Proteomic analysis of circulating monocytes in Chinese premenopausal females with extremely discordant bone mineral density.

Osteoporosis (OP) is a major public health problem, mainly characterized by low bone mineral density (BMD). Circulating monocytes (CMCs) may serve as progenitors of osteoclasts and produce a wide variety of factors important to bone metabolism. However, the specific action mechanism of CMCs in the pathogenesis of OP is far from clear. We performed a comparative protein expression profiling study of CMCs in Chinese premenopausal females with extremely discordant BMD, identified a total of 38 differentially expressed proteins, and confirmed with Western blotting five proteins: ras suppressor protein1 (RSU1), gelsolin (GSN), manganese-containing superoxide dismutase (SOD2), glutathione peroxidase 1(GPX1), and prolyl 4-hydroxylase beta subunit (P4HB). These proteins might affect CMCs' trans-endothelium, differentiation, and/or downstream osteoclast functions, thus contribute to differential osteoclastogenesis and finally lead to BMD variation. The findings promote our understanding of the role of CMCs in BMD determination, and provide an insight into the pathogenesis of human OP.

Comparative proteomics analysis of human lung squamous carcinoma.

Two-dimensional polyacrylamide gel electrophoresis (2-DE) profiles of human lung squamous carcinoma tissue and paired surrounding normal bronchial epithelial tissue were compared. Selected differential protein-spots were identified with peptide mass fingerprinting based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and database searching. Well-resolved and reproducible 2-DE patterns of both the tumor and the normal tissues were acquired. The average deviations of spot position were 0.873+/-0.125mm in IEF direction and 1.025+/-0.213mm in SDS-PAGE direction, respectively. For the tumor tissues, a total of 1349+/-67 spots were detected and 1235+/-48 spots were matched with an average matching rate of 91.5%. For the corresponding normal tissues, a total of 1297+/-73 spots were detected and 1183+/-56 spots were matched with an average matching rate of 91.2%. A total of 1069+/-45 spots were matched between the tumor and the normal tissues. Forty differential proteins between tumor and normal tissues were characterized. Some proteins were the products of oncogenes and others were involved in the regulation of cell cycle and signal transduction. These data are valuable for mass identification of differentially expressed proteins involved in lung carcinogenesis, establishing human lung cancer proteome database and screening molecular marker to further study human lung squamous carcinoma.

[Differential proteomic analysis of human lung adenocarcinoma cell line A-549 and of normal cell line HBE].

To explore the differential proteomic expressions between human lung adenocarcinoma cell line A-549 and normal cell line HBE, a series of methods, including immobilized pH gradient-two dimensional polyacrylamide gel electrophoresis, silver staining, PDQuest 2-DE software analysis, peptide mass fingerprinting based on matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) and SWISS-PROT database searching, were used to separate and identify the differential proteomic expressions between A-549 and HBE. The results showed that the good 2-DE pattern including high resolution and reproducibility was obtained. After silver staining, the 2-DE image analysis by PDQuest 2-DE software detected average (890 +/- 38) spots in A-549, and (757 +/- 27) spots in HBE. The average positional deviation of the matched spots between A-549 and HBE 2-DE maps was (2.85 +/- 0.48) mm in IEF direction, and (2.69 +/- 0.37) mm in SDS-PAGE direction. The differential proteomic expression analysis found that there were 535 matched spots between A-549 and HBE 2-DE maps, 355 spots that were not matched in A-549, 222 spots that were not matched in HBE. 18 differential spots (8 spots in A-549 and 10 spots in HBE) were cut off from silver staining gel at random, digested in gel with TPCK-trypsin, measured with MALDI-TOF-MS and searched in the SWISS-PROT database with PeptIdent software. 18 protein were preliminarily identified. These proteins were related to cell signal transduction, cell metabolism, proliferation and differentiation etc. There was a significant difference at protein level between human lung adenocarcinoma cell line A-549 and normal cell line HBE. It suggests that the differential expression analysis of proteomes may be useful to further study of the related proteins and the molecular markers of lung adenocarcinoma.

Identification of Protein Spots in Silver-stained Two-dimensional Gels by MALDI-TOF Mass Peptide Map Analysis.

Protein spots in silver-stained two dimensional gels were analyzed and identified by employing an improved procedure of mass spectrometric peptide mapping, including i) In-gel reduction, alkylation and enzymatic digestion ii) Extraction and desalting by using a pipette tip containing a small C18 micro-column (ZipTip(TM)) iii) Direct MALDI-TOF mass analysis and protein database searching. The results demonstrate that single silver-stained protein spots in a 2-DE-gel could be identified rapidly by this procedure and the use of the ZipTip(TM) pipette tip could increase evidently the sensitivity of the MALDI-TOF analysis. By using this the procedure, 10 protein spots in a silver-stained gel of 2-DE of crude venom of the spider S.Huwena were analyzed and identified.