Decitabine enhances stem cell antigen-1 expression in cigarette smoke extract-induced emphysema in animal model

Abstract

Stem cell antigen-1 (Sca-1) is a mouse glycosyl phosphatidylinositol-anchored protein and a cell surface marker found on hematopoietic stem cells (HSCs). Despite decades of study, its biological functions remain little known. Sca-1 is a typical marker of bone marrow-derived HSCs, it is also expressed by a mixture of tissue-resident stem, progenitor cells in nonhematopoietic organs. Endothelial progenitor cell (EPC) is a subtype of HSC and contributes to endothelial repair by homing in on locations of injury. Abnormal genetic methylation has been detected in smoking-related diseases. The present study aimed to investigate the lung function and histomorphology, the expression of Sca-1 gene in lung tissues, and bone marrow-derived EPCs in cigarette smoke extract (CSE)-induced emphysema mice, and to further determine whether Decitabine (Dec), the most widely used inhibitor of DNA methylation, could protect against the damages caused by CSE. The results of the present study demonstrated that Dec could partly protect against CSE-induced emphysema in mice, enhance Sca-1 expression in lung tissue, and bone marrow-derived EPCs. The results suggested that the depletion of the progenitor cell pool and DNA methylation of Sca-1 gene may be involved in the progression of emphysema in mice.

Keywords: Cigarette smoke; Decitabine; emphysema; endothelial progenitor cells; stem cell antigen-1.

Figure 1

Maximal expiratory flow volume curve. Of each picture, upper part is expiratory phase and lower part is inspiratory phase. Compared with controls (a) and Dec group (b), Dec + CSE group (c), and CSE group (d) have a abrupt ascending and the descending limb concave to the axis showing prolonged expiratory phase. Dec, Decitabine; CSE, cigarette smoke extract

Figure 2

Histomorphological changes of lung tissues. Lung tissues of Dec + CSE group (c) and CSE group (d) exhibited enlarged alveolar space, thinner alveolar septum, and destroyed alveolar wall when compared with controls (a) and Dec group (b). Alveolar space of Dec + CSE group (c) seems to be smaller than that of CSE group (d). Scale bar represents 50 µm. Dec, Decitabine; CSE, cigarette smoke extract

Figure 3

The morphological changes of bone marrow-derived EPCs during culture. On day 1 of the culture, the cells formed round, the sizes of cells were almost same, and the cells were suspended in the culture medium (a). On day 4 of the culture, the cells attempted to attach each other, and their sizes were getting enlarged, and their shapes became oval, spindle or polygonal (b). On day 7 of the culture, the cells shaped fusiform or polygon patterns and contacted each other to attempt to form capillary structure (c). Scale bar represents 50 µm. EPCs, endothelial progenitor cells

Figure 4

The identification of EPCs by double positive staining with Dil-acLDL and FITC-UEA-1. The laser scanning confocal microscope (LSCM) illustrated that the cells on day 7 of the culture displayed red cytoplasm when uptaking Dil-acLDL (a), green cytomembrane when binding FITC-UEA-1 (b), orange when double positively staining with Dil-acLDL and FITC-UEA-1) (c), and blue when staining with DAPI in nuclear localization (d). Scale bar represents 10 µm. Dil, 1,1’-dioctadecyl-3,3,3’,3-tetramethylindocarbocyanine perchlorate; acLDL, acetylated low density lipoprotein; FITC, fluorescein isothiocyanate; UEA-1, ulex europaeus agglutinin-1; DAPI, 4',6-diamidino-2-phenylindole

Figure 5

Expressions of Sca-1 protein in lung tissues and EPCs. There were pictures of expressions of Sca-1 protein in lung tissues (a) and EPCs (b), and corresponding quantitative analysis (c). Sca-1, stem cell antigen-1; N, normal controls; D, Dec, Decitabine; C, CSE, cigarette smoke extract; EPCs, endothelial progenitor cells. ⋆P < 0.05 vs. group N; ★P < 0.01 vs. group N; △P < 0.05 vs. group Dec; ▴P < 0.01 vs. group Dec; ♦P < 0.01 vs. group Dec + CSE

Figure 6

Expressions of Sca-1 mRNA in lung tissues and EPCs. Sca-1, stem cell antigen-1; EPCs, endothelial progenitor cells; N, normal control; Dec, Decitabine; CSE, cigarette smoke extract. ★P < 0.01 vs. group N; ▴P < 0.01 vs. group Dec; ♦P < 0.01 vs. group Dec + CSE