Induction of cell death in RAW 264.7 cells by alpha-lactalbumin.
Alpha-lactalbumin (alpha-LA), a major human milk whey protein, has been reported to exhibit bactericidal properties, immune suppressive effects, anti-proliferation and apoptosis in transformed cells; however, little is known about its anti-inflammation and related molecular mechanism. In this study we investigated the effects of alpha-LA on macrophages. We found that treatment with high concentration alpha-LA (> or = 100 microg/ml) could result in a time- and dose-dependent decrease in growth activity, morphological changes, increase in hypodiploid DNA population, and DNA fragmentation in RAW 264.7 cells. We also found that high dose alpha-LA could induce cellular apoptosis and necrosis, as determined by Annexin V binding assay. The alpha-LA could enhance the expression levels of cytochrome c, active caspase 3, active caspase 8, extracellular signal-regulated kinase (ERK1/2) and c-Jun N-terminal kinase (JNK) activation without changing the protein levels, but suppress the protein level of Bcl-2. The broad-spectrum caspase inhibitor, Boc-D-fmk, failed to block cell death, indicating that alpha-LA-induced cell death was modulated in a caspase-independent manner. In addition, the ERK1/2 inhibitor, PD98059, could partially rescue alpha-LA-induced cell death, while the JNK inhibitor, SP600125, could weakly protect cells from death. Our results suggested that activation of ERK1/2 might mediate alpha-LA-induced cell death in RAW 264.7 cells.
Lin
I-Chun
IC
Biophysics Laboratory, Department of Research and Education, Taipei Veterans General Hospital, Taipei, Taiwan.
Su
Shu-Li
SL
Kuo
Cheng-Deng
CD
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D020287:In Situ Nick-End Labeling; D008264:Macrophages / Q000187:drug effects; D051379:Mice