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IgE to the Mammalian Oligosaccharide Galactose-α-1,3-Galactose Is Associated With Increased Atheroma Volume and Plaques With Unstable Characteristics—Brief Report

Supplemental Digital Content is available in the text.

I nflammation is integral to the pathogenesis of coronary artery disease (CAD), and a role for food-specific antibodies and mast cells has long been a subject of inquiry. [1][2][3] Recent reports have confirmed a role for mast cells and demonstrated an association between levels of total serum IgE and cardiovascular disease, though no specific allergens have been identified. [4][5][6] Galactose-α-1,3-galactose (α-Gal) is a blood group-like oligosaccharide of nonprimate mammals and is recognized in humans as a target of natural IgM, IgA, and IgG antibodies. 7,8 In 2008, IgE specific to α-Gal was identified as the cause of anaphylactic reactions to cetuximab. 9 Subsequently, it became clear that this IgE antibody is also the primary cause of delayed anaphylaxis to red meat (α-Gal syndrome). 10 In the United States, sensitization to α-Gal is recognized as a consequence of bites of the tick Amblyomma americanum, which is in keeping with a prevalence of sensitization to α-Gal as high as 20% in areas of the southeast. 11 Although α-Gal is present in mammalian meat as both glycolipids and glycoproteins, the delay in reactions to red meat is best explained by the time taken for processing of lipids into chylomicrons and smaller lipoprotein particles, such as LDL (low-density lipoprotein) or HDL (high-density lipoprotein). 12,13 Taken together, α-Gal represents a foreign epitope, which is regularly consumed as glycolipids and to which a subset of the population has a strikingly different type of immune response. Thus, it is possible that specific IgE (sIgE) on mast cells, including those in coronary arteries, could increase the inflammatory response to dietary glycolipids. Here, we sought to use intravascular ultrasound imaging in a population where sensitization is prevalent to investigate whether the presence of IgE sensitization to α-Gal is related to the burden of CAD. 14

Methods
Data are available on request from the authors.

Study Cohort
The studied population consisted of subjects between the ages of 30 to 80 years who were undergoing medically warranted cardiac catheterization and were enrolled into ongoing studies at the University of Virginia as described previously. 15 Written consent was obtained from all subjects, and the studies were approved by the Institutional Review Board at the University of Virginia.

Intravascular Ultrasound
Intravascular ultrasound was conducted as previously described and in accordance with the standards of the American College of Cardiology for image acquisition on a noninfarct-related artery. 14,15 Analysis was conducted by an investigator (A.T.N. or A.M.T.) who was blind to IgE assay results. The fraction of atherosclerotic plaque that was fibrous, fibrofatty, necrotic, or calcified (as determined by intravascular ultrasound) was calculated as a percentage.

Statistics
Prism 7.0 (GraphPad, San Diego, CA) was used for graphs and statistics. Categorical variables were compared with Fisher exact test. Continuous data were compared by the Mann-Whitney U test. Spearman rank correlation was used to compare the relationship between α-Gal IgE and total IgE. Regression modeling was performed with IBM SPSS statistics, version 24.0 (IBM Corp, Armonk, NY).

Results
Using a cutoff of ≥0.1 kU/L, 26.3% of the subjects had detectable levels of IgE to α-Gal, and, not unexpectedly, there was higher total IgE in the sensitized group. There was no difference in established risk factors or markers of CAD, such as hypertension, lipid levels, or diabetes mellitus, though the sensitized group had a higher percentage of males and black subjects ( Table I in  adults, we stratified the cohort into 3 age groups with roughly equal numbers of subjects. This stratification revealed a striking difference between subjects of relatively younger versus older age. The significant association of atheroma burden and volume in α-Gal-sensitized compared with nonsensitized subjects was predominantly driven by results in subjects ≤65 years of age (Figure [B]), so additional analysis focused on this group (n=79). The demographic characteristics and risk factors in these subjects (≤65 years) were similar to the cohort as a whole ( Table I in the online-only Data Supplement), and atherosclerotic plaques had significantly more fibrofatty, necrotic, and calcified content but were less fibrous in α-Galsensitized than nonsensitized subjects (Figure [C]). IgE to a panel of inhalant allergens (including dust mite, oak, timothy grass, and ragweed) and IgE to peanut, in contrast to IgE to α-Gal, did not have a significant relationship with atheroma burden, atheroma volume, or maximal stenosis (Table 1). To model the association of CAD with α-Gal sIgE status, we conducted linear regression of atheroma burden as a continuous dependent variable with the independent variable of presence or absence of α-Gal sIgE. Although there was a significant association between α-Gal sIgE and total IgE ( Figure I in the online-only Data Supplement), in regression modeling, the strength of the relationship with atheroma burden was stronger for IgE to α-Gal than for high levels of total IgE (using 100 kU/L as a cutoff; Table 2). Moreover, total IgE >100 kU/L did not have a significant association with atheroma volume or maximal stenosis (Table 1). Collectively, these results support the specificity of the relationship between α-Gal sIgE and CAD. Finally, the relationship between α-Gal IgE status and atheroma burden was preserved in regression analysis adjusted for sex, diabetes mellitus, hypertension, statin use, and total IgE (regression coefficient, 12.2; SE, 5.2; P=0.02; Table 2).

Discussion
To our knowledge, this is the first report that has described an association between the IgE response to α-Gal, or any specific allergen, and the burden of CAD. A link between a food allergen and CAD may not seem a priori obvious, but there are several elements of the immune response to α-Gal that argue for biological plausibility. Because many, or perhaps most, subjects with IgE to α-Gal do not have allergic symptoms, an implication is that many people, despite mounting a unique and potentially detrimental immune response to α-Gal, continue to consume foods that contain the oligosaccharide allergen. 16 In regard to the cause-effect relationship, we are not aware of any evidence that dietary exposure to mammalian products can promote IgE to α-Gal, and it is similarly unlikely that atherosclerosis itself could contribute to this response. 11 The study design limits mechanistic insight, but we hypothesize that chronic ingestion of α-Gal-glycosylated mammalian products, particularly the glycolipid form of the allergen, can lead to recurrent release of inflammatory products from mast cells bearing sIgE. 4 A role for other cells, which express members of the Fcε or Fcγ family of receptors, such as macrophages or basophils, cannot be discounted, especially given that IgG 1 often increases in parallel with IgE. 17 The induction of sIgE (and IgG 1 ) to the same antigen that is the target of abundant natural antibodies, as occurs with α-Gal, is also interesting because it is in keeping with prior reports that natural antibodies to oxidation-specific epitopes are protective in atherosclerosis, whereas class-switched antibodies can be proatherogenic. 18 Relatedly, an inverse association between levels of α-Gal-specific IgM and IgG 2 antibodies and CAD has been reported previously. 19 There are several limitations to consider. We do not know the allergic histories or dietary habits of the participating subjects. Our assumption, based on clinical experience and other cohorts, is that most subjects in this study did not have a history of symptomatic α-Gal syndrome (ie, urticaria or anaphylaxis) and consequently are regularly consuming mammalian products. 16 The known association of total IgE with atherosclerosis is a potential confounder; however, in this cohort, the association with CAD is stronger for α-Gal sIgE than total IgE. Moreover, in areas where delayed anaphylaxis to red meat is common, it is likely that elevated total IgE levels are often directly attributable to the same environmental trigger that generates IgE to α-Gal (ie, tick bites). 11,20 The specificity of the relationship with α-Gal sIgE is also supported by the finding that sIgE antibodies to a panel of inhalants and peanut did not have a significant relationship with CAD. Finally, our current cohort is relatively small, and the effect was predominantly in subjects ≤65 years of age.
In summary, we report here that a specific antibody response to the dietary antigen α-Gal is associated with an increased burden of atherosclerosis and plaques with less stable characteristics. As the causal agent of a delayed allergic reaction to mammalian meat, which is IgE mediated, the glycolipid form of α-Gal may be particularly relevant to understanding its role in atherosclerosis. Mechanistic studies in animal models are suggested, but ultimately, efforts to translate this finding into clinical relevance will benefit from analysis of larger cohorts, including disparate geographic areas and prospective studies of adult subjects, which include information on diet, allergic history, and detailed investigation into sIgE antibodies. Regression modeling is reported where α-Gal sIgE status, sex, diabetes mellitus, hypertension, statin use, and total IgE >100 kU/L are categorical variables. α-Gal indicates galactose-α-1,3-galactose; and sIgE, specific IgE.