Myocardial Infarction Does Not Accelerate Atherosclerosis in a Mouse Model of Type 1 Diabetes
Total Page：16
File Type：pdf, Size：1020Kb
Diabetes Volume 69, October 2020 2133 Myocardial Infarction Does Not Accelerate Atherosclerosis in a Mouse Model of Type 1 Diabetes Farah Kramer,1 Amy M. Martinson,2 Thalia Papayannopoulou,3 and Jenny E. Kanter1 Diabetes 2020;69:2133–2143 | https://doi.org/10.2337/db20-0152 In addition to increasing the risk of an initial myocardial observed in response to the acute ischemic event was pos- infarction (MI), diabetes increases the risk of a recur- tulated to be driven by enhanced extramedullary hemato- rent MI. Previous work suggests that an experimental poiesis resulting in increased levels of circulating monocytes MI can accelerate atherosclerosis via monocytosis. To available for recruitment into the nascent atherosclerotic test whether diabetes and experimental MI synergize to lesion, thereby accelerating atherosclerosis (9). accelerate atherosclerosis, we performed ligation of Diabetes accelerates atherosclerosis lesion initiation and the left anterior descending coronary artery to induce progression and hinders lesion regression in response to experimental MI or sham surgery in nondiabetic and dramatic lipid lowering (10–13). Changes in monocyte and diabetic mice with preexisting atherosclerosis. All mice macrophage phenotype are believed to contribute to the COMPLICATIONS fi subjected to experimental MI had signi cantly reduced acceleration of atherosclerosis in diabetes. In both mouse left ventricular function. In our model, in comparisons and human studies, diabetes results in increased macro- with nondiabetic sham mice, neither diabetes nor MI phage accumulation within the artery wall (10,14,15). For resulted in monocytosis. Neither diabetes nor MI led to example, autopsy and atherectomy samples from humans increased atherosclerotic lesion size, but diabetes ac- celerated lesion progression, exemplified by necrotic have shown that lesions from subjects with diabetes have core expansion. The necrotic core expansion was de- increased macrophage accumulation compared with speci- pendent on monocyte recruitment, as mice with mye- mens from patients without diabetes (16). Studies of mouse loid cells deficient in the adhesion molecule integrin a4 models of diabetes-accelerated atherosclerosis have revealed were protected from necrotic core expansion. In sum- that arterial accumulation of macrophages is accelerated by mary, diabetes, but not MI, accelerates lesion progres- diabetes (11,17). What is driving the increase in macrophage sion, suggesting that the increased risk of recurrent MI accumulation is not fully understood. Increased monocyte in diabetes is due to a higher lesional burden and/or recruitment appears to be involved, at least in models of elevated risk factors rather than the acceleration of the lesion regression (18). Consistent with the notion that underlying pathology from a previous MI. recruitment is increased, diabetes is often associated with increased endothelial adhesion molecule expression (19,20), which in part could be explained by reduced insulin signal- Diabetes increases the risk of atherosclerotic cardiovascu- ing in endothelial cells (21). In addition to changes in the lar disease (1–4). In addition to increasing the risk of an endothelium, monocytes isolated from humans with di- initial myocardial infarction (MI), diabetes also increases abetes adhere better to aortic endothelial cells (22). the risk of a recurrent MI (5–8). The mechanisms whereby Monocyte recruitment involves initial rolling, adhesion, this happens are unknown. and, finally, transmigration, all of which are highly regulated In experimental model systems, an acute experimental steps. One of the critical interactions is between endothelial MI under nondiabetes conditions can accelerate the un- VCAM-1, which is upregulated in diabetes (19,20), and derlying atherosclerotic process (9). Accelerated atherosclerosis monocyte very-late antigen 4 (VLA4). VLA4 consists of 1Division of Metabolism, Endocrinology and Nutrition, Department of Medicine, Received 12 February 2020 and accepted 17 July 2020 University of Washington Medicine Diabetes Institute, University of Washington This article contains supplementary material online at https://doi.org/10.2337/ School of Medicine, Seattle, WA figshare.12670325. 2Department of Laboratory Medicine and Pathology, University of Washington © 2020 by the American Diabetes Association. Readers may use this article as School of Medicine, Seattle, WA long as the work is properly cited, the use is educational and not for profit, and the 3Division of Hematology, Department of Medicine, University of Washington School work is not altered. More information is available at https://www.diabetesjournals of Medicine, Seattle, WA .org/content/license. Corresponding author: Jenny E. Kanter, [email protected] 2134 MI, Diabetes, and Atherosclerosis Diabetes Volume 69, October 2020 integrin a4 (also known as CD49D) and integrin b1. Block- to expose the heart, and the left coronary artery was ing VLA4 using monoclonal antibodies and blocking pep- permanently ligated with an 8-0 suture. The chest was tides reduces monocyte and macrophage recruitment to the then closed in layers, and the animal was allowed to recover. artery wall in atherosclerosis-prone mice (23,24), and de- The mice in the sham groups were subjected to the same letion of integrin a4 results in a reduced recruitment of procedure of bar ligation of the artery. The survival rates monocytes to an inflamed peritoneum (25,26). were ;80% in both the diabetic and nondiabetic groups, The current study was designed to investigate whether with no difference in survival within the groups subjected to diabetes and experimental MI synergize to accelerate permanent ligation. All mice received buprenorphine anal- atherosclerosis, potentially explaining the increased risk of gesia and were monitored regularly until end point analysis. recurrent MI in people with diabetes. We show that neither Mice were euthanized 3.5 weeks after the surgery. Four days diabetes nor MI results in monocytosis but that diabetes before euthanasia, the mice were injected retro-orbitally alone and in the presence of MI results in an expansion of with 250 mL yellow-green (YG) latex microparticles (di- the necrotic core, a hallmark of advanced, unstable lesion luted 1:4 in sterile PBS) for monocyte tracing (Fluoresbrite phenotypes. Furthermore, the development of the necrotic YG Microspheres 0.5 mm; Polysciences, via Thermo Fisher cores is dependent on integrin a4–mediated monocyte Scientific) (see Supplementary Table 1). For the analysis of recruitment, highlighting the importance of monocytes monocyte recruitment, the number of beads per lesion was and macrophage in diabetes-accelerated atherosclerosis. normalized to the number of labeled monocytes in circu- lation. At the end of the experiments, macrophages were RESEARCH DESIGN AND METHODS isolated from the peritoneal cavity, using ice-cold PBS 1 Mouse Models 5 mmol/L EDTA. The macrophages were allowed to adhere 2 2 The T cell–induced LDL receptor–deficient (Ldlr / ) GpTg for 1 h before they were harvested for either RNA isolation mouse model of type 1 diabetes–accelerated atherosclero- (NucleoSpin RNA Plus; MACHEREY-NAGEL, Bethlehem, sis has previously been described (11). These mice express PA) or measurements of total cholesterol, free cholesterol, the lymphocytic choriomeningitis virus (LCMV) glycopro- and cholesteryl esters (Amplex Red Cholesterol Assay Kit; tein under control of the insulin promoter, allowing reli- Thermo Fisher Scientific, Grand Island, NY). Additional 1 able induction of diabetes due to CD8 T cell–mediated control groups included mice that were not bone marrow b-cell destruction following LCMV injection. In the current transplanted and had not had prior fat feeding, mice that 2 2 study (Fig. 1A), female Ldlr / ;GpTg mice on the C57BL/6J did not express the viral glycoprotein but were injected background, 6–8 weeks of age, mice were fed a high-fat, with LCMV, and a comparison cohort of male and female semipurified diet (40% calories from fat, 1.25% choles- nondiabetic and diabetic mice that had been fat fed but terol) to initiate preexisting lesions for 12 weeks (11,13) not bone marrow transplanted. and then switched to standard mouse chow to reduce plasma cholesterol to allow for successful bone marrow Echocardiogram reconstitution. The mice were lethally irradiated (10 Gy) and One day prior to euthanasia, mice were subjected to 1 fl fl given back either Lyz2-Cre /wt;Itga4 / (myeloid-selective integrin echocardiography using a VisualSonics Vevo 2100. The left 1 a–deficient) (knockout [KO]) or LyzM-Cre /wt;Itga4wt/wt (WT) ventricular function was assessed in M-mode at the level bone marrow and allowed to recover for 5 weeks before where the papillary muscle could be observed during iso- one-half of the cohort was rendered diabetic with LCMV. flurane sedation (mean heart rate 430 bpm). Data were Nondiabetic littermates were injected with saline. At this analyzed by an investigator blinded to the study groups. point, a subset of animals was euthanized, and baseline atherosclerosis was analyzed. At the onset of diabetes Analysis of Glucose and Lipid Metabolism (defined as blood glucose levels .250 mg/dL) 10 days Blood glucose was measured in the saphenous vein blood after LCMV injection, the nondiabetic and diabetic mice by stick tests (OneTouch Ultra). As the glucometer does were fed a low-fat semipurified diet without added choles- not go beyond 600 mg/dL, values above that are set to terol as previously described (11). The low-fat