A Novel Model: Microenvironmentally-induced Carcinogenesis For Screening Bone Regeneration Scaffolds

The aim of this study is to develop a subcutaneous model in NOD/SCID IL2 R gamma null (NOG) mice to evaluate the tumorigenic potential of functionalized/bioactive scaffolds designed for the use in bone tissue engineering.

Method:

Dysplastic (precancerous) oral keratinocytes (DOK cell line) known not to be tumorigenic in NUDE mice were injected subcutaneously in NOG mice alone (n=6) or together with carcinoma associated fibroblasts (CAF) (n=12) in order to develop an experimental model of microenvironmentally-induced carcinogenesis. DOK were then transfected to express luciferase such that bioluminescence in vivo imaging of cells/scaffold construct could be carried out in parallel with classical morphometric analysis. They were then cultured alone (n=6) or together with CAF (n=6) on poly(L-lactide-co-e-caprolactone) scaffolds and implanted subcutaneously in NOG mice to optimize the model for future use in screening of various scaffolds intended for bone regeneration.

Result:

When 10³ DOK cells were xenotransplanted alone they did not form tumors, while 66.66% of the mice injected with 10³ DOK and 10⁵ CAF formed tumors. Total photon count from in vivo imaging using luciferase-transfected DOK with different densities showed significantly higher bioluminescence intensity of scaffolds xenotransplanted with DOK and CAFs than of DOK alone for all seeding densities, which was just above the threshold one week after xenotransplantation and throughout the whole period (12 weeks) of in vivo imaging.

Conclusion:

An experimental model of microenvironmentally-induced carcinogenesis has been established, with DOK injected alone as a negative control and DOK co-transplanted with CAFs as a positive control. By use of bioluminescence, this model provides also an easier and more reliable monitoring bioassay for the tumorigenic potential of functionalized scaffolds used for bone regeneration. 

Key words: Tumorigenicity, scaffolds, functionalized scaffolds, bone tissue engineering