title: The development of a rat in vitro model of spinal cord injury demonstrating the additive effects of Rho and ROCK inhibitors on neurite outgrowth and myelination.
abstract: It is currently thought that treatment for spinal cord injury (SCI) will involve a combined pharmacological and biological approach; however, testing their efficacy in animal models of SCI is time-consuming and requires large animal cohorts. For this reason we have modified our myelinating cultures as an in vitro model of SCI and studied its potential as a prescreen for combined therapeutics. This culture comprises dissociated rat embryonic spinal cord cells plated onto a monolayer of astrocytes, which form myelinated axons interspaced with nodes of Ranvier. After cutting the culture, an initial cell-free area appears persistently devoid of neurites, accompanied over time by many features of SCI, including demyelination and reduced neurite density adjacent to the lesion, and infiltration of microglia and reactive astrocytes into the lesioned area. We tested a range of concentrations of the Rho inhibitor C3 transferase (C3) and ROCK inhibitor Y27632 that have been shown to promote SCI repair in vivo. C3 promoted neurite extension into the lesion and enhanced neurite density in surrounding areas but failed to induce remyelination. In contrast, while Y27632 did not induce significant neurite outgrowth, myelination adjacent to the lesion was dramatically enhanced. The effects of the inhibitors were concentration-dependent. Combined treatment with C3 and Y27632 had additive affects with an enhancement of neurite outgrowth and increased myelination adjacent to the lesion, demonstrating neither conflicting nor synergistic effects when coadministered. Overall, these results demonstrate that this culture serves as a useful tool to study combined strategies that promote CNS repair.
authors.affiliation: Institute of Infection, Immunity and Inflammation, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8TA, United Kingdom.
authors.forename: Stephanie D
authors.initials: SD
authors.lastname: Boomkamp
authors.forename: Mathis O
authors.initials: MO
authors.lastname: Riehle
authors.forename: Jenifer
authors.initials: J
authors.lastname: Wood
authors.forename: Michael F
authors.initials: MF
authors.lastname: Olson
authors.forename: Susan C
authors.initials: SC
authors.lastname: Barnett
mesh_terms: D000577:Amides / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D001253:Astrocytes / Q000187:drug effects*; D039481:CD11b Antigen / Q000378:metabolism; D002478:Cells, Cultured; D003176:Complement C3 / Q000627:therapeutic use; D003711:Demyelinating Diseases / Q000188:drug therapy / Q000209:etiology / Q000378:metabolism; D004622:Embryo, Mammalian; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins / Q000378:metabolism; D020690:GTPase-Activating Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D005911:Gliosis / Q000188:drug therapy / Q000209:etiology / Q000473:pathology; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D018991:Myelin Proteolipid Protein / Q000378:metabolism; D009186:Myelin Sheath / Q000187:drug effects / Q000502:physiology*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D016501:Neurites / Q000187:drug effects*; D009474:Neurons / Q000166:cytology / Q000187:drug effects; D019081:O Antigens / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013116:Spinal Cord / Q000166:cytology*; D013997:Time Factors; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D000577:Amides; D039481:CD11b Antigen; D003176:Complement C3; D004791:Enzyme Inhibitors; C490174:Fabp7 protein, rat; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D020690:GTPase-Activating Proteins; D005904:Glial Fibrillary Acidic Protein; D008869:Microtubule-Associated Proteins; D018991:Myelin Proteolipid Protein; D009419:Nerve Tissue Proteins; D019081:O Antigens; D011725:Pyridines; C085379:rho GTPase-activating protein; C108830:Y 27632; D054460:rho-Associated Kinases