Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh, India.
PLoS One. 2013 Sep 11;8(9):e68390. doi: 10.1371/journal.pone.0068390. eCollection 2013.
A chance discovery of the tumoricidal action of a human milk fraction led to the characterization of the active component as oleic acid complex of the α-lactalbumin, which was given the acronym HAMLET. We report in this study that the oleic acid complex of bovine α-lactalbumin (BAMLET) is hemolytic to human erythrocytes as well as to those derived from some other mammals. Indirect immunofluorescence analysis suggested binding of BAMLET to erythrocytes prior to induction of hemolysis. Free OA was hemolytic albeit at higher concentrations, while sodium oleate caused hemolysis at far lower concentrations. Amiloride and BaCl2 offered protection against BAMLET-induced hemolysis suggesting the involvement of a cation leak channel in the process. BAMLET coupled to CNBr-activated Sepharose was not only hemolytic but also tumoricidal to Jurkat and MCF-7 cells in culture. The Sepharose-linked preparation was however not toxic to non-cancerous peritoneal macrophages and primary adipocytes. The tumoricidal action was studied using the MTT-assay while apoptosis induction measured by the annexin V-propidium iodide assay. Repeated incubation of the immobilized BAMLET with erythrocytes depleted oleic acid and decreased the hemolytic activity of the complex. Incubation of MCF-7 and Jurkat cells with OA, soluble or immobilized BAMLET resulted in increase in the uptake of Lyso Tracker Red and Nile red by the cells. The data presented support the contention that oleic acid plays the key role, both in BAMLET-induced hemolysis and tumoricidal action.
人乳某部分的抑瘤活性的偶然发现导致了活性成分被鉴定为α-乳白蛋白的油酸复合物，其缩写为 HAMLET。我们在这项研究中报告，牛α-乳白蛋白的油酸复合物（BAMLET）对人红细胞以及一些其他哺乳动物的红细胞具有溶血作用。间接免疫荧光分析表明，BAMLET 在诱导溶血之前与红细胞结合。游离 OA 虽然在较高浓度下也具有溶血作用，但油酸钠在低得多的浓度下即可引起溶血。阿米洛利和 BaCl2 提供了对 BAMLET 诱导的溶血的保护作用，表明该过程涉及阳离子泄漏通道。与 CNBr 激活的琼脂糖偶联的 BAMLET 不仅具有溶血作用，而且对培养的 Jurkat 和 MCF-7 细胞也具有杀瘤作用。然而，与非癌细胞腹膜巨噬细胞和原代脂肪细胞相比，连接到琼脂糖上的制剂没有毒性。使用 MTT 测定法研究了杀瘤作用，而通过 Annexin V-propidium iodide 测定法测量了细胞凋亡的诱导。用红细胞耗尽油酸重复孵育固定化 BAMLET 会降低复合物的溶血活性。用 OA、可溶性或固定化 BAMLET 孵育 MCF-7 和 Jurkat 细胞会导致细胞内 Lyso Tracker Red 和 Nile red 的摄取增加。所提供的数据支持油酸在 BAMLET 诱导的溶血和杀瘤作用中起关键作用的论点。
