Mesenchymal Stem Cell-Conditioned Medium Induces Neutrophil Apoptosis Associated with Inhibition of the NF-κB Pathway in Endotoxin-Induced Acute Lung Injury.

Vincent Yi-Fong Su,Shih-Chieh Hung,Chi-Shiuan Lin,Kuang-Yao Yang

doi:10.3390/ijms20092208

Abstract

The immunomodulatory effects of mesenchymal stem cells (MSCs) are established. However, the effects of MSCs on neutrophil survival in acute lung injury (ALI) remain unclear. The goal of this study was to investigate the effect of an MSC-conditioned medium (MSC-CM) on neutrophil apoptosis in endotoxin-induced ALI. In this study, an MSC-CM was delivered via tail vein injection to wild-type male C57BL/6 mice 4 h after an intratracheal injection of lipopolysaccharide (LPS). Twenty-four hours later, bronchoalveolar lavage fluid (BALF) and lung tissue were collected to perform histology, immunohistochemistry, apoptosis assay of neutrophil, enzyme-linked immunosorbent assays, and an electrophoretic mobility shift assay. Human neutrophils were also collected from patients with sepsis-induced acute respiratory distress syndrome (ARDS). Human neutrophils were treated in vitro with LPS, with or without subsequent MSC-CM co-treatment, and were then analyzed. Administration of the MSC-CM resulted in a significant attenuation of histopathological changes, the levels of interleukin-6 and macrophage inflammatory protein 2, and neutrophil accumulation in mouse lung tissues of LPS-induced ALI. Additionally, MSC-CM therapy enhanced the apoptosis of BALF neutrophils and reduced the expression of the anti-apoptotic molecules, Bcl-xL and Mcl-1, both in vivo and in vitro experiments. Furthermore, phosphorylated and total levels of nuclear factor (NF)-κB p65 were reduced in lung tissues from LPS + MSC-CM mice. Human MSC-CM also reduced the activity levels of NF-κB and matrix metalloproteinase-9 in the human neutrophils from ARDS patients. Thus, the results of this study suggest that the MSC-CM attenuated LPS-induced ALI by inducing neutrophil apoptosis, associated with inhibition of the NF-κB pathway.

Highlights

Acute lung injury (ALI) is an inflammatory disease which can lead to the development of acute respiratory distress syndrome (ARDS) and significant morbidity and mortality [1]
The induced lung injury was confirmed histopathologically and characterized by lung edema, alveolar spaces filled with mononuclear/neutrophilic infiltrates, and thickening of the alveolar walls and interstitium (Figure 1A)
When the induction of acute lung injury (ALI) was followed by a tail vein injection of the mesenchymal stem cells (MSCs)-CM 4 h later, histological evaluations showed that the numbers of infiltrative neutrophils and injured areas were significantly reduced after 24 h compared to the lung tissues with ALI that did not receive MSC-conditioned medium (MSC-CM) treatment (Figure 1A)

Summary

Introduction

Acute lung injury (ALI) is an inflammatory disease which can lead to the development of acute respiratory distress syndrome (ARDS) and significant morbidity and mortality [1]. Neutrophils are the major inflammatory cells that are recruited to inflamed lungs and they play a central role in the pathogenesis of ALI. Pulmonary edema and endothelial and epithelial cell injury are observed in cases of ALI [2]. Lipopolysaccharide (LPS) has been shown to cause excessive activation and/or delayed apoptosis in neutrophils, thereby contributing to tissue damage [3]. It is hypothesized that neutrophil apoptosis is needed for the resolution of ARDS and restoration of tissue homeostasis [4,5]. Nuclear factor (NF)-κB is a transcription factor that regulates the expression of proinflammatory and anti-apoptotic genes during the development of ALI and ARDS [6]

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Results