Co-expression of mRNA for Neurotrophic Factors in Human Neurons and Glial Cells in Culture

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Abstract

Enriched populations of neurons and astrocytes of 93-99% purity were obtained from mixed cultures of four human fetal brains. Total cellular RNA was extracted from cell pellets and reverse transcribed into cDNA. Five ^1 of cDNA were subjected to PCR amplification using primers specific for sequences of NGF, BDNF, NT-3 and CNTF. PCR products were separated through 5% acrylamide gel and identified by DNA sequencing. Results showed that neurons expressed detectable levels of mRNA for NGF in all four cultures; BDNF and NT-3 mRNA was seen only in two cultures; CNTF mRNA was not detected in all four cultures. Astrocytes expressed mRNA for NGF, BDNF, and NT-3 but not for CNTF in all cultures examined. Astrocytic expression of mRNA for NGF, BDNF and NT-3 was found during the active cell proliferation as well as at a phase of mitotic quiescence. This study provides evidence that dissociated cell cultures of human neurons produce NGF, BDNF and NT-3 in early stages of their development and that astrocytes are constitutively committed to synthesize neurotrophic factors, NGF, BDNF and NT-3. The active synthesis of selected neurotrophic factors by neurons and astrocytes is relevant in supporting migration, survival and differentiation of developing neurons in vivo.

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