5A and B), and exhibited an elongated fibroblast-like morphology after 3 d, which indicated a viable state. A continuous increase in cell number was visually observed for both fibers. After 21 d, LIVE/DEAD? staining Baricitinib cost determined the presence of 96.5 �� 2.5% and 97.1 �� 1.5% live cells for PCL and PCL-��-CD fibers, respectively. Figure 5. hADSCs cultured on either PCL or PCL-��-CD fibers are stained for (A) LIVE/DEAD? Viability/Cytotoxicity (InvitrogenTM, Life Technologies, Grand Island, NY) using calcein AM, ethidium homodimer-1 and Hoechst dye; (B) F-actin … Positive staining with alizarin red and ALP (Fig. 5C) was observed on both fibers, which confirmed calcium deposition and mineralization. A substantial increase in the intensity of alizarin red staining was observed from day 14 to day 21 on both fibers, suggesting that by day 21, mineral deposition was greatly enhanced (Fig.
5C). We then performed quantitative analysis of osteogenic gene-expression. Four osteogenic markers were selected for this study: osteogenesis transcription factor Runx2, and three bone collagen structural proteins: osteopontin, collagen type I and collagen type X. In general, PCL-��-CD fibers induced greater amounts of osteogenic gene expression compared with PCL fibers (Fig. 6A�CD). Similarly, relatively higher collagen deposition was obtained on PCL-��-CD fibers (Fig. 6E and F). In summary, ADSCs proliferated at a similar rate on both types of fibers, while PCL-��-CD fibers enhanced osteogenesis. Figure 6.
The relative gene expression of some osteogenic markers during osteogenesis of hADSCs seeded on PCL and PCL-��-CD fibers, including RunX2 (A), osteopontin (B), collagen type I (C) and collagen type X (D); biochemical assays showing … Discussion Scaffolds based on aliphatic polyesters, such as PCL nanofibers, have been successfully used in biomedical applications, including stem cell culture and differentiation.12,15,16,48 However, depending on the desired cell function and tissue formation outcomes, aliphatic polyester nanofibers are often chemically modified with bioactive molecules and cell-recognizable ligands by mimicking natural ECM��s chemical and biological cues. In this context, our reported PCL-��-CD-based electrospun nanofibrous scaffold has unique advantages: first, it is as easy to fabricate as PCL fibers; second, it has multiple functional sites for further conjugation and third, it is independent of the PCL-main chain modification as ��-CD physically threads onto PCL chains.
The ease of conjugation of various Drug_discovery chemical and biological components to create user-specific unique cell environments without PCL modification, makes these nanofibers a powerful biomaterial tool for tissue engineering. For example, we showed the utility of the hydroxyl groups of the ��-CD on the fiber surface by conjugating a fluorescent small molecule, fluorescamine, and a polystyrene nanobead (Figs. 3 and and4).4).