Eosinophils are the major cellular effectors of allergic inflammation and represent an important therapeutic target. Although the genesis and activation of eosinophils have been extensively explored, little is known about their intravascular kinetics or physiological fate. This study was designed to determine the intravascular life span of eosinophils, their partitioning between circulating and marginated pools, and sites of disposal in healthy persons. Using autologous, minimally manipulated 111-Indium-labeled leukocytes with blood sampling, we measured the eosinophil intravascular residence time as 25.2 hours (compared with 10.3 hours for neutrophils) and demonstrated a substantial marginated eosinophil pool. ␥ camera imaging studies using purified eosinophils demonstrated initial retention in the lungs, with early redistribution to the liver and spleen, and evidence of recirculation from a hepatic pool. This work provides the first in vivo measurements of eosinophil kinetics in healthy volunteers and shows that 111-Indium-labeled eosinophils can be used to monitor the fate of eosinophils noninvasively. (Blood. 2012;120(19):4068-4071) IntroductionEosinophils play a key role in allergic inflammation 1 and represent an important therapeutic target in asthma and other allergic diseases. They have the capacity to release histotoxic substances, including granule proteins, inflammatory cytokines, and reactive oxygen metabolites, which cause bronchoconstriction, epithelial damage, hyper-responsiveness, and airway remodeling. [2][3][4][5][6] Much is known about the cellular mechanisms regulating the development and maturation of eosinophils, their release from the bone marrow, and the processes involved in their recruitment, activation, and clearance during allergic inflammation. 7-11 By contrast, very little is known about the physiology of circulating eosinophils in humans. Because of the relative scarcity of eosinophils in the blood of healthy persons (range, 0.0-0.4 ϫ 10 9 /L), previous attempts to study eosinophil kinetics have been restricted to patients with hypereosinophilia, 12-14 hampered by label reuse after pulse injection of 3 H-thymidine, 15 or relied on autoradiographs developed Ͼ 500 days. 16 We have used 111-Indiumlabeled mixed leukocytes with postinjection isolation of eosinophils to ascertain their intravascular life span, and subsequently purified 111-Indium-labeled autologous eosinophils with ␥ camera imaging to assess organ-specific trafficking in vivo. We have demonstrated an intravascular lifespan for circulating eosinophils exceeding 24 hours and revealed extensive intravascular margination of these cells, together with evidence of recirculation from a hepatic pool. Methods ParticipantsHealthy male and female adults with normal lung function and eosinophil counts (range, 0.02-0.38 ϫ 10 9 /L) gave written informed consent in accordance with the Declaration of Helsinki. The study was approved by Cambridgeshire Research Ethics Committee (09/H0308/119) and the Administration of Radioactive Substan...

We have demonstrated for the first time in humans that IL-6R blockade affects neutrophil trafficking to the bone marrow without influencing neutrophil functional capacity.

Eosinophils are mediators of allergic inflammation and are implicated in the pathogenesis of numerous conditions including asthma, parasitic infections, neoplasms, hyper-eosinophilic syndromes, vasculitic disorders, and organ-specific conditions. Assessing eosinophilic inflammation is therefore important in establishing a diagnosis, in monitoring and assessing response to treatment, and in testing novel therapeutics. Clinical markers of atopy and eosinophilic inflammation include indirect tests such as lung function, exhaled breath condensate analysis, fractional exhaled nitric oxide, serum immunoglobulin E levels and serum periostin. Direct measures, which quantify but do not anatomically localise inflammation include blood eosinophil counts, serum or plasma eosinophil cationic protein and sputum eosinophil levels. Cytology from bronchoalveolar lavage and histology from endobronchial and transbronchial biopsies are better at localising inflammation but are more invasive. Novel approaches using radiolabelled eosinophils with single-photon emission computed tomography, offer the prospect of non-invasive methods to localise eosinophilic inflammation.

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However, the additional flush after EVLP is expected to clear the medication from the vasculature. Furthermore, the accumulation of the drug in the extravascular space has been shown to be minimal and would have been rapidly dissipated once in vivo status had been reestablished (10). In our case there were no signs of fibrinolytic complication such as bleeding.With the success obtained in the Human Ex vivo Lung Perfusion trial and evidence from research studies demonstrating stable pulmonary function over prolonged normothermic perfusions over 12 hours, we believe that advanced lung preservation techniques should involve EVLP as a platform for accurate diagnosis and directed therapy (16). As opposed to our ongoing preclinical research using advanced molecular therapeutics such as adenoviral IL-10 gene therapy (17), in this case we used readily available clinical diagnostic techniques and a clinically proven, readily available medication to treat donor lungs in an almost guideline-based fashion, but in the ex vivo setting. Thus, this case exemplifies how the concept of personalized medicine for the donor organ can be applied in the field of lung transplantation.

The detection of focal eosinophilic inflammation by non-invasive means may aid the diagnosis and follow-up of a variety of pulmonary pathologies. All current methods of detection involve invasive sampling, which may be contraindicated or too high-risk to be performed safely. The use of injected autologous technetium-99m (Tc-99m)-labelled eosinophils coupled to single-photon emission computed tomography (SPECT) has been demonstrated to localise eosinophilic inflammation in the lungs of a patient with antineutrophil cytoplasmic antibody-positive vasculitis. Here, we report on the utility of this technique to detect active eosinophilic inflammation in a patient with focal lung inflammation where a biopsy was contraindicated.

Radiolabelled leukocyte scans are widely used in nuclear medicine to locate sites of infection and inflammation. Radiolabelling of leukocyte subpopulations can also yield valuable information on cell trafficking and kinetics in vivo, but care must be taken to minimize inadvertent cell activation ex vivo. Here, we describe the use of autologous indium(111)-labelled eosinophils to measure eosinophil intravascular life-span and monitor their distribution and fate using gamma camera imaging in healthy non-atopic individuals.

Neutrophils play an important role in the lung tumour microenvironment. We hypothesised that radiolabelled neutrophils coupled to single-photon emission CT (SPECT) may non-invasively quantify neutrophil uptake in tumours from patients with non-small cell lung cancer. We demonstrated increased uptake of radiolabelled neutrophils from the blood into tumours compared with non-specific uptake using radiolabelled transferrin. Moreover, indium-111-neutrophil activity in the tumour biopsies also correlated with myeloperoxidase (MPO)-positive neutrophils. Our data support the utility of imaging with In-111-labelled neutrophils and SPECT-CT to quantify neutrophil uptake in lung cancer.