PROX1 Inhibits PDGF-B Expression to Prevent Myxomatous Degeneration of Heart Valves.

BACKGROUND: Cardiac valve disease is observed in 2.5% of the general population and 10% of the elderly people. Effective pharmacological treatments are currently not available, and patients with severe cardiac valve disease require surgery. PROX1 (prospero-related homeobox transcription factor 1) and FOXC2 (Forkhead box C2 transcription factor) are transcription factors that are required for the development of lymphatic and venous valves. We found that PROX1 and FOXC2 are expressed in a subset of valvular endothelial cells (VECs) that are located on the downstream (fibrosa) side of cardiac valves. Whether PROX1 and FOXC2 regulate cardiac valve development and disease is not known. METHODS: We used histology, electron microscopy, and echocardiography to investigate the structure and functioning of heart valves from Prox1ΔVEC mice in which Prox1 was conditionally deleted from VECs. Isolated valve endothelial cells and valve interstitial cells were used to identify the molecular mechanisms in vitro, which were tested in vivo by RNAScope, additional mouse models, and pharmacological approaches. The significance of our findings was tested by evaluation of human samples of mitral valve prolapse and aortic valve insufficiency. RESULTS: Histological analysis revealed that the aortic and mitral valves of Prox1ΔVEC mice become progressively thick and myxomatous. Echocardiography revealed that the aortic valves of Prox1ΔVEC mice are stenotic. FOXC2 was downregulated and PDGF-B (platelet-derived growth factor-B) was upregulated in the VECs of Prox1ΔVEC mice. Conditional knockdown of FOXC2 and conditional overexpression of PDGF-B in VECs recapitulated the phenotype of Prox1ΔVEC mice. PDGF-B was also increased in mice lacking FOXC2 and in human mitral valve prolapse and insufficient aortic valve samples. Pharmacological inhibition of PDGF-B signaling with imatinib partially ameliorated the valve defects of Prox1ΔVEC mice. CONCLUSIONS: PROX1 antagonizes PDGF-B signaling partially via FOXC2 to maintain the extracellular matrix composition and prevent myxomatous degeneration of cardiac valves.

Benchtop von Willebrand Factor Testing: Comparison of Commercially Available Ventricular Assist Devices and Evaluation of Variables for a Standardized Test Method.

Gastrointestinal bleeding occurs in 20-30% of patients receiving ventricular assist devices (VADs) due, in part, to acquired von Willebrand syndrome. We examined factors to optimize a benchtop method to quantify changes in von Willebrand Factor (VWF) multimer distribution and function in VADs, then applied them to evaluate commercially available devices. Human plasma was circulated through flow loops with VADs. Several experimental conditions were examined, including temperature, viscosity, and enzyme inhibition. Samples were analyzed for VWF collagen-binding activity (VWF:CB) and VWF antigen level. von Willebrand Factor multimer profiles were quantified using gel electrophoresis, near-infrared in-gel visualization, and densitometric analysis. The VWF:CB/antigen ratio in the HeartMate II, CentriMag, and HVAD exhibited average decreases of 46%, 44%, and 36% from baseline after 360 minutes of operation. High molecular weight (hVWF) multimer loss occurred within 30 minutes, although the Levacor and control loop profiles were unchanged. Varying temperature and viscosity altered hVWF degradation rate, but not the final results. Inhibition of a disintegrin and metalloprotease with thrombospondin type 1 repeats, member 13 (ADAMTS13) can potentially distinguish mechanoenzymatic cleavage of VWF from mechanical degradation. We developed a repeatable benchtop method to evaluate VWF compatibility of VADs similar to hemolysis testing that can be adopted for preclinical VAD evaluation.