Serum amyloid A induces expression of secretory phospholipase A2 in vascular smooth muscle cells: A role in cholesterol trafficking.

摘要

Atherosclerosis is a multifactorial vascular disease characterized by inflammatory lesion formation. Acute phase serum amyloid A (SAA) accumulates in atherosclerotic lesions, but its function remains unclear. SAA causes movement of cholesterol to the endoplasmic reticulum in neonatal rat aortic smooth muscle cells. Data in this thesis support a role for SAA-induced secretory phospholipase A2, group IIA (sPLA2) expression in cholesterol trafficking. Increased sPLA2 activity was detected in media and cell layer samples from SAA-treated smooth muscle cells. SAA treatment increased in both sPLA2 mRNA and heteronuclear RNA indicating that sPLA2 expression increases at the level of transcription. To test this, transient transfections were performed using a plasmid containing a 488 bp rat sPLA2 promoter fragment and SAA increased promoter activity. Previous reports in the literature showed that interleukin-1beta (IL-1beta) upregulates sPLA2 gene expression via CAAT-enhancer binding protein beta (C/EBPbeta) and nuclear factor kappa B (NFkappaB). Therefore, the role of these transcription factors in SAA-induced sPLA2 gene expression was explored. C/EBPbeta mRNA and protein expression increased in SAA-treated smooth muscle cells. NFkappaB accumulated in the nucleus in cells treated with SAA. Transfection and electrophoretic mobility shift assays confirmed the involvement of C/EBP13 and NFkappaB in SAA-induced sPLA2 gene expression. Finally, the hypothesis that increased sPLA2 activity contributes to SAA-induced trafficking of cholesterol through activation of sphingomyelinase was tested. Sphingomyelinase was activated in SAA-treated cells and its inhibition blocked cholesterol trafficking. Moreover, inhibition of sPLA2 activity decreased the SAA-induced movement of cholesterol to the endoplasmic reticulum. Interestingly, although SAA upregulated expression of IL-1beta, SAA-induced sPLA2 activity and movement of cholesterol to the endoplasmic reticulum were not mediated via the IL-1 receptor. In conclusion, these studies suggest that SAA upregulates sPLA2 gene expression, that transcriptional upregulation is mediated by C/EBPbeta and NFkappaB and that increased sPLA2 contributes to SAA-induced cholesterol trafficking from the plasma membrane to the endoplasmic reticulum through the activation of sphingomyelinase. These findings suggest that SAA is an important factor in smooth muscle cell lipid homeostasis.