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Protection against vascular leak in neprilysin transgenic mice with complex overexpression pattern

Abstract

Neprilysin (NEP) is a cell surface metallopeptidase found in many tissues. Based mostly on pharmacological manipulations, NEP has been thought to protect blood vessels from plasma extravasation. We have suggested that NEP may protect against pulmonary vascular injury. However, these prior studies did not utilize mice which overexpress NEP. The aims of the present investigation were to develop and characterize doubly transgenic (DT) mice that overexpress NEP universally and conditionally, and to investigate the protective effect that overexpressed NEP may have against plasma extravasation in the vasculature. The duodenum, which is often used to assess vascular permeability, and in which the NEP protein was overexpressed in our DT mice two-fold, was selected as our experimental preparation. We found that substance P-induced plasma extravasation was decreased substantially (3.5-fold) in the duodenums of our doxycycline-treated DT mice, giving independent evidence of NEP’s protective effects against plasma extravasation. Transgenic lung NEP protein was not stably expressed in the DT mice, so we were not able to test the effect of NEP overexpression in the lung. Although initially overexpressed nearly nine-fold at that site, pulmonary NEP protein overexpression eventually dissipated. Surprisingly, at a time when there was no lung transgenic NEP protein overexpression, lung NEP mRNA expression was still increased 23-fold, indicating that the expression defect probably is not transcriptional. These studies help to characterize our complex transgenic model of NEP overexpression and further demonstrate NEP’s protective effects against plasma extravasation.

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Acknowledgments

Supported by Grants received from the National Heart, Lung and Blood Institute (NHLBI RO1 HL078929, PPG HL14985 and RO3 HL095439) and the Department of Veterans’ Affairs (Merit Review to ECD). YEM is supported by NCI P50 CA058187 and NCI P30 CA046934. The authors wish to thank Dr. James West (Vanderbilt University) for the FVB/N Rosa26- rtTA2-M2 singly transgenic mice, Dr. Lou Hersh (University of Kentucky) for the NEP-pCSC-SP-PW lentiviral vector, and Dr Jeffrey Whitsett (Cincinnati Children’s Hospital) for the reengineered Tet-O-CMV vector and FVB/N SP-C-rtTA transgenic mice. The authors also wish to thank Sandra Walchak and Andy Poczobutt for expert technical and administrative assistance.

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Correspondence to Marilee J. Wick.

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Wick, M.J., Loomis, Z.L., Harral, J.W. et al. Protection against vascular leak in neprilysin transgenic mice with complex overexpression pattern. Transgenic Res 25, 773–784 (2016). https://doi.org/10.1007/s11248-016-9969-x

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  • DOI: https://doi.org/10.1007/s11248-016-9969-x

Keywords

  • Neprilysin
  • Transgenic mouse
  • Vascular leak
  • Evans blue
  • Duodenum
  • Rosa26 promoter