טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentMichaeli-Geller Gal
SubjectExtra-Cortical Bone Augmentation Using Human Peripheral
Blood Derived Endothelial Progenitor Cells (hEPC)
DepartmentDepartment of Medicine
Supervisors Professor Eli Machtei
Dr. Hadar Zigdon
Full Thesis textFull thesis text - English Version


Abstract

Alveolar bone atrophy results from tooth extraction, trauma and diseases. Rehabilitation of resorbed bone is necessary to allow restoration mastication, phonetics and aesthetics. Current techniques for vertical bone augmentation are unpredictable and limited. This is associated with low amount of osteoprogenitor cells and inadequate vascularization at bone regeneration site. Peripheral blood comprises of Endothelial Progenitor Cells (EPC) which participate in neovascularization and angiogenesis, a crucial process for bone formation.  Previous studies in our laboratory, managed to demonstrate vertical bone augmentation via transplantation of autologous rat EPC.

Thus, the aim of the present study was to evaluate whether local transplantation of human peripheral blood derived endothelial progenitor cells (hEPC) combined with β-Tricalcium Phosphate (βTCP) and coupled with barrier domes in the athymic rat calvaria model may stimulate vascularization and consequently accelerate extra-cortical vertical bone augmentation.

Materials and methods included isolation of hEPC from peripheral blood of volunteers. They were cultured, expanded and characterized flow cytometry. Calvaria of 24 athymic rats were exposed and gold domes, serving for cell exclusion, were filled with βTCP alone (control) or βTCP loaded with 106 hEPC (test). Rats were sacrificed following three and five months. Bone augmentation was analyzed histologically and via micro-computed tomography (µCT). Histomorphometric measurements of newly regenerated bone were performed on Masson-Trichrome stained sections: gained vertical bone height (gVBH), bone area (BA) and blood vessel density (BVD). Immuno-histological staining for human specific antigens was performed to localize hEPC in newly formed bone.

The results revealed that hEPC were CD31 positive, illustrated high proliferation rate and cobble stone morphology. Macroscopic view revealed, in all rats, hard augmented tissue firmly attached to the original calvaria. Histologically, this tissue was composed of bone, βTCP and vascular connective tissue. The coupling between osteogenesis and angiogenesis was demonstrated. Histomorphometric analyses revealed a significant increase in gVBH (~5mm) and BA following three and five months of hEPC transplantation (p≤0.05).Additionally, hEPC continued to enhance bone augmentation between three and five months (p≤0.05).

 µCT revealed no changes in the mineral density of the newly regenerated bone compared to the native bone. Similarly, BVD was 7-9 times higher in test group compared to control at both time points (p≤0.000). Moreover, hEPC expressing human specific CD31 were integrated into blood vessel walls adjacent to newly formed bone.

In conclusion, transplantation of hEPC in athymic rat calvaria model, combined with dome barrier significantly enhanced angiogenesis and extra-cortical vertical bone formation without affecting bone quality.