Amelioration of atherosclerosis in apolipoprotein E-deficient mice by combined RNA interference of lipoprotein-associated phospholipase A2 and YKL-40.

Hui Zhang,Wenping Zhou,Wenjing Zhang,Chang Cao,Gangqiong Liu,Jinying Zhang,Junming Yue

doi:10.1371/journal.pone.0202797

Abstract

To test the hypothesis that combined RNA interference (RNAi) of lipoprotein-associated phospholipase A2 (Lp-PLA2) and YKL-40 is superior to RNAi of Lp-PLA2 or YKL-40 alone in ameliorating atherosclerosis. A total of 120 apolipoprotein E-deficient mice (apoE-/- mice) were randomly divided into five groups, including the vehicle alone, scrambled RNAi, Lp-PLA2 RNAi, YKL-40 RNAi, and combined Lp-PLA2 and YKL-40 RNAi groups. Constrictive collars were used to induce plaque formation. Lp-PLA2 RNAi and YKL-40 RNAi viral suspensions were transduced into carotid plaques of the mice. Carotid plaques were harvested for histological analysis four weeks after viral vector transduction. Inflammatory gene expression in the plasma and atherosclerotic plaques was determined by ELISA and real-time PCR. Four weeks after RNAi, the serum concentration and plaque mRNA expression of Lp-PLA2 and YKL-40 were remarkably attenuated, leading to reduced inflammatory gene expression. Plaques from the Lp-PLA2 or YKL-40 RNAi group showed lower lipid content, higher collagen content, increased fibrous cap thickness, and lower mRNA expressions of MCP-1 and MMP-8 than than those in the vehicle and scramble groups. When compared with the isolated Lp-PLA2 or YKL-40 RNAi group, the combined Lp-PLA2 and YKL-40 RNAi group exhibited higher collagen content and fibrous cap thickness, and lower lipid content and local inflammation. The beneficial effects of RNAi were independent of the plasma lipoprotein profile. Combined RNAi of Lp-PLA2 and YKL-40 is superior to RNAi of Lp-PLA2 or YKL-40 alone in ameliorating atherosclerosis.

Highlights

Atherosclerosis and its clinical complications are the leading causes of death and disability in the western world [1]
Our previous work has shown that Lp-PLA2 RNA interference (RNAi) inhibited the expression of monocyte chemotactic protein-1 (MCP-1) and matrix metalloproteinase-8 (MMP-8) in RAW264.7 cells [4,5], and the expression of MCP-1 and MMP-8 was sharply reduced after YKL-40 RNAi (Fig 1B and 1C)
One major finding of the current investigation was that the expression of both Lp-PLA2 and YKL-40 in collar-induced atherosclerotic plaques was remarkably attenuated by RNAi

Summary

Introduction

Atherosclerosis and its clinical complications are the leading causes of death and disability in the western world [1]. Described inflammation mediators, such as lipoprotein-associated phospholipase A2 (Lp-PLA2) and YKL-40, are highly expressed in atherosclerotic plaques and contribute significantly to the progression of atherosclerosis [2]. Plasma YKL-40 is associated with cardiovascular and all-cause mortality [7, 8] Increased concentrations of both Lp-PLA2 and YKL-40 have been reported in patients with atherosclerosis [2,3,4,5,6,7,8]. It is likely that both Lp-PLA2 and YKL-40 may contribute significantly to the formation and progression of atherosclerosis, and we hypothesized that simultaneously down-regulating the expression of LpPLA2 and YKL-40 may ameliorate atherosclerotic plaques more efficiently than knockdown of Lp-PLA2 or YKL-40 alone. We constructed two lentiviral vectors to down-regulate the expression of Lp-PLA2 or YKL-40 following collar-induced atherosclerosis in apolipoprotein E-deficient (apoE-/-) mice and tested the hypothesis that knockdown of both YKL-40 and Lp-PLA2 together is more effective than knockdown of either alone in ameliorating atherosclerosis in apoE-/- mice

Methods

Results

Discussion