Autor:innen:
Dr. med. Paolo Valente | Universitätsspital Basel | Switzerland
Alessandro Certelli | Universitätsspital Basel | Switzerland
Andrea Uccelli | Universitätsspital Basel | Switzerland
Andrea Grosso | Universitätsspital Basel | Switzerland
Dr. Nunzia Di Maggio | Universitätsspital Basel | Switzerland
PD Dr. med. Thomas Wolff | Universitätsspital Basel | Switzerland
Prof. Dr. med. Lorenz Gürke | Universitätsspital Basel | Switzerland
Dr. Roberto Gianni-Barrera | Universitätsspital Basel | Switzerland
PD Dr. Andrea Banfi | Universitätsspital Basel | Switzerland
Background. There is the unmet clinical need to investigate pro-angiogenic treatments to increase therapeutic flow to the superficial wound bed from the deeper circulation through the recruitment of new arterial connections (therapeutic arteriogenesis). Here we aimed at establishing the angiogenic and arteriogenic potential of fibrin-based co-delivery of VEGF and PDGF-BB in a pre-clinical animal model of diabetic skin.
Methods. 10-week old diabetic mice received intracutaneous injections (in the dermal layer) of fibrin hydrogels (20 µl) containing VEGF alone (100 µg/ml) or together with PDGF-BB (10 µg/ml), or no factors as negative control (6 injections/animal; n=6-9/condition and time-point). Tissues were harvested at 7- and 28-day time-points after intravascular perfusion of fixative (1% paraformaldehyde), frozen and cryosectioned. Vascular growth was imaged by immunofluorescence staining and confocal microscopy. The amount and size of induced vessels were quantified with the Olympus Cell Sense software.
Results. By 7 days fibrin was almost consumed, but some was still present in all tissues. Both VEGF alone (V) or with PDGF-BB (VP) induced a comparable increase in vascular density (Vessel Length Density, VLD: V=24,1±0,9 mm/mm2; VP=25.9±0,9 mm/mm2; control=11,9±0,9 mm/mm2; p < 0.0001). The total amount of induced vessels (vessel density multiplied by the total area of angiogenic effect) was also similar in V and VP tissues. However, V-induced new vessels were significantly larger than those induced by VP (V=11,7±0,77 µm; VP= 8,9±0,72 µm; control= 5,6±0,34 µm; p < 0.05 V vs VP), confirming the ability of PDGF-BB to prevent aberrant vascular enlargement by high-dose VEGF. After 28 days fibrin was no longer detectable in any condition. Vessels induced by VEGF alone showed partial regression compared to 7 days (VLD V=19,1±1,5 mm/mm2), which was prevented by addition of PDGF-BB (VP=23,6±1,1 mm/mm2). Definitively remodeled vessels showed similar capillary-size diameters in all conditions by 28 days (V = 4±0,15 µm; VP= 3,9±0,12 µm; control= 4,2±0,12 µm; p n.s.), suggesting a preferential regression of the more aberrant structures induced by high-dose VEGF alone.
Conclusions and perspectives. These results establish the feasibility of fibrin-based delivery of angiogenic factors in the skin of diabetic mice. Co-delivery of PDGF-BB both prevented aberrant angiogenesis and ensured persistence of new vessels despite transient factor delivery. Although spontaneous angiogenesis is impaired in diabetic mice, high-dose delivery of fibrin-bound VEGF and PDGF-BB was effective to induce robust growth of stable and physiological vascular networks. The functional efficacy of this strategy will be investigated in a wound healing model.