Protocol: Analysis of gene expression of aldose reductase and sorbitol dehydrogenase in rat Schwann cells by competitive RT-PCR method using non-homologous DNA standards

• Published in: Brain research. Brain research protocols 2001-12, Vol.8 (3), p.219-227
• Main Authors: Maekawa, Keiko, Tanimoto, Tsuyoshi, Okada, Satoshi, Suzuki, Takeshi, Suzuki, Tsunemasa, Yabe-Nishimura, Chihiro
• Format: Article
• Language: English
• ISSN: 1385-299X
• DOI: 10.1016/S1385-299X(01)00121-0

Aldose reductase (AR) and sorbitol dehydrogenase (SDH) are the enzymes constituting the polyol pathway, an alternate route of glucose metabolism. A wealth of experimental data has indicated the involvement of the polyol pathway in the pathogenesis of diabetic complications. However, there has been surprisingly little research on the relative abundance of SDH to AR in the tissues affected in diabetes. We therefore developed a competitive RT-PCR system to simultaneously determine the mRNA levels of these two enzymes in small amounts of samples, and studied their expression in Schwann cells isolated from adult rat sciatic nerves. Although both AR and SDH mRNA were expressed in the Schwann cells, the levels of SDH cDNA were much lower than those of AR cDNA. The induction of AR mRNA expression in the Schwann cells under hyperosmotic conditions was similarly detected by Northern blot analysis and our competitive RT-PCR method. The RT-PCR system developed in this study may be a useful tool in ascertaining the relative contributions of AR and SDH to the metabolic derangements resulting from the acceleration of polyol pathway activity in the target organ of diabetic complications.