Differential response of human basophil activation markers: a multi-parameter flow cytometry approach

<p>Abstract</p> <p>Background</p> <p>Basophils are circulating cells involved in hypersensitivity reactions and allergy but many aspects of their activation, including the sensitivity to external triggering factors and the molecular aspects of cell responses, are still to be focused. In this context, polychromatic flow cytometry (PFC) is a proper tool to investigate basophil function, as it allows to distinguish the expression of several membrane markers upon activation in multiple experimental conditions. </p> <p>Methods</p> <p>Cell suspensions were prepared from leukocyte buffy coat of K2-EDTA anticoagulated blood specimens; about 1500-2500 cellular events for each tested sample, gated in the lymphocyte CD45dim area and then electronically purified as HLADRnon expressing/CD123bright, were identified as basophilic cells. Basophil activation with fMLP, anti-IgE and calcium ionophore A23187 was evaluated by studying up-regulation of the indicated membrane markers with a two-laser six-color PFC protocol.</p> <p>Results</p> <p>Following stimulation, CD63, CD13, CD45 and the ectoenzyme CD203c up-regulated their membrane expression, while CD69 did not; CD63 expression occurred immediately (within 60 sec) but only in a minority of basophils, even at optimal agonist doses (in 33% and 14% of basophils, following fMLP and anti-IgE stimulation respectively). CD203c up-regulation occurred in the whole basophil population, even in CD63non expressing cells. Dose-dependence curves revealed CD203c as a more sensitive marker than CD63, in response to fMLP but not in response to anti-IgE and to calcium ionophore.</p> <p>Conclusion</p> <p>Use of polychromatic flow cytometry allowed efficient basophil electronic purification and identification of different behaviors of the major activation markers. The simultaneous use of two markers of activation and careful choice of activator are essential steps for reliable assessment of human basophil functions.</p

US perspective on gluten-related diseases

Maureen M Leonard,1 Brintha Vasagar1,2 1Center for Celiac Research, Massachusetts General Hospital for Children, Boston, MA, 2Department of Family Medicine, Spartanburg Regional Healthcare System, Spartanburg, SC, USA Abstract: The incidence of allergy and autoimmune disease in the US and other industrialized nations is increasing, and gluten-related disorders are no exception. The US has documented a profound rise in celiac disease that cannot be fully explained by improved serological techniques or better recognition by physicians. Non-celiac gluten sensitivity, a condition only recently recognized by the medical community, has become a commonly diagnosed entity. Proteins, including gluten are increasingly being identified as a source of wheat allergy. Although the gluten free diet represents a safe and effective treatment for these conditions, there is still much to be learned about the development of gluten-related disorders and the apparent increase in incidence within the US. In this article, we present a review of current knowledge on the epidemiology of gluten-related disorders within a global context, with a focus on diagnostic trends and the evaluation of potential risk factors. Keywords: celiac disease, non-celiac gluten sensitivity, wheat allergy, risk factors, review, epidemiolog