scholarly journals Bactericidal/Permeability-Increasing Protein Preeminently Mediates Clearance of Pseudomonas aeruginosa In Vivo via CD18-Dependent Phagocytosis

2021 ◽  
Vol 12 ◽  
Author(s):  
Jomkuan Theprungsirikul ◽  
Sladjana Skopelja-Gardner ◽  
Ashley S. Burns ◽  
Rachel M. Wierzbicki ◽  
William F. C. Rigby
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Critical Role ◽  
Chronic Obstructive ◽  
Dependent Manner ◽  
Opsonic Activity ◽  
Obstructive Pulmonary Disease ◽  
Neutrophil Dysfunction ◽  
Chronic Lung Infection

Chronic Pseudomonas aeruginosa infection mysteriously occurs in the airways of patients with cystic fibrosis (CF), bronchiectasis (BE), and chronic obstructive pulmonary disease (COPD) in the absence of neutrophil dysfunction or neutropenia and is strongly associated with autoimmunity to bactericidal permeability-increasing protein (BPI). Here, we define a critical role for BPI in in vivo immunity against P. aeruginosa. Wild type and BPI-deficient (Bpi-/-) mice were infected with P. aeruginosa, and bacterial clearance, cell infiltrates, cytokine production, and in vivo phagocytosis were quantified. Bpi-/- mice exhibited a decreased ability to clear P. aeruginosa in vivo in concert with increased neutrophil counts and cytokine release. Bpi-/- neutrophils displayed decreased phagocytosis that was corrected by exogenous BPI in vitro. Exogenous BPI also enhanced clearance of P. aeruginosa in Bpi-/- mice in vivo by increasing P. aeruginosa uptake by neutrophils in a CD18-dependent manner. These data indicate that BPI plays an essential role in innate immunity against P. aeruginosa through its opsonic activity and suggest that perturbations in BPI levels or function may contribute to chronic lung infection with P. aeruginosa.

scholarly journals Aryl Hydrocarbon Receptor Defect Attenuates Mitogen-Activated Signaling Through Leucine-Rich Repeats and Immunoglobulin-Like Domains 1 (LRIG1)-Dependent EGFR Degradation

2021 ◽  
Vol 22 (18) ◽  
pp. 9988
Author(s):  
Han-Lin Hsu ◽  
Hong-Kai Chen ◽  
Chi-Hao Tsai ◽  
Po-Lin Liao ◽  
Yen-Ju Chan ◽  
...  
Keyword(s):  
Aryl Hydrocarbon Receptor ◽  
Egf Receptor ◽  
Chronic Obstructive ◽  
Dependent Manner ◽  
Obstructive Pulmonary Disease ◽  
Aryl Hydrocarbon ◽  
Copd Patients ◽  
Leucine Rich Repeats

Aryl hydrocarbon receptor (AHR) genomic pathway has been well-characterized in a number of respiratory diseases. In addition, the cytoplasmic AHR protein may act as an adaptor of E3 ubiquitin ligase. In this study, the physiological functions of AHR that regulate cell proliferation were explored using the CRISPR/Cas9 system. The doubling-time of the AHR-KO clones of A549 and BEAS-2B was observed to be prolonged. The attenuation of proliferation potential was strongly associated with either the induction of p27Kip1 or the impairment in mitogenic signal transduction driven by the epidermal growth factor (EGF) and EGF receptor (EGFR). We found that the leucine-rich repeats and immunoglobulin-like domains 1 (LRIG1), a repressor of EGFR, was induced in the absence of AHR in vitro and in vivo. The LRIG1 tends to degrade via a proteasome dependent manner by interacting with AHR in wild-type cells. Either LRIG1 or a disintegrin and metalloprotease 17 (ADAM17) were accumulated in AHR-defective cells, consequently accelerating the degradation of EGFR, and attenuating the response to mitogenic stimulation. We also affirmed low AHR but high LRIG1 levels in lung tissues of chronic obstructive pulmonary disease (COPD) patients. This might partially elucidate the sluggish tissue repairment and developing inflammation in COPD patients.

scholarly journals Cross-talk between PKA-Cβ and p65 mediates synergistic induction of PDE4B by roflumilast and NTHi

2015 ◽  
Vol 112 (14) ◽  
pp. E1800-E1809 ◽  
Author(s):  
Seiko Susuki-Miyata ◽  
Masanori Miyata ◽  
Byung-Cheol Lee ◽  
Haidong Xu ◽  
Hirofumi Kai ◽  
...  
Keyword(s):  
Cross Talk ◽  
Nuclear Factor Κb ◽  
Airway Epithelial Cells ◽  
Pde4 Inhibitor ◽  
Chronic Obstructive ◽  
Dependent Manner ◽  
Obstructive Pulmonary Disease ◽  
Synergistic Induction

Phosphodiesterase 4B (PDE4B) plays a key role in regulating inflammation. Roflumilast, a phosphodiesterase (PDE)4-selective inhibitor, has recently been approved for treating severe chronic obstructive pulmonary disease (COPD) patients with exacerbation. However, there is also clinical evidence suggesting the development of tachyphylaxis or tolerance on repeated dosing of roflumilast and the possible contribution of PDE4B up-regulation, which could be counterproductive for suppressing inflammation. Thus, understanding how PDE4B is up-regulated in the context of the complex pathogenesis and medications of COPD may help improve the efficacy and possibly ameliorate the tolerance of roflumilast. Here we show that roflumilast synergizes with nontypeable Haemophilus influenzae (NTHi), a major bacterial cause of COPD exacerbation, to up-regulate PDE4B2 expression in human airway epithelial cells in vitro and in vivo. Up-regulated PDE4B2 contributes to the induction of certain important chemokines in both enzymatic activity-dependent and activity-independent manners. We also found that protein kinase A catalytic subunit β (PKA-Cβ) and nuclear factor-κB (NF-κB) p65 subunit were required for the synergistic induction of PDE4B2. PKA-Cβ phosphorylates p65 in a cAMP-dependent manner. Moreover, Ser276 of p65 is critical for mediating the PKA-Cβ–induced p65 phosphorylation and the synergistic induction of PDE4B2. Collectively, our data unveil a previously unidentified mechanism underlying synergistic up-regulation of PDE4B2 via a cross-talk between PKA-Cβ and p65 and may help develop new therapeutic strategies to improve the efficacy of PDE4 inhibitor.

OsaR (PA0056) functions as a repressor of the gene fleQ encoding an important motility regulator in Pseudomonas aeruginosa

2021 ◽  
Author(s):  
Yibing Ma ◽  
Yujie Liu ◽  
Yutong Bi ◽  
Xiao Han ◽  
Yongxin Jin ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Binding Site ◽  
Biofilm Formation ◽  
Transcriptional Regulator ◽  
Regulatory Mechanisms ◽  
Chronic Obstructive ◽  
Transcriptional Level ◽  
Obstructive Pulmonary Disease

FleQ plays a crucial role in motility and biofilm formation by regulating flagellar and exopolysaccharide biosynthesis in Pseudomonas aeruginosa . It has been reported that the expression of FleQ is transcriptionally downregulated by the virulence factor regulator Vfr. Herein we demonstrated that a LysR-type transcriptional regulator, OsaR, is also capable of binding to the promoter region of fleQ and repressing its transcription. Through gel shift and DNase I footprinting assays, the OsaR binding site was identified and characterized as a dual LysR-type transcriptional regulator box (AT-N 11 -AT-N 7 -A-N 11 -T). Mutation of the A-T palindromic base pairs in fleQ promoter not only reduced the binding affinity of OsaR in vitro , but also de-repressed fleQ transcription in vivo . The OsaR binding site was found to cover the Vfr binding site; knockout of osaR or vfr separately exhibited no effect on the transcriptional level of fleQ ; however, fleQ expression was repressed by overexpression of osaR or vfr . Furthermore, simultaneously deleting both osaR and vfr resulted in an upregulation of fleQ , but it could be complemented by the expression of either of the two repressors. In summary, our work revealed that OsaR and Vfr function as two transcriptional repressors of fleQ that bind to the same region of fleQ but work separately. IMPORTANCE Pseudomonas aeruginosa is a widespread human pathogen, which accounts for serious infections in the hospital, especially for lung infection in cystic fibrosis and chronic obstructive pulmonary disease patients. P. aeruginosa infection is closely associated with its motility and biofilm formation, which are both under the regulation of the important transcription factor FleQ. However, the upstream regulatory mechanisms of fleQ have not been fully elucidated. Therefore, our research identifying a novel regulator of fleQ as well as new regulatory mechanisms controlling its expression will be significant for better understanding the intricate gene regulatory mechanisms related to P. aeruginosa virulence and infection.

Allyl isothiocyanate upregulates MRP1 expression through Notch1 signaling in human bronchial epithelial cells

2020 ◽  
Vol 98 (5) ◽  
pp. 324-331
Author(s):  
Ni-ni Li ◽  
Yan Guo ◽  
Cheng-jun Jiang ◽  
Yuan-yuan Zhou ◽  
Chen-hui Li ◽  
...  
Keyword(s):  
Critical Role ◽  
Allyl Isothiocyanate ◽  
Chronic Obstructive ◽  
Therapeutic Effects ◽  
Mobility Shift ◽  
Obstructive Pulmonary Disease ◽  
Suppression Effect ◽  
Notch1 Signaling ◽  
Mrp1 Expression

Multidrug resistance associated protein-1 (MRP1) and Notch signaling are closely related and both play a critical role in chronic obstructive pulmonary disease (COPD) establishment and progression. The aim of our work was to test whether Notch1 is involved in allyl isothiocyanate (AITC) induced MRP1 expression. We used cigarette smoke extract (CSE) to simulate the smoking microenvironment in vitro. The results demonstrated that CSE led to apoptosis as well as reduced the expression of Notch1, Hes1, and MRP1, while AITC significantly reversed this downregulation. Transfected with Notch1 siRNA downregulated MRP1 expression and activity, aggravated the suppression effect by CSE, and abolished the AITC-induced Notch1, Hes1, and MRP1 expression. Validation of the correlation between Notch1 and MRP1 was implemented by gel-shift assays (electrophoretic mobility shift assay). The result revealed an interaction between a specific promoter region of MRP1 and the intracellular domain of Notch1. In conclusion, Notch1 signaling positively regulated MRP1 in 16HBE cells and AITC induced MRP1 expression and function may be attributed to Notch1 signaling. These findings show that Notch1 and MRP1 might have a potential protective effect in the COPD process and become a new therapeutic target for COPD or other lung diseases. It also provides a theoretical basis for the therapeutic effects of AITC.

Interaction and Regulation of HSC with Osteopontin Is Mediated through α4β1 and α9β1Integrins, Which Are Differentially Expressed in the HSC Pool.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1330-1330
Author(s):  
David N. Haylock ◽  
Genevieve A. Whitty ◽  
Brenda Williams ◽  
Melonie J. Storan ◽  
Susie K. Nilsson
Keyword(s):  
Cord Blood ◽  
Critical Role ◽  
Negative Regulator ◽  
Hemopoietic Stem Cell ◽  
Null Mouse ◽  
Dependent Manner ◽  
Independent Manner ◽  
Hsc Niche

Abstract Osteoblasts are a key cellular component of the hemopoietic stem cell (HSC) niche and directly regulate the HSC pool. Molecules synthesised by osteoblasts both promote or inhibit HSC proliferation. Osteopontin (Opn) is an osteoblast produced, RGD containing protein with roles in cell adhesion and migration. Until recently, the role of Opn in hemopoiesis was seen as restricted to the regulation of bone turnover. However, from analysis of hemopoiesis in the Opn null mouse, we have demonstrated that Opn plays a critical role in regulating the HSC pool. Furthermore Opn is critical in trans-marrow migration and lodgement of HSC within the BM after transplantation. When added to in vitro HSC cultures, exogenous thrombin-cleaved Opn also inhibits cell proliferation and potently suppresses HSC differentiation. We have now demonstrated that this interaction occurs in an RGD-independent manner via the cryptic SVVYGLR epitope revealed on the N-terminal fragment of Opn following thrombin cleavage. This epitope has previously been shown to bind to α4β1 and α9β1. HSC are known to express α4β1, but we have now shown that within the HSC pool this occurs in a differential manner, mimicking that of CD38, with more committed CD34+CD38+ cord blood progenitors having the highest levels of expression. In addition, we have shown the previously unrecognised characteristic of human marrow and cord blood HSC, the expression of α9β1, which also occurs in a differential manner, but mimicking CD34. Expression of α9β1 is highest on cord blood CD34+CD38− cells, a population highly enriched for HSC. Using the synthetic SVVYGLR peptide in culture, we re-capitulated the thrombin-cleaved Opn induced suppression of HSC differentiation in a dose dependent manner. Antibody blocking experiments demonstrated that binding to this peptide was occurring through both α4β1 and α9β1. In contrast, suppression of HSC proliferation and differentiation did not occur through the upstream alternate α4β1 binding site. Furthermore, we have now demonstrated endogenous binding of Opn to α4β1 and α9β1 to cord blood HSC in vivo. Together, these data provide strong evidence that Opn is an important component of the HSC niche which acts as a physiological negative regulator. Furthermore, our studies identify the previously unrecognised characteristic of HSC, the expression of α9β1, which together with α4β1 provides two receptors on HSC with differing expression signatures and potentially a mechanism for fine tunning the physiological effects of Opn.

Quantitative Phosphoproteomics Identified a New Syk-Lyn-Axl Signalling Pathway Involved In Resistance to Nilotinib In Chronic Myeloid Leukemia Cells.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3376-3376
Author(s):  
Romain Gioia ◽  
Cedric Leroy ◽  
Claire Drullion ◽  
Valérie Lagarde ◽  
Serge Roche ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Tyrosine Kinase ◽  
Myeloid Leukemia ◽  
Conflicts Of Interest ◽  
Critical Role ◽  
Dependent Manner ◽  
Stable Isotope Labelling ◽  
Resistant Cells

Abstract Abstract 3376 Nilotinib has been developed to overcome resistance to imatinib, the first line treatment of chronic myeloid leukemia (CML). To anticipate resistance to nilotinib, we generate nilotinib resistant CML cell lines in vitro to characterize mechanisms and signaling pathways that may contribute to resistance. Among the different mechanisms of resistance identified, the overexpression of the Src-kinase Lyn was involved in resistance both in vitro, in a K562 cell line (K562-rn), and in vivo, in nilotinib-resistant CML patients. To characterize how Lyn mediates resistance, we performed a phosphoproteomic study using SILAC (Stable Isotope Labelling with Amino acid in Cell culture). Quantification and identification of phosphotyrosine proteins in the nilotinib resistant cells point out two tyrosine kinases, the spleen tyrosine kinase Syk and the UFO receptor Axl. The two tyrosine kinase Syk and Axl interact with Lyn as seen by coimmunopreciptation. Syk is phosphorylated on tyrosine 323 and 525/526 in Lyn dependent manner in nilotinib resistant cells. The inhibition of Syk tyrosine kinase by R406 or BAY31-6606 restores sensitivity to nilotinib in K562-rn cells. In parallel, the inhibition of Syk expression by ShRNA in K562-rn cells abolishes Lyn and Axl phosphorylation and then interaction between Lyn and Axl leading to a full restoration of nilotinib efficacy. In the opposite, the coexpression of Lyn and Syk in nilotinib sensitive K562 cells induced resistance to nilotinib whereas a Syk kinase dead mutant did not. These results highlight for the first time the critical role of Syk in resistance to tyrosine kinase inhibitors in CML disease emphasizing the therapeutic targeting of this tyrosine kinase. Moreover, Axl, which is already a target in solid tumor, will be also an interesting pathway to target in CML. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Critical role for Epac1 in inflammatory pain controlled by GRK2-mediated phosphorylation of Epac1

2016 ◽  
Vol 113 (11) ◽  
pp. 3036-3041 ◽  
Author(s):  
Pooja Singhmar ◽  
XiaoJiao Huo ◽  
Niels Eijkelkamp ◽  
Susana Rojo Berciano ◽  
Faiza Baameur ◽  
...  
Keyword(s):  
Chronic Pain ◽  
Molecular Mechanism ◽  
Inflammatory Pain ◽  
Critical Role ◽  
Specific Inhibitor ◽  
Mechanical Hyperalgesia ◽  
Dependent Manner ◽  
Phosphorylation Event

cAMP signaling plays a key role in regulating pain sensitivity. Here, we uncover a previously unidentified molecular mechanism in which direct phosphorylation of the exchange protein directly activated by cAMP 1 (EPAC1) by G protein kinase 2 (GRK2) suppresses Epac1-to-Rap1 signaling, thereby inhibiting persistent inflammatory pain. Epac1−/− mice are protected against inflammatory hyperalgesia in the complete Freund’s adjuvant (CFA) model. Moreover, the Epac-specific inhibitor ESI-09 inhibits established CFA-induced mechanical hyperalgesia without affecting normal mechanical sensitivity. At the mechanistic level, CFA increased activity of the Epac target Rap1 in dorsal root ganglia of WT, but not of Epac1−/−, mice. Using sensory neuron-specific overexpression of GRK2 or its kinase-dead mutant in vivo, we demonstrate that GRK2 inhibits CFA-induced hyperalgesia in a kinase activity-dependent manner. In vitro, GRK2 inhibits Epac1-to-Rap1 signaling by phosphorylation of Epac1 at Ser-108 in the Disheveled/Egl-10/pleckstrin domain. This phosphorylation event inhibits agonist-induced translocation of Epac1 to the plasma membrane, thereby reducing Rap1 activation. Finally, we show that GRK2 inhibits Epac1-mediated sensitization of the mechanosensor Piezo2 and that Piezo2 contributes to inflammatory mechanical hyperalgesia. Collectively, these findings identify a key role of Epac1 in chronic inflammatory pain and a molecular mechanism for controlling Epac1 activity and chronic pain through phosphorylation of Epac1 at Ser-108. Importantly, using the Epac inhibitor ESI-09, we validate Epac1 as a potential therapeutic target for chronic pain.

scholarly journals Neutrophil proteases alter the interleukin-22-receptor-dependent lung antimicrobial defence

2015 ◽  
Vol 46 (3) ◽  
pp. 771-782 ◽  
Author(s):  
Antoine Guillon ◽  
Youenn Jouan ◽  
Deborah Brea ◽  
Fabien Gueugnon ◽  
Emilie Dalloneau ◽  
...  
Keyword(s):  
Chronic Obstructive ◽  
Dependent Lung ◽  
Obstructive Pulmonary Disease ◽  
Copd Exacerbations ◽  
Interleukin 22 ◽  
Copd Patients ◽  
Acute Exacerbations ◽  
Underlying Mechanisms

Chronic obstructive pulmonary disease (COPD) is punctuated by episodes of infection-driven acute exacerbations. Despite the life-threatening nature of these exacerbations, the underlying mechanisms remain unclear, although a high number of neutrophils in the lungs of COPD patients is known to correlate with poor prognosis. Interleukin (IL)-22 is a cytokine that plays a pivotal role in lung antimicrobial defence and tissue protection. We hypothesised that neutrophils secrete proteases that may have adverse effects in COPD, by altering the IL-22 receptor (IL-22R)-dependent signalling.Using in vitro and in vivo approaches as well as reverse transcriptase quantitative PCR, flow cytometry and/or Western blotting techniques, we first showed that pathogens such as the influenza virus promote IL-22R expression in human bronchial epithelial cells, whereas Pseudomonas aeruginosa, bacterial lipopolysaccharide or cigarette smoke do not. Most importantly, neutrophil proteases cleave IL-22R and impair IL-22-dependent immune signalling and expression of antimicrobial effectors such as β-defensin-2. This proteolysis resulted in the release of a soluble fragment of IL-22R, which was detectable both in cellular and animal models as well as in sputa from COPD patients with acute exacerbations.Hence, our study reveals an unsuspected regulation by the proteolytic action of neutrophil enzymes of IL-22-dependent lung host response. This process probably enhances pathogen replication, and ultimately COPD exacerbations.

scholarly journals Defining tissue- and disease-associated macrophages using a transcriptome-based classification model

2019 ◽  
Author(s):  
Hung-Jen Chen ◽  
Andrew Y.F. Li Yim ◽  
Guillermo R. Griffith ◽  
Wouter J. de Jonge ◽  
Marcel M.A.M. Mannens ◽  
...  
Keyword(s):  
Expression Profiles ◽  
Interleukin 10 ◽  
Interferon Γ ◽  
Classification Model ◽  
Chronic Obstructive ◽  
Activated Macrophages ◽  
Obstructive Pulmonary Disease ◽  
Disease Specific

AbstractMacrophages are heterogeneous multifunctional leukocytes which are regulated in a tissue-and disease-specific context. Many different studies have been published using in vitro macrophage models to study disease. Here, we aggregated public expression data to define consensus expression profiles for eight commonly-used in vitro macrophage models. Altogether, we observed well-known but also novel markers for different macrophage subtypes. Using these data we subsequently built the classifier macIDR, capable of distinguishing macrophage subsets with high accuracy (>0.95). This classifier was subsequently applied to transcriptional profiles of tissue-isolated and disease-associated macrophages to specifically define macrophage characteristics in vivo. Classification of these in vivo macrophages showed that alveolar macrophages displayed high resemblance to interleukin-10 activated macrophages, whereas macrophages from patients with chronic obstructive pulmonary disease patients displayed a drop in interferon-γ signature. Adipose tissue-derived macrophages were classified as unstimulated macrophages, but resembled LPS-activated macrophages more in diabetic-obese patients. Finally, rheumatoid arthritic synovial macrophages showed characteristics of both interleukin-10 or interferon-γ signatures. Altogether, our results suggest that macIDR is capable of identifying macrophage-specific changes as a result of tissue-and disease-specific stimuli and thereby can be used to better define and model populations of macrophages that contribute to disease.

Sign in / Sign up

Export Citation Format

Share Document