Author + information
- Cong Guangzhi,
- Ru Yan,
- Kai Wang,
- Hui Huang and
- Shaobin Jia
Hyperhomocysteinemia(Hhcy) promotes unstable atherosclerotic plaque formation. Macrophage apoptosis and plaque necrosis are the two hallmarks of advanced plaque. Endoplasmic reticulum stress plays a critical role in macrophage apoptosis and plaque instability induced by homocysteine. The present study was sight to explore the mechanisms macrophage apoptosis mediated by endoplasmic reticulum stress in the the pathogenesis of unstable plaque in Hhcy ApoE double knock (ApoE-/-) mice.
Four week old C57BL/6J mice and ApoE-/- mice were received a control (n= 12) or methionine (2%) diet (n= 12). After 16 weeks on the diets, mice were sacrificed, and the levels of total plasma homocysteine, plasma cholesterol and triglycerides were analyzed. Parameters of advanced plaque progression including necrotic core size, collagen content and macrophage apoptosis in both en face aortae and aortic valve sections were quantified. Meanwhile, the makers of endoplasmic reticulum stress were detected in aortas of mice. Furthermore, Raw264.7 macrophage were challenged with homocysteine (50, 100,200 or 500 μmol/L) in the absence or presence of ox-LDL (50ug/ml) or PBA(an ER stress inhibitor). The apoptosis and makers of endoplasmic reticulum stress were comprehensively accessed.
Hhcy was observed in all groups supplemented with methionine, compared to diet-matched control groups(P＜0.05). The differences of plasma total cholesterol levels or lipid profiles were not observed between methionine-supplemented groups and the diet-matched control groups (P＞0.05). Atherosclerotic lesions were only observed in both en face aortae and aortic valve sections in ApoE-/- mice mice but not C57BL/6J mice even though the methionine diet. The methionine-supplemented ApoE-/- mice showed significantly increased atherosclerotic lesions area (P＜0.05). Necrotic core area and lesion apoptotic cells was significantly increased and collagen content reduced in plaques of ApoE-/- mice supplemented with methionine (P＜0.05). The expression of endoplasmic reticulum stress makers included BIP, ERO1 α and IRE1α were also increased in the methionine-supplemented ApoE-/- mice (P＜0.05). Homocysteine enhanced foam cell formation with or without OX-LDL in RAW264.7 cell lines in a dose dependent manner (P＜0.05). But Homocysteine induced RAW264.7 cell apoptosis only in the presence of ox-LDL (50ug/ml) by AV/PI and Tunnel assays (P＜0.05). The expression of endoplasmic reticulum stress makers included BIP, ERO1 α and IRE1α were also increased in the homocysteine challenged RAW264.7 cell in the absence or presence of ox-LDL (50ug/ml) (P＜0.05). PBA inhibited foam cell formation, attenuated apoptosis and inhibited expression of ER stress marker (P＜0.05).
Hhcy promotes unstable plaque formation in ApoE-/-mice. ER stress may play an important role in macrophage apoptosis and plaque instability induced by Hhcy.