Resolution of the pathway taken by implanted Schwann cells to a spinal cord lesion by prior infection with a retrovirus encoding beta-galactosidase

Abstract

This series of experiments is designed to follow the fate of implanted Schwann cells by first labeling them with a recombinant retrovirus encoding the bacterial beta-galactosidase gene, then injecting them into the spinal cord after a demyelinating lesion has been produced. The label provides a means of distinguishing the exogenous Schwann cells from endogenous ones and of determining their travel pattern and myelinating or ensheathing behavior in the central nervous system (CNS). Neonatal rat primary Schwann cells were stimulated to divide by administering glial growth factor and forskolin. Fresh virus-containing supernatant from Psi2 cells producing retrovirus LZ1 was placed in cell culture to label the cells. The capacity of infected Schwann cells to form myelin was verified by coculturing in vitro with neurons from embryonic dorsal root ganglia. Infected cells were injected into the right side of adult syngenic rat spinal cords after a lysolecithin-induced demyelinating lesion had been produced 1 cm caudal on the left side. After 3 weeks the animals were killed, perfused for electron microscopy, and spinal cord sections histochemically stained for beta-galactosidase activity using the chromogenic substrate 5-bromo-4-chloro-3-indoyl-beta-D-galactosidase (X-Gal) which forms a blue precipitate in infected cells. The labeled cells, easily recognized macro- and microscopically, were clustered at the cell injection site, in the dorsal meninges and, at the area of demyelination, bilaterally in the superficial aspect of the dorsal funiculi. Labeled cells were not evident in the neuropil midway between the injection and demyelination sites.(ABSTRACT TRUNCATED AT 250 WORDS)