Surface modification of PHBV nanofiber mats for rapid cell cultivation and harvesting.

To maintain the original function of a specific tissue for therapeutic tissue engineering, an advanced cell culture surface for repeat cell proliferation is necessary. We designed a novel cell proliferation and rapid harvesting surface by combining nonwoven nanofiber mat and a thermo-responsive polymer. Nanofibrous poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) mats were fabricated by the electrospinning technique. A poly(N-isopropylacrylamide) (PNIPAM) thermo-responsive layer was grafted on the PHBV nanofiber mat by electron beam irradiation. The average diameter of the PNIPAM-grafted PHBV nanofibers was determined by SEM. ATR-FTIR and ESCA were used to confirm the grafting of PNIPAM onto the PHBV nanofiber surface. Water contact angles on the mats were measured in response to temperature changes. Human adipose-derived stem cells (ADSCs) were cultured on the PNIPAM-grafted PHBV nanofiber mat to investigate cell proliferation, harvesting, and functionality during repeat subculture. Detached ADSCs from each surface by low temperature treatment and trypsin-EDTA were compared by a fluorescence-activated cell sorter (FACS) using expression of stem cell membrane-specific markers such as CD-13 PE, CD-29 PE, and CD-90 FITC. The mass cultivation and intact harvesting of stem cells by low temperature treatment using a thermo-responsive PHBV nanofiber mat is a promising technique for use in regenerative medicine and stem cell therapy.

Fabrication and characterization of thermoresponsive polystyrene nanofibrous mats for cultured cell recovery.

Rapid cell growth and rapid recovery of intact cultured cells are an invaluable technique to maintain the biological functions and viability of cells. To achieve this goal, thermoresponsive polystyrene (PS) nanofibrous mat was fabricated by electrospinning of PS solution, followed by the graft polymerization of thermoresponsive poly(N-isopropylacrylamide)(PIPAAm) on PS nanofibrous mats. Image analysis of the PS nanofiber revealed a unimodal distribution pattern with 400 nm average fiber diameter. Graft polymerization of PIPAAm on PS nanofibrous mats was confirmed by spectroscopic methods such as ATR-FTIR, ESCA, and AFM. Human fibroblasts were cultured on four different surfaces, PIPAAm-grafted and ungrafted PS dishes and PIPAAm-grafted and ungrafted PS nanofibrous mats, respectively. Cells on PIPAAm-grafted PS nanofibrous mats were well attached, spread, and proliferated significantly much more than those on other surfaces. Cultured cells were easily detached from the PIPAAm-grafted surfaces by decreasing culture temperature to 20 °C, while negligible cells were detached from ungrafted surfaces. Moreover, cells on PIPAAm-grafted PS nanofibrous mats were detached more rapidly than those on PIPAAm-grafted PS dishes. These results suggest that thermoresponsive nanofibrous mats are attractive cell culture substrates which enable rapid cell growth and recovery from the culture surface for application to tissue engineering and regenerative medicine.