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Apoptosis of Leukemia Cells Induced by Valine-Deficient Medium

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Apoptosis of Leukemia Cells Induced by Valine-Deficient Medium Leukemia (1998) 12, 1651–1656 1998 Stockton Press All rights reserved 0887-6924/98 $12.00 http://www.stockton-press.co.uk/leu CORRESPONDENCE Apoptosis of leukemia cells induced by valine-deficient medium TO THE EDITOR pase-3 activity of L1210 cells treated with valine, isoleucine and asparagine is shown by curves a, b and c, respectively. l-Asparaginase from Escherichia coli, which catalyzes the From the results obtained above, it can be concluded that hydrolysis of l-asparagine to l-asparatic acid and ammonia, DNA fragmentation of L5178Y or L1210 cells is closely asso- has been used as an effective therapeutic agent for the treat- ciated with the enhancement of caspase-3 activity. In human ment of acute lymphoblastic leukemia and lymphosarcoma.1,2 leukemia cells (THP-1 and U937 cells), apoptosis was effec- However, only limited types of leukemia cells are sensitive to tively induced by valine-deficient medium in comparison with l-asparaginase treatment. Murine leukemia cells treated with other amino acid deficiency (isoleucine or asparagine). l-asparaginase undergo apoptosis,3 which is closely associa- Experiments are now in progress not only to clarify the ted with the cell cycle arrest in G1 phase.4 Apoptosis of leuke- mechanism of apoptosis induced by valine-deficient medium mia cells is also induced by lacking l-asparagine from the cul- but also to apply to the therapy of human leukemia using ture medium.4 Apoptosis is closely associated with the valine-degrading enzyme. activation of caspases, mitochondrial permeability transition, cell volume loss, chromatin condensation and nucleosomal DNA fragmentation.5–8 The present study deals with the frag- Acknowledgements mentation of chromosomal DNA and activation of caspase-3 (CPP32, Yama or apopain) in murine and human leukemia We thank Dr T Shirai (Juntendo University, Tokyo, Japan) and cells treated with amino acid-deficient medium. Dr J Takagi (Tokyo Institute of Technology, Tokyo, Japan) for First, we examined the fragmentation of chromosomal DNA providing murine leukemia cells (L1210) and human leukemia in murine leukemia cells (L5178Y and L1210) induced by cells (U937 and THP-1), respectively. treatment with one amino acid-deficient medium out of total 20 amino acids in the RPMI 1640 medium containing 10% K Ohtawa11Toin Human Science dialyzed fetal bovine serum, penicillin (50 U/ml) and strepto- T Ueno1 and Technology Center, mycin (50 ␮g/ml) for 24 h. Among them, valine- and iso- K Mitsui1,2 Department of Materials Science leucine-deficient media induced the fragmentation of chromo- Y Kodera1 and Technology, somal DNA within 24 h incubation. Then, DNA fragmentation M Hiroto1 Toin University of Yokohama, and caspase-3 activity of murine leukemia cells (L5178Y and A Matsushima1 Yokohama, Japan; and L1210) treated with one of the amino acid-deficient media (l- H Nishimura1,2 2CREST, Japan Science and valine, l-isoleucine and l-asparagine) was tested (Figure 1). Y Inada1 Technology Corporation (JST) The treatment of L5178Y or L1210 cells with valine-deficient Saitama, Japan medium for 8, 12, 16, 20 and 24 h causes the fragmentation of chromosomal DNA into the size equivalent to single and multiple nucleosomes (Figure 1a). A similar phenomenon is References also observed for L5178Y and L1210 cells treated with iso- leucine-deficient medium for 12, 16, 20 and 24 h incubation. 1 Kamisaki Y, Wada H, Yagura T, Nishimura H, Matsushima A, l DNA fragmentation was also observed in L5178Y cells treated Inada Y. Increased antitumor activity of Escherichia coli -asparag- inase by modification with monomethoxypolyethylene glycol. with asparagine-deficient medium for 20 and 24 h incubation Gann 1982; 73: 470–474. but was not observed in L1210 cells within 24 h incubation 2 Ertel IJ, Nesbit ME, Hammond D, Weiner J, Sather H. Effective (data not shown). dose of l-asparaginase for induction of remission in previously Figure 1b shows the caspase-3 activity of L5178Y and treated children with acute lymphocytic leukemia: a report from a L1210 cells treated with one amino acid-deficient medium children’s cancer study group. Cancer Res 1979; 39: 3893–3896. l (valine, isoleucine or asparagine) during the course of incu- 3 Story MD, Voehringer DW, Stephens LC, Meyn RE. -asparaginase kills lymphoma cells by apoptosis. Cancer Chemother Pharmacol bation (0–24 h). In the case of L5178Y cells, the caspase-3 1993; 32: 129–133. activity of valine-deficient medium increases with incubation 4 Ueno T, Ohtawa K, Mitsui K, Kodera Y, Hiroto M, Matsushima A, time and becomes maximal for 12 h followed by decreasing Inada Y, Nishimura H. Cell cycle arrest and apoptosis of leukemia activity (curve a). In the treatment with isoleucine-deficient cells induced by l-asparaginase. Leukemia 1997; 11: 1858–1861. medium, the activity appears at more than 12 h incubation 5 Arends MJ, Morris RG, Wyllie AH. Apoptosis: the role of the and increases linearly with time (curve b). In asparagine- endonuclease. Am J Pathol 1990; 136: 593–608. 6 Matthew GVH, Navdeep SC, Edward KW, Paul TS, Craig BT. Bcl- deficient medium, the caspase-3 activity was little observed xL regulates the membrane potential and volume homeostasis of (curve c). A similar result was obtained for L1210 cells as mitochondria. Cell 1997; 91: 627–637. shown by the lower column of Figure 1b in which the cas- 7 Sellins KS, Cohen JJ. Gene induction by ␥-irradiation leads to DNA fragmentation in lymphocytes. J Immunol 1987; 139: 3199–3206. 8 Enari M, Talanian RV, Wong WW, Nagata S. Sequential activation Correspondence: Y Inada, Toin Human Science and Technology of ICE-like and CPP32-like proteases during Fas-mediated Center, Department of Materials Science and Technology, Toin Uni- apoptosis. Nature 1996; 380: 723–726. versity of Yokohama, 1614 Kurogane-cho, Aoba-ku, Yokohama 225- 8502, Japan; Fax: 81 45 972 5972 Received 26 March 1998; accepted 10 June 1998 Correspondence 1652 Figure 1 DNA fragmentation and caspase-3 activity of L5178Y or L1210 cells induced by treatment with one amino acid-deficient medium (valine, isoleucine or asparagine). (a) DNA fragmentation of L5178Y cells or L1210 cells treated with valine- or isoleucine-deficient medium for 0, 4, 8, 12, 16, 20 and 24 h. (b) Caspase-3 activity of L5178Y cells or L1210 cells treated with one amino acid-deficient medium for various times. Curves a, b and c: valine-, isoleucine- and asparagine-deficiency, respectively. The fragmentation of chromosomal DNA was analyzed by the method of Sellins et al.7 The protease activity of caspase-3 was determined by the method of Enari et al.8.
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