Investigating the role of platelets and platelet-derived transforming growth factor-ß in idiopathic pulmonary fibrosis.

Transforming growth factor-ß1 (TGFß1) is the key profibrotic cytokine in idiopathic pulmonary fibrosis (IPF), but the primary source of this cytokine in this disease is unknown. Platelets have abundant stores of TGFß1, although the role of these cells in IPF is ill-defined. In this study, we investigated whether platelets, and specifically platelet-derived TGFß1, mediate IPF disease progression. Patients with IPF and non-IPF patients were recruited to determine platelet reactivity, and separate cohorts of patients with IPF were followed for mortality. To study whether platelet-derived TGFß1 modulates pulmonary fibrosis (PF), mice with a targeted deletion of TGFß1 in megakaryocytes and platelets (TGFß1fl/fl.PF4-Cre) were used in the well-characterized bleomycin-induced pulmonary fibrosis (PF) animal model. In a discovery cohort, we found significantly higher mortality in patients with IPF who had elevated platelet counts within the normal range. However, our validation cohort did not confirm this observation, despite significantly increased platelets, neutrophils, active TGFß1, and CCL5, a chemokine produced by inflammatory cells, in the blood, lung, and bronchoalveolar lavage (BAL) of patients with IPF. In vivo, we showed that despite platelets being readily detected within the lungs of bleomycin-treated mice, neither the degree of pulmonary inflammation nor fibrosis was significantly different between TGFß1fl/fl.PF4-Cre and control mice. Our results demonstrate for the first time that platelet-derived TGFß1 does not significantly mediate inflammation or fibrosis in a PF animal model. Furthermore, our human studies revealed blood platelet counts do not consistently predict mortality in IPF but other platelet-derived mediators, such as C-C chemokine ligand 5 (CCL5), may promote neutrophil recruitment and human IPF.NEW & NOTEWORTHY Platelets are a rich source of profibrotic TGFß; however, the role of platelets in idiopathic pulmonary fibrosis (IPF) is unclear. We identified that patients with IPF have significantly more platelets, neutrophils, and active TGFß in their airways than control patients. Using an animal model of IPF, we demonstrated that platelet-derived TGFß does not significantly drive lung fibrosis or inflammation. Our findings offer a better understanding of platelets in both human and animal studies of IPF.

Clinical and Radiological Phenotypes and Endotypes.

Bronchiectasis is a heterogenous disease with multiple etiologies and associated comorbidities. As bronchiectasis is a complex disease, it is unsound to think of it as a single disease particularly when the differing etiologies are likely to be driving bronchiectasis through initial divergent molecular pathways, known as endotypes, that phenotypically present as the same disease due to protracted airway inflammation, but revealing potential differing underlying mechanisms that may have disparity of drug responses. Improved understanding of the cellular immune, inflammatory, and microbiological milieu associated with clinical and radiological features of bronchiectasis has resulted in the recognition of important endotypes and phenotypes that will allow for personalized treatments to improve quality of life and outcomes of patients with bronchiectasis. Here we discuss clinical and radiological phenotypes, as well as emerging molecular endotypes that are possible treatable traits in bronchiectasis.

Correction: Naturally Acquired Human Immunity to Pneumococcus Is Dependent on Antibody to Protein Antigens.

[This corrects the article DOI: 10.1371/journal.ppat.1006137.].

Naturally Acquired Human Immunity to Pneumococcus Is Dependent on Antibody to Protein Antigens.

Naturally acquired immunity against invasive pneumococcal disease (IPD) is thought to be dependent on anti-capsular antibody. However nasopharyngeal colonisation by Streptococcus pneumoniae also induces antibody to protein antigens that could be protective. We have used human intravenous immunoglobulin preparation (IVIG), representing natural IgG responses to S. pneumoniae, to identify the classes of antigens that are functionally relevant for immunity to IPD. IgG in IVIG recognised capsular antigen and multiple S. pneumoniae protein antigens, with highly conserved patterns between different geographical sources of pooled human IgG. Incubation of S. pneumoniae in IVIG resulted in IgG binding to the bacteria, formation of bacterial aggregates, and enhanced phagocytosis even for unencapsulated S. pneumoniae strains, demonstrating the capsule was unlikely to be the dominant protective antigen. IgG binding to S. pneumoniae incubated in IVIG was reduced after partial chemical or genetic removal of bacterial surface proteins, and increased against a Streptococcus mitis strain expressing the S. pneumoniae protein PspC. In contrast, depletion of type-specific capsular antibody from IVIG did not affect IgG binding, opsonophagocytosis, or protection by passive vaccination against IPD in murine models. These results demonstrate that naturally acquired protection against IPD largely depends on antibody to protein antigens rather than the capsule.

Review of the British Thoracic Society Winter Meeting 2016, 7-9 December, London, UK.

This article reviews the British Thoracic Society Winter Meeting 2016 and highlights the new developments in scientific and clinical research across the breadth of respiratory medicine.

Evidence for chemokine synergy during neutrophil migration in ARDS.

BACKGROUND: Acute respiratory distress syndrome (ARDS) is a life-threatening condition characterised by pulmonary oedema, respiratory failure and severe inflammation. ARDS is further characterised by the recruitment of neutrophils into the lung interstitium and alveolar space. OBJECTIVES: The factors that regulate neutrophil infiltration into the inflamed lung and our understanding of the pathomechanisms in ARDS remain incomplete. This study aimed at determining the role of the chemokine (C-C motif) ligand (CCL)2 and CCL7 in ARDS. METHODS: CCL2 and CCL7 protein levels were measured in bronchoalveolar lavage (BAL) fluid obtained from lipopolysaccharide(LPS)-challenged human volunteers and two separate cohorts of patients with ARDS. Neutrophil chemotaxis to ARDS BAL fluid was evaluated and the contribution of each was assessed and compared with chemokine (C-X-C motif) ligand 8 (CXCL8). Chemokine receptor expression on neutrophils from blood or BAL fluid of patients with ARDS was analysed by flow cytometry. RESULTS: CCL2 and CCL7 were significantly elevated in BAL fluid recovered from LPS-challenged volunteers and patients with ARDS. BAL fluid from patients with ARDS was highly chemotactic for human neutrophils and neutralising either CCL2 or CCL7 attenuated the neutrophil chemotactic response. Moreover, CCL2 and CCL7 synergised with CXCL8 to promote neutrophil migration. Furthermore, neutrophils isolated from the blood or BAL fluid differentially regulated the cell surface expression of chemokine (C-X-C motif) receptor 1 and C-C chemokine receptor type 2 during ARDS. CONCLUSION: This study highlights important inflammatory chemokines involved in regulating neutrophil migration, which may have potential value as therapeutic targets for the treatment of ARDS.

Adult pneumococcal vaccination: advances, impact, and unmet needs.

PURPOSE OF REVIEW: Preventing pneumonia in the elderly and individuals with comorbidities is an unmet clinical need. Streptococcus pneumoniae is the commonest bacterial cause of pneumonia, and we summarize recent findings regarding current S. pneumoniae vaccines, and debate their efficacy and cost-effectiveness in risk groups. We also discuss potential future vaccine strategies such as protein antigen vaccines. RECENT FINDINGS: Current vaccination with pneumococcal polysaccharide vaccine does not prevent S. pneumoniae pneumonia. Vaccination with pneumococcal conjugated vaccine (PCV) prevents nasopharyngeal colonization, but although PCV13 has recently been shown to prevent S. pneumoniae pneumonia in adults, its overall efficacy was relatively low. The results of cost-effectiveness studies of PCV vaccination in adults are variable with some showing this is a cost-effective strategy, whereas others have not. The lack of cost-effectiveness is predominantly because of the current cost of the PCV vaccine and the existing herd immunity effect from childhood PCV vaccination on vaccine serotypes. SUMMARY: S. pneumoniae pneumonia is a vaccine-preventable disease but remains a common cause of morbidity and mortality. Advances in vaccination using approaches that induce serotypes-independent immunity and are immunogenic in high-risk groups are required to reduce the burden of disease because of S. pneumoniae.

Regulation of neutrophilic inflammation by proteinase-activated receptor 1 during bacterial pulmonary infection.

Neutrophils are key effector cells of the innate immune response to pathogenic bacteria, but excessive neutrophilic inflammation can be associated with bystander tissue damage. The mechanisms responsible for neutrophil recruitment to the lungs during bacterial pneumonia are poorly defined. In this study, we focus on the potential role of the major high-affinity thrombin receptor, proteinase-activated receptor 1 (PAR-1), during the development of pneumonia to the common lung pathogen Streptococcus pneumoniae. Our studies demonstrate that neutrophils were indispensable for controlling S. pneumoniae outgrowth but contributed to alveolar barrier disruption. We further report that intra-alveolar coagulation (bronchoalveolar lavage fluid thrombin-antithrombin complex levels) and PAR-1 immunostaining were increased in this model of bacterial lung infection. Functional studies using the most clinically advanced PAR-1 antagonist, SCH530348, revealed a key contribution for PAR-1 signaling in influencing neutrophil recruitment to lung airspaces in response to both an invasive and noninvasive strain of S. pneumoniae (D39 and EF3030) but that PAR-1 antagonism did not impair the ability of the host to control bacterial outgrowth. PAR-1 antagonist treatment significantly decreased pulmonary levels of IL-1ß, CXCL1, CCL2, and CCL7 and attenuated alveolar leak. Ab neutralization studies further demonstrated a nonredundant role for IL-1ß, CXCL1, and CCL7 in mediating neutrophil recruitment in response to S. pneumoniae infection. Taken together, these data demonstrate a key role for PAR-1 during S. pneumoniae lung infection that is mediated, at least in part, by influencing multiple downstream inflammatory mediators.

Macrophage migration inhibitory factor-CXCR4 is the dominant chemotactic axis in human mesenchymal stem cell recruitment to tumors.

Mesenchymal stromal cells (MSCs) are inherently tumor homing and can be isolated, expanded, and transduced, making them viable candidates for cell therapy. This tumor tropism has been used to deliver anticancer therapies to various tumor models. In this study, we sought to discover which molecules are the key effectors of human MSC tumor homing in vitro and using an in vivo murine model. In this study, we discover a novel role for macrophage migration inhibitory factor (MIF) as the key director of MSC migration and infiltration toward tumor cells. We have shown this major role for MIF using in vitro migration and invasion assays, in presence of different receptor inhibitors and achieving a drastic decrease in both processes using MIF inhibitor. Additionally, we demonstrate physical interaction between MIF and three receptors: CXCR2, CXCR4, and CD74. CXCR4 is the dominant receptor used by MIF in the homing tumor context, although some signaling is observed through CXCR2. We demonstrate downstream activation of the MAPK pathway necessary for tumor homing. Importantly, we show that knockdown of either CXCR4 or MIF abrogates MSC homing to tumors in an in vivo pulmonary metastasis model, confirming the in vitro two-dimensional and three-dimensional assays. This improved understanding of MSC tumor tropism will further enable development of novel cellular therapies for cancers.

Exposure to welding fumes and lower airway infection with Streptococcus pneumoniae.

BACKGROUND: Welders are at increased risk of pneumococcal pneumonia. The mechanism for this association is not known. The capacity of pneumococci to adhere to and infect lower airway cells is mediated by host-expressed platelet-activating factor receptor (PAFR). OBJECTIVE: We sought to assess the effect of mild steel welding fumes (MS-WF) on PAFR-dependent pneumococcal adhesion and infection to human airway cells in vitro and on pneumococcal airway infection in a mouse model. METHODS: The oxidative potential of MS-WF was assessed by their capacity to reduce antioxidants in vitro. Pneumococcal adhesion and infection of A549, BEAS-2B, and primary human bronchial airway cells were assessed by means of quantitative bacterial culture and expressed as colony-forming units (CFU). After intranasal instillation of MS-WF, mice were infected with Streptococcus pneumoniae, and bronchoalveolar lavage fluid (BALF) and lung CFU values were determined. PAFR protein levels were assessed by using immunofluorescence and immunohistochemistry, and PAFR mRNA expression was assessed by using quantitative PCR. PAFR was blocked by CV-3988, and oxidative stress was attenuated by N-acetylcysteine. RESULTS: MS-WF exhibited high oxidative potential. In A549 and BEAS-2B cells MS-WF increased pneumococcal adhesion and infection and PAFR protein expression. Both CV-3988 and N-acetylcysteine reduced MS-WF-stimulated pneumococcal adhesion and infection of airway cells. MS-WF increased mouse lung PAFR mRNA expression and increased BALF and lung pneumococcal CFU values. In MS-WF-exposed mice CV-3988 reduced BALF CFU values. CONCLUSIONS: Hypersusceptibility of welders to pneumococcal pneumonia is in part mediated by the capacity of welding fumes to increase PAFR-dependent pneumococcal adhesion and infection of lower airway cells.

Review of the British Thoracic Society Winter Meeting 2015, 2-4 December, London, UK.

The British Thoracic Society Winter Meeting 2015 is reviewed in this article. Over 3 days in December, this annual scientific meeting attracted over 2300 delegates and up-to-date respiratory research was presented by leading UK and international speakers. This article reviews a number of symposia and selected abstract presentations from the meeting.

Enhanced inflammation in aged mice following infection with Streptococcus pneumoniae is associated with decreased IL-10 and augmented chemokine production.

Streptococcus pneumoniae is the most common cause of severe pneumonia in the elderly. However, the impact of aging on the innate inflammatory response to pneumococci is poorly defined. We compared the innate immune response in old vs. young adult mice following infection with S. pneumoniae. The accumulation of neutrophils recovered from bronchoalveolar lavage fluid and lung homogenates was increased in aged compared with young adult mice, although bacterial outgrowth was similar in both age groups, as were markers of microvascular leak. Aged mice had similar levels of IL-1ß, TNF, IFN-γ, IL-17, and granulocyte colony-stimulating factor following S. pneumoniae infection, compared with young mice, but increased levels of the chemokines CXCL9, CXCL12, CCL3, CCL4, CCL5, CCL11, and CCL17. Moreover, levels of IL-10 were significantly lower in aged animals. Neutralization of IL-10 in infected young mice was associated with increased neutrophil recruitment but no decrease in bacterial outgrowth. Furthermore, IL-10 neutralization resulted in increased levels of CCL3, CCL5, and CXCL10. We conclude that aging is associated with enhanced inflammatory responses following S. pneumoniae infection as a result of a compromised immunomodulatory cytokine response.

Review of the British Thoracic Society Winter Meeting 2014, 3-5 December, London, UK.

The British Thoracic Society Winter Meeting 2014 is reviewed in this article. This annual scientific meeting attracted its largest number of delegates ever and over 3 days in December up-to-date respiratory research was presented and current thinking in respiratory science and clinical academia was discussed. This article reviews a number of symposia and selected abstract presentations from the meeting.

Community-acquired pneumonia.

PURPOSE OF REVIEW: Community-acquired pneumonia (CAP) is the most common infectious disease cause of death. We summarize recent findings regarding the epidemiology of CAP in adults, efficacy of vaccines against Streptococcus pneumoniae, diagnostics, and discuss the current controversy between CAP and healthcare-associated pneumonia (HCAP). RECENT FINDINGS: The emergence of the Middle East respiratory syndrome coronavirus and the avian influenza A strain H7N9 are of concern but still these are infrequent causes of CAP. Recent data indicate that vaccinating children also protects adults against CAP by generating significant herd immunity, and that the conjugated pneumococcal vaccine in adults may offer some efficacy in preventing CAP caused by vaccine serotypes. The immunochromotagraphic urinary antigen test has improved the diagnostic yield for the aetiology of CAP, and initial data demonstrate that a novel multiplex urinary antigen test will further increase the sensitivity for detection of S. pneumoniae. There has been significant concern that a relatively recently described pneumonia category, HCAP, requires empirical treatment for potentially multidrug-resistant organisms (MDRO). However, new evidence shows that (at least in Europe) pneumonia caused by MDRO remains uncommon even in HCAP category patients. SUMMARY: CAP remains a major cause of morbidity and mortality. Advances in vaccination and diagnosis should help reduce the amount of disease due to S. pneumoniae, the commonest cause of CAP. Outside of the United States, MDRO are relatively uncommon causes of CAP, and the increased mortality of HCAP category patients seems to be related to their comorbidities and age rather than microbial aetiology.

Proteinase-activated receptor-1, CCL2, and CCL7 regulate acute neutrophilic lung inflammation.

PAR1 plays a central role in mediating the interplay between coagulation and inflammation, but its role in regulating acute neutrophilic inflammation is unknown. We report that antagonism of PAR1 was highly effective at reducing acute neutrophil accumulation in a mouse model of LPS-induced lung inflammation. PAR1 antagonism also reduced alveolar-capillary barrier disruption in these mice. This protection was associated with a reduction in the expression of the chemokines, CCL2 and CCL7, but not the proinflammatory cytokines, TNF and IL-6, or the classic neutrophil chemoattractants, CXCL1 and CXCL2. Antibody neutralization of CCL2 and CCL7 significantly reduced LPS-induced total leukocyte and neutrophil accumulation, recovered from the bronchoalveolar lavage fluid of challenged mice. Immunohistochemical analysis revealed that CCL2 predominantly localized to alveolar macrophages and pulmonary epithelial cells, whereas CCL7 was restricted to the pulmonary epithelium. In keeping with these observations, the intranasal administration of recombinant CCL2 (rCCL2) and rCCL7 led to the accumulation of neutrophils within the lung airspaces of naive mice in the absence of any underlying inflammation. Flow cytometry analysis further demonstrated an increase in Ly6G(hi) neutrophils expressing the chemokine receptors, CCR1 and CCR2, isolated from mouse lungs compared with circulating neutrophils. Conversely, the expression of CXCR2 decreased on neutrophils isolated from the lung compared with circulating neutrophils. Furthermore, this switch in chemokine receptor expression was accentuated after acute LPS-induced lung inflammation. Collectively, these findings reveal a novel role for PAR1 and the chemokines, CCL2 and CCL7, during the early events of acute neutrophilic inflammation.

Protective role of the capsule and impact of serotype 4 switching on Streptococcus mitis.

The polysaccharide capsule surrounding Streptococcus pneumoniae is essential for virulence. Recently, Streptococcus mitis, a human commensal and a close relative of S. pneumoniae, was also shown to have a capsule. In this study, the S. mitis type strain switched capsule by acquisition of the serotype 4 capsule locus of S. pneumoniae TIGR4, following induction of competence for natural transformation. Comparison of the wild type with the capsule-switching mutant and with a capsule deletion mutant showed that the capsule protected S. mitis against phagocytosis by RAW 264.7 macrophages. This effect was enhanced in the S. mitis strain expressing the S. pneumoniae capsule, which showed, in addition, increased resistance against early clearance in a mouse model of lung infection. Expression of both capsules also favored survival in human blood, and the effect was again more pronounced for the capsule-switching mutant. S. mitis survival in horse blood or in a mouse model of bacteremia was not significantly different between the wild type and the mutant strains. In all models, S. pneumoniae TIGR4 showed higher rates of survival than the S. mitis type strain or the capsule-switching mutant, except in the lung model, in which significant differences between S. pneumoniae TIGR4 and the capsule-switching mutant were not observed. Thus, we identified conditions that showed a protective function for the capsule in S. mitis. Under such conditions, S. mitis resistance to clearance could be enhanced by capsule switching to serotype 4, but it was enhanced to levels lower than those for the virulent strain S. pneumoniae TIGR4.

Proteinase-activated receptors in fibroproliferative lung disease.

The coagulation cascade plays a central role in the pathogenesis of fibroproliferative lung diseases such as the acute respiratory distress syndrome (ARDS) and idiopathic pulmonary fibrosis (IPF) through multifaceted effects on haemostasis, inflammation and tissue repair. However, targeting the coagulation cascade using traditional anticoagulant approaches has not resulted in improved outcomes for these patients. The cellular effects of the coagulation cascade are mediated via a family of four proteinase-activated receptors (PAR(1-4)). PARs are G protein-coupled receptors that have a unique method of activation involving proteolytic cleavage. They play key roles in mediating the interplay between coagulation and inflammation and tissue repair and fibrosis. Current evidence suggests a central role for PAR(1) and PAR(2) in influencing these responses, although data from animal models suggest that their contribution is highly dependent on both the nature of the insult and disease status. Nonetheless, these receptors may represent important targets in conditions associated with uncontrolled coagulation signalling responses including IPF, ARDS, asthma and chronic obstructive pulmonary disease.

Review of the British Thoracic Society Winter Meeting 2013, 4-6 December, London, UK.

This article reviews the British Thoracic Society Winter Meeting 2013, the annual scientific meeting attended by over 2000 delegates and representing the depth and breadth of UK respiratory medicine. This year's meeting from 4 to 6 December in London featured cutting-edge research alongside keynote symposia from international experts in respiratory science, epidemiology and clinical trials. This article reviews the key symposia and selected abstract sessions from the 2013 meeting.

Anesthesia for bronchoscopy.

PURPOSE OF REVIEW: To discuss the recent advances in sedation and anesthesia for the practice of both flexible and rigid bronchoscopy, which are increasingly performed outside of the operating room by interventional pulmonologists and thoracic surgeons. RECENT FINDINGS: Studies have recently documented the advantages of pharmacological sedatives and anesthetics for use in bronchoscopy. In particular, we review the increasing body of literature highlighting the advantages and benefits of propofol anesthesia for both flexible and rigid bronchoscopy. SUMMARY: As our practice expands, relocation of appropriately triaged pulmonary interventional procedures including rigid bronchoscopy that were previously assigned to a traditional operating room setting improves provider flexibility, presents more cost-effective options while maintaining patient safety and satisfaction and reducing the time to recovery. Anesthesia practice has, therefore, shifted to caring for these sick patients outside the operating room and increasingly cooperation between anesthesiologist and proceduralist is required.

Emerging therapies and respiratory infections: Focus on the impact of immunosuppressants and immunotherapies.

Host defences to infection are based upon an integrated system of physical and biochemical barriers, innate and adaptive immunity. Weakness in any of these defensive elements leads to increased susceptibility to specific pathogens. Understanding how medical therapies disrupt host defences is key to the successful prevention, diagnosis and management of respiratory infection in the immunocompromised host.