Identification and early characterization of genetically modified NGF-producing neural stem cells grafted into the injured adult rat brain.
OBJECTIVE
To investigate whether genetically modified mouse neural stem cells (NSC) expressing recombinant human nerve growth factor (rhNGF) and transplanted in chemically injured rat brain, can survive and eventually acquire phenotypic characteristics of early nerve cells.


METHODS
Stably high expression of rhNGF in NSC was obtained by a new lentivirus-mediated expression system. To test the effectiveness of hNGF secreted by rhNGF-NSC, hereby we performed either a bioassay for neurite outgrowth in PC12 rat cells or immunoblot analysis for TrkA, the high-affinity NGF receptor, from engineered NSC. rhNGF and mock-NSC were grafted into adult injured rats striatum and 3 days later, animals were killed, and brains were removed and examined by immunohistochemical analysis.


RESULTS
The results showed that rhNGF-producing NSC cultured for extended period of time release bioactive hNGF in the culture media which promotes PC12 neuronal differentiation and correlates with the up-regulation of TrkA. rhNGF-NSC transplanted into the injured brain can survive, produce hNGF and induce the expression of NGF receptors, p75(NTR) and TrkA.


CONCLUSIONS
In vitro and in vivo experiments confirmed the ability of rhNGF-NSC to secrete bioactive hNGF. Our data provide by means of genetically modified rhNGF-producing NSC, a useful experimental tool to test the potential clinical effectiveness of trophic factors relevant to central nervous system (CNS).
Pagani
Lucia
L
Consiglio Nazionale delle Ricerche, Istituto di Neurobiologia e Medicina Molecolare, CNR, NGF Section, Via del Fosso di Fiorano 64, 00143 Roma, Italy.
Cenciarelli
Carlo
C
Casalbore
Patrizia
P
Budoni
Manuela
M
Sposato
Valentina
V
Aloe
Luigi
L
D000704:Analysis of Variance; D000818:Animals; D000831:Animals, Newborn; D001930:Brain Injuries / Q000473:pathology / Q000628:therapy*; D002452:Cell Count / Q000379:methods; D002478:Cells, Cultured; D003342:Corpus Striatum / Q000378:metabolism; D017077:Culture Media, Conditioned / Q000494:pharmacology; D049452:Green Fluorescent Proteins / Q000096:biosynthesis; D006801:Humans; D051379:Mice; D020932:Nerve Growth Factor / Q000235:genetics* / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D016501:Neurites / Q000187:drug effects / Q000502:physiology; D009474:Neurons / Q000166:cytology; D051381:Rats; D020917:Receptor, trkA / Q000378:metabolism; D033581:Stem Cell Transplantation / Q000379:methods*; D013234:Stem Cells / Q000166:cytology* / Q000502:physiology; D014161:Transduction, Genetic
D017077:Culture Media, Conditioned; D009419:Nerve Tissue Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D020932:Nerve Growth Factor; D020917:Receptor, trkA