Muristerone A-induced nerve growth factor release from genetically engineered human dermal fibroblasts for peripheral nerve tissue engineering

In this study, human dermal fibroblasts (hDFBs) were genetically modified t o release human nerve growth factor (NGF) using an ecdysone-inducible syste m. NGF cDNA was inserted into the pIND vector and then hDFBs were cotransfe cted with pIND-NGF and pVgRXR. Muristerone A, an analog of ecdysone, was us ed as the inducing agent. NGF release from transfected hDFBs was assessed i n vitro and in vivo. Transfected hDFBs in the presence of Muristerone A pos sessed a maximal in vitro release of 8.5 +/- 0.4 pg of NGF/mL per 10(3) cel ls, demonstrating significantly higher NGF levels compared to control hDFBs . The in vitro release rate curve for transfected hDFBs in the presence of Muristerone A exhibited a maximum of 5.1 +/- 0.2 ng NGF/10(6) cells/day. A PC-12 bioassay demonstrated that the in vitro NGF released is bioactive. Wh en transfected hDFBs in the presence of Muristerone A were placed in vivo i n nude rats, NGF levels reach 2074 +/- 257 pg/mL and 1620 +/- 132 pg/mL at 24 and 48 h, respectively. These levels were significantly higher than nega tive control and wound fluid levels. Results support further in vivo invest igation of this molecular "on" switch for peripheral nerve regeneration.