不同濃度沙利度胺對K562細胞凋亡及血管內皮生長因子分泌的影響_《華西醫學》

作者：

張玉高 ¹ ,  韓麗英 ¹ , 陳楓 ² , 趙華 ³

瀘州醫學院附屬醫院（四川瀘州，646000） 1 風濕血液科，2 傳染科，3 檢驗科;

關鍵詞：

慢性粒細胞白血病沙利度胺凋亡 K562細胞血管內皮生長因子

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【摘要】　目的　研究沙利度胺對人慢性粒細胞白血病急變株K562細胞凋亡及血管內皮生長因子（vascular endothelia growth factor，VEGF）分泌的影響。　方法　采用不同濃度的沙立度胺（0.5、1.0、2.0 mmol/L）作用于K562細胞24、48、72、96 h，瑞-姬（Wright-Giemsa）染色法觀察細胞形態；四甲基偶氮唑鹽（methylthiazolyl tetrozolium，MTT）法檢測細胞增殖；流式細胞儀膜聯蛋白V-異硫氰酸熒光素/碘化丙啶雙染法檢測凋亡率；瓊脂糖凝膠電泳法檢測脫氧核糖核酸梯狀條帶；酶聯免疫吸附法檢測VEGF濃度。　結果　培養24、48 h后，沙立度胺對K562細胞生長無抑制作用；作用72 h后，1.0、 2.0 mmol/L濃度組開始出現對K562細胞生長的抑制（P lt;0.001）；作用96 h后，0.5 mmol/L濃度組也產生對K562細胞生長的抑制（P lt;0.001），呈一定的濃度和時間依賴性。沙立度胺處理72 h后，K562細胞出現形態學改變，其體積縮小，出現空泡化，邊緣出現突起，染色質濃縮、邊集，核固縮、出現凋亡小體。經沙立度胺處理后，流式分析結果顯示K562細胞凋亡率增加（P lt;0.001）。沙立度胺作用72 h后，瓊脂糖凝膠電泳可見典型的DNA梯狀條帶。K562細胞培養48 h后，沙立度胺抑制VEGF的分泌（P lt;0.001），并且VEGF濃度與凋亡率呈負相關（r=-0.789）。　結論　沙利度胺抑制K562細胞的增殖，呈一定的濃度和時間依賴性；沙利度胺對K562細胞凋亡有明顯誘導作用；沙利度胺抑制K562細胞VEGF的分泌。
【Abstract】　Objective　 To investigate the effect of thalidomide on apoptosis of k562 cells and its vascular endothelial growth factor secretion. 　Methods　 K562 cells were cultured in vitro with 0.5, 1.0, and 2.0 mmol/L thalidomide for 24, 48, 72, and 96 hours. Morphology of the K562 cells was observed by the Wright-Giemsa staining method. Methylthiazolyl tetrozolium （MTT） assay was used to determine the cell growth. The rate of apoptosis was analyzed by flow cytometry (FCM) with annexin V-fluorescein isothiocyanate/propidium iodide （AnnexinV-FITC/PI）double-staining method. Agarose gel electrophoresis was used to detected Deoxyribonucleic acid Ladder（DNA Ladder）. The concentration of VEGF was quantified by the enzyme-linked immunosorbent assay (ELISA).　Results　 Cultured for 24 or 48 hours, thalidomide had no effect on the proliferation of the K562 cells. But after cultured for 72 hours, thalidomide began to inhibit the growth of the K562 cells at the concentration of 1.0 and 2.0 mmol/L （P lt;0.001）. After cultured for 96 hours, the proliferation of the K562 cells was inhibited too at the concentration of 0.5 mmol/L thalidomide （P lt;0.001）. Thus, thalidomide inhibited the growth of the K562 cells with a dose-and time-dependent manner to some extent. After exposure to thalidomide for 72 hours, K562 cells underwent typical morphological changes of apoptosis such as vaculization, the budding of cytoplasm, chromatin condensation, margination, shrunken nucleus and apoptotic body. The results of flow cytometry showed that thalidomide could obviously increase the rates of the apoptosis of K562 cells （P lt;0.001）. After treated with thalidomide for 72 hours, DNA was extracted for Agarose gel electrophoresis and typical DNA ladder strips were observed. The secretion of VEGF was inhibited when exposure to thalidomide for 48 hours（P lt;0.001）, and there was negative correlation between VEGF concentrations and apoptotic rates（r=-0.789）.　Conclusions　 Thalidomide could inhibite the growth of the K562 cells with a dose-and time-dependent manner to some extent. Thalidomide could obviously induce the apoptosis of the K562 cells and inhibit its secretion of VEGF.

引用本文： 張玉高,韓麗英,陳楓,趙華. 不同濃度沙利度胺對K562細胞凋亡及血管內皮生長因子分泌的影響. 華西醫學, 2011, 26(9): 1298-1302. doi: 復制

1.	Katouli AA, Komarova NL. Optimizing combination therapies with existing and future CML drugs[J]. Plos One, 2010, 5(8): 1-12.
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3.	Venepalli N, Rezvani K, Mielke S, et al. Role of allo-SCT for CML in 2010[J]. Bone Marrow Transpl, 2010, 45(11): 1579-1586.
4.	Volpe G, Panuzzo C, Ulisciani S, et al. Imatinib resistance in CML[J]. Cancer Lett, 2009, 274(1): 1-9.
5.	Quentmeier H, Eberth S, Romani J, et al. BCR-ABL1-independent PI3 Kinase activation causing imatinib-resistance[J]. J Hematol Oncol, 2011, 4(1): 6-15.
6.	韓麗英, 陳楓, 張玉高. 大蒜素對K562細胞增殖抑制和誘導凋亡作用的實驗研究[J]. 臨床血液學雜志, 2006, 19(4): 239-240.
7.	韓麗英, 陳楓, 張玉高. 他莫昔芬誘導K562細胞凋亡的實驗研究[J]. 白血病·淋巴瘤, 2006, 15(1): 13-15.
8.	Du GJ, Lin HH, Xu QT, et al. Thalidomide inhibits growth of tumors through COX-2 degradation independent of antiangiogenesis[J]. Vasc Pharmacol, 2005, 43(2): 112-119.
9.	Lima LM, Manssour Fraga CA, Gonalves Koatz VL, et al. Thalidomide and analogs as anti-Inflammatory and immunomodulator drug candidates[J]. Anti-Inflammatory & Anti-Aller, 2006, 5(1): 79-95.
10.	Sissung TM, Thordardottir S, Gardner ER, et al. Current Status of Thalidomide and CC-5013 in the treatment of metastatic prostate cancer[J]. Anti-Cancer Agent Me, 2009, 9(10): 1058-1069.
11.	Kastritis E, Palumbo A, Dimopoulos MA. Treatment of relapsed/refractory multiple myeloma[J]. Semin Hematol, 2009, 46(2): 143-157.
12.	Laubach JP, Schlossman RL, Mitsiades CS, et al. Thalidomide, lenalidomide and bortezomib in the management of newly diagnosed multiple myeloma[J]. Expert Rev Hematol, 2011, 4(1): 51-60.
13.	張玉高, 韓麗英. 反應停的抗腫瘤作用機制及臨床應用[J]. 醫學綜述, 2007, 13(3): 200-202.
14.	Ergun MA, Konac E, Erbas D, et al. Apoptosis and nitric oxide release induced by thalidomide, gossypol and dexamethasone in cultured human chronic myelogenous leukemic K-562 cells[J]. Cell Bio Int, 2004, 28(3): 237-242.
15.	Ayala F, Dewar R, Kieran M, et al. Contribution of bone microenvironment to leukemogenesis and leukemia progression[J]. Leukemia, 2009, 23(12): 2233-2241.
16.	Fragoso R, Elias AP, Dias S. Autocrine VEGF loops, signaling pathways, and acute leukemia regulation[J]. Leukemia Lymphoma, 2007, 48(3): 481-488.

1. 　Katouli AA, Komarova NL. Optimizing combination therapies with existing and future CML drugs[J]. Plos One, 2010, 5(8): 1-12.
2. 　Hehlmann R, Hochhous A, Baccarani M. Chronic myeloid leukaemia[J]. Lancet, 2007, 370(9584): 342-350.
3. 　Venepalli N, Rezvani K, Mielke S, et al. Role of allo-SCT for CML in 2010[J]. Bone Marrow Transpl, 2010, 45(11): 1579-1586.
4. 　Volpe G, Panuzzo C, Ulisciani S, et al. Imatinib resistance in CML[J]. Cancer Lett, 2009, 274(1): 1-9.
5. 　Quentmeier H, Eberth S, Romani J, et al. BCR-ABL1-independent PI3 Kinase activation causing imatinib-resistance[J]. J Hematol Oncol, 2011, 4(1): 6-15.
6. 　韓麗英, 陳楓, 張玉高. 大蒜素對K562細胞增殖抑制和誘導凋亡作用的實驗研究[J]. 臨床血液學雜志, 2006, 19(4): 239-240.
7. 　韓麗英, 陳楓, 張玉高. 他莫昔芬誘導K562細胞凋亡的實驗研究[J]. 白血病·淋巴瘤, 2006, 15(1): 13-15.
8. 　Du GJ, Lin HH, Xu QT, et al. Thalidomide inhibits growth of tumors through COX-2 degradation independent of antiangiogenesis[J]. Vasc Pharmacol, 2005, 43(2): 112-119.
9. 　Lima LM, Manssour Fraga CA, Gonalves Koatz VL, et al. Thalidomide and analogs as anti-Inflammatory and immunomodulator drug candidates[J]. Anti-Inflammatory & Anti-Aller, 2006, 5(1): 79-95.
10. 　Sissung TM, Thordardottir S, Gardner ER, et al. Current Status of Thalidomide and CC-5013 in the treatment of metastatic prostate cancer[J]. Anti-Cancer Agent Me, 2009, 9(10): 1058-1069.
11. 　Kastritis E, Palumbo A, Dimopoulos MA. Treatment of relapsed/refractory multiple myeloma[J]. Semin Hematol, 2009, 46(2): 143-157.
12. 　Laubach JP, Schlossman RL, Mitsiades CS, et al. Thalidomide, lenalidomide and bortezomib in the management of newly diagnosed multiple myeloma[J]. Expert Rev Hematol, 2011, 4(1): 51-60.
13. 　張玉高, 韓麗英. 反應停的抗腫瘤作用機制及臨床應用[J]. 醫學綜述, 2007, 13(3): 200-202.
14. 　Ergun MA, Konac E, Erbas D, et al. Apoptosis and nitric oxide release induced by thalidomide, gossypol and dexamethasone in cultured human chronic myelogenous leukemic K-562 cells[J]. Cell Bio Int, 2004, 28(3): 237-242.
15. 　Ayala F, Dewar R, Kieran M, et al. Contribution of bone microenvironment to leukemogenesis and leukemia progression[J]. Leukemia, 2009, 23(12): 2233-2241.
16. 　Fragoso R, Elias AP, Dias S. Autocrine VEGF loops, signaling pathways, and acute leukemia regulation[J]. Leukemia Lymphoma, 2007, 48(3): 481-488.

《華西醫學》

不同濃度沙利度胺對K562細胞凋亡及血管內皮生長因子分泌的影響

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《華西醫學》

不同濃度沙利度胺對K562細胞凋亡及血管內皮生長因子分泌的影響

摘要 全文 圖表 視頻 參考文獻 施引文獻 補充材料

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Content

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