Analysis of TGF-beta 1 gene expression in contused rat spinal cord using quantitative RT-PCR.

ABSTRACT

We have used northern blot analysis and quantitative reverse transcription polymerase chain reaction (RT-PCR) to determine the postinjury expression profile of the transforming growth factor-beta 1 (TGF-beta 1) gene in the contused rat spinal cord. Spectrophotometric estimates of total sample RNA and quantitative analyses of cyclophilin mRNA using RT-PCR served as controls for comparisons between samples. No changes in cyclophilin gene expression were found at any postinjury survival times. The results of the TGF-beta 1 analyses, which were carried out on spinal cord samples taken at postinjury intervals ranging from 6 h to 10 days, show that the amount of TGF-beta 1 mRNA present in spinal cord increases rapidly following injury, reaching maximum levels 7 days postinjury. Unoperated control samples contained approximately 2 x 10(8) molecules of TGF-beta 1 mRNA/0.5 microgram total RNA. By 1 day postinjury, the amount of TGF-beta 1 mRNA in the cord had increased by a factor of 2.5 to 5 x 10(8) molecules/0.5 microgram total RNA. At 7 days postinjury, there were approximately 15 x 10(8) molecules of TGF-beta 1 mRNA/0.5 microgram total RNA. By 10 days postinjury the amount of TGF-beta 1 mRNA present in the spinal cord had declined to 8 x 10(8) molecules of TGF-beta 1 mRNA/0.5 microgram total RNA, a value similar to that observed at 3 days postinjury. The roles that TGF-beta 1 might play in modifying cellular responses in injured spinal cord are discussed.