Intestinal fatty acid-binding protein mediates atherosclerotic progress through increasing intestinal inflammation and permeability

Abstract

Atherosclerosis is one of leading phenotypes of cardiovascular diseases, featured with increased vascular intima-media thickness (IMT) and unstable plaques. The interaction between gastrointestinal system and cardiovascular homeostasis is emerging as a hot topic. Therefore, the present study aimed to explore the role of an intestinal protein, intestinal fatty acid-binding protein (I-FABP/FABP2) in the atherosclerotic progress. In western diet-fed ApoE^-/- mice, FABP2 was highly expressed in intestine. Silence of intestinal Fabp2 attenuated western diet-induced atherosclerotic phenotypes, including decreasing toxic lipid accumulation, vascular fibrosis and inflammatory response. Mechanistically, intestinal Fabp2 knockdown improved intestinal permeability through increasing the expression of tight junction proteins. Meanwhile, intestinal Fabp2 knockdown mice exhibited down-regulation of intestinal inflammation in western diet-fed ApoE^-/- mice. In clinical patients, the circulating level of FABP2 was obviously increased in patients with cardiovascular disease and positively correlated with the value of carotid intima-media thickness, total cholesterol and triglyceride. In conclusion, FABP2-induced intestinal permeability could address a potential role of gastrointestinal system in the development of atherosclerosis, and targeting on intestinal FABP2 might provide a therapeutic approach to protect against atherosclerosis.

Keywords: atherosclerosis; inflammation; intestinal fatty acid-binding protein; intestinal permeability.

FIGURE 1

High‐fat high‐cholesterol diet increases the level of FABP2 in ApoE^−/− mice. A, The distribution of Fabp2 in aorta, heart, kidney, visceral fat and small intestine of ApoE^−/− mice fed with standard chow for 12 wk. B, Male ApoE^−/− mice were fed with high‐fat high‐cholesterol diet (HFHC) for 0, 4, 12 and 20 wk. The Fabp2 expression in small intestine. Data are shown as mean ± SEM (**P < .01 and ***P < .001, anova analysis was used for multiple group comparison, n = 5 mice/group)

FIGURE 2

Blockage of FABP2 attenuates HFHC diet–induced atherosclerotic progress in ApoE^−/− mice. Male ApoE^−/− mice were fed with HFHC for 12 wk and then treated with 1 × 10⁹ lentiviral particles encoding villin‐drived Fabp2 siRNA or control siRNA (Veh) for 8 wk. A‐B, Representative images of Oil Red O staining of En Face aorta (A) and quantitative analysis of relative density (B). C‐D, Oil red O staining of aorta root (C) and quantitative analysis of relative lipid deposits (D, red colour area). Scale bar = 100 μm. E‐H, Slides of aorta root were stained with moma‐2 and α‐sma antibodies, or trichrome staining solution. Representative images of staining (E), and quantitative analysis of relative density of aorta moma‐2 (F), α‐sma (G) and collagen (H). Scale bar = 50 μm. I‐M, Real‐time PCR analysis of aorta gene expression of Tnf‐α (I), Il‐1β (J), Mcp‐1 (K), Vcam‐1 (L) and Icam‐1 (M). Data are shown as mean ± SEM (**P < .01 and ***P < .001, Student's t test was used for 2‐group comparison, n = 5‐7 mice/group)

FIGURE 3

Blockage of FABP2 suppresses HFHC diet–induced intestinal permeability in ApoE^−/− mice. Male ApoE^−/− mice were fed with HFHC for 12 wk, then treated with 1 × 10⁹ lentiviral particles encoding villin‐drived Fabp2 siRNA or control siRNA (Veh) for 8 wk. A, Haematoxylin and eosin staining of small intestine. Scale bar = 100 μm. B‐C, Real‐time PCR analysis of intestinal zonula occludens (ZO)‐1 (B) and occludin (C) levels. D‐F, Western blot analysis of ZO‐1 and occluding (D), and quantitative analysis of relative density of ZO‐1/Tubulin (E) and occluding/Tubulin (F). G, Serum levels of lipopolysaccharides (LPS). H, The circulating concentration of DX‐4000‐FITC after mice was orally administrated with FITC‐labelled dextran. Data are shown as mean ± SEM (*P < .05, **P < .01 and ***P < .001, Student's t test was used for 2‐group comparison, n = 6‐7 mice/group)

FIGURE 4

Blockage of FABP2 inhibits intestinal inflammatory response in ApoE^−/− mice. Male ApoE^−/− mice were fed with HFHC for 12 wk and then treated with 1 × 10⁹ lentiviral particles encoding villin‐drived Fabp2 siRNA or control siRNA (Veh) for 8 wk. A‐E, Real‐time PCR analysis of inflammatory gene expression on small intestine. F‐H, Intestinal production of TNF‐α, IL‐1β and MCP‐1. Small intestine was collected and homogenized, and the levels of cytokines TNF‐α (F), IL‐1β (G) and MCP‐1 (H) were measured by ELISA. Data are shown as mean ± SEM (*P < .05, **P < .01 and ***P < .001, Student's t test was used for 2‐group comparison, n = 6‐7 mice/group)

FIGURE 5

Circulating FABP2 is closely correlated with atherosclerotic parameters in clinical patients. A, The plasma FABP2 levels in 18 patients with carotid intima‐media thickness (IMT) ≥ 0.85 mm and 24 patients with IMT < 0.85 mm. B‐D, The correlation between circulating levels of FABP2 and IMT (B), total cholesterol (C) and triglyceride (D). Data are shown as mean ± SEM (**P < .01, Student's t test was used for 2‐group comparison, and Pearson's analysis was used for the correlation)