DNase-1 Treatment Exerts Protective Effects in Neurogenic Pulmonary Edema via Regulating the Neutrophil Extracellular Traps after Subarachnoid Hemorrhage in Mice

Abstract

It has been reported that neutrophil extracellular traps (NETs) involve inflammation, coagulation and cell death. Acute lung injury is also considered to be connected with NETs. Deoxyribonuclease I (DNase-1), a clinical medication for the respiratory system, has been reported to degrade cell-free DNA (cfDNA), which is the main component of NETs. Herein, we did research to clarify the therapeutic value of DNase-1 in NPE after SAH. In this model, we found that the treatment of DNase-1 remarkably decreased lung water, neutrophilic infiltration and inflammation. In addition, DNase-1 inhibited the NETs and proinflammatory subtype transition of the macrophages. Moreover, the depletion of neutrophil also verified the role of NETs in NPE. Our results suggest that DNase-1 has the potential to effectively relieve the NPE after SAH and to be a clinical drug for use after SAH.

Keywords: DNase-1; inflammation; neurogenic pulmonary edema; neutrophil extracellular traps; subarachnoid hemorrhage.

Figure 1

The protective effect of DNase-1 for neurogenic pulmonary edema (NPE) after subarachnoid hemorrhage (SAH). (A) The quantification of the SAH grade; n = 18/group. (B) The quantification of the modified Garcia scale; n = 18/group. (C) Quantification of the lung water; n = 6/group. (D) Representative lung histopathology at 24 h after SAH. Scale bar = 50 μm; n = 5/group. (E–H) Quantitative qPCR analysis of the relative expression of TNF-α, IL-1β, IL-6 and IL-10; n = 5/group. * p < 0.05 versus the sham and # p < 0.05 versus the SAH + vehicle group.

Figure 2

DNase-1 inhibits the formation of NETs in the NPE after SAH. (A) Representative photograph of the colocalization of CitH3 (red) with Ly6G (green) among the groups. The nuclei were stained with DAPI (blue). Scale bar = 50 μm. (B) Representative photograph of the colocalization of NE (red) with Ly6G (green) among the groups. The nuclei were stained with DAPI (blue). Scale bar = 50 μm. (C) Quantitative analysis of CitH3-positive neutrophils; n = 5/group. (D) Quantitative analysis of NE-positive neutrophils; n = 5/group. * p < 0.05 versus the sham and # p < 0.05 versus the SAH + vehicle group.

Figure 3

DNase-1 inhibits the proinflammatory transition in NPE after SAH. (A) Representative photograph and quantitative analysis of CD16-positive cells (red) among the groups. The nuclei were stained with DAPI (blue). Scale bar = 50 μm; n = 5/group. (B) Representative photograph and quantitative analysis of CD86-positive cells (red) among the groups. The nuclei were stained with DAPI (blue). Scale bar = 50 μm; n = 5/group. * p < 0.05 versus the sham and # p < 0.05 versus the SAH + vehicle group.

Figure 4

Ly6G depletion decreases the formation of NETs in NPE after SAH. (A) Representative flow cytometry analysis of the blood from SAH mice in different groups. (B) Representative photograph of the colocalization of CitH3 (red) with Ly6G (green) in different groups. The nuclei were stained with DAPI (blue). Scale bar = 50 μm. (C) Representative photograph of the colocalization of NE (red) with Ly6G (green) in different groups. The nuclei were stained with DAPI (blue). Scale bar = 50 μm. (D) Quantitative analysis of CitH3-positive neutrophils; n = 5/group. (E) Quantitative analysis of NE-positive neutrophils; n = 5/group. * p < 0.05 versus the SAH + vehicle group.