P38 mitogen‐activated protein kinase signal transduction in the diagnosis and follow up of immunotherapy of wasp venom allergy

Abstract

Background: P38 mitogen-activated protein kinase (MAPK) is known to govern IgE-mediated basophil activation. Intracellular phosphorylated p38 MAPK (Pp38 MAPK) in IgE-activated basophils can be quantified flow cytometrically.Objectives: To study whether Pp38 MAPK constitutes a potential novel read-out for flow-assisted diagnosis of hymenoptera venom allergy and to investigate whether this marker allows follow-up of successful venom immunotherapy (VIT).Methods: Fifty-two patients with documented wasp venom allergy and seven wasp-stung asymptomatic control individuals were enrolled. Wasp venom-induced basophil activation was analyzed flow cytometrically with anti-IgE, anti-CD63, and anti-Pp38 MAPK to assess their activation status before starting immunotherapy. To assess whether p38 MAPK constitutes a candidate marker for monitoring VIT, we repeated the basophil activation test (BAT) in 25 patients on the fifth day of a build-up immunotherapy. In addition, we investigated whether the Pp38 MAPK-based BAT could contribute in the decision of discontinuing VIT in a cross-sectional analysis in 13 patients receiving treatment for 3 years and 14 patients receiving treatment for 5 years.Results: Patients exhibited a dose-dependent basophil activation with phosphorylation of p38 MAPK and upregulation of downstream CD63. In contrast, stung controls demonstrated a dose-dependent but ''abrogated'' signal transduction in basophils with less and shorter duration of the phosphorylation of p38 MAPK and without subsequent upregulation of CD63. When repeated after 5 days of VIT and when investigated cross-sectionally after 3 years or 5 years of maintenance therapy, no effect of VIT on the phosphorylation of p38 MAPK was demonstrable.Conclusions: This study discloses that not only basophils from patients, but also from the stung control individuals, respond to wasp venom stimulation with phosphorylation of p38 MAPK, although to a lesser extend. No clear effect of VIT on the phosphorylation of p38 MAPK was shown. Thus, although p38 MAPK provides an additional tool in the diagnosis of wasp venom allergy, it does not contribute to the decision whether to stop successful VIT.

Basophilic histamine content and release during venom immunotherapy: Insights by flow cytometry

“…This parallels the recent observations of Maintz et al(15), who reported normal mRNA levels of L‐histidine decarboxylase (the enzyme responsible for histamine synthesis) at the end of early VIT. Finally, our experiments confirm that build‐up VIT does not affect the in vitro responsiveness of basophils (20, 23–25) or the median release of histamine per cell. Thus, the effect of early VIT is likely to be related, at least partially, to a reduction of circulating basophils.…”

STAT5 in human basophils: IL‐3 is required for its FcεRI‐mediated phosphorylation

“…Recently, it has been demonstrated that flow cytometry can generate qualitative results concerning dose responses and kinetics of signaling molecules similar and complementary to the data obtained with blotting techniques (19,(26)(27)(28). Flow cytometry does not require a purification of the basophils.…”

Basophil Activation Test in Allergy: Time for an Update?

“…In drug allergy, by combining BAT with in vitro passive sensitization of donor basophils with patients’ serum, one can prove the IgE dependence of a drug reaction [13]. In allergy immunotherapy, BAT furthermore represents a powerful tool to follow up patients with allergy manifestations [51,102,107,127]. Improvements in BAT methodology to achieve the best performance have also been made [107], but follow-up of allergy still represents a major concern, for which BAT seems to be a promising approach.…”

“…In this context, phosphorylation of p38-MAP kinase (p38MAPK) represents a crucial step in IgE receptor signaling in basophils. The relationship between p38MAPK and the well-validated diagnostic cell surface marker CD63 has recently been evaluated in a clinical allergy model [106], although criticism of the use of intracellular p38MAPK in monitoring therapy has recently been raised [107]. …”