Objectives: A brushite-forming calcium phosphate cement (CPC) stabilized by poly (l-lactide-co-glycolide) acid (PLGA) fibers (≤ 10% w/w) showed high biocompatibility and its fiber component enhanced bone formation in sheep lumbar vertebroplasty. However, CPC effects on the osteogenic differentiation of resident mesenchymal stem cells remained unexplored. Thus, the present study analyzed the influence of a CPC scaffold with a low PLGA proportion (as a mimicry of natural bone) on pluripotency, and osteogenic, chondrogenic, and adipogenic differentiation of human adipose tissue-derived mesenchymal stem cells (hASCs).
Methods: hASCs were cultivated on CPC discs with/without PLGA fibers (5 and 10%) in the absence of osteogenic medium for 3, 7, and 14 days. Gene expression of pluripotency (Nanog, Tert, Rex), osteogenic (Runx2, osterix, alkaline phosphatase, collagen I, osteonectin, osteopontin, osteocalcin), chondrogenic (collagen II, Sox9, aggrecan), and adipogenic markers (PPARG, Leptin, and FABP4) was analyzed by RT-PCR, alkaline phosphatase (ALP) activity by enzme assay, and cell number and viability by fluorescein diacetate/propidium iodide staining.
Results: Compared to pure CPC, hASCs cultured on fiber-reinforced CPC showed transiently induced gene expression of the osteogenic transcription factors Runx2 and osterix (day 3), and long-lastingly augmented expression of the bone markers alkaline phosphatase (including its activity), collagen I, and osteonectin (until day 14). In contrast, augmented expression of all chondrogenic, adipogenic and pluripotency markers was limited to day 3, followed by a significant downregulation on days 7 and 14. Cultivation of hASCs on fiber-reinforced CPC reduced the cell number, but not the proportion of viable cells (viability > 95%).
Conclusions: The PLGA component of fiber-reinforced, brushite-forming CPC supports long-lasting osteogenic differentiation of hASCs, whereas chondrogenesis, adipogenesis, and pluripotency are only transiently augmented. In parallel to the augmentation of bone formation by fiber addition in vivo, PLGA fibers may represent an interesting clinical target for future improvement of CPC-based bone regeneration.