title: Human Adipose-Derived Micro-vessel Constructs: a potential method for experimental bone regeneration
Creators: name: Boston, Bridget A
Publisher: name: Griffith University
Year: 2021
Subjects: subject: medication-related osteonecrosis of the jaws (MRONJ)
subject: human-adipose derived microvessel (HAMV)
subject: bone
subject: regeneration and healing
subject: 3-dimensional (3D) collagen hydrogel

description: Bisphosphonates are a group of drugs widely used in medical practice to prevent bone destruction, occurring in diseases like osteoporosis and bone cancer. An unfortunate sideeffect of these potent drugs is the subsequent death of areas of bone in the jaw following minor local trauma, commonly occurring during dental treatment. This unique, but welldocumented, adverse effect is termed medication-related osteonecrosis of the jaws (MRONJ). While the precise cause of MRONJ is still unclear, research suggests the inhibition of new blood vessel formation, limiting the capacity for bone healing, may be a significant factor. Unfortunately, treatment options for MRONJ are very limited, and despite successful development of bone mimetic materials, most available materials do not induce sufficient formation of blood vessels. Adequate vascularisation is crucial for timely and adequate transport of nutrients and waste removal, and the provision of progenitor cells for tissue remodeling and repair. This study provides a rationale for a new innovative approach using a human-adipose derived microvessel (HAMV)-osteoblast construct, which will potentially support local bone regeneration and healing. This project takes the important first steps in vitro, investigating the growth, migration, and viability of the HAMV within a 3D hydrogel construct and assessing the influence of HAMV development on osteogenesis, in order to provide the data necessary for subsequent testing in a pre-clinical animal model. In this study, HAMV were seeded into tissue culture plates and cultured over 28 days, to evaluate the proliferation potential of the HAMV in a 3-dimensional (3D) collagen hydrogel construct. Growth, proliferation and morphological changes, including cell death, of HAMV were also visualised and were examined via LIVE/DEAD® assay with the aid of confocal microscopy. The human osteoblast cell line, MG63, was cultured in conditioned media derived from the HAMV cultures to understand the influence of HAMV on bone cells. qPCR was employed to assess the changes in the relative expression of osteoblast differentiation markers during culture with HAMV conditioned media. Cell viability and morphological changes were recorded using a common cell-culture inverted microscope. Initial proliferation assay and confocal microscopy showed robust proliferation and outgrowth of HAMV from two weeks of HAMV culture. qPCR analysis demonstrated the ability of the HAMVconditioned medium to trigger the expression of osteogenic differentiation marker/genes of bone cells by day 14, compared to cells that did not receive HAMV-conditioned media. Overall, this study demonstrated HAMV can be incorporated into 3D collagen hydrogel and make viable deliverable constructs. This study showed that HAMV-conditioned medium has the ability to modulate the osteogenic potential of osteoblasts, suggesting that HAMV could be used as a viable tool in various regenerative medicine experiments. When considering clinical treatment aspects, this study indicates that a microvessel encapsulated in collagen hydrogel construct, derived from adipose tissue, may be an efficient therapy of MRONJ, promoting osteogenesis and vascularisation.