Fig 2: In vitro characterization of the receptor binding profiles of VEGF-B isoforms(A) All the VEGF-B isoforms were detected from the HeLa cell medium after adenoviral transduction.(B) VEGF-B immunoprecipitation with sVEGFR-1-Fc and sNrp-1-Fc from HeLa cell medium after adenoviral transduction. VEGF-B186 bound to VEGFR-1 and its proteolytically processed form (∼18 kDa) to VEGFR-1 and Nrp-1, whereas VEGF-B186R127S only bound to VEGFR-1. VEGF-B127, resembling the proteolytically processed form, was confirmed to bind both VEGFR-1 and Nrp-1, whereas VEGF-B109 bound solely to VEGFR-1. The C-terminal fragment did not bind to VEGFR-1 or Nrp-1.

Fig 3: Study protocol and timepointsPart A evaluated angiogenic effects of different VEGF-B isoforms in the normoxic heart. In Part B, the primary endpoints were myocardial perfusion reserve and histological quantification of the capillary area in ischemic hearts. In Part B, ischemia was induced by bottleneck stent placement to LAD two weeks before the gene transfer. 15O-H2O PET imaging was performed for nine animals one day before the gene transfer and one day before the sacrifice.

Fig 4: Characterization of the designed proteolytically resistant VEGF-B186R127S. (A) Graphical representation of VEGF-B186R127S. (B) Representative image of VEGF-B protein expression following HeLa cell transduction. (C) BaF3-VEGFR1 cell survival and proliferation assay. Each dot represents mean ± SD from two technical replicates from a representative experiment. (D) Receptor binding properties of different VEGF-B forms. Each lane represents the precipitation of ligands with hVEGFR1, hNRP1, hNRP2, or no receptor (blank). Red arrow signs indicate the full-length and proteolytically processed VEGF-B186, VEGF-B186R127S, or VEGF-A165.