C5a anaphylatoxin is a major regulator of activating versus inhibitory FcγRs in immune complex–induced lung disease

Diverse clinical factors, including intestinal ischemia, contribute to acute lung injury (ALI), which has up to a 40% mortality rate. During the development of lung injury an immune response is elicited that exacerbates the lung insult. Neutrophils have been well studied in mediating the pulmonary insults through an assortment of mechanisms, such as release of granule contents and production of proinflammatory cytokines due to the overactivation of complement and cytokines. In this study, we found that enhanced endoplasmic reticulum (ER) stress was observed in infiltrated neutrophils in the early stage of an ALI mice model. In neutrophils, complement 5a (C5a) inspires strong ER stress through inositol-requiring kinase 1a and, to a less extent, the protein kinase R–like ER kinase signaling pathway. The granule release induced by C5a was ER stress mediated. Knowkdown of X-box–binding protein 1, a downstream signaling molecule of inositol-requiring kinase 1a, impaired granule release, based on myeloperoxidase production. Further analysis revealed that C5a induced ER stress by binding to C5a receptor in neutrophils. Using xbpf/f MRP8-cre mice in which X-box–binding protein 1 is deficient specifically in neutrophils and ER stress is deprived, we confirmed that ER stress in neutrophils was required for granule release in vivo and led to ALI, whereas dampening ER stress in neutrophils substantially alleviated ALI. Taken together, our results demonstrated that C5a receptor–mediated ER stress induced granule release in neutrophils, contributing to the development of ALI. This novel mechanism suggests a new potential therapeutic target in autophagy regulation for ALI.

Section: Discussionmentioning

confidence: 99%

Endoplasmic Reticulum Stress of Neutrophils Is Required for Ischemia/Reperfusion–Induced Acute Lung Injury

Chen

Jia

et al. 2015

“…Monocyte/macrophage cell counts and flow cytometry CD11b-positive liver macrophages as well as lavaged macrophages from lung and peritoneum were isolated and quantitated as described previously (26)(27)(28). Blood was obtained from anesthetized mice by puncture of the retro-orbital plexus, transferred to EDTA-coated tubes, and counted for blood monocytes by the automate Animal Blood Counter (Scil Animal Care, Viernheim, Germany).…”

Section: Micementioning

confidence: 99%

Defective Macrophage Migration in Gαi2- but Not Gαi3-Deficient Mice

Wiege

Syed

et al. 2012

Self Cite

Various heterotrimeric Gi proteins are considered to be involved in cell migration and effector function of immune cells. The underlying mechanisms, how they control the activation of myeloid effector cells, are not well understood. To elucidate isoform-redundant and -specific roles for Gαi proteins in these processes, we analyzed mice genetically deficient in Gαi2 or Gαi3. First, we show an altered distribution of tissue macrophages and blood monocytes in the absence of Gαi2 but not Gαi3. Gαi2-deficient but not wild-type or Gαi3-deficient mice exhibited reduced recruitment of macrophages in experimental models of thioglycollate-induced peritonitis and LPS-triggered lung injury. In contrast, genetic ablation of Gαi2 had no effect on Gαi-dependent peritoneal cytokine production in vitro and the phagocytosis-promoting function of the Gαi-coupled C5a anaphylatoxin receptor by liver macrophages in vivo. Interestingly, actin rearrangement and CCL2- and C5a anaphylatoxin receptor-induced chemotaxis but not macrophage CCR2 and C5a anaphylatoxin receptor expression were reduced in the specific absence of Gαi2. Furthermore, knockdown of Gαi2 caused decreased cell migration and motility of RAW 264.7 cells, which was rescued by transfection of Gαi2 but not Gαi3. These results indicate that Gαi2, albeit redundant to Gαi3 in some macrophage activation processes, clearly exhibits a Gαi isoform-specific role in the regulation of macrophage migration.

“…Cellfree supernatants were collected after 15, 30, and 60 min and were analyzed for myeloperoxidase (MPO) activity as described (30). Briefly, 20 ml supernatant was incubated with 230 ml 0.167 mg/ml o-dianisidine dihydrochloride solution (Sigma-Aldrich, Munich, Germany) with 160 mM hydrogen peroxide (Sigma-Aldrich, Munich, Germany) for 15 min at room temperature.…”

Section: Animalsmentioning

confidence: 99%

Protein Kinase C-θ Is Required for Murine Neutrophil Recruitment and Adhesion Strengthening under Flow

Bertram

Zhang

Vietinghoff

et al. 2012

Self Cite

Protein kinase C (PKC)-θ is involved in T cell activation via regulating the avidity of the β2 integrin LFA-1 in the immunological synapse. LFA-1 also mediates leukocyte adhesion. To investigate the role of PKC-θ in neutrophil adhesion, we performed intravital microscopy in cremaster venules of mice reconstituted with bone marrow from LysM-GFP+ (wild-type [WT]) and PKC-θ gene-deficient (Prkcq−/−) mice. Following stimulation with CXCL1, both WT and Prkcq−/− cells became adherent. Although most WT neutrophils remained adherent for at least 180 s, 50% of Prkcq−/− neutrophils were detached after 105 s and most by 180 s. Upon CXCL1 injection, rolling of all WT neutrophils stopped for 90 s, but rolling of Prkcq−/− neutrophils started 30 s after CXCL1 stimulation. A similar neutrophil adhesion defect was seen in vitro, and spreading of Prkcq−/− neutrophils was delayed. Prkcq−/− neutrophil recruitment was impaired in fMLP-induced transmigration into the cremaster muscle, thioglycollate-induced peritonitis, and LPS-induced lung injury. We conclude that PKC-θ mediates integrin-dependent neutrophil functions and is required to sustain neutrophil adhesion in postcapillary venules in vivo. These findings suggest that the role of PKC-θ in outside–in signaling following engagement of neutrophil integrins is relevant for inflammation in vivo.