Regulation of cytokine receptors and activity markers in eosinophilic inflammatory processes

Abstract

The eosinophil is an important effector cell in allergic inflammation and asthma and an increased number of eosinophils and eosinophil-derived cytotoxic proteins have been found in blood and airways of allergic patients. The life cycle of the eosinophil starts with differentiation of bone marrow progenitor cells. The eosinophils then proliferate and are recruited into the blood stream. They are activated, and migrate into tissues, where they are further activated and degranulate, and then die and are removed. The cytokines IL-5, IL-3 and GM-CSF are of crucial importance for this course, and increased levels have been detected in serum and airways of asthmatic patients.

The main topic for this thesis was to investigate the role and regulation of eosinophil-specific cytokines and cytokine receptors in inflammatory processes involving eosinophils. An additional purpose was to find new relevant risk markers to predict the clinical course of allergic inflammation. We have demonstrated an increased number of peripheral blood eosinophils (PBE) and lymphocytes (PBL) with intracellularly stored IL-5 in allergic asthmatics, but not in healthy subjects, after allergen inhalation provocation. This suggests that IL-5 containing leukocytes provide a readily accessible pool of IL-5. These cells increase in number during allergen challenge and IL-5 positive cells may thus serve as a risk marker for asthma development. In asthmatic children withdrawn from budesonide treatment, we demonstrated that the eosinophil count in blood and levels of ECP and EPO in serum increased and was higher as compared to asthmatics continuously treated with budesonide.

We also showed that the levels of these markers could be used to predict the development of symptoms and need for corticosteroid treatment. In addition, budesonidetreatment induced a downregulation of the IL-5 receptor alpha (IL-5R) but not GM-CSF receptor alpha (GM-CSFR), concomitant with a reduced in vitro responsiveness towards IL-5 but not to GM-CSF, when compared to asthmatics not on steroid treatment and healthy controls. This budesonide-related downregulation of the IL-5R might be a mechanism by which steroid treatment inhibits the action of IL-5 on eosinophil accumulation and activation in vivo. Tissue eosinophils recovered from peritoneal dialysis fluid (PF) from Continuous Ambulatory Peritoneal Dialysis (CAPD)-patients with peritoneal fluid eosinophilia (PFE) were activated as shown by the expression of the EG2-epitope on intracellular ECP, CD9, CD11b and CD69. Moreover, the proportion of IL-5R positive eosinophils in PF was downregulated as compared to eosinophils in peripheral blood.

In PF from a PFE patient we found ECP, IL-5 and eotaxin whereas only eotaxin was detected in PF from a patient with bacterial peritonitis and neutrophilia. The concomitant finding of activated eosinophils, ECP, IL-5 and eotaxin indicate that the recruitment of eosinophils observed in PFE involves mechanisms similar to those involved in allergic inflammation. When purified PBE from healthy individuals were incubated with PF from a patient with PFE, the cells were activated as indicated by CD69-upregulation, and an intense downregulation of the IL-5R was obtained. This downregulation was almost abolished when IL-5, but not when GM-CSF, was neutralised in the fluid. Moreover, incubation of PBE with recombinant IL-5 downregulated the IL-5R extensively while incubation with GM-CSF only markedly downregulated the IL-5R. The results suggest that activated tissue eosinophils have a downregulated IL-5R as compared to PBE and that IL5, counter to its proposed action on eosinophil progenitors, downregulates the IL-5R on mature eosinophils.

In conclusion, the number of IL-5 positive eosinophils, eosinophil count, ECP and EPO levels in blood are useful markers to follow the clinical course of allergic inflammation. The differentiation stage and localisation of the eosinophil, as well as the different actions of IL-5 on IL-5R expression, have to be taken into account when evaluating the biological consequences of IL-5 in eosinophilic inflammatory responses.