Role of group II secretory phospholipase A₂ in atherosclerosis: 2. Potential involvement of biologically active oxidized phospholipids

Secretory nonpancreatic phospholipase A₂ (group II sPLA₂) is induced in inflammation and present in atherosclerotic lesions. In an accompanying publication we demonstrate that transgenic mice expressing group II sPLA₂ developed severe atherosclerosis. The current study was undertaken to determine whether 1 mechanism by which group II sPLA₂ might contribute to the progression of inflammation and atherosclerosis is by increasing the formation of biologically active oxidized phospholipids. In vivo measurements of bioactive lipids were performed, and in vitro studies tested the hypothesis that sPLA₂ can increase the accumulation of bioactive phospholipids. We have shown previously that 3 oxidized phospholipids derived from the oxidation of 1-palmitoyl-2-arachidonoyl-sn-glycero-3- phosphorylcholine (PAPC) stimulated endothelial cells to bind monocytes, a process that is known to be an important step in atherogenesis. We now show that these 3 biologically active phospholipids are significantly increased in livers of sPLA₂ transgenic mice fed a high-fat diet as compared with nontransgenic littermates. We present in vitro evidence for several mechanisms by which these phospholipids may be increased in sPLA₂ transgenics. These studies demonstrated that polyunsaturated free fatty acids, which are liberated by sPLA₂, increased the formation of bioactive phospholipids in LDL, resulting in increased ability to stimulate monocyte- endothelial interactions. Moreover, sPLA₂-treated LDL was oxidized by cocultures of human aortic endothelial cells and smooth muscle cells more efficiently than untreated LDL. Analysis by electrospray ionization-mass spectrometry revealed that the bioactive phospholipids, compared with unoxidized PAPC, were less susceptible to hydrolysis by human recombinant group II sPLA₂. In addition, HDL from the transgenic mice and human HDL treated with recombinant sPLA₂ in vitro failed, in the coculture system, to protect against the formation of biologically active phospholipids in LDL. This lack of protection may in part relate to the decreased levels of paraoxonase seen in the HDL isolated from the transgenic animals. Taken together, these studies show that levels of biologically active oxidized phospholipids are increased in sPLA₂ transgenic mice; they also suggest that this increase may be mediated by effects of sPLA₂ on both LDL and HDL.