Analysis of the expression protein profiles of lung squamous carcinoma cell using shot-gun proteomics strategy

Abstract

The aim of this study is to globally screen and identify the expression protein profiles of lung squamous carcinoma cell (SqCC) using shot-gun proteomics strategy and to further analyze function of individual proteins by bioinformatics, which may likely result in the identification of new biomarkers and provide helpful clues for pathogenesis, early diagnosis, and progression of lung SqCC. The specific tumor cells were isolated and collected from the tissues of six patients with lung SqCC by laser capture microdissection (LCM). Total proteins from the LCM cells were extracted, digested with trypsin. The sequence information of resulting peptides was acquired by high-performance liquid chromatography (HPLC) and tandem mass spectrometry (TMS). The global protein profiles of lung SqCC cell were identified with BioworksTM software in IPI human protein database. Cellular component, molecular function, and biological process of the all proteins were analyzed using gene ontology (GO). About 720,000 tumor cells were satisfactorily collected from tissues of six patients with lung SqCC by LCM and the homogeneities of cell population were estimated to be over 95% as determined by microscopic visualization. The high resolution profiles including HPLC, full mass spectrum, and tandem mass spectrum were successfully obtained. Database searching of the resulting bimolecular sequence information identified 1982 proteins in all samples. The bioinformatics of these proteins, including amino acids sequence, fraction of coverage, molecular weight, isoelectric point, etc., were analyzed in detail. Among them, the function of most proteins was recognized by using GO. Five candidate proteins, Prohibitin (PHB), Mitogen-activated protein kinase (MAPK), Heat shock protein27 (HSP27), Annexin A1(ANXA1), and High mobility group protein B1 (HMGB1), might play an important role in SqCC genesis, progression, recurrence, and metastasis according to relative literatures. We have successfully isolated the interesting cells and effectively solved the heterogeneous problem of lung SqCC using LCM. The globally expressional proteins of lung SqCC cell were identified by shot-gun proteomics strategy. The five proteins might be hopefully used as markers of lung SqCC.

Prohibitin in squamous cell carcinoma of the lung: its expression and possible clinical significance

“…One recent study using a shot-gun proteomics strategy showed that PHB is a candidate protein for a biomarker of lung SqCC [28]. A few other proteomics analysis studies focused on the correlation between altered expression of proteins and drug resistance in lung cancer cell lines [29,30].…”

Microproteomics: Quantitative Proteomic Profiling of Small Numbers of Laser-Captured Cells

“…Recently, 2D-DIGE and quantitative gel-free mass spectrometry approaches have been coupled to LCM for proteomic analyses of distinct, pure cell populations (Shekouh et al 2003; Li et al 2004; Zang et al 2004; Haqqani et al 2005; Bagnato et al 2007; Mustafa et al 2008; Patel et al 2008; Wang et al 2008; Zhang and Koay 2008; Nan et al 2009; Scicchitano et al 2009; Waanders et al 2009; Yao et al 2009; Asomugha et al 2010; Zhang et al 2010). Other methods such as punch biopsy can be used to microdissect tissues for subsequent proteomic analyses (Folli et al 2010), but because it is difficult to see the three-dimensional boundaries of structures from the surface of the tissue, punch biopsies can sample adjacent regions that are not of interest.…”

ANXA1 (annexin A1)

“…Prognosis The ANXA1 might play an important role in lung squamous cell carcinoma genesis, progression, recurrence, and metastasis and might be used as markers of this carcinoma (Nan et al, 2009). …”