Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Lung injury induced by common bile duct ligation in mice

ABSTRACT: Liver dysfunction and cirrhosis affect vasculature in several organ systems and cause impairment of organ functions, thereby increasing morbidity and mortality. If a mouse model of hepatopulmonary syndrome (HPS) could be established, greater insight into the genetic basis of the disease would be gained. Our objectives were to establish a mouse model of lung injury after common bile duct ligation (CBDL) and to investigate pulmonary pathogenesis for application in future therapeutic approaches. Balb/c mice were subjected to CBDL. Immunohistochemical analyses and real-time quantitative reverse transcriptional polymerase chain reaction were performed on pulmonary tissues. The presence of HPS markers were detected by western blot and microarray analyses. We observed extensive proliferation of CD31-positive pulmonary vascular endothelial cells 2 weeks after CBDL, and identified 11 up-regulated and 8 down-regulated proteins that were associated with angiogenesis. MMP-9 protein was highly expressed at 3 weeks after CBDL, and less expressed in lungs of the control group. Contrary to our expectation, lung pathology in our mouse model exhibited differences from that of rat models, and the mechanisms responsible for these differences are unknown. This phenomenon may be explained by contrasting processes related to TNF induction of angiogenic signaling pathways in the inflammatory phase; thus, we suggest that our mouse model can be applied to pulmonary pathological analyses in the inflammatory phase, i.e., to systemic inflammatory response syndrome, acute lung injury, and MOD syndrome. After induction of anesthesia, a median abdominal incision was made and the common bile duct was identified. The duct was dissected carefully under a microscope, and doubly ligated with 7-0 Prolene and transected. In the sham operation (control) group, the duct was dissected without common bile duct ligation. Mice were sacrificed at 2 and 3 weeks after surgery. CD31-positive cells were assembled from three mice in each group.

ORGANISM(S): Mus musculus

SUBMITTER: Koh-ichi Nakashiro

PROVIDER: E-GEOD-50088 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Early postnatal life is considered as a critical time window for determination of long-term metabolic states and organ functions. Extrauterine growth restriction (EUGR) causes the development of adult onset chronic diseases, including pulmonary arterial hypertension (PAH). However, the effects of nutritional disadvantages during early postnatal period on pulmonary vascular consequences in later life are not fully understood. Our study was designed to test whether epigentic dysregulation mediates the cellular memory of this early postnatal event. To test this hypothesis, we isolated pulmonary vascular endothelial cells (PVEC) by magnetic-activated cell sorting (MACS) from EUGR and control rats. A postnatal insult, nutritional restriction-induced EUGR caused development of an increased pulmonary artery pressure at 9-week of age in male rats. MeDIP-chip (Methyl-DNA immune precipitation chip), genome-scale mapping studies to search for differentially methylated loci between control and EUGR rats revealed significant difference in cytosine methylation between EUGR and control rats. We validated candidate dysregulated loci with quantitative assays of cytosine methylation and gene expressions. EUGR changes cytosine methylation at ~500 loci in male rats at 9 weeks of age, preceding the development of PAH and these represent candidate loci for mediating the pathogenesis of pulmonary vascular disease that occurs later in life. These results demonstrate that epigenetic dysregulation is a strong mechanism for propagating the cellular memory of early postnatal events, causing changes in expression of genes and long term susceptibility to PAH, and further providing a new insight into prevention and treatment of EUGR-related PAH. MeDIP together with microarray analysis demonstrated that significant differences in cytosine methylation between EUGR and control rats. Comparison of EUGR(n=3) vs Control (n=3) male rats' pulmonary vascular endothelial cells in 9-week age old rats

Project description:Disordered angiogenesis is implicated in the pulmonary vascular remodeling secondary to congenital heart disease (CHD). However, the underlying genes are not well delineated. We have previously shown that an ovine model of CHD with increased pulmonary blood flow (PBF, Shunt) has an M-bM-^@M-^\angiogenesis burstM-bM-^@M-^] between 1-4 weeks of age. Thus, we hypothesized that the increased pulmonary blood flow elicited a pro-angiogenic gene expression profile prior to the onset of vessel growth. To test this we utilized microarray analysis to identify genes that could be responsible for the angiogenic response. Total RNA was isolated from the lungs of Shunt and Control lambs at 3 days of age and hybridized to Affymetrix GeneChips for microarray analyses (n=8 for each group). A total of 89 angiogenesis-related genes were found to be up-regulated and 26 angiogenesis-related genes down-regulated in Shunt lungs compared to control (cutting at 1.2 fold difference, P<0.05). We then confirmed the up-regulation of pro-angiogenic genes: FGF2, Angiopoietin2 (Angpt2), and Birc5 at both mRNA and protein levels and the up-regulation of ccl2 at mRNA level in the 3-day shunt lungs. Further, we found that pulmonary arterial endothelial cells (PAEC) isolated from juvenile lambs exhibited increased expression of FGF2, Angpt2, Birc5, and ccl2 as well as enhanced angiogenesis when exposed to elevated shear stress (35 dyn/cm2) compared to cells exposed to more physiological level shear stress (20 dyn/cm2). Finally, we demonstrated that blocking FGF2, Angpt2, or Birc 5 signaling, using neutralizing antibodies, significantly decreased the angiogenic response induced by shear stress. In conclusion, we have identified a pro-angiogenic gene expression profile in a lamb model of CHD with increased PBF that precedes the onset of pulmonary vascular remodeling. Further, our data indicate that FGF2, Angiopoietin2, Birc5, and ccl2 may play important roles in the angiogenic response. Total RNA was isolated from the lungs of Shunt and Control lambs at 3 days of age and hybridized to Affymetrix GeneChips for microarray analyses. Eight chips were employed for both control and shunt groups.

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Lung injury induced by common bile duct ligation in mice

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