Laboratorio de Bioquímica-Genética, Instituto Nacional de Pediatría, Insurgentes Sur 3700-C, Col. Insurgentes Cuicuilco, Alcaldía Coyoacán, Ciudad de México, CP 04530, México.
Red de Apoyo a la Investigación, Coordinación de la Investigación Científica, Universidad Nacional Autónoma de México-Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán, Ciudad de México 14080, México.
J Proteomics. 2023 Feb 20;273:104809. doi: 10.1016/j.jprot.2022.104809. Epub 2022 Dec 29.
Immunological mechanisms of non-IgE-mediated cow's milk protein allergy (CMPA) are not well understood. Such a circumstance requires attention with the aim of discovering new biomarkers that could lead to better diagnostic assays for early treatment. Here, we sought both to investigate the mechanism that underlies non-IgE-mediated CMPA and to identify cow's milk immunoreactive proteins in a Mexican pediatric patient group (n = 34). Hence, we determined the IgE and IgG subclass antibody levels against cow's milk proteins (CMP) by ELISA. Then, we performed 2D-Immunoblots using as first antibody immunoglobulins in the patients'serum that bound specifically against CMP together with CMP enrichment by ion-exchange chromatography. Immunoreactive proteins were identified by mass spectrometry-based proteomics. The serological test confirmed absence of specific IgE in the CMPA patients but showed significant increase in antigen-specific IgG. Additionally, we identified 11 proteins that specifically bound to IgG. We conclude that the detection of specific IgG together with an immunoproteomics approach is highly relevant to the understanding of CMPA's physiopathology and as a possible aid in making a prognosis since current evidence indicates IgG occurrence as an early signal of potential risk toward development of IgE-mediated food allergy. SIGNIFICANCE: Allergies are one of the most studied topics in the field of public health and novel protein allergens are found each year. Discovery of new principal and regional allergens has remarkable repercussions in precise molecular diagnostics, prognostics, and more specific immunotherapies. In this context, specific IgE is widely known to mediate physiopathology; however, allergies whose mechanism does not involve this immunoglobulin are poorly understood although their incidence has increased. Therefore, accurate diagnosis and adequate treatment are delayed with significant consequences on the health of pediatric patients. The study of type and subtypes of immunoglobulins associated with the immunoreactivity of cow's milk proteins together with an immunoproteomics approach allows better comprehension of physiopathology, brings the opportunity to discover new potential cow's milk protein allergens and may help in prognosis prediction (IgG occurrence as an early signal of possible risk toward development of IgE-mediated food allergy).
非 IgE 介导的牛奶蛋白过敏 (CMPA) 的免疫机制尚不清楚。这种情况需要引起重视，目的是发现新的生物标志物，从而为早期治疗提供更好的诊断检测方法。在这里，我们既旨在研究非 IgE 介导的 CMPA 的发病机制，又旨在鉴定一个墨西哥儿科患者群体（n = 34）中的牛奶免疫反应性蛋白。因此，我们通过 ELISA 测定了针对牛奶蛋白（CMP）的 IgE 和 IgG 亚类抗体水平。然后，我们使用患者血清中的免疫球蛋白作为第一抗体进行了 2D-免疫印迹，这些免疫球蛋白特异性地与 CMP 结合，并通过离子交换色谱法对 CMP 进行富集。通过基于质谱的蛋白质组学鉴定免疫反应性蛋白。血清学测试证实 CMPA 患者中不存在特异性 IgE，但显示出针对抗原的 IgG 显著增加。此外，我们还鉴定了 11 种特异性结合 IgG 的蛋白。我们得出结论，特异性 IgG 的检测与免疫蛋白质组学方法的结合对于理解 CMPA 的病理生理学非常重要，并且可能有助于预测预后，因为目前的证据表明 IgG 的发生是 IgE 介导的食物过敏潜在风险的早期信号。
过敏是公共卫生领域研究最多的课题之一，每年都会发现新的蛋白过敏原。新的主要和区域过敏原的发现对精确的分子诊断、预后和更具体的免疫疗法具有重要意义。在这种情况下，广泛认为特异性 IgE 介导病理生理学；然而，其机制不涉及这种免疫球蛋白的过敏反应却知之甚少，尽管它们的发病率有所增加。因此，准确的诊断和适当的治疗会被延迟，这对儿科患者的健康有重大影响。研究与牛奶蛋白免疫反应性相关的免疫球蛋白类型和亚型以及免疫蛋白质组学方法可以更好地理解病理生理学，有机会发现新的潜在牛奶蛋白过敏原，并有助于预测预后（IgG 的发生是 IgE 介导的食物过敏发展的可能风险的早期信号）。
