Edward Pickering

Full-Length IL-33 Promotes Inflammation but not Th2 Response In Vivo in an ST2-Independent Fashion

Expression of IL-33 is elevated in patients with pulmonary diseases, and full-length (not proteolytically processed) IL-33 is the predominant form in the lungs in health and disease. To determine whether activation of IL-33 is needed for functional effects, activities of full-length mouse and mature mouse (mm) forms of IL-33 were compared in vivo. Replication-deficient adenoviral constructs were used for gene delivery. Both isoforms caused pulmonary infiltration of lymphocytes and neutrophils, whereas mmIL-33 also caused pulmonary eosinophilia and goblet cell hyperplasia and increased expression of IL-4, IL-5, IL-13, IL-17, MCP-1, and KC. The different effects were not associated with differential release from IL-33–producing cells or by differences in subcellular distributions of IL-33 isoforms. Germline deficiency of the cell surface receptor chain ST2 abrogated the mmIL-33–induced Th2-associated effects (pulmonary eosinophilia, goblet cell hyperplasia, and increased IL-4 and IL-5), yet the lymphocytic infiltration induced by full-length mouse IL-33 or mmIL-33 was not fully abrogated by the absence of ST2. The similar effects of IL-33 isoforms were associated with comparable regulation of gene expression, notably matrix metalloproteinases 3, 10, and 13. Thus, full-length IL-33 is functionally active in vivo in an ST2-independent fashion, and its effects are partially different from those of mature IL-33. The different effects of these isoforms, particularly the pro-Th2 effects of mature IL-33, are due to differential utilization of the IL-33R chain ST2, whereas their similar effects result from regulation of gene expression.

Natural production and functional effects of alternatively spliced interleukin-4 protein in asthma

We have previously described an alternatively spliced isoform of IL-4 mRNA that omits exon 2 and is termed IL-4δ2. However, the natural production of IL-4δ2 protein and its association with disease have not been previously assessed due to unavailability of an antibody that interacts with IL-4δ2 without cross-reactivity with full length IL-4. We used a unique monoclonal antibody (mAb) that reacts with IL-4δ2, but not with IL-4, and observed that IL-4δ2 is naturally produced by T cells from patients with asthma, but not from healthy controls. The kinetics of IL-4δ2 and IL-4 production by phorbol myristate acetate (PMA)/ionomycin-activated cells differed, with IL-4δ2 increasing at 48-72h and IL-4 peaking at 24h. The steady-state levels of IL-4δ2 mRNA varied significantly among the donors and were discordant with the corresponding protein levels, suggesting post-transcriptional regulation of protein production. Polarized Th1 or Th2 lymphocytes were not a major source of IL-4δ2. Stimulation of cultured T lymphocytes with IL-4δ2 caused elevated production of IFN-γ, IL-10, IL-6, MCP-1, and TNF-α, with notable differences between patients and controls in the production of IFN-γ, IL-10, and IL-6. Thus, IL-4δ2 is natively produced not only as mRNA but also as a protein by cells other than Th1 or Th2. It is regulated post-transcriptionally, is associated with allergic asthma, and regulates production of other cytokines by primary T lymphocytes. Alternatively spliced interleukin-4 may be a new biomarker, a pathophysiological player, and possibly a molecular target for future therapies in asthma.

IFN-γ Directly Controls IL-33 Protein Level through a STAT1- and LMP2-dependent Mechanism

Background: IL-33 levels are regulated through poorly understood cytokine-dependent mechanisms. Results: IFN-γ but not IL-4 down-regulates IL-33 protein by activating STAT1 and LMP2 proteasome, without engaging caspase-1, -3, or -8. Conclusion: Down-regulation of IL-33 protein by IFN-γ requires STAT1 and non-canonical involvement of LMP2 proteasome. Significance: Understanding the mechanisms of IL-33 regulation is important for the development of IL-33-targeting therapies. IL-33 contributes to disease processes in association with Th1 and Th2 phenotypes. IL-33 mRNA is rapidly regulated, but the fate of synthesized IL-33 protein is unknown. To understand the interplay among IL-33, IFN-γ, and IL-4 proteins, recombinant replication-deficient adenoviruses were produced and used for dual expression of IL-33 and IFN-γ or IL-33 and IL-4. The effects of such dual gene delivery were compared with the effects of similar expression of each of these cytokines alone. In lung fibroblast culture, co-expression of IL-33 and IFN-γ resulted in suppression of the levels of both proteins, whereas co-expression of IL-33 and IL-4 led to mutual elevation. In vivo, co-expression of IL-33 and IFN-γ in the lungs led to attenuation of IL-33 protein levels. Purified IFN-γ also attenuated IL-33 protein in fibroblast culture, suggesting that IFN-γ controls IL-33 protein degradation. Specific inhibition of caspase-1, -3, and -8 had minimal effect on IFN-γ-driven IL-33 protein down-regulation. Pharmacological inhibition, siRNA-mediated silencing, or gene deficiency of STAT1 potently up-regulated IL-33 protein expression levels and attenuated the down-regulating effect of IFN-γ on IL-33. Stimulation with IFN-γ strongly elevated the levels of the LMP2 proteasome subunit, known for its role in IFN-γ-regulated antigen processing. siRNA-mediated silencing of LMP2 expression abrogated the effect of IFN-γ on IL-33. Thus, IFN-γ, IL-4, and IL-33 are engaged in a complex interplay. The down-regulation of IL-33 protein levels by IFN-γ in pulmonary fibroblasts and in the lungs in vivo occurs through STAT1 and non-canonical use of the LMP2 proteasome subunit in a caspase-independent fashion.

From electrocautery, balloon dilatation, neodymium-doped:yttrium-aluminum-garnet (Nd:YAG) laser to argon plasma coagulation and cryotherapy

Over the past decade, there has been significant advancement in the development/application of therapeutics in thoracic diseases. Ablation methods using heat or cold energy in the airway is safe and effective for treating complex airway disorders including malignant and non-malignant central airway obstruction (CAO) without limiting the impact of future definitive therapy. Timely and efficient use of endobronchial ablative therapies combined with mechanical debridement or stent placement results in immediate relief of dyspnea for CAO. Therapeutic modalities reviewed in this article including electrocautery, balloon dilation (BD), neodymium-doped:yttrium-aluminum-garnet (Nd:YAG) laser, argon plasma coagulation (APC), and cryotherapy are often combined to achieve the desired results. This review aims to provide a clinically oriented review of these technologies in the modern era of interventional pulmonology (IP).

Nonpharmacologic approach to minimizing shivering during surface cooling: A proof of principle study☆

PURPOSE This study had 2 objectives: (1) to quantify the metabolic response to physical cooling in febrile patients with systemic inflammatory response syndrome (SIRS) and (2) to provide proof for the hypothesis that the efficiency of external cooling and the subsequent shivering response are influenced by site and temperature of surface cooling pads. METHODS To quantify shivering thermogenesis during surface cooling for fever, we monitored oxygen consumption (VO(2)) in 6 febrile patients with SIRS during conventional cooling with cooling blankets and ice packs. To begin to determine how location and temperature of surface cooling influence shivering, we compared 5 cooling protocols for inducing mild hypothermia in 6 healthy volunteers. RESULTS In the patients with SIRS, core temperature decreased 0.67 °C per hour, all patients shivered, VO(2) increased 57.6%, and blood pressure increased 15% during cooling. In healthy subjects, cooling with the 10 °C vest was most comfortable and removed heat most efficiently without shivering or VO(2) increase. Cooling with combined vest and thigh pads stimulated the most shivering and highest VO(2) and increased core temperature. Reducing vest temperature from 10 °C to 5 °C failed to increase heat removal secondary to cutaneous vasoconstriction. Capsaicin, an agonist for the transient receptor potential cation channel subfamily V member 1 (TRPV1) warm-sensing channels, partially reversed this effect in 5 subjects. CONCLUSIONS Our results identify the hazards of surface cooling in febrile critically ill patients and support the concept that optimization of cooling pad temperature and position may improve cooling efficiency and reduce shivering.

Feasibility, safety, and utility of bronchoscopy in patients with ARDS while in the prone position

Prone positioning (PP) was shown to reduce mortality in mechanically ventilated (MV) patients with severe ARDS [1]. Despite its common use, safety concerns inhibit use of flexible bronchoscopy (FB) in patients with ARDS, and there are few reports of FB performed in PP [2]. We reviewed all adults receiving FB in PP in one institution between April 2016 and September 2017. The study was approved by the institutional review board. Four men and three women were identified (Table 1). In five patients, FB was indicated for clearance of thick secretions, and in two patients for microbial analysis. The mode of mechanical ventilation was not changed for FB, but FIO2 was universally set to 100%. All subjects had invasive hemodynamic and pulse oximetry monitoring. End-tidal carbon dioxide (EtCO2) was monitored in 3/7 subjects. With the subject’s head tilted to the side, the bronchoscope was advanced into the airways, repeatedly, and in short cycles, allowing time for oxygenation, ventilation, and lung recruitment between insertions. Therapeutic aspiration was performed in 6/7 subjects. Bronchoalveolar lavage was performed in two subjects. No significant hemodynamic compromise was observed during any of the procedures. Significant oxygen desaturation and rising EtCO2 were observed in one case (patient 4). Both derangements resolved with withdrawal of the bronchoscope and recruitment. No additional complications were documented. Figure 1 illustrates evolution of the PaO2:FIO2 ratio over time for each subject. Six subjects had antibiotics modified based on FB-obtained cultures. Consistent with previous data [3], 4/7 subjects survived 30 days following discharge from the ICU. Although PP is lung-protective, it may result in mobilization of secretions into the airways, impairing oxygenation and providing nidus for infection [4]. Despite documented risks [5], FB may be beneficial in this situation. Several limitations need to be addressed when interpreting our data. This is a retrospective analysis. Although physiologic monitoring was automatically captured, ventilator data were not and ventilator output during FB could not be accurately analyzed. Additionally, EtCO2 was not measured in all cases during FB. Finally, PP was shown to reduce mortality in patients with moderate to severe ARDS, however, our study subjects’ oxygenation had started to improve by the time FB was performed (Fig. 1, T1). This likely reflects reluctance to perform FB in subjects with severe hypoxemia due to excessive risks. Our report demonstrates the feasibility of FB performed in brief increments in carefully monitored patients with ARDS ventilated in PP. Further studies are needed to better delineate optimal ventilator management during FB in PP.

Ice cream cone sign: reversible ballooning of the trachea due to tracheostomy tube cuff overinflation

A woman in her 80s with ventilator-dependent respiratory failure due to severe cardiomyopathy and critical illness myopathy was transferred to our hospital after experiencing recurrent episodes of ventricular fibrillation cardiac arrest requiring cardiopulmonary resuscitation. On arrival, there was a significant air leak despite tracheostomy tube (TT) cuff pressures of 50 cm H2O (normal <25 cm H2O). Chest radiography revealed what we term the ‘ice cream cone sign’ (figure 1 and video 1), consistent with proximal trachea …