Identification of the 11 beta-hydroxysteroid dehydrogenase type 1 mRNA and protein in human mononuclear leukocytes.
The enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) catalyzes the interconversion between inactive 11-ketoglucocorticoids and their active 11beta-hydroxy derivatives, such as cortisol and corticosterone. We have investigated the expression of 11beta-HSD1 in freshly isolated human peripheral mononuclear leukocytes (MNL). The presence of 11beta-HSD1 mRNA was demonstrated in total RNA by RT-PCR using specific primers designed on the 4th and 5th exons of the human 11beta-HSD1 gene. Fragments of the expected size were consistently detected on agarose gels, and sequencing showed complete identity with the corresponding sequence deposited in GenBank. The occurrence of 11beta-HSD1 protein was established by Western immunoblot analysis with a specific polyclonal antibody. Enzyme oxo-reductase activity was investigated by incubating 12 samples of MNL isolated from from 8 subjects with [3H]cortisone and formation of cortisol was established only in 4 subjects (yield range: 0.15-1.3%) after acetylation and TLC, blank subtraction and correction for losses. 18beta-Glycyrrhetinic acid, an inhibitor of 11 beta-HSD1, reduced cortisol production below detection limit. Dehydrogenase activity could not be demonstrated. It is suggested that, although enzyme activity of 11beta-HSD1 in circulating MNL is low, it is apparently ready for enhancement after MNL migration to sites of inflammation.
Fiore
C
C
Department of Medical and Surgical Sciences, Endocrinology- University of Padua, Padua 35128, Italy.
Nardi
A
A
Dalla Valle
L
L
Pellati
D
D
Krozowski
Z
Z
Colombo
L
L
Armanini
D
D
D043205:11-beta-Hydroxysteroid Dehydrogenase Type 1 / Q000235:genetics / Q000378:metabolism*; D000328:Adult; D015153:Blotting, Western; D003348:Cortisone / Q000378:metabolism; D005260:Female; D006801:Humans; D006854:Hydrocortisone / Q000096:biosynthesis; D007963:Leukocytes, Mononuclear / Q000378:metabolism*; D057230:Limit of Detection; D008297:Male; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
D012333:RNA, Messenger; D043205:11-beta-Hydroxysteroid Dehydrogenase Type 1; D003348:Cortisone; D006854:Hydrocortisone