The regrowth of axons within tissue defects in the CNS is promoted by implanted hydrogel matrices that contain BDNF and CNTF producing fibroblasts.
In this study we demonstrate the potential for combining biocompatible polymers with genetically engineered cells to elicit axon regrowth across tissue defects in the injured CNS. Eighteen- to 21-day-old rats received implants of poly N-(2-hydroxypropyl)-methacrylamide (HPMA) hydrogels containing RGD peptide sequences that had been infiltrated with control (untransfected) fibroblasts (n = 8), fibroblasts engineered to express brain-derived neurotrophic factor (BDNF) (n = 5), ciliary neurotrophic factor (CNTF) (n = 5), or a mixture of BDNF and CNTF expressing fibroblasts (n = 11). Fibroblasts were prelabeled with Hoechst 33342. Cell/polymer constructs were inserted into cavities made in the left optic tract, between thalamus and superior colliculus. After 4-8 weeks, retinal projections were analyzed by injecting right eyes with cholera toxin (B-subunit). Rats were perfused 24 h later and sections were immunoreacted to visualize retinal axons, other axons (RT97 antibody), host astrocytes and macrophages, donor fibroblasts, and extracellular matrix molecules. The volume fraction (VF) of each gel that was occupied by RT97(+) axons was quantified. RT-PCR confirmed expression of the transgenes prior to, and 5 weeks after, transplantation. Compared to control rats (mean VF = 0.02 +/- 0.01% SEM) there was increased ingrowth of RT97(+) axons into implants in CNTF (mean VF = 0.33 +/- 0.19%) and BDNF (mean VF = 0.62 +/-0.19%) groups. Axon growth into hydrogels in the mixed BDNF/CNTF group (mean VF = 3.58 +/- 0.92%) was significantly greater (P < 0.05) than in the BDNF or CNTF fibroblast groups. Retinal axons exhibited a complex branching pattern within gels containing BDNF or BDNF/CNTF fibroblasts; however, they regrew the greatest distances within implants containing both BDNF and CNTF expressing cells.
Loh
N K
NK
Department of Anatomy and Human Biology, The University of Western Australia, Crawley, Perth, WA 6009, Australia.
Woerly
S
S
Bunt
S M
SM
Wilton
S D
SD
Harvey
A R
AR
D000818:Animals; D000943:Antigens, Differentiation / Q000378:metabolism; D001369:Axons / Q000187:drug effects / Q000378:metabolism*; D001930:Brain Injuries / Q000473:pathology / Q000628:therapy*; D019208:Brain-Derived Neurotrophic Factor / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D020934:Ciliary Neurotrophic Factor / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology; D004195:Disease Models, Animal; D004343:Drug Implants; D005109:Extracellular Matrix / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism* / Q000637:transplantation; D005353:Fibronectins / Q000378:metabolism; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000378:metabolism / Q000494:pharmacology; D051381:Rats; D011916:Rats, Inbred F344; D012160:Retina / Q000166:cytology; D013477:Superior Colliculi / Q000166:cytology; D013788:Thalamus / Q000166:cytology; D019076:Transgenes; D014795:Visual Pathways / Q000187:drug effects / Q000473:pathology
D000943:Antigens, Differentiation; D019208:Brain-Derived Neurotrophic Factor; D020934:Ciliary Neurotrophic Factor; D004343:Drug Implants; D005353:Fibronectins; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate