LPS induces permeability injury in lung microvascular endothelium via AT1 receptor
Section snippets
Reagents
LPS (Escherichia coli 0111:B4), Ang II, and [Sar1,Ile8]Ang II were all purchased from Sigma (Sigma–Aldrich, St. Louis, MO). Polyclonal antisera against the AT1 receptor and AT2 receptor were purchased from Santa Cruz Biotechnology (Delaware, CA, USA). Tripure Isolation Reagent was from Roche Molecular Biochemicals (Sandhofer Strasse, Mannheim, Germany). PC12W (a subline of PC12 rat pheochromocytoma cell line that expresses high levels of AT2 receptor, but no AT1 receptor) were purchased from
The presence of Ang II receptor in RPMVECs (Figs. 1 and 2)
To determine whether RPMVECs expressed the AT1 receptor or AT2 receptor, total RNA was isolated, RT-PCR analysis demonstrates that only AT1 mRNA was detected in RPMVECs. On the other hand, only AT2 mRNA was detected in PC12W cells. RPMVECs did not express the AT2 receptor mRNA (Fig. 1).
To further validate the expression of AT1 receptor on RPMVECs, these cells were lysed, immunoprecipitated, and detected with AT1 receptor or AT2 receptor specific antibody. Western blot analysis demonstrated that
Discussion
The results of the present study have clearly demonstrated the expression of functional Ang II receptor subunits in RPMVECs. RT-PCR and Western blot analysis both demonstrated that RPMVECs only express AT1 receptor.
Pulmonary endothelium serves as a semi-selective barrier between the plasma and interstitial to circulatory cells, micromolecules, and bioactive agents. Maintenance of this semi-selective barrier represents an important physiological process for vessel wall homeostasis and lung
Acknowledgment
This study was supported by the National Natural Science Foundation of China, No. 30070334.
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