Vertical Sleeve Gastrectomy Offers Protection against Disturbed Flow-Induced Atherosclerosis in High-Fat Diet-Fed Mice

Abstract

Bariatric surgery reduces body weight, enhances metabolic and diabetic control, and improves outcomes on obesity-related comorbidities. However, the mechanisms mediating this protection against cardiovascular diseases remain unclear. We investigated the effect of sleeve gastrectomy (SG) on vascular protection in response to shear stress-induced atherosclerosis using an overweighted and carotid artery ligation mouse model. Eight-week-old male wild-type mice (C57BL/6J) were fed a high-fat diet (HFD) for two weeks to induce weight gain and dysmetabolism. SG was performed in HFD-fed mice. Two weeks after the SG procedure, partial carotid-artery ligation was performed to promote disturbed flow-induced atherosclerosis. Compared with the control mice, HFD-fed wild-type mice exhibited increased body weight, total cholesterol level, hemoglobin A1c, and enhanced insulin resistance; SG significantly reversed these adverse effects. As expected, HFD-fed mice exhibited greater neointimal hyperplasia and atherosclerotic plaques than the control group, and the SG procedure attenuated HFD-promoted ligation-induced neointimal hyperplasia and arterial elastin fragmentation. Besides, HFD promoted ligation-induced macrophage infiltration, matrix metalloproteinase-9 expression, upregulation of inflammatory cytokines, and increased vascular endothelial growth factor secretion. SG significantly reduced the above-mentioned effects. Moreover, HFD restriction partially reversed the intimal hyperplasia caused by carotid artery ligation; however, this protective effect was significantly lower than that observed in SG-operated mice. Our study demonstrated that HFD deteriorates shear stress-induced atherosclerosis and SG mitigates vascular remodeling, and this protective effect was not comparable in HFD restriction group. These findings provide a rationale for using bariatric surgery to counter atherosclerosis in morbid obesity.

Keywords: atherosclerosis; bariatric surgery; carotid artery ligation; insulin resistance; obesity; type 2 diabetes; vertical sleeve gastrectomy.

Figure 1

Effects of SG on glucose and cholesterol metabolism in HFD-fed mice. Timeline for the experimental procedures, blood sampling, and tissue processing during the study (A). The SG and carotid artery ligation procedures are illustrated in (B,C). Changes of glucose, insulin, and homeostatic assessment model for insulin resistance (HOMI-IR) were assessed (D–H). The cholesterol level in mice increased significantly after HFD and decreased significantly after SG (I). The intraperitoneal glucose tolerance test and area under curve (IPGTT-AUC), hemoglobin A1c (HbA1c), HOMA-IR for glucose tolerance and insulin resistance survey were measured (G–I) (n = 12–15 per group). The p for the trend of IPGTT with one-way ANOVA and Scheffee post-hoc test was all significant with a p = 0.003. The differences between chow diet, HFD, and HFD with SG were significant. Significant differences were not observed in the above-mentioned groups that did/did not undergo carotid artery ligation. The difference in IPGTT in HFD-fed mice that underwent SG was significant compared with the HFD-fed mice without SG. A significant difference was not observed upon comparison with control chow-fed mice. Chow diet: Picolab Rodent Diet 20; 4 kcal/g, 2% cholesterol, 5.7% fat; High fat diet: Teklad diet TD 88137; 4.5 kcal/g, 0.2% cholesterol, 21.2% fat; Harlan Tackle Co. * High-fat diet (HFD) compared with chow diet (CD) in mice that underwent carotid artery ligation or those that did not. * p < 0.05, ** p < 0.005, ^† Mice that underwent SG compared to those that did not undergo SG with or without carotid ligation. ^† p < 0.05, ^†† p < 0.005. The p-value for trend and one-way ANOVA were estimated for diet intake and serial changes of glucose in IPGTT with Scheffee and Tukey post hoc tests.

Figure 2

SG alleviated the levels of circulating HFD-induced inflammatory cytokines. The circulating concentrations of TNF-α (A), IL-6 (B), MMP-9 (C), and VEGF (D) as estimated by ELISA in mice. (n = 12 per group), Bar graphs indicate mean ± SD; High-fat diet (HFD) compared with chow diet (CD) in mice with or without carotid ligation, ** p < 0.005, ^†† p < 0.005. Two-way analysis of variance was performed with one treatment parameter being normal chow, HFD, and HFD with SG, and the other parameter being carotid ligation/no carotid ligation. A p-value < 0.05 was considered to be statistically significant.

Figure 3

SG attenuated HFD-induced intimal thickening. Representative light micrographs from the ligated left common carotid artery of the mice. H&E and VVG staining were performed (A,B) Intima, media areas, intima/media area ratio, and arterial fragmentation counts in all groups were measured. Two-way analysis of variance was performed with one treatment parameter being normal chow, HFD, and HFD with SG, and the other parameter being carotid ligation/no carotid ligation. A p-value < 0.05 was considered to be statistically significant. (C–E) (n = 6 per group), Scale bars = 100 μm. * High-fat diet (HFD) compared with chow diet (CD) in mice with or without carotid ligation, * p < 0.05, ** p < 0.005, ^†† p < 0.005.

Figure 4

SG ameliorated HFD induced inflammatory reaction and protein expression in the ligated carotid artery. Representative light micrographs from the ligated left common carotid artery of the mice. Carotid artery immunostaining revealed ox-LDL lesions, macrophage infiltration, and MMP-9 expression (A,B and Supplementary Figure S3A through Figure S3D). The right panels are the quantified immunohistochemical results by densitometry analysis (B and Supplementary Figure S3B,D). Scale bars: 100 μm. (n = 6 per group). The proteins expression of TNF-α, MMP-9, and VEGF were determined in the carotid artery (C–F). Data are illustrated as the mean ± SD. (n = 4 per group), * High-fat diet (HFD) compared with chow diet (CD) in mice with or without carotid ligation, Two-way analysis of variance was also performed with one treatment parameter being normal chow, HFD, and HFD with SG, and the other parameter being carotid ligation/no carotid ligation. A p-value < 0.05 was considered to be statistically significant. * p < 0.05, ** p < 0.005, ^† Mice with SG with or without carotid ligation, ^† p < 0.05, ^†† p < 0.005.