Bronchodilators in Tobacco-Exposed Persons with Symptoms and Preserved Lung Function.

BACKGROUND: Many persons with a history of smoking tobacco have clinically significant respiratory symptoms despite an absence of airflow obstruction as assessed by spirometry. They are often treated with medications for chronic obstructive pulmonary disease (COPD), but supporting evidence for this treatment is lacking. METHODS: We randomly assigned persons who had a tobacco-smoking history of at least 10 pack-years, respiratory symptoms as defined by a COPD Assessment Test score of at least 10 (scores range from 0 to 40, with higher scores indicating worse symptoms), and preserved lung function on spirometry (ratio of forced expiratory volume in 1 second [FEV1] to forced vital capacity [FVC] ≥0.70 and FVC ≥70% of the predicted value after bronchodilator use) to receive either indacaterol (27.5 µg) plus glycopyrrolate (15.6 µg) or placebo twice daily for 12 weeks. The primary outcome was at least a 4-point decrease (i.e., improvement) in the St. George's Respiratory Questionnaire (SGRQ) score (scores range from 0 to 100, with higher scores indicating worse health status) after 12 weeks without treatment failure (defined as an increase in lower respiratory symptoms treated with a long-acting inhaled bronchodilator, glucocorticoid, or antibiotic agent). RESULTS: A total of 535 participants underwent randomization. In the modified intention-to-treat population (471 participants), 128 of 227 participants (56.4%) in the treatment group and 144 of 244 (59.0%) in the placebo group had at least a 4-point decrease in the SGRQ score (difference, -2.6 percentage points; 95% confidence interval [CI], -11.6 to 6.3; adjusted odds ratio, 0.91; 95% CI, 0.60 to 1.37; P = 0.65). The mean change in the percent of predicted FEV1 was 2.48 percentage points (95% CI, 1.49 to 3.47) in the treatment group and -0.09 percentage points (95% CI, -1.06 to 0.89) in the placebo group, and the mean change in the inspiratory capacity was 0.12 liters (95% CI, 0.07 to 0.18) in the treatment group and 0.02 liters (95% CI, -0.03 to 0.08) in the placebo group. Four serious adverse events occurred in the treatment group, and 11 occurred in the placebo group; none were deemed potentially related to the treatment or placebo. CONCLUSIONS: Inhaled dual bronchodilator therapy did not decrease respiratory symptoms in symptomatic, tobacco-exposed persons with preserved lung function as assessed by spirometry. (Funded by the National Heart, Lung, and Blood Institute and others; RETHINC ClinicalTrials.gov number, NCT02867761.).

The SARS-CoV-2 pandemic and asthma: What we have learned and what is still unknown.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has brought new insights into the immunologic intricacies of asthma. In this review, we discuss the epidemiology of asthma in patients infected with SARS-CoV-2 and the risk of severe infection. Type 2 inflammation had an overall protective effect against SARS-CoV-2 infection by various mechanisms summarized in this review. Asthma, intranasal, and inhaled corticosteroids decreased the angiotensin-converting enzyme 2 receptor, an important receptor for SARS-CoV-2 entry into host cells. We summarize the nuances of the treatment of type 2 inflammation despite its underlying protective effects. Research to date has shown that patients on various allergen immunotherapies and biologics do benefit from being vaccinated.

Dupilumab reduced impact of severe exacerbations on lung function in patients with moderate-to-severe type 2 asthma.

BACKGROUND: Severe asthma exacerbations increase the risk of accelerated lung function decline. This analysis examined the effect of dupilumab on forced expiratory volume in 1 s (FEV1 ) in patients with moderate-to-severe asthma and elevated type 2 biomarkers from phase 3 LIBERTY ASTHMA QUEST (NCT02414854). METHODS: Changes from baseline in pre- and post-bronchodilator (BD) FEV1 and 5-item Asthma Control Questionnaire (ACQ-5) scores were assessed in patients with elevated type 2 biomarkers at baseline (type 2-150/25: eosinophils ≥150 cells/µl and/or fractional exhaled nitric oxide [FeNO] ≥25 ppb; type 2-300/25: eosinophils ≥300 cells/µl and/or FeNO ≥25 ppb), stratified as exacerbators (≥1 severe exacerbation during the study) or non-exacerbators. RESULTS: In exacerbators and non-exacerbators, dupilumab increased pre-BD FEV1 by Week 2 vs placebo; differences were maintained to Week 52 (type 2-150/25: LS mean difference (LSMD) vs placebo: 0.17 L (95% CI: 0.10-0.24) and 0.17 L (0.12-0.23); type 2-300/25: 0.22 L (0.13-0.30) and 0.21 L (0.15-0.28)), in exacerbators and non-exacerbators, respectively (p < .0001). Similar trends were seen for post-BD FEV1 . Dupilumab vs placebo also showed significantly greater improvements in post-BD FEV1 0-42 days after first severe exacerbation in type 2-150/25 (LSMD vs placebo: 0.13 L [0.06-0.20]; p = .006) and type 2-300/25 (0.14 L [0.06-0.22]; p = .001) patients. ACQ-5 improvements were greater with dupilumab vs placebo in both groups. CONCLUSION: Dupilumab treatment led to improvements in lung function independent of exacerbations and appeared to reduce the impact of exacerbations on lung function in patients who experienced a severe exacerbation during the study.

Age of asthma onset does not impact the response to omalizumab.

TRIAL REGISTRATION: These studies were conducted before clinical trial registration was required; therefore, clinical trial registration numbers are not available.

Comparison and validation of algorithms for asthma diagnosis in an electronic medical record system.

BACKGROUND: Asthma is one of the most common chronic health conditions, and to leverage the wealth of data in the electronic medical record (EMR), it is important to be able to accurately identify asthma diagnosis. OBJECTIVE: We aimed to determine the rule-based algorithm with the most balanced performance for sensitivity and positive predictive value of asthma diagnosis. METHODS: We performed a diagnostic accuracy study of multiple rule-based algorithms intended to identify asthma diagnosis in the EMR. Algorithm performance was validated by manual chart review of 795 charts of patients seen in a multisite, tertiary-level, pulmonary specialty clinic using explicit diagnostic criteria to distinguish asthma cases from controls. RESULTS: An asthma diagnosis anywhere in the medical record had a 97% sensitivity and a 77% specificity for asthma (F-score 80) when tested on a validation set of asthma cases and nonasthma respiratory disease from a pulmonary specialty clinic. The most balanced performance was seen with asthma diagnosis restricted to an encounter, hospital problem, or problem list diagnosis with a sensitivity of 94% and specificity of 85% (F-score 84). High sensitivity was achieved with the modified Health Plan Employer Data and Information Set criteria and high specificity was achieved with the NUgene algorithm, an algorithm developed for identifying asthma cases by EMR for genome-wide association studies. CONCLUSION: Asthma diagnosis can be accurately identified for research purposes by restricting to encounter, hospital problem, or problem list diagnosis in a pulmonary specialty clinic. Additional rules lead to steep drop-offs in algorithm sensitivity in our population.

Ambient Air Pollutants and Traffic Factors Were Associated with Blood and Urine Biomarkers and Asthma Risk.

The UK Biobank (UKBB) is a large population-based cohort that provides a unique opportunity to study the association between environmental exposure and biomarkers and to identify biomarkers as potential instruments for assessing exposure dose, health damage, and disease risks. On 462 063 participants of European ancestry, we characterized the relationship of 38 disease-relevant biomarkers, asthma diagnosis, ambient pollution, traffic factors, and genetic background. The air pollutant exposure on the UKBB cohort was fairly low (e.g., mean PM2.5 concentration at 10.0 µg/m3). Nevertheless, 30 biomarkers were in association with at least one environmental factor; e.g., C-reactive protein levels were positively associated with NO (padj = 2.99 × 10-4), NO2 (padj = 4.15 × 10-4), and PM2.5 (padj = 1.92 × 10-6) even after multiple testing adjustment. Asthma diagnosis was associated with four pollutants (NO, NO2, PM2.5, and PM10). The largest effect size was observed in PM2.5, where a 5 µg/m3 increment of exposure was associated with a 1.52 increase in asthma diagnosis (p = 4.41 × 10-13). Further, environmental exposure and genetic predisposition influenced biomarker levels and asthma diagnosis in an additive model. The exposure-biomarker associations identified in this study could serve as potential indicators for environmental exposure induced health damages. Our results also shed light on possible mechanisms whereby environmental exposure influences disease-causing biomarkers and in turn increases disease risk.

The relationship between asthma, eosinophilia, and outcomes in coronavirus disease 2019 infection.

BACKGROUND: The impact of asthma diagnosis and asthma endotype on outcomes from coronavirus disease 2019 (COVID-19) infection remains unclear. OBJECTIVE: To describe the association between asthma diagnosis and endotype and clinical outcomes among patients diagnosed as having COVID-19 infection. METHODS: Retrospective multicenter cohort study of outpatients and inpatients presenting to 6 hospitals in the Mount Sinai Health System New York metropolitan region between March 7, 2020, and June 7, 2020, with COVID-19 infection, with and without a history of asthma. The primary outcome evaluated was in-hospital mortality. Secondary outcomes included hospitalization, intensive care unit admission, mechanical ventilation, and hospital length of stay. The outcomes were compared in patients with or without asthma using a multivariate Cox regression model. The outcomes stratified by blood eosinophilia count were also evaluated. RESULTS: Of 10,523 patients diagnosed as having COVID-19 infection, 4902 were hospitalized and 468 had a diagnosis of asthma (4.4%). When adjusted for COVID-19 disease severity, comorbidities, and concurrent therapies, patients with asthma had a lower mortality (adjusted odds ratio [OR], 0.64 (0.53-0.77); P < .001) and a lower rate of hospitalization and intensive care unit admission (OR, 0.43 (0.28-0.64); P < .001 and OR, 0.51 (0.41-0.64); P < .001, respectively). Those with blood eosinophils greater than or equal to 200 cells/µL, both with and without asthma, had lower mortality. CONCLUSION: Patients with asthma may be at a reduced risk of poor outcomes from COVID-19 infection. Eosinophilia, both in those with and without asthma, may be associated with reduced mortality risk.

Clinicians' Use of Intravaginal Boric Acid Maintenance Therapy for Recurrent Vulvovaginal Candidiasis and Bacterial Vaginosis.

A retrospective chart review characterized clinicians' use of maintenance intravaginal boric acid for women with recurrent vulvovaginal candidiasis or bacterial vaginosis. Average length of use was 13 months with high patient satisfaction and few adverse events. Prospective studies are needed to evaluate the efficacy of maintenance boric acid for these conditions.

Bedside medication delivery programs: suggestions for systematic evaluation and reporting.

PURPOSE: Several factors lead to medication non-adherence after hospital discharge. Hospitals and pharmacies have implemented bedside medication delivery (BMD) programs for patients, in an attempt to reduce barriers and improve medication adherence. Here, we provide a critical review of the literature on these programs. DATA SOURCES: We conducted a literature search on BMD programs in PubMed, Google Scholar, Scopus and a general Google search using these keywords: 'medication delivery bedside', 'discharge medication delivery', 'meds to bedside' and 'meds to beds'. STUDY SELECTION: We identified 10 reports and include data from all reports. DATA EXTRACTION: Data on study characteristics and settings were extracted along with four outcomes: medication error, patient satisfaction, 30-day hospital readmission and visits to the emergency department. RESULTS OF DATA SYNTHESIS: Of the 10 reports, only 4 were peer-reviewed publications; others were reported in the lay press. Outcomes were reported in both qualitative and quantitative terms. Less than half of reports provided quantitative data on 30-day readmission and patient satisfaction. Others suggested qualitative improvement in these outcomes but did not provide data or specific details. None reported outcomes of their programs beyond 30 days. CONCLUSION: We highlight the need for increased use of optimal program design and more rigorous evaluations of the impact of BMD programs. We also provide guidelines on the types of evaluations that are likely needed and encourage improved reporting.

Models for the bystander effect in gradient radiation fields: Range and signalling type.

Bystander responses to radiation are responsible for a significant fraction of cell death, but are not included in the conventional linear-quadratic (LQ) radiobiological model. Strong dose gradients in radiation fields affect the distribution of bystander signals and can be used to decrease the survival of cancer cells. Predictive models incorporating bystander effects are needed to design the dose gradients in modulated fields to improve cancer treatments. Fundamental questions concern the nature and range of bystander signalling. Some authors propose bystander signals are carried by diffusing molecular factors expressed into the extracellular medium and that strong dose gradients drive their diffusion. Others propose bystander effects occur between neighbouring cells through gap-junctions, leaving no universal agreement. Here we test three assumptions concerning the effective range of bystander signals using both average and local measures of survival. Model 1 assumes short range signalling (e.g. gap-junction mediated) proportional to the local dose gradient, without relying on diffusion across the extracellular medium; Model 2 assumes metabolite diffusion governed by Fick's second law with either negative or both signs of bystander effect; Model 3 assumes that the extent of signal production is dependent on the average of the dose gradient over the field and that the signals have long range distribution. A single bystander parameter for each model was fitted to observed average survival of cancer cells in uniform and modulated fields. All models gave better fits than the classical LQ model. Model 2 fitted best with one sign of bystander effect on survival. Model 3 gave the best overall fit of average survival. The models were then used to predict local survival and survival as a function of dose in modulated fields, using independent datasets, without changing the bystander parameter. Model 3 gave the best overall prediction. This study demonstrates that the bystander effect can be controlled by design of the radiation field modulation.

Evolutionary Paths That Expand Plasmid Host-Range: Implications for Spread of Antibiotic Resistance.

The World Health Organization has declared the emergence of antibiotic resistance to be a global threat to human health. Broad-host-range plasmids have a key role in causing this health crisis because they transfer multiple resistance genes to a wide range of bacteria. To limit the spread of antibiotic resistance, we need to gain insight into the mechanisms by which the host range of plasmids evolves. Although initially unstable plasmids have been shown to improve their persistence through evolution of the plasmid, the host, or both, the means by which this occurs are poorly understood. Here, we sought to identify the underlying genetic basis of expanded plasmid host-range and increased persistence of an antibiotic resistance plasmid using a combined experimental-modeling approach that included whole-genome resequencing, molecular genetics and a plasmid population dynamics model. In nine of the ten previously evolved clones, changes in host and plasmid each slightly improved plasmid persistence, but their combination resulted in a much larger improvement, which indicated positive epistasis. The only genetic change in the plasmid was the acquisition of a transposable element from a plasmid native to the Pseudomonas host used in these studies. The analysis of genetic deletions showed that the critical genes on this transposon encode a putative toxin-antitoxin (TA) and a cointegrate resolution system. As evolved plasmids were able to persist longer in multiple naïve hosts, acquisition of this transposon also expanded the plasmid's host range, which has important implications for the spread of antibiotic resistance.

Evolved plasmid-host interactions reduce plasmid interference cost.

Antibiotic selection drives adaptation of antibiotic resistance plasmids to new bacterial hosts, but the molecular mechanisms are still poorly understood. We previously showed that a broad-host-range plasmid was poorly maintained in Shewanella oneidensis, but rapidly adapted through mutations in the replication initiation gene trfA1. Here we examined if these mutations reduced the fitness cost of TrfA1, and whether this was due to changes in interaction with the host's DNA helicase DnaB. The strains expressing evolved TrfA1 variants showed a higher growth rate than those expressing ancestral TrfA1. The evolved TrfA1 variants showed a lower affinity to the helicase than ancestral TrfA1 and were no longer able to activate the helicase at the oriV without host DnaA. Moreover, persistence of the ancestral plasmid was increased upon overexpression of DnaB. Finally, the evolved TrfA1 variants generated higher plasmid copy numbers than ancestral TrfA1. The findings suggest that ancestral plasmid instability can at least partly be explained by titration of DnaB by TrfA1. Thus under antibiotic selection resistance plasmids can adapt to a novel bacterial host through partial loss of function mutations that simultaneously increase plasmid copy number and decrease unfavorably high affinity to one of the hosts' essential proteins.

Characterisation of asthma subgroups associated with circulating YKL-40 levels.

The chitinase-like protein YKL-40 mediates airway inflammation and serum levels are associated with asthma severity. However, asthma phenotypes associated with YKL-40 levels have not been precisely defined.We conducted an unsupervised cluster analysis of asthma patients treated at the Yale Center for Asthma and Airways Disease (n=156) to identify subgroups according to YKL-40 level. The resulting YKL-40 clusters were cross-validated in cohorts from the Severe Asthma Research Programme (n=167) and the New York University/Bellevue Asthma Repository (n=341). A sputum transcriptome analysis revealed molecular pathways associated with YKL-40 subgroups.Four YKL-40 clusters (C1-C4) were identified. C3 and C4 had high serum YKL-40 levels compared with C1 and C2. C3 was associated with earlier onset and longer duration of disease, severe airflow obstruction, and near-fatal asthma exacerbations. C4 had the highest serum YKL-40 levels, adult onset and less airflow obstruction, but frequent exacerbations. An airway transcriptome analysis in C3 and C4 showed activation of non-type 2 inflammatory pathways.Elevated serum YKL-40 levels were associated with two distinct clinical asthma phenotypes: one with irreversible airway obstruction and another with severe exacerbations. The YKL-40 clusters are potentially useful for identification of individuals with severe or exacerbation-prone asthma.

Grid therapy using high definition multileaf collimators: realizing benefits of the bystander effect.

BACKGROUND: In microbeam radiotherapy (MRT), parallel arrays of high-intensity synchrotron x-ray beams achieve normal tissue sparing without compromising tumor control. Grid-therapy using clinical linacs has spatial modulation on a larger scale and achieves promising results for palliative treatments of bulky tumors. The availability of high definition multileaf collimators (HDMLCs) with 2.5 mm leaves provides an opportunity for grid-therapy to more closely approach MRT. However, challenges to the wider implementation of grid-therapy remain because spatial modulation of the target volume runs counter to current radiotherapy practice and mechanisms for the beneficial effects of MRT are not fully understood. Without more knowledge of cell dose responses, a quantitative basis for planning treatments is difficult. The aim of this study is to determine if therapeutic benefits of MRT can be achieved using a linac with HDMLCs and if so, to develop a predictive model to support treatment planning. MATERIAL AND METHODS: HD120-MLCs of a Varian Novalis TXTM were used to generate grid patterns of 2.5 and 5.0 mm spacing, which were characterized dosimetrically using GafchromicTM EBT3 film. Clonogenic survival of normal (HUVEC) and cancer (NCI-H460, HCC-1954) cell lines following irradiation under the grid and open fields using a 6 MV photon beam were compared in-vitro for the same average dose. RESULTS AND CONCLUSIONS: Relative to an open field, survival of normal cells in a 2.5 mm striped field was the same, while the survival of both cancer cell lines was significantly lower. A mathematical model was developed to incorporate dose gradients of the spatial modulation into the standard linear quadratic model. Our new bystander extended LQ model assumes spatial gradients drive the diffusion of soluble factors that influence survival through bystander effects, successfully predicting the experimental results that show an increased therapeutic ratio. Our results challenge conventional radiotherapy practice and propose that additional gain can be realized by prescribing spatially modulated treatments to harness the bystander effect.

Predictors of asthma exacerbation among patients with poorly controlled asthma despite inhaled corticosteroid treatment.

BACKGROUND: Asthma exacerbations are associated with decreased quality of life and increased health care usage. Identification of characteristics that predict increased risk of future exacerbations in patients with suboptimal control of asthma could guide treatment decisions. OBJECTIVE: To examine patient characteristics associated with risk of asthma exacerbations in patients with uncontrolled persistent asthma. METHODS: A retrospective analysis of adults and children with inadequately controlled asthma despite asthma controller therapy and enrolled in 2 randomized trials was conducted. Baseline characteristics of subjects who experienced an asthma exacerbation during the treatment period were compared with those of subjects who did not experience an exacerbation. RESULTS: Of 718 subjects (402 adults and 295 children), 108 adults (27%) and 110 children (37%) experienced an asthma exacerbation during the study period. Unscheduled health care visits for asthma or use of oral corticosteroids in the previous year were significantly associated with asthma exacerbation during the study period (P < .01). Adult subjects who experienced an exacerbation had significantly lower forced expiratory volume in 1 second compared with those who did not (2.3 vs 2.5 L, respectively, P = .02). Children who experienced an exacerbation had lower baseline pre- and post-bronchodilator ratios of forced expiratory volume in 1 second to forced vital capacity (77% vs 81%, P < .01; 82% vs 86%, P < .001, respectively). Symptom scores on validated questionnaires were significantly worse in adults but not in children who developed an exacerbation. CONCLUSION: Spirometric measurements can help identify adults and children at increased risk for asthma exacerbation. Symptom scores could be helpful in identifying adults who are at high risk for exacerbations but could be less helpful in children.

Role of anticholinergics in asthma management: recent evidence and future needs.

PURPOSE OF REVIEW: Anticholinergic antimuscarinic bronchodilators play a major role in the treatment of chronic obstructive pulmonary disease, but their role in asthma has long been limited to acute management. More recently, the role of long-acting antimuscarinic bronchodilators (LAMAs) in chronic asthma management has been explored. This review will examine the pharmacological rationale for use of inhaled anticholinergics in the treatment of asthma, and provide an overview of the current literature supporting this use, as well as describe future research needs in this area. RECENT FINDINGS: Short-acting anticholinergic bronchodilators have a role as add-on agents in the treatment of acute asthma. Preliminary clinical studies suggest that inhaled LAMAs may be comparable to long-acting beta2-agonists (LABAs) as an add-on therapy in patients not controlled by inhaled corticosteroids (ICS) alone, and may also have added benefit in patients not controlled on combined ICS-LABA. Mechanistic studies suggest that apart from their bronchodilator activity, LAMAs may have anti-inflammatory and antiremodeling influences on the airways. Further research is needed to clarify the clinical relevance of these experimental observations. SUMMARY: Accumulating evidence supports the use of inhaled LAMAs as an add-on therapy in patients with asthma, who remain symptomatic despite guideline-based therapy with ICS with or without LABAs. Further studies are warranted to help define mechanisms of action of LAMAs, apart from their role as bronchodilators, and determine how these other actions impact asthma outcomes over time. Furthermore, future studies need to examine the long-term efficacy and safety of LAMAs in asthma and identify a subgroup of patients who would benefit from such therapies to facilitate early, personalized therapy.

The alternative complement component factor B regulates UV-induced oedema, systemic suppression of contact and delayed hypersensitivity, and mast cell infiltration into the skin.

Ultraviolet (UV) wavelengths in sunlight are the prime cause of skin cancer in humans with both the UVA and UVB wavebands making a contribution to photocarcinogenesis. UV has many different biological effects on the skin that contribute to carcinogenesis, including suppression of adaptive immunity, sunburn and altering the migration of mast cells into and away from irradiated skin. Many molecular mechanisms have been identified as contributing to skin responses to UV. Recently, using gene set enrichment analysis of microarray data, we identified the alternative complement pathway with a central role for factor B (fB) in UVA-induced immunosuppression. In the current study we used mice genetically deficient in fB (fB-/- mice) to study the functional role of the alternative complement pathway in skin responses to UV. We found that fB is required for not only UVA but also UVB-induced immunosuppression and solar-simulated UV induction of the oedemal component of sunburn. Factor B-/- mice had a larger number of resident skin mast cells than control mice, but unlike the controls did not respond to UV by increasing mast cell infiltration into the skin. This study provides evidence for a function role for fB in skin responses to UV radiation. Factor B regulates UVA and UVB induced immunosuppression, UV induced oedema and mast cell infiltration into the skin. The alternative complement pathway is therefore an important regulator of skin responses to UV.

Understanding the challenges of palliative care in everyday clinical practice: an example from a COPD action research project.

Palliative care seeks to improve the quality of life for patients suffering from the impact of life-limiting illnesses. Palliative care encompasses but is more than end-of-life care, which is defined as care during the final hours/days/weeks of life. Although palliative care policies increasingly require all healthcare professionals to have at least basic or non-specialist skills in palliative care, international evidence suggests there are difficulties in realising such policies. This study reports on an action research project aimed at developing respiratory nursing practice to address the palliative care needs of patients with advanced chronic obstructive pulmonary disease (COPD). The findings suggest that interlevel dynamics at individual, team, interdepartmental and organisational levels are an important factor in the capacity of respiratory nurses to embed non-specialist palliative care in their practice. At best, current efforts to embed palliative care in everyday practice may improve end-of-life care in the final hours/days/weeks of life. However, embedding palliative care in everyday practice requires a more fundamental shift in the organisation of care.

Profiling of the secretome of human cancer cells: preparation of supernatant for proteomic analysis.

Secretomic analysis requires removal of serum proteins from cell-culture media. We evaluate the proteins washed from cells prepared in bovine serum-supplemented medium. PBS and serum-free-medium (SFM) were the washing solutions. A Bradford assay was used for total protein concentration and a 1D gel and LC-MS/MS, to assign the protein to human or bovine origin. For both wash solutions, all bovine protein had been removed by the third wash, without compromising the number of living cells. Further washes reduced the number of living cells, especially when using PBS. Proteomic analysis of wash supernatant showed that SFM induced greater lysis of dead cells. Three washes were sufficient to minimize the effects on cell viability, while still removing serum proteins. Washing in SFM resulted in contamination of the wash supernatant with lysed dead cell proteins. Washed cells were incubated in SFM and exposed to ionizing radiation. Analysis of the supernatant showed an increase in human cytoplasmic, plasma membrane, and nuclear protein following irradiation. Secreted proteins were also detected, but in smaller quantities. The significance of these findings extend to in vitro studies of bystander phenomena, since the proteins of lysed dead cells may participate in driving bystander responses.