Wang Y, Zhang Y, Dudley C, Fitzgerald K, Carson AS, et al.
The Journal of allergy and clinical immunology •
In alpha-gal syndrome (AGS), IgE antibodies form against the glycan galactose-α-1,3-galactose (alpha-gal) in mammal products rather than food proteins. Alpha-gal glycolipids and glycoproteins activate human basophils sensitized with AGS plasma in an IgE-dependent fashion. However, it is unknown whether alpha-gal specific IgE (sIgE), independent of other blood proteins, is sufficient for mediating basophil and mast cell activation. We sought to determine whether alpha-gal antigens could activate passively sensitized rat basophilic leukemia (RBL) SX-38 cells, which express human IgE receptors and are commonly used to model allergen/IgE-mediated mast cell activation in food protein allergy. Using the clustered regularly interspaced short palindromic repeats technology, we created a novel, alpha-gal-deficient RBL cell line, alpha-gal knockout RBL SX-38, passively sensitizing cells with sera from AGS donors or with novel alpha-gal sIgE clones, and then stimulated with alpha-gal glycoproteins. To assess effector cell activation, we measured cell surface expression of activation marker CD63 by flow cytometry and mediator release through β-hexosaminidase release assays. After alpha-gal antigen stimulation, the percentage of CD63 alpha-gal knockout RBL SX-38 cells sensitized with AGS sera increased 3-fold compared with cells sensitized with control serum. Select human AGS IgE clones facilitated alpha-gal antigen-dependent and antigen-independent CD63 upregulation. Cells sensitized with pooled AGS sera released β-hexosaminidase in an alpha-gal-independent fashion. We saw no β-hexosaminidase release above background in cells sensitized with alpha-gal sIgE clones. Certain alpha-gal-specific human IgE clones may partially activate allergic effector cells independent of antigen, potentially lowering thresholds for subsequent alpha-gal-induced or antigen-independent allergic effector cell degranulation. This may affect duration and severity of allergic symptoms in patients with AGS.
Kepley CL, Wang Y, Yelton A, Siebert ER, Iweala OI
Current allergy and asthma reports •
Alpha-gal syndrome (AGS) is a novel allergic disease characterized by hypersensitivity responses following exposure to the glycan galactose-alpha-1,3-galactose (alpha-gal or α-Gal) attached to mammalian proteins and fats in food, supplements, and medications. Bites from hard-bodied ticks, including Amblyomma americanum (Aa or lone star tick) in the United States, have been identified as drivers of allergic sensitization to alpha-gal. Here, we review clinical presentation, epidemiology, diagnosis, and current understanding of the mechanistic drivers of AGS, with particular focus on the roles of allergic effector cells - mast cells (MCs) and basophils. We explore unique clinical characteristics of AGS through the lens of alpha-gal-specific IgE and MC activation in AGS. We propose potential explanations for delayed symptom onset, inconsistent symptom development, and persistence of allergic symptoms in some AGS patients despite removing all mammal products from the diet. Current evidence implicate bites from hard-bodied ticks as the primary sensitizing agent in AGS. However, there is emerging evidence that other ecto- and endoparasites may also induce alpha-gal-specific IgE. Multiparameter flow and mass cytometry and RNA sequencing have demonstrated an enrichment of unique populations of T, B, invariant natural killer T (iNKT), natural killer B (NKB) and MC progenitor cells in human volunteers with AGS. Recently developed mouse models of AGS will support future studies to identify which cells are critical for the development of AGS. In vitro models of the allergic effector phase of AGS using human sera and novel human alpha-gal-specific IgE monoclonal antibodies in humanized rat allergic effector cell lines, human basophils, human MC lines, and primary human MC cultures, confirm alpha-gal-induced allergic effector cell activation. They also provide a system to study potential alpha-gal-antigen-independent drivers of MC activation in AGS. Efforts are ongoing to understand the epidemiology and immune mechanisms of AGS. Novel reagents (e.g. alpha-gal-specific monoclonal antibodies) and murine AGS models will facilitate deeper investigation of tick-driven, alpha-gal-specific IgE and allergic effector cell-induced pathology in AGS.
Alpha-gal syndrome (AGS) is an IgE-mediated allergy to the oligosaccharide galactose-alpha-1,3-galactose (alpha-gal). Alpha-gal is found in the tissues of non-catarrhine mammals, and the characteristic delayed reactions are caused by the consumption of red meat, visceral organs, dairy, gelatin, and other products, including medications sourced from non-primate mammals. Although the syndrome is not nationally notifiable, it is estimated that 450,000 cases exist, making AGS the tenth-most common food allergy in the US. The syndrome is profoundly influenced by geographic locale, reflecting the important role of tick bites in sensitization. However, the specific immune cells, their interactions, and the downstream signaling cascades triggered by tick bites are not well understood. To address this gap, we conducted a comprehensive study to analyze the immune cells in the blood of AGS subjects compared to those of healthy controls. Peripheral blood mononuclear cell preparations were enriched for B cells from the same patient and sequentially labeled with sample tags and BD AbSeq oligo-conjugated antibodies. Sequencing libraries were prepared to targeted mRNA, antibody-oligonucleotides, and sample tags from AGS and control subjects. Multimodal analysis of both transcriptomes and surface marker expression of immune cells at the single-cell level was used to profile the immune response in AGS. Several clusters of cells were unique to AGS subjects, including natural killer B (NKB) cells, natural killer T (NKT) cells and most notably, a circulating mast cell progenitor population. In addition, subjects with AGS had increased expression of several genes involved in immunomodulation and type 2 immunity. Although rare, we identified and characterized alpha-gal-specific memory B cells and alpha-gal-specific IgE-secreting cells. Our findings also revealed the presence of IgE-secreting transcripts, along with other classes of immunoglobulins (Ig), in a single cell, suggesting a unique pattern of Ig gene arrangements and class switching in B cells. Tick bites appear to induce a population of circulating mast cell progenitors and innate-like cells, such as NKT and NKB cells, that may play a critical role in sensitization to alpha-gal. Furthermore, alpha-gal-specific IgE is secreted by a heterogeneous population of B cells, including CCR6-proficient B cells and CCR6-deficient plasmablast/plasma cells.
Keet C, McGowan EC, Jacobs D, Post WS, Richards NE, et al.
The Journal of allergy and clinical immunology •
In individuals without symptomatic food allergy, food-specific IgE is considered clinically irrelevant. However, recent studies have suggested that galactose-α-1,3-galactose (alpha-gal) IgE is associated with cardiovascular (CV) disease. We sought to determine whether sensitization to common food allergens is associated with CV mortality. The association between IgE sensitization to foods and CV mortality ascertained to 2019 was examined in the National Health and Examination Survey (NHANES) 2005-2006 and the Wake Forest site of the Multi-Ethnic Study of Atherosclerosis (MESA) cohort; MESA enrolled adults without baseline clinical CV diseases between 2000 and 2002. Total and specific IgE was measured to cow's milk, egg, peanut, shrimp, and a panel of aeroallergens (NHANES), and to cow's milk, alpha-gal, peanut, dust mite, and timothy grass (MESA). Cox proportional hazard models were constructed, adjusting for sex, age, race/ethnicity, smoking, education, and asthma. A total of 4414 adults from NHANES (229 CV deaths) and 960 from MESA (56 CV deaths) were included. In NHANES, sensitization to at least 1 food was associated with higher CV mortality (hazard ratio [HR], 1.7 [95% confidence interval (CI), 1.2-2.4], P = .005). Milk sensitization was particularly associated (HR, 2.0 [95% CI, 1.1-3.8], P = .026), a finding replicated in MESA (HR, 3.8 [95% CI, 1.6-9.1], P = .003). Restricting analyses in NHANES to consumers of the relevant allergen strengthened food sensitization relationships, unmasking shrimp and peanut sensitization as additional risk factors for CV mortality. The finding that food sensitization is associated with increased risk of CV mortality challenges the current paradigm that sensitization without overt allergy is benign. Further research is needed to clarify mechanisms of this association.
Alpha-gal syndrome (AGS) is characterized by delayed hypersensitivity to non-primate mammalian meat in people having specific immunoglobulin E (sIgE) to the oligosaccharide galactose-alpha-1,3-galactose. AGS has been linked to tick bites from Amblyomma americanum (Aa) in the U.S. A small animal model of meat allergy is needed to study the mechanism of alpha-gal sensitization, the effector phase leading to delayed allergic responses and potential therapeutics to treat AGS. Eight- to ten-weeks old mice with a targeted inactivation of alpha-1,3-galactosyltransferase (AGKO) were injected intradermally with 50 μg of Aa tick salivary gland extract (TSGE) on days 0, 7, 21, 28, 42, and 49. Total IgE and alpha-gal sIgE were quantitated on Day 56 by enzyme-linked immunosorbent assay. Mice were challenged orally with 400 mg of cooked pork kidney homogenate or pork fat. Reaction severity was assessed by measuring a drop in core body temperature and scoring allergic signs. Compared to control animals, mice treated with TSGE had 190-fold higher total IgE on Day 56 (0.60 ± 0.12 ng/ml vs. 113.2 ± 24.77 ng/ml; p < 0.001). Alpha-gal sIgE was also produced in AGKO mice following TSGE sensitization (undetected vs. 158.4 ± 72.43 pg/ml). Further, sensitized mice displayed moderate clinical allergic signs along with a drop in core body temperature of ≥2°C as an objective measure of a systemic allergic reaction. Interestingly, female mice had higher total IgE responses to TSGE treatment but male mice had larger declines in mean body temperature. TSGE-sensitized AGKO mice generate sIgE to alpha-gal and demonstrate characteristic allergic responses to pork fat and pork kidney. In keeping with the AGS responses documented in humans, mice reacted more rapidly to organ meat than to high fat pork challenge. This mouse model establishes the central role of tick bites in the development of AGS and provides a small animal model to mechanistically study mammalian meat allergy.
Wilson JM, Schuyler AJ, Workman L, Gupta M, James HR, et al.
The journal of allergy and clinical immunology. In practice •
Red meat allergy has historically been understood as a rare disease of atopic children, but the discovery of the "α-Gal syndrome," which relates to IgE to the oligosaccharide galactose-α-1,3-galactose (α-Gal), has challenged that notion. To describe the clinical and immunologic characteristics of a large group of subjects with self-reported allergy to mammalian meat. This was an observational study of 261 children and adults (range, 5-82 years) who presented for evaluation for allergic reactions to mammalian meat. Results were based on serum assays and a detailed questionnaire. α-Gal specific IgE ≥ 0.35 IU/mL was detected in 245 subjects and symptom onset occurred ≥2 hours after eating mammalian meat in 211 (81%). Component testing supported a diagnosis of α-Gal syndrome in 95%, pork-cat syndrome in 1.9%, and primary beef allergy in 1.1%. Urticaria was reported by 93%, anaphylaxis by 60%, and gastrointestinal symptoms by 64%. Levels of IgE and IgG specific to α-Gal were similar in subjects who reported early- or delayed-onset symptoms, and in those with and without anaphylaxis. Levels of α-Gal specific IgE and severity of reactions were similar among those with and without traditional atopy, and among children (n = 35) and adults (n = 226). Blood group B trended toward being under-represented among α-Gal-sensitized subjects; however, α-Gal specific IgE titers were high in symptomatic cases with B-antigen. The α-Gal syndrome is a regionally common form of food allergy that has a characteristic but not universal delay in symptom onset, includes gastrointestinal symptoms, can develop at any time in life, and is equally common in otherwise nonatopic individuals.
The syndrome of delayed allergic reactions to the carbohydrate galactose-alpha-1,3-galactose ("alpha-gal") has become increasingly recognized in allergy and immunology clinics regionally throughout the southeastern USA. Due to the increasing awareness of this unique food allergy, cases have been identified in the northeastern and central USA as well as in Central and South America, Europe, Asia, Scandinavia, and Australia. Clinically, alpha-gal allergy is characterized by reactions to non-primate mammalian meat (e.g., beef, pork, lamb) that occur 3-6 h following exposure. The IgE response to alpha-gal is thought to develop after tick bites and can result in the loss of tolerance to foods that have been safely consumed for years. Although the initial description of alpha-gal allergy in 2009 was limited to red meat, this epitope is now identified in an expanded number of products, medications and foods-both labeled and unlabeled. Moreover, we are beginning to recognize that alpha-gal food allergy is the tip of the iceberg for this immune response.
Food allergy is defined as an IgE-mediated hypersensitivity response to ingested food with allergic symptoms ranging from urticaria to life-threatening anaphylaxis. Food allergy is thought to develop because of (1) failed induction of tolerance upon initial exposure to food antigen or (2) breakdown of established tolerance to food antigen. We review current understanding of the pathogenesis, epidemiology, and natural history of food allergy, including the unconventional IgE-mediated food allergy to mammalian meat known as alpha-gal food allergy. We highlight emerging data on food allergy treatment and prevention, emphasizing the growing appeal of manipulating the gut microenvironment using probiotics and helminth products to blunt systemic allergic responses to food.
