Mirela Hasu

Deficiency of invariant Vα14 natural killer T cells decreases atherosclerosis in LDL receptor null mice

Abstract Aims CD1d-restricted natural killer T (NKT) cells function by regulating numerous immune responses during innate and adaptive immunity. Depletion of all populations of CD1d-dependent NKT cells has been shown by several groups to reduce atherosclerosis in two different mouse models of the disease. In this study, we determined if removal of a single (Vα14) NKT cell population protects mice from the disease. Methods and results Targeted deletion of the Jα18 gene results in selective depletion of CD1d-dependent Vα14 NKT cells in C57BL/6 mice without affecting the population of other NKT, NK, and conventional T cells. Therefore, to study the effect of Vα14 NKT cell depletion on the progression of atherosclerosis, we examined the extent of lesion formation using paired littermate LDL receptor null mice that were either +/+ or −/− for the Jα18 gene following the feeding of these mice a cholesterol- and fat-enriched diet for 8 weeks. At the end of the study, we found no difference in either serum total- or lipoprotein-cholesterol distributions between groups. However, quantification of atherosclerosis revealed that Vα14 NKT cell deficiency significantly decreased lesion size in the aortic root (20–28%) and arch (28–38%) in both genders of mice. By coupling the techniques of laser capture microdissection with quantitative real-time RT–PCR, we found that expression of the proatherogenic cytokine interferon (IFN)-γ was significantly reduced in lesions from Jα18−/− mice. Conclusion This study is the first to identify a specific subpopulation of NKT cells that promotes atherosclerosis via a mechanism appearing to involve IFN-γ expression.

Caspase-1 Deficiency Decreases Atherosclerosis in Apolipoprotein E-Null Mice

BACKGROUND Caspase-1 is a cysteine protease that contributes to mammalian immunity through proteolytic activation of the proinflammatory cytokines, interleukin (IL)-1β and IL-18. METHODS To determine if caspase-1 deficiency can protect apolipoprotein E-null (Apoe(-/-)) mice from atherosclerosis, gender-matched, paired-littermate Apoe(-/-) mice with (Casp1(+/+)Apoe(-/-)) or without (Casp1(-/-)Apoe(-/-)) a functional caspase-1 (Casp1) gene were fed either a low fat diet for 26 weeks, or a saturated fat and cholesterol-enriched diet for 8 weeks. Plasma lipids and lipoproteins were determined and atherosclerosis was quantified in the aortic sinus and aortic arch. RESULTS On either diet, caspase-1 deficiency did not affect total serum cholesterol concentrations and lipoprotein-cholesterol distributions. However, caspase-1 deficiency significantly decreased atherosclerosis in the ascending aorta by 35%-45% in both sexes of mice fed either diet. We further examined atherosclerotic lesions for 2 indices of immune cell activation: Major Histocompatibility Complex (MHC) class II and interferon (IFN)-γ expression. There was a 40%-50% reduction in the number of lesion-associated cells expressing MHC class II from both sexes of Casp1(-/-)Apoe(-/-) mice compared with Casp1(+/+)Apoe(-/-) mice and, a significant reduction in lesion-associated IFN-γ in female Casp1(-/-)Apoe(-/-) compared with their Casp1(+/+)Apoe(-/-) counterparts. CONCLUSIONS We conclude that caspase-1 promotes atherosclerosis by enhancing the inflammatory status of the lesion through a mechanism likely involving activation of lesion-associated immune cells and IFN-γ expression.

Insulin-degrading enzyme deficiency in bone marrow cells increases atherosclerosis in LDL receptor-deficient mice.

BACKGROUND Insulin-degrading enzyme (IDE), a protease implicated in several chronic diseases, associates with the cytoplasmic domain of the macrophage Type A scavenger receptor (SR-A). Our goal was to investigate the effect of IDE deficiency (Ide(-/-)) on diet-induced atherosclerosis in low density lipoprotein-deficient (Ldlr(-/-)) mice and on SR-A function. METHODS Irradiated Ldlr(-/-) or Ide(-/-)Ldlr(-/-) mice were reconstituted with wild-type or Ide(-/-) bone marrow and, 6 weeks later, were placed on a high-fat diet for 8 weeks. RESULTS After 8 weeks on a high-fat diet, male Ldlr(-/-) recipients of Ide(-/-) bone marrow had more atherosclerosis, higher serum cholesterol and increased lesion-associated β-amyloid, an IDE substrate, and receptor for advanced glycation end products (RAGE), a proinflammatory receptor for β-amyloid, compared to male Ldlr(-/-) recipients of wild-type bone marrow. IDE deficiency in male Ldlr(-/-) recipient mice did not affect atherosclerosis or cholesterol levels and moderated the effects of IDE deficiency of bone marrow-derived cells. No differences were seen between Ldlr(-/-) and Ide(-/-)Ldlr(-/-) female mice reconstituted with Ide(-/-) or wild-type bone marrow. IDE deficiency in macrophages did not alter SR-A levels, cell surface SR-A, or foam cell formation. CONCLUSION IDE deficiency in bone marrow-derived cells results in larger atherosclerotic lesions, increased lesion-associated Aβ and RAGE, and higher serum cholesterol in male, Ldlr(-/-) mice.

Loss of Cellular Inhibitor of Apoptosis Protein 2 Reduces Atherosclerosis in Atherogenic apoE / C57BL/6 Mice on High-Fat Diet

Background Cellular inhibitor of apoptosis protein 2 (cIAP2) is predicted to participate in atherosclerosis; however, its direct role in atherosclerosis development has not been investigated. We aimed to examine and assess the loss of cIAP2 on atherosclerosis lesion development. Methods and Results We used apoE−/− C57BL/6 male mice, either cIAP2−/− or cIAP2+/+. At 8 weeks, mice were fed a high‐fat diet (HFD) for 4 and 12 weeks. Aortic root was serially sectioned and stained with Sudan IV, CD68, α‐actin, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL). cIAP2−/− mice displayed a significant decrease in atherosclerotic lesions macrophage number after 4 weeks of HFD. Similarly, decrease in lesion area at 4 and 12 weeks HFD was detected by use of en face analysis (cIAP2−/− 0.58±0.37% versus cIAP2+/+ 1.51±0.79% [P=0.0056]); (cIAP2−/− 9.34±4.88% versus cIAP2+/+ 17.65±6.24% [P=0.0019]). Aortic root lesion area after 4 and 12 weeks of HFD also decreased (cIAP2−/− 0.0328±0.014 mm2 versus cIAP2+/+ 0.0515±0.021 mm2 [P=0.022]); (cIAP2−/− 0.3614±0.1157 mm2 versus cIAP2+/+ 0.4901±0.125 mm2 [P=0.065]). TUNEL analysis after 4 and 12 weeks of HFD showed a 2.5‐fold increase in TUNEL+ cells (cIAP2−/− 4.47±2.26% versus cIAP2+/+ 1.74±0.98% [P=0.036]); (cIAP2−/− 2.39±0.75% versus cIAP2+/+ 1.29±0.47% [P=0.032]). Smooth muscle cell content in cIAP2−/− mice was 3.075±3.3% compared with cIAP2+/+ with 0.085±0.1% (P=0.0071). Conclusions Results uncover a key role for cIAP2 in atherosclerotic lesion development, and targeting it may represent a novel therapeutic strategy.