells and increased the percentage of apoptotic cells stained by Hoechest 33258 fluorescence stain. The typical ladders of DNA fragments in gel electrophoresis could be seen in each experimental group of HL-60 cells, while the control showed no ladder. In order to investigate the mechanism of the effects of Tβ4 and AcSDKP, the fluorescent intracellular indicator, fura-2AM, was used to determine the effects of Tβ4 and AcSDKP on the intracellular free Ca2 concentration ([Ca2 ]i) of HL-60 leukemic cells. It was shown that Tβ4 and AcSDKP increased [Ca2 ]i of HL-60 cells in calcium-free medium. Moreover, AcSDKP increased [Ca2"]i of HL-60 cells in calcium-contained medium further. But it had no distinct difference for Tβ4 in calcium-free medium or calcium-contained medium. So it implied that Tβ4 and AcSDKP could increase [Ca2 ]i by stimulating the release of Ca2 from intracellular Ca2 pool. Moreover, AcSDKP could also elicit a potent extracellular calcium influx in HL-60 cells. It was shown by the Atlas CdnA expression array that when HL-60 cells were cultured with Tβ4 for 3 days, the expression level of myc proto-oncogene protein, fos-related antigen 1, Apopain, DNA -binding protein inhibitor ID-2, transcriptional represser protein YY1, ICE-LAP6 and MCH4 was increased significantly as compared with the control. These results strongly suggested that Tβ4 could change apoptotic-related gene expression in leukemic cells, and result in the inhibition of proliferation and induction of differentiation and apoptosis of leukemic cells.

Original languageEnglish (US)
JournalBlood
Volume96
Issue number11 PART II
StatePublished - Dec 1 2000

ASJC Scopus subject areas

  • Hematology

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