A possible injectable tissue engineered nucleus pulposus constructed with platelet-rich plasma and ADSCs in vitro

Abstract

BackgroundInjectable tissue engineered nucleus pulposus is a new idea for minimally invasive repair of degenerative intervertebral disc. The platelet-rich plasma (PRP) and adipose-derived stromal cells (ADSCs) could be harvested from autologous tissue easily. PRP contains numerous autologous growth factors and has reticulate fibrous structure which may have the potential to make ADSCs differentiate into nucleus pulposus-like cells. The goal of this study was to explore the feasibility of constructing a possible injectable tissue engineered nucleus pulposus with PRP gel scaffold and ADSCs.MethodsAfter identification with flow cytometry, the rabbit ADSCs were seeded into PRP gel and cultured in vitro. At the 2nd, 4th, and 8th week, the PRP gel/ADSCs complex was observed by macroscopy, histological staining, BrdU immunofluorescence, and scanning electron microscopy. The glycosaminoglycans (GAG) in the PRP gel/ADSCs complex were measured by safranin O staining with spectrophotometry. In PRP gel/ADSCs complex, gene expression of HIF-1α, aggrecan, type II collagen were tested by RT-PCR. The injectability of this complex was evaluated.ResultsMacroscopically, the complex was solidified into gel with smooth surface and good elasticity. The safranin O dye was almost no positive staining at 2nd week; however, the positive staining of extracellular matrix was enhanced obviously at 4th and 8th week. The HE staining and SEM demonstrated that the cells were well-distributed in the reticulate scaffold. BrdU immunofluorescence showed that ADSCs can survive and proliferate in PRP gel at each time points. The level of GAG at 4th week was higher than those at 2nd week (P < 0.05), and significant difference was also noted between 4th and 8th week (P < 0.05). HIF-1α, aggrecan, type II collagen gene expression at 4th week were much more than those at 2nd week (P < 0.05), and significant differences were also noted between 4th and 8th week (P < 0.05). The flow rate of complex was 0.287 mL/min when passed through the 19-gauge needle with the 100 mmHg injection pressure.ConclusionsOur preliminary findings suggest that the PRP gel make it possible for rabbit ADSCs differentiated into nucleus pulposus-like cells after coculture in vitro. According to the results, it is a better feasible method for construction of autologous injectable tissue engineered nucleus pulposus.