Stimulation of Ly-6G on neutrophils in LPS-primed mice induces platelet-activating factor (PAF)-mediated anaphylaxis-like shock

Abstract

Previously, two anti-Ly-6G mAb-RB6-8C5 and 1A8-have been used to deplete neutrophils in mice and to clarify their involvement in immune responses. During the course of experiments on neutrophil depletion, we noticed that i.v. injection of RB6-8C5 or 1A8 induced anaphylaxis-like shock in mice pretreated i.v. with LPS. Signs of shock, such as hypothermia, appeared within a few minutes, and the mice died of shock within 20 min of the antibody injection. In vivo experiments, including depletion of various cell types, indicated that neutrophils and macrophages (but not platelets, basophils, or mast cells) are involved in the shock. Experiments using various drugs and gene-targeted mice demonstrated that PAF is the central mediator of the shock. Optimal LPS priming required at least 1 h, and the priming was associated with neutrophil accumulation within pulmonary and hepatic blood vessels. Consistently, following 1A8 injection into LPS-pretreated mice, the mRNA for LysoPAFAT (a PAF biosynthetic enzyme) was markedly up-regulated in neutrophils accumulated in the lung but not in macrophages. These results suggest that (1) stimulation of Ly-6G on LPS-primed neutrophils induces PAF-mediated anaphylaxis-like shock in mice, (2) neutrophils are primed by LPS during and/or after their accumulation in lung and liver to rapidly induce LysoPAFAT, and (3) macrophages may play a pivotal role in the priming phase and/or in the challenge phase by unknown mechanisms. These findings may be related to adult respiratory distress syndrome, although the natural ligand for Ly-6G remains to be identified.

Anti‐interleukin‐33 inhibits cigarette smoke‐induced lung inflammation in mice

“…Neutrophils and macrophages are the key cell components in CS‐induced airway inflammation and play a critical role in the pathogenesis of COPD . We therefore further assess the effect of anti‐IL‐33 on the CS‐induced neutrophil and macrophage infiltration in lung tissue by immunohistochemical staining of Ly‐6G and Mac‐3, surface markers for tissue neutrophils and macrophages, respectively …”

“…1,2,5,8 We therefore further assess the effect of anti-IL-33 on the CSinduced neutrophil and macrophage infiltration in lung tissue by immunohistochemical staining of Ly-6G and Mac-3, surface markers for tissue neutrophils and macrophages, respectively. 27,28 . Ly-6G staining showed a significant increase in the levels of Ly-6G + neutrophils in the lung tissue of CS-exposed mice compared with controls (p < 0Á01).…”

Involvement of IL-1 in the Maintenance of Masseter Muscle Activity and Glucose Homeostasis

“…It is a well-known phenomenon that typical IL-1 producing cells such as neutrophils and monocytes are recruited into skeletal muscle tissues during the exercise [ 44 ]. Herein, Gr-1, a major marker for granulocytes, could be clearly distinguished in IL-1α/β-producing cells; IL-1β-positive cells co-expressing Gr-1 signals were concluded to be migrating neutrophils [ 38 ]. However, IL-1α-positive cells acted as other immune cells like mast cells, but not neutrophils, owing to their absence of Gr-1 signals [ 38 , 53 ].…”

“…Herein, Gr-1, a major marker for granulocytes, could be clearly distinguished in IL-1α/β-producing cells; IL-1β-positive cells co-expressing Gr-1 signals were concluded to be migrating neutrophils [ 38 ]. However, IL-1α-positive cells acted as other immune cells like mast cells, but not neutrophils, owing to their absence of Gr-1 signals [ 38 , 53 ]. Considering the above observations, it would be of great interest to further identify the phenotype(s) of the IL-1-positive cells and the mechanism underlying their recruitment, as well as the precise role of the IL-1-positive cells in muscle physiology in future studies.…”

“…Next, the sections were incubated with a secondary biotinylated goat anti-rabbit antibody (1: 1,000 dilution; Vector Laboratories, Tokyo, Japan), and then visualized using the Vectastain ABC kit (Vector Laboratories). Fluorescent double-labeled IHC was carried out using the primary antibody for granulocyte-differentiation antigen (Gr)-1 (1:500 dilution; BioLegend, San Diego, CA, USA, clone RB6-8C5), a major marker for granulocytes and partially for subsets of endothelial cells [ 38 , 39 ], and DAPI counterstaining. The secondary antibodies used were Alexa 488 -conjugated goat-anti rabbit and Alexa 555 -conjugated goat anti-rat (1:750 dilution; Molecular Probes, Eugene, OR, USA).…”