Inactivation of Rab11a GTPase in Macrophages Facilitates Phagocytosis of Apoptotic Neutrophils

Abstract

The timely and efficient clearance of apoptotic neutrophils by macrophages (efferocytosis) is required for the resolution of inflammation and tissue repair, but the regulatory mechanisms remain unclear. Here, we investigated the role of the small GTPase Rab11a in regulating efferocytosis, and on this basis the resolution of inflammatory lung injury. We observed that apoptotic neutrophil feeding induced a rapid loss of Rab11a activity in bone marrow-derived macrophages and found that depletion of Rab11a in macrophages by siRNA dramatically increased the phagocytosis of apoptotic neutrophils compared with control cells. In addition, overexpression of wild type Rab11a inhibited macrophage efferocytosis, whereas overexpression of dominant negative Rab11a (Rab11a S25N) increased the clearance of apoptotic neutrophils. Rab11a knockdown also increased the surface level of CD36 in macrophages, but reduced cell surface expression of a disintegrin and metalloproteinase (ADAM) 17. Depletion of ADAM17 rescued the decreased surface CD36 expression found in macrophages over-expressing wild type Rab11a. Also, blockade of CD36 abolished the augmented efferocytosis seen in Rab11a-depleted macrophages. In mice challenged with endotoxin, intratracheal instillation of Rab11a-depleted macrophages reduced neutrophil count in bronchoalveolar lavage fluid, increased the number of macrophages containing apoptotic neutrophils, and prevented inflammatory lung injury. Thus, Rab11a inactivation in macrophages as result of apoptotic cell binding initiates phagocytosis of apoptotic neutrophils via the modulation of ADAM17-mediated CD36 cell surface expression. Our results raise the possibility that inhibition of Rab11a activity in macrophages is a promising strategy for activating the resolution of inflammatory lung injury.

The GTPase Rab1 Is Required for NLRP3 Inflammasome Activation and Inflammatory Lung Injury

“…Using an established in vivo model (27, 43), we next determined whether macrophage Rab1a activation plays an essential role in lung inflammation and injury. Mice were depleted of alveolar macrophages (Supplemental Fig.…”

“…instillation of clodronate liposome (100 μl, 1:2 dilution in PBS) to anesthetized (ketamine 100 mg/kg, i.p.) mice (27). PBS or BMDMs transfected with a vector, Rab1a WT, or Rab1a N124I cDNA (2×10 6 cells, 200 μl of total volume each) were given to alveolar macrophage–depleted mice via i.t.…”

“…The lungs were cut into 4–5 μm thick sections, stained with H&E, and determined by light microscopy. Lung injury scores were evaluated by an investigator blinded to the treatment groups using the histological semiquantitative scoring system as described previously (27).…”

Emodin alleviates LPS-induced inflammatory response in lung injury rat by affecting the function of granulocytes

“…The resolution of an acute inflammatory response requires subsequent phagocytosis of apoptotic granulocytes by macrophages. Delayed apoptosis of activated granulocytes can lead to persistent acute lung inflammation and can eventually develop into one of the mechanisms of ARDS [35]. In this experiment, we mainly compared the effects of different doses of Emo on LPS-stimulated granulocyte apoptosis.…”

Emodin alleviates LPS-induced inflammatory response in lung injury rat by affecting the function of neutrophils

“…The resolution of an acute inflammatory response requires subsequent phagocytosis of apoptotic neutrophils by macrophages. Delayed apoptosis of activated neutrophils can lead to persistent acute lung inflammation and can eventually develop into one of the mechanisms of ARDS [35] . In this experiment, we mainly compared the effects of different doses of Emo on LPS-stimulated neutrophil apoptosis.…”