Expression of cytochrome P450 side-chain cleavage enzyme and 3 beta-hydroxysteroid dehydrogenase in the rat central nervous system: a study by polymerase chain reaction and in situ hybridization.
In examining steroid synthesis in the CNS, expression of the mRNAs encoding for cytochrome P450 side-chain cleavage enzyme (P450scc) and 3 beta-hydroxy-steroid dehydrogenase/delta 5-delta 4 isomerase (3 beta-HSD) has been studied in the rat brain. P450scc transforms cholesterol into pregnenolone and 3 beta-HSD transforms pregnenolone into progesterone. PCR was used to amplify cDNA sequences from total RNA extracts. Classical steroidogenic tissues, like adrenal and testis, as well as the non-steroidogenic tissue lung have been used as controls. The expression of P450scc and 3 beta-HSD have been demonstrated by PCR in cortex, cerebellum, and spinal cord. In addition, primary cultures of rat cerebellar glial cells and rat cerebellar granule cells were found to express P450scc and 3 beta-HSD at comparable levels. Furthermore, three of the four known isoenzymes of 3 beta-HSD were identified, as determined using selective PCR primers coupled with discriminative restriction enzymes and sequencing analysis of the amplified brain products. Using RNA probes, in situ hybridization indicated that P450scc and 3 beta-HSD are expressed throughout the brain at a low level and mainly in white matter. Enrichment of glial cell cultures in oligodendrocytes, however, does not increase the relative abundance of P450scc and 3 beta-HSD mRNA detected by PCR. This discrepancy suggests that the developmental state of cultured cells and their intercellular environment may be critical for regulating the expression of these enzymes. These findings support the proposal that the brain apparently has the capacity to synthesize progesterone from cholesterol, through pregnenolone, but that the expression of these enzymes appears to be quite low.(ABSTRACT TRUNCATED AT 250 WORDS)
Sanne
J L
JL
Fidia-Gerogetown Institute for the Neurosciences, Georgetown University School of Medicine, Washington, D.C. 20007, USA.
Krueger
K E
KE
D015096:3-Hydroxysteroid Dehydrogenases / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000378:metabolism*; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics; D017403:In Situ Hybridization; D015335:Molecular Probes / Q000235:genetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014018:Tissue Distribution
D018076:DNA, Complementary; D015335:Molecular Probes; D012333:RNA, Messenger; D015096:3-Hydroxysteroid Dehydrogenases; D002786:Cholesterol Side-Chain Cleavage Enzyme