Interleukin‐33 Dependent Endothelial Cell Hyperproliferation and Vascular Remodeling in Sugen/Hypoxia Mice
Authors: Indralingam Cynthia , Johns Scott C. , Fuster Mark M. , Jennings Patricia A. , Breen Ellen C. 
Published: April 2020

Pulmonary arterial hypertension (PAH) is a devastating disease that is often misdiagnosed and has a high mortality rate. It is characterized by remodeling of the small resistance arteries in the lungs that leads to elevated right ventricular pressure (RVP) and cardiac failure. Interleukin‐33 (IL‐33) is a cytokine that functions as an alarmin to sense danger or damage to endothelial cells, signal proliferation and restore barrier integrity. However, uncontrolled, active forms of IL‐33 could initiate endothelial hyperproliferation, a putative first step in the remodeling process. We hypothesize that IL‐33 initiates the hyperproliferative response of pulmonary artery endothelial cells and contributes to the wall remodeling and increased RVP observed in PAH. To test this, we induced pulmonary hypertension of C57Bl/6J (wild‐type), IL‐33 receptor knock‐out (ST2KO) and the adaptor protein, MyD88 knock‐out (MyD88KO) mice by exposure to 10% O2 and weekly SU5416 injections (SuHx). Hypertension was assessed by measuring RVP and arterial wall thickness (ratio of wall cross‐sectional area/total vessel cross‐sectional area). Proliferating pulmonary endothelial (CD31+/BrdU+) cells were detected by flow cytometry. Processed IL‐33 forms in the lung were analyzed by western blot. RVPs increased in wild‐type mice in response to SuHx (C57Bl/6J male, RA/DMSO 27.0 ± 3.0 mmHg, SuHx 40.5 ± 6.3 mmHg, p < 0.01; female, RA/DMSO 25.8 ± 3.3 mmHg, SuHx 43.8 ± 7.4 mmHg, p < 0.01) and, this response was attenuated in ST2KO mice (ST2KO male, RA/DMSO 25.0 ± 4.3 mmHg, SuHx 32.8 ± 6.9 mmHg, p<0.01; female, RA/DMSO 25.3 ± 3.5 mmHg, SuHx 32.8 ± 4.9 mmHg, p<0.01) and completely blocked in MyD88KO mice (MyD88KO male, RA/DMSO 19 ± 2.9 mmHg, SuHx 19.7 ± 5.9 mmHg, NS; female, RA/DMSO 20.2 ± 2.5 mmHg, SuHx 24.5 ± 7.1 mmHg, NS). Similarly, wall thickness increased in wild‐type mice, but not ST2KO and MyD88KO mice, under SuHx conditions (C57Bl/6J Male, RA/DMSO 0.35 ± 0.07, SuHx 0.53 ± 0.12, p < 0.01; Female RA/DMSO 0.34 ± 0.05, SuHx 0.41 ± 0.1, p=0.4; ST2KO Male, RA/DMSO 0.37 ± 0.07, SuHx 0.38 ± 0.13, NS; Female RA/DMSO 0.31 ± 0.02, SuHx 0.34 ± 0.03, NS; MyD88KO Male, RA/DMSO 0.37 ± 0.05, SuHx 0.41 ± 0.10, NS; Female RA/DMSO 0.34 ± 0.07, SuHx 0.38 ± 0.03, NS). The percentage of proliferating endothelial cells was increased in wild‐type mice under SuHx conditions (C57Bl/6J Male, RA/DMSO 1.9 ± 0.4, SuHx 4.8 ± 2.6, p< 0.01; Female RA/DMSO 0.3 ± 0.05, SuHx 0.4 ± 0.1, p= 0.07). This endothelial proliferative response was absent in ST2KO and Myd88KO mice (ST2KO Male, RA/DMSO 0.35 ± 0.07, SuHx 0.53 ± 0.12, NS; Female RA/DMSO 0.34 ± 0.05, SuHx 0.41 ± 0.1, NS). Proliferating (Ki‐67+) cells were detected in the intima layer of small arteries. The C‐terminal and caspase cleaved form of IL‐33 is increased in wild‐type and ST2KO mice but not in MyD88KO mice under SuHx conditions. IL‐33 signaling through the ST2 receptor is important for endothelial cell proliferation in the SuHx model. This response is associated with increased levels of both the C‐terminal IL‐1 like cytokine domain and caspase cleaved fragment of IL‐33 in lung tissue.Support or Funding InformationDepartment of Veteran Affairs