04 February 2019 by Rafael Soares Godoy

Balance between Endothelium-Pericyte Injury and Repair Determines Pulmonary Hypertension in Mouse Model for Targeted Diphtheria Toxin-Induced Cell Death.

Endothelial cell (EC) apoptosis is a critical trigger for pulmonary arterial hypertension (PAH), resulting the loss of fragile distal lung arterioles either directly, by a degenerative mechanism, and/or indirectly, by promoting the emergence of growth dysregulated vascular cells.

We developed a murine model for EC apoptosis via diphtheria toxin (DT) exposure in order to study its downstream effects.

48-72 hours after the intra-tracheal  dministration of 10ng DT, binary transgenic mice (CRE+DTR+) harbouring the human DT receptor (hDTR) targeted to ECs showed an increase in lung endothelial TUNEL and activated caspase-3 staining associated with an incrrase lung vascular permeability (Evans-Blue assay). Flow-cytometric analysis showed a >50% loss of hDTR+, CD144+, CD31+ and CD34+ ECs at 72 hrs, with an 80% decrease in NG5+pericytes, leading to 35% decrease in total lung microvascular volume by Micro-CT, mainly in vessels <150um in diameter. Right ventricular systolic pressure (RVSP) (n=19) was elevated at 72 hours after a single dose of DT compared with saline (33.7±1 vs. 23.9±0.35 mmHg, respectively; p<0.0001), consistent with pulmonary hypertension (PH), returning to control levels by 1 week (n=20).

In contrast, right ventricular hypertrophy (RVH) was only seen at 1 week post DT (n=15) (0.27±0.006) vs. control (0.24±0.004; p=0.001). Resolution of PH was accompanied by recovery in lung vascular permeability and volume, as well lung EC and pericyte populations. Interestingly, a marked increase in CD11b+Ly6G+ neutrophils was seen at 72 hours post DT (1414±293.5) vs. saline (109±29; p=0.004), with no change in M1 or M2 macrophage numbers, returning to control levels at 1 week, suggesting a potential role of inflammatory cells in the regenerative/recovery process in this model.

These data suggest that sustained loss of ECs is needed to maintain a PAH-phenotype and that the balance between injury and repair may be an important determinant in the pathogenesis of PAH.

Key Contributors

Rafael Soares Godoy 1, Mohammed Taha 1,2, Yupu Deng 1, Katelynn Rowe 1, Duncan J Stewart 1,2 : 1.Sinclair Center for Regenerative Medicine, Ottawa Hospital Research Institute, Canada 2.University of Ottawa, Faculty of Medicine, Ottawa, Canada

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