Prina Ruparelia

Is the neutrophil the key effector cell in severe asthma

The importance of the neutrophil as the dominant inflammatory cell in many of the non-atopic and more severe phenotypes of asthma is now clear Eosinophilic inflammation has long been considered one of the most distinctive pathological hallmarks of asthma1 and features in many contemporary definitions of this disease. A plethora of studies published from the mid 1990s onwards have suggested, however, that airway eosinophilia is not a universal finding. This has fuelled debate that discrete pathological phenotypes of asthma may exist, with the neutrophil—rather than the eosinophil—dominating in certain circumstances.2–4 We present data that support the current renewed interest in the neutrophil as a primary driver of airways inflammation, particularly in the most severe forms of asthma. There are also some intriguing data to suggest that, when the eosinophil has been “red carded” and disappears from the inflamed airway, the neutrophil may be drawn in and act as the substitute granulocyte. The hypothesis that the eosinophil is the key effector cell involved in the pathogenesis of asthma has run into trouble for several reasons: (1) eosinophilic inflammation is present in the airway lumen of only 50% of asthmatic subjects;4 (2) even intense eosinophilic inflammation, as occurs in eosinophilic bronchitis, fails to induce asthma;5 (3) many asthma exacerbations occur …

Use of 111-Indium-labelled autologous eosinophils to establish the in vivo kinetics of human eosinophils in healthy subjects

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. γ 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.

Measuring whole-body neutrophil redistribution using a dedicated whole-body counter and ultra-low doses of 111Indium

Eur J Clin Invest 2010; 41 (1): 77–83

Pulmonary elimination rate of inhaled 99mTc-sestamibi radioaerosol is delayed in healthy cigarette smokers

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT * Very little is known about the physiology of P-glycoprotein (P-gp) expression in the lungs. * Ex vivo evidence based on resected lung tissue suggests that pulmonary P-gp is upregulated by cigarette smoke, but there are no in vivo studies to date. WHAT THIS STUDY ADDS * The novel observation that healthy cigarette smokers have a delayed pulmonary elimination rate of inhaled (99m)Tc-sestamibi, a P-gp substrate, provides for the first time a potential method for quantifying functional pulmonary P-gp expression that may inform about drug therapy by inhalation as well as provide a non-invasive, quantitative, human biomarker for assessing P-gp modulators. AIM To explore inhaled technetium-99m-labelled hexakis-methoxy-isobutyl isonitrile ((99m)Tc-sestamibi) for quantifying pulmonary P-glycoprotein (P-gp) expression. METHODS The elimination rate from the lungs of (99m)Tc-sestamibi was recorded scintigraphically for 30 min following inhalation as an aerosol in healthy smokers, nonsmokers and patients with lung disease. RESULTS (99m)Tc-sestamibi elimination rates [% min(-1) (SD; P vs. healthy nonsmokers)] were: healthy nonsmokers, 0.43 (0.083); healthy smokers, 0.19 (0.056; P < 0.001); chronic obstructive pulmonary disease patients, 0.26 (0.077; P < 0.001). Elimination rates in three patients with interstitial lung disease were not accelerated. CONCLUSION Cigarette smoke upregulates lung P-gp. (99m)Tc-sestamibi elimination in normal smokers could be used to test new P-gp modulators. The findings also have implications for inhaled drug delivery.

Use of Radiolabelled Leukocytes for Drug Evaluation in Man

The endeavour to radiolabel or simply “label” autologous leukocytes has been a major clinical need. The endeavour to improve the labelling conditions and minimise interventional stresses and maintain cell functionality has been the driving objective. This chapter focuses on novel techniques used in this laboratory to radiolabel leukocytes, examples of the clinical indications that such labelled products might be informative, and how we can use these labelled cells in clinical situations to describe the life cycle behaviour (transit times and migratory capacity) of these labelled cells. Labelling techniques which preserve leukocyte functionality will assist in the development of new anti-inflammatory agents, anti-infectives, and indeed any drug or biologic where their clinical use may have an effect on these cells. Examples of labelling methods and clinical scenarios with imaging are described for conditions such as abscesses, ARDS, COPD, rheumatoid arthritis, inflammatory bowel disease, and vasculitis.

Does P-glycoprotein have a role in the lung clearances of inhaled 99mTc-sestamibi and 99mTc-tetrofosmin?

ObjectiveThe clearance rate of inhaled 99mTc-sestamibi from the lungs of healthy nonsmoking individuals is much slower than would be expected from its physical properties. The clearance rate is even slower in healthy cigarette smokers. As 99mTc-sestamibi is a substrate for P-glycoprotein (P-gp), pulmonary P-gp may be influential in 99mTc-sestamibi clearance and may be upregulated in smokers. 99mTc-tetrofosmin is also a substrate for P-gp, therefore we hypothesized that it would display similar kinetics to 99mTc-sestamibi and support a role for P-gp. We also hypothesized that administration of P-gp modulators would accelerate clearance of 99mTc-sestamibi. MethodsWe measured clearance rates of 99mTc-tetrofosmin in four healthy smokers and four healthy nonsmokers and of 99mTc-sestamibi in six otherwise healthy patients with psoriasis before and after 2 weeks of therapy with cyclosporine A (2.5–5 mg/kg/day) and two healthy women taking the oral contraceptive pill, as both cyclosporine and steroids are known to be P-gp modulators. ResultsThe clearance rate of 99mTc-tetrofosmin in nonsmokers ranged from 0.38 to 0.63%/min, similar to the previously recorded rate for 99mTc-sestamibi [0.43 (SD 0.083)%/min], but it was not delayed in smokers (range 0.42–0.97%/min). Cyclosporine had no significant effect on 99mTc-sestamibi clearance, although clearance rates in the two women taking the oral contraceptive pill were both fast (0.58 and 0.62%/min). ConclusionAlthough the role of P-gp expression in the clearance of 99mTc-sestamibi remains unproven, we conclude that 99mTc-tetrofosmin is not as P-gp-avid as 99mTc-sestamibi. A role for P-gp expression in the clearance of 99mTc-sestamibi remains unproven. Higher doses of P-gp inhibitors will be required and clearance rates correlated with immunohistochemical expression of P-gp.

99mTechnetium-labelled neutrophil scanning in pneumonia.

A 70-year-old man with severe smoking-related chronic obstructive pulmonary disease (COPD) was recruited to a research study to examine the trafficking of 99m-technetium (99mTc)-labelled neutrophils. These studies were undertaken in patients with stable moderate to severe COPD and designed to establish novel methodology (using autologous 99mTc-labelled neutrophils and single photon emission computed tomography (SPECT)) to quantify neutrophil trafficking and accumulation within the lungs of patients with COPD. At the time of scanning he was clinically stable with a C-reactive …

P86 Optimising patient flow and use of resources in the two week wait pathway

Introduction Lung cancer is the most common cause of cancer death in the UK. Survival is improving but is worse than in some European countries and North America.¹ NICE guidelines recommend that patients with suspected malignancy are seen within two weeks of referral. An earlier local audit found that 22% of patients referred on the pathway had lung cancer and identified a need to streamline the service. Referrals are triaged as high, intermediate and low risk by a respiratory Consultant based on chest X-ray and clinical details. High-risk patients are prioritised for CT imaging and lung clinical nurse specialist (CNS) time. All patients remain on the two week pathway regardless of triage status. This study reviewed whether this triage system is accurate in identifying patients with malignancy, thereby improving resource utilisation. Method Data was collected retrospectively on two week wait referrals during June and July 2015, using referral forms and electronic medical records. Data included key dates in the pathway, triage status and diagnosis. Results 25 of 60 patients were triaged as high risk. Thirteen of these patients had cancer. Two patients with lung cancer were not triaged as high risk. Triaging a patient as high risk had a sensitivity for lung malignancy of 86.7% and specificity of 71.4%. Positive predictive value was 0.52. Nineteen patients triaged as high risk had a CT prior to clinician review and the remaining three had a CT within 3 days. This was three times higher than in the low/intermediate group. The CNS attended the majority of initial clinic appointments in prioritised patients, unless CT showed benign disease. Conclusion The triage method correctly identified patients with malignancy in the majority of cases. This led to efficient use of resources. Patients with lung cancer had earlier imaging and access to the CNS. Lung cancer symptoms can overlap with other respiratory conditions and following the initial clinical review, the respiratory clinician may decide to investigate some patients less urgently. In future, this method could help stratify urgency of referral. Abstract P86 Figure 1 Number of patients triages high/intermediate/low with benign or malignant disease Reference Global surveillance of cancer survival 1995–2009: (CONCORD-2). Lancet 2015;385:977–1010.

A16 Using whole-body counting to determine neutrophil trafficking and loss in COPD

COPD is an inflammatory condition of the airways characterized by airway neutrophilia. Using a whole-body-counter (WBC) we have assessed neutrophil trafficking and loss in COPD using small activities of In-labelled neutrophils. Patients were scanned in the WBC 45 min, 1, 2, 4, 7 and 10 days after injection of autologous In-labelled ‘pure’ neutrophils and whole-body In retention recorded. Using a single-slit collimator, biodistribution was also recorded from the posterior projection as a count profile from head to toe. SPECTon another occasion with Tc neutrophils was used to validate bio-distribution. Mean retention at 10 days was: controls 89.3% (SEM 0.78) (n = 12), COPD 88.9% (1.00) (n = 10) and bronchiectasis 85% (2.69) (n = 10). Profile data showed that the neutrophils initially localized to the liver/spleen region in all patients and thereafter redistributed to the pelvic and thoracic areas. SPECT (n = 3) demonstrated that 19.6% of the thoracic uptake was in the lungs with the remaining signal in the rib and vertebral bone marrow. Whilst it is known that In-labelled neutrophils migrate to the lungs in COPD, mobilization into the airways and subsequent loss from the body is insufficient to quantify by WBC. This technique does provide accurate monitoring of neutrophil trafficking to the liver/spleen and BM compartments, however.