Regulation of Platelet-activating Factor Acetylhydrolase by Oxidized Phospholipids and Proinflammatory Cytokines
Author: Muralikrishna Mukkamala
Publisher:
Published: 2008
Total Pages:
ISBN-13:
DOWNLOAD EBOOKPlatelet-Activating Factor Acetylhydrolase (PAFAH) is a monocyte-derived phospholipase A2 that catalyzes the hydrolysis of platelet-activating factor (PAF) and has been implicated in atherosclerosis. Although PAF and other proinflammatory stimuli are postulated to induce the enzyme, mechanisms controlling PAFAH expression are largely unknown at present. We have shown that PAFAH induction in monocytes is increased in response to oxidized phospholipids. The PAFAH 5' flanking region has at least 10 putative Stat elements, and IL-6 has been shown to be downstream from the prostaglandin receptor, EP2, which has been shown to bind oxidized phospholipids, prompting the hypothesis that Stat proteins might regulate its expression. To test this hypothesis, we treated human monocytes with IL-6, a monocyte-derived cytokine that activates Stat3, IL-8, a monocyte-derived cytokine induced by Stat3, and oxidized 1-palmitoyl-2-arachidonoyl-sn-3-phosphocholine (oxPAPC), a major component of the oxidized LDL particle. Two monocyte-derived cell types, macrophages (MO) and dendritic cells (DC) were prepared from primary human monocytes. The cells were treated with various doses of IL-6, IL-8, or oxPAPC for various time frames in the absence of serum. Culture supernatants from the cytokine-treated cells were harvested and screened for PAFAH protein and activity and cell monolayers were assessed for PAFAH mRNA by quantitative real time PCR (qPCR). Cells treated with oxPAPC were further analyzed for secreted IL-6 using ELISA and activation of Stat3 using Western Blot. Both IL-6 and IL-8 induced PAFAH expression in a dose-dependent manner. Although both MO and DC responded to the cytokines, preliminary experiments suggested that induction of PAFAH is more robust in DC than MO. Cytokine-treated cells exhibited increased PAFAH activity in their culture supernatants that correlated with increased PAFAH protein levels. Treatment with oxPAPC induced IL-6 secretion and subsequent Stat3 activation in DC. Together, these data support the hypothesis that PAFAH expression is regulated by oxidized phospholipids and proinflammatory cytokines in developing atheromas.
