Abstract:Aim To investigate the effect of curcumin on the progression of atherosclerosis and macrophage polarization in vivo. Methods The apolipoprotein E deficient (ApoE－/－) mice were fed with high fat diet to establish atherosclerosis model. The aortas were isolated for haematoxylin and eosin, masson trichrome, picrosirus red, oil red O and immunofluorescence staining. Inducible nitric oxide synthase (iNOS) and CD206 were utilized as biomarkers of M1 and M2 phenotypes, respectively. Quantitative real-time polymerase chain reaction (PCR) was carried out to examine the gene expression of inflammatory factors. Results Curcumin significantly decreased atherosclerotic burden and plaque vulnerability in the experimental atherosclerosis model. Furthermore, curcumin decreased the ratio of M1/M2 macrophages together with the gene expressions of pro-inflammatory interleukin 1beta (IL-1β), inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF-α), but promoted the levels of anti-inflammatory cytokines IL-10, Ym1, Fizz1 in the atherosclerotic plaque. Conclusion Curcumin could alleviate the progression of atherosclerosis by inhibiting macrophage polarizing towards M1 phenotype as well as inflammatory response.

Abstract:Aim In order to evaluate the instability of atherosclerotic plaque more comprehensively and objectively, the computational method of plaque vulnerability index based on lipid accumulation, which was put forward by Shiomi was improved in this study. Methods The experimental was divided into the control group (C57BL/6 mice) and the model group (LDLR－/－ mice and ApoE－/－ mice), with 6 mice in each group. The control group was given normal diet, while the model group was given high-fat diet. After 25 weeks, the mice were sacrificed and the aortic root and aortic arch were isolated and sectioned after OCT embedding. Serial sections were stained by hematoxylin and eosin (H&E), picrosirius red, oil red O, alcian blue, and quantitatively studied the amount of endothelial cell and smooth muscle cell by immunofluorescence, as well as matrix metalloproteinase and macrophage with immunohistochemistry. The calculation formula of plaque vulnerability index proposed by Shiomi was perfected through adding the factors affecting the plaque instability to the formula numerator and the stability index of the plaque to the denominator. The stability of aorta root and innominate artery plaque in LDLR－/－ mice was compared by two formulas, and the reliability of the optimized formula was evaluated. Finally, the stability of atherosclerotic plaques in different parts of aorta of ApoE－/－ and LDLR－/－ mice was evaluated by the optimized formula, and the difference of atherosclerosis progression between two genetically engineered mice was compared. Results The plaque damage area, lipid deposition, the distribution of collagen, proteoglycan and matrix metalloproteinase, macrophage infiltration, the endothelial cell coverage and smooth muscle cell coverage of fibrous cap in the model group were significantly changed compared with the control group. On this basis, the formula for calculating plaque instability based on multi index pathological staining was optimized. Through calculation, it was found that Plaque vulnerability index of innominate artery was significantly larger than that of aortic root in ApoE－/－ and LDLR－/－ mice. In the same site, plaque vulnerability index in ApoE－/－ mice was significantly higher than that in LDLR－/－ mice. Conclusion Optimized calculation formula of plaque vulnerability index based on multi index pathological staining covers a number of important factors that affect plaque instability, and can accurately assess plaque instability. The method is simple, comprehensive, reliable and of great practical value.

Abstract:Aim To explore the effect and significance of mammalian sterile 20-like kinase 1 (MST1) and its DNA methylation in the kidney damage of apolipoprotein E gene knocked-out (ApoE－/－) mice. Methods The experimental animals were divided into 2 groups:(1)ApoE－/－ group (n＝10):male ApoE－/－ mice were fed with high methionine diet; (2)Control group (n＝10):male C57BL/6J mice were fed with high methionine diet. After 14 weeks of feeding, serum creatinine and urea nitrogen levels in mice were determined by full automatic biochemical analyzer. PAS staining and transmission electron microscope were used to observe the renal tissue damage in mice. Real-time quantitative PCR and Western blot were used to detect the expression levels of MST1 mRNA and protein in mice kidney. MST1 DNA methylation level in mice kidney was detected by nested methylation specific PCR (nMS-PCR). Results Compared with the control group, serum levels of creatinine and urea nitrogen were increased by 1.1 times and 1.6 times in ApoE－/－ group (P＜0.01). PAS staining and transmission electron microscopy showed that the kidney was obviously damaged in ApoE－/－ group compared with control group. In ApoE－/－ mouse kidney, MST1 expression was positively correlated with serum creatinine and urea nitrogen levels (R2＝0.7571, P＝0.0012; R2＝0.7342, P＝0.0015). The result of nMS-PCR showed that renal MST1 DNA methylation level in ApoE－/－ group was significantly lower than that in control group (P＜0.01). Conclusions The upregulation of MST1 expression may play an important role in kidney damage of ApoE－/－ mice. DNA low methylation in MST1 promoter region may be an important mechanism of MST1 expression upregulation.