TMT-Based proteomics analysis of LPS-induced acute lung injury

Abstract

Purpose:

The proteome during lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice is unclear.

Materials and Methods:

In this study, eight-week-old male C57BL/6 mice were intraperitoneally injected with LPS and sacrificed 18 hours after LPS administration to identify protein expression levels in lung tissue using tandem mass tag (TMT) analysis for relative quantification. Hematoxylin-eosin (HE) staining was used to evaluate lung injury in mice. Immunohistochemical staining was used to calculate the production of myeloperoxidase (MPO) and TUNEL staining was performed to detect apoptosis. GO functional clustering and KEGG pathway enrichment analyses were performed to determine functions of differentially expressed proteins (DEPs) and transduction pathways. Domain annotation and subcellular localization analysis of the DEPs were also performed. Furthermore, parallel reaction monitoring (PRM) analysis was used to verify the top 30 DEPs.

Results:

A total of 5188 proteins were found to be expressed in lung tissues from LPS- and saline-treated mice. Among these proteins, 293 were differentially expressed between the two groups; 255 proteins were upregulated in the LPS-treated ALI mice, while 38 were downregulated. GO analysis showed that the DEPs are mainly extracellular, and KEGG analysis suggested that the DEPs are mainly enriched in the NOD-like receptor signaling pathway, complement and coagulation cascades and natural killer cell-mediated cytotoxicity. Enrichment of the DEPs is mainly peptidase S1A, serine proteases, peptidase S1, and the serpin domain. 26.6% of the DEPs are in the nucleus, 24.6% are in the cytosol, 19.1% are in the extracellular space, and 18.8% are in the plasma membrane. PRM validation showed that the trend of 30 DEPs was same with TMT analysis. Among these, Cytochrome b-245 heavy chain (Cybb), Monocyte differentiation antigen CD14 (Cd14) and Neutrophil gelatinase-associated lipocalin (NGAL) were the most obvious change.

Conclusions:

Our results may help to identify markers and therapeutic targets for LPS-induced ALI.

Keywords:

• acute lung injury
• proteomics analysis
• tandem mass tag

Disclosure statement

No potential conflict of interest was reported by the authors.

Availability of data and material

All data generated or analyzed during this study are included in this published article.

Author contributions

All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Sheng-Song Chen and Qing-Yuan Zhan. The first draft of the manuscript was written by Sheng-Song Chen and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

Ethical statement

All animal experiments had been approved by the Experimental Animal Ethics Committee of China-Japan Friendship Hospital.

Correction Statement

This article has been republished with minor changes. These changes do not impact the academic content of the article.

Additional information

Funding

This study was supported by the National Natural Science Foundation of China (No. 81870072).