Hoque Mehboob, Nanduri Ravikanth, Gupta Jyoti, Mahajan Sahil, Gupta Pawan, Saleemuddin M
Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh 202002, India.
CSIR-Institute of Microbial Technology, Chandigarh 160036, India.
Biochim Biophys Acta. 2015 Sep;1850(9):1729-39. doi: 10.1016/j.bbagen.2015.04.009. Epub 2015 Apr 23.
Complexes of oleic acid (OA) with milk α-lactalbumin, received remarkable attention in view of their selective toxicity towards a spectrum of tumors during the last two decades. OA complexes of some structurally related/unrelated proteins are also tumoricidal. Erythrocytes are among the few differentiated cells that are sensitive and undergo hemolysis when exposed to the complexes.
The effects of OA complex of bovine α-lactalbumin (Bovine Alpha-lactalbumin Made LEthal to Tumor cells, BAMLET) on human, goat and chicken erythrocytes on calcein leakage, phosphatidylserine exposure, morphological changes and hemolysis were studied by confocal microscopy, FACS analysis, scanning electron microscopy and measuring hemoglobin release.
Erythrocytes exposed to BAMLET undergo eryptosis-like alterations as revealed by calcein leakage, surface phosphatidylserine exposure and transformation to echinocytes at low concentrations and hemolysis when the concentration of the complex was raised. Ca(2+) was not essential and restricted the alterations when included in the medium. The BAMLET-induced alterations in human erythrocytes were prevented by the cation channel inhibitors, amiloride and BaCl2 but not by inhibitors of thiol proteases, sphingomyelinase and by the antioxidant N-acetyl cysteine.
The work shows for the first time that low concentrations of BAMLET induces eryptosis in erythrocytes by a novel mechanism not requiring Ca(2+) and hemolysis by detergent-like action by the released OA at higher concentrations.
The study points out to the need for a comprehensive evaluation of the toxicity of OA complexes of α-lactalbumin and other proteins towards erythrocytes and other differentiated cells before being considered for therapy.
油酸（OA）与牛奶α-乳白蛋白的复合物在过去二十年中因其对一系列肿瘤的选择性毒性而备受关注。一些结构相关/不相关蛋白质的OA复合物也具有杀肿瘤作用。红细胞是少数几种分化细胞，当暴露于这些复合物时会敏感并发生溶血。
通过共聚焦显微镜、流式细胞术分析、扫描电子显微镜和测量血红蛋白释放，研究了牛α-乳白蛋白的OA复合物（牛α-乳白蛋白使肿瘤细胞致死，BAMLET）对人、山羊和鸡红细胞的钙黄绿素泄漏、磷脂酰丝氨酸暴露、形态变化和溶血的影响。
暴露于BAMLET的红细胞会发生类似细胞凋亡的改变，如低浓度时钙黄绿素泄漏、表面磷脂酰丝氨酸暴露并转变为棘状细胞，当复合物浓度升高时则发生溶血。Ca(2+)并非必需，且当培养基中含有Ca(2+)时会限制这些改变。阳离子通道抑制剂阿米洛利和BaCl2可预防BAMLET诱导的人红细胞改变，但巯基蛋白酶抑制剂、鞘磷脂酶抑制剂和抗氧化剂N-乙酰半胱氨酸则不能。
该研究首次表明，低浓度的BAMLET通过一种不需要Ca(2+)的新机制诱导红细胞发生细胞凋亡，而高浓度时释放的OA通过类似去污剂的作用导致溶血。
该研究指出，在考虑将α-乳白蛋白和其他蛋白质的OA复合物用于治疗之前，需要对其对红细胞和其他分化细胞的毒性进行全面评估。
