VEGF Over-Expression by Engineered BMSC Accelerates Functional Perfusion, Improving Tissue Density and In-Growth in Clinical-Size Osteogenic Grafts

Rene' D Largo; Maximilian G Burger; Oliver Harschnitz; Conny F Waschkies; Andrea Grosso; Celeste Scotti; Alexandre Kaempfen; Sinan Gueven; Gernot Jundt; Arnaud Scherberich; Dirk J Schaefer; Andrea Banfi; Nunzia Di Maggio

doi:10.3389/fbioe.2020.00755

VEGF Over-Expression by Engineered BMSC Accelerates Functional Perfusion, Improving Tissue Density and In-Growth in Clinical-Size Osteogenic Grafts

Front Bioeng Biotechnol. 2020 Jul 3:8:755. doi: 10.3389/fbioe.2020.00755. eCollection 2020.

Authors

Rene' D Largo^{1

2}, Maximilian G Burger^{1

2}, Oliver Harschnitz^{1

2}, Conny F Waschkies^{3

4}, Andrea Grosso^{1

2}, Celeste Scotti⁵, Alexandre Kaempfen², Sinan Gueven⁵, Gernot Jundt⁶, Arnaud Scherberich⁵, Dirk J Schaefer², Andrea Banfi^{1

2}, Nunzia Di Maggio¹

Affiliations

¹ Cell and Gene Therapy, Department of Biomedicine, >Basel University Hospital and University of Basel, Basel, Switzerland.
² Plastic and Reconstructive Surgery, Department of Surgery, Basel University Hospital and University of Basel, Basel, Switzerland.
³ Institute for Biomedical Engineering, ETH and University of Zurich, Zurich, Switzerland.
⁴ Department of Surgical Research, University Hospital Zurich, Zurich, Switzerland.
⁵ Tissue Engineering, Department of Biomedicine, University Hospital of Basel, University of Basel, Basel, Switzerland.
⁶ Institute of Pathology, University Hospital of Basel, Basel, Switzerland.

Abstract

The first choice for reconstruction of clinical-size bone defects consists of autologous bone flaps, which often lack the required mechanical strength and cause significant donor-site morbidity. We have previously developed biological substitutes in a rabbit model by combining bone tissue engineering and flap pre-fabrication. However, spontaneous vascularization was insufficient to ensure progenitor survival in the core of the constructs. Here, we hypothesized that increased angiogenic stimulation within constructs by exogenous VEGF can significantly accelerate early vascularization and tissue in-growth. Bone marrow stromal cells from NZW rabbits (rBMSC) were transduced with a retroviral vector to express rabbit VEGF linked to a truncated version of rabbit CD4 as a cell-surface marker. Autologous cells were seeded in clinical-size 5.5 cm³ HA scaffolds wrapped in a panniculus carnosus flap to provide an ample vascular supply, and implanted ectopically. Constructs seeded with VEGF-expressing rBMSC showed significantly increased progenitor survivival, depth of tissue ingrowth and amount of mineralized tissue. Contrast-enhanced MRI after 1 week in vivo showed significantly improved tissue perfusion in the inner layer of the grafts compared to controls. Interestingly, grafts containing VEGF-expressing rBMSC displayed a hierarchically organized functional vascular tree, composed of dense capillary networks in the inner layers connected to large-caliber feeding vessels entering the constructs at the periphery. These data constitute proof of principle that providing sustained VEGF signaling, independently of cells experiencing hypoxia, is effective to drive rapid vascularization and increase early perfusion in clinical-size osteogenic grafts, leading to improved tissue formation deeper in the constructs.

Keywords: VEGF; bone marrow stromal cells; gene therapy; osteogenic grafts; vascularization.