The use of genetically altered astrocytes to provide nerve growth factor to adrenal chromaffin cells grafted into the striatum

Abstract

Transplantation of adrenal chromaffin cells into the striatum of Parkinson's disease patients is unlikely to become a reliable therapy unless techniques are devised to improve cell survival. To address this issue, we investigated the use of genetically altered astrocytes that constitutively secrete beta-nerve growth factor (NGF) to provide trophic support for adrenal chromaffin cells grafted into the dopamine-denervated striatum of the rat. Primary rat astrocytes were altered genetically in vitro by infection with a retroviral vector harboring a mouse beta-NGF transgene under constitutive long terminal repeat transcriptional control. Confluent cultures of these genetically altered astrocytes secrete NGF into their culture medium at a rate of approximately 9 pg/10(5) cells/h. This rate of NGF secretion is at least 10-fold higher than that of confluent sister cultures of uninfected astrocytes. The effects of the NGF-secreting astrocytes on the survival and neuronal transformation of dissociated adrenal chromaffin cells were assessed in vitro and following transplantation into the dopamine-denervated striatum of the adult rat. In vitro experiments demonstrated that neuritic outgrowth is stimulated when postnatal day 12 chromaffin cells are grown on a monolayer of the genetically altered astrocytes. When co-grafted with genetically altered astrocytes, young postnatal chromaffin cells displayed extensive neuritic outgrowth within the host brain 2 weeks postimplantation, whereas chromaffin cells grafted alone or with normal astrocytes retain an endocrine-like morphology. Survival of the chromaffin cells is also enhanced 3-6-fold when co-grafted with the genetically altered astrocytes. In addition, the neuronally transformed chromaffin cells appear to lose adrenergic properties as assessed by diminished immunoreactivity to the adrenergic marker, phenylethanolamine-N-methyltransferase. Although their survival is also enhanced approximately 4-fold relative to controls, adult chromaffin cells do not convert to a neuronal morphology when co-grafted with the genetically altered astrocytes. These studies demonstrate that rat astrocytes carrying a mouse NGF transgene provide trophic support for intrastriatal chromaffin cell grafts.