| Objective In order to provide us new clues to induce some endogenous protective molecular mechanisms, the changes in gene expression profile induced by ischemia-reperfusion in pulmonary tissues of rats were investigated and the dynamic mechanism of pulmonary ischemia-reperfusion injury was elucidated.Methods Thirty male SD rats were randomly divided into 6 groups:5 ischemia-reperfusion (I/R) groups (I/R 0-h, I/R 1-h, I/R 3-h, I/R 6-h, I/R 24-h) and control group (n=6 in each). An in situ ischemia-reperfusion lung injury rat model was established by occluded hilus of lung. The RatRef-12 Expression Beadchip (22 226 gene probes per array) was used to analyze the pattern of gene expression in all groups.Results The results showed that 648,340,711,1279 and 641 genes were differentially expressed in I/R 0-,1-,3-,6- and 24-h groups respectively. The differentially expressed genes were classified as following 7 functional categories:cytokine, growth factor and apoptosis-related factor, oxidation and antioxidation molecule, metabolic enzyme, ion channel and aquaporin, signal transduction molecule and complement. Clusters analysis identified 12 clusters of genes in which each cluster had similar expression pattern.Conclusions It was suggested that gene chip technology was an effective and quick method for screening differentially expressed genes. Many differentially expressed genes with different functions interacted each other to result in pulmonary ischemia-reperfusion injury. Genes with similar expression pattern could have the similar regulated mechanism. Objective The multilineage capability of adult bone marrow mesenchymal stem cells (MSCs) supports the feasibility of tissue engineering multiphasic constructs using a single cell source. Therefore, it is important to establish a method of isolation and culture of the rat mesenchymal stem cells, make a study on the differentiation character and pluripotency of MSCs under different conditions and to investigate the expression of green fluorescent protein (GFP) gene carried by lentiviral vectors into MSCs.Methods MSCs, which were initially isolated from the bone marrow of rats, were cultured in vitro, isolated by trypsin digestion method, purified by adherence method and tested by flow cytomertry. After MSCs were transferred to osteogenic, adipogenic or chondrogenic differentiation medium respectively, the morphological characterization of induced cells was observed. The expression of marker genes was measured by RT-PCR analysis. Then MSCs were infected with lentiviral vectors. The results of the expression of GFP and infection efficiency were observed by fluorescence microscope.Results MSCs of high purity were obtained from bone marrow using adherence screening method. The stable pH of culture medium was important for cell growth and subculture. MSCs typically expressed the antigens CD44 (94.81%), CD90 (99.53%), CD106 (76.34%). They were negative for typical lymphocytic markers like CD45 (1.94%) and CD11b (1.42%) and for the early hematopoietic markers CD34 (0.04%). MSCs can differentiate to adipocyte, osteocyte and chondrocyte in vitro. The results of fluorescence microscopic imaging proved that the high level and stability of GFP expression in MSCs infected with lentiviral vector. So the exogenous GFP and multilineage potential of MSCs had no severe influences on each other.Conclusions Since the MSCs can be easily obtained and abundant, it is proposed that they may be promising candidate cells for further studies on tissue engineering. Imaging with expression of GFP facilitates the research on MSCs physiological behavior and application in tissue engineering during differentiation both in vitro and in vivo. Objective To elucidate effect of adult bone marrow mesenchymal stem cells (MSCs) on pulmonary ischemia-reperfusion (I/R) injury and make a research on differentiaon and regulatory mechanism of MSCs in I/R injury.Methods MSCs were initially isolated from the bone marrow of rats and cultured in vitro. Then MSCs were infected with lentiviral vectors to obtain EGFP mark. MSCs were injected into rats of I/R injury model and control groups. Pathological changes and function alterations of lung were observed at 6 h,1 d,3 d and 7 d after I/R injury. Migration to lung and persistence in lung of MSCs were also observed. Immunohistochemisty method was used to investigate the differentiation of MSCs. Real-time RT-PCR was used to monitor gene expression changes of TNF-αand complement 5 (C5) in all groups. ELISA was used to detect concentration alterations of IL-1β, IL-10, IL-17, KGF and MIP-1αin pulmonary tissue.Results MSCs could decrease pulmonary ratio of wet to dry and MPO activity. MSCs could migrate to lung and the number of cells at 24 h was larger than those at 7 d. However, the double positive of EGFP and SP-C in MSCs was found at 7 d after I/R injury. Concertaion of pro-inflammatory cytokines auch as TNF-α, C5, IL-17 and MIP-1αwere decreased and expression of antiinflammatory cytokines such as IL-10 and KGF were upregulated.Conclusions MSCs have beneficial effects in experimental pulmonary I/R injury model. The beneficial effect of MSCs derives from their differentiation to typeⅡalveolar cells and more from their capacity to secrete paracrine soluble factors that modulate immune responses.
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