Glucocorticoids regulate glutaminase gene expression in human intestinal epithelial cells.
Glutamine is essential for intestinal metabolism and function, but its circulating and luminal availability to the mucosa may be diminished during critical illness. We hypothesized that glucocorticoids, which are produced in increased amounts during critical illness, accelerate mucosal glutamine metabolism. We studied intestinal glutamine utilization by examining the regulation of glutaminase in vitro, the enterocyte's principal enzyme of glutamine metabolism. Differentiated confluent human enterocytic cells (Caco-2 cells) were incubated with dexamethasone. Glutaminase activity was assayed and mRNA was extracted. Glutaminase transcripts were labeled with a 32P-labeled glutaminase cDNA probe, quantitated by phosphoimaging, and normalized to beta-actin. Dose- and time-response studies were performed. Dexamethasone-treated cells were also incubated with actinomycin D and cycloheximide. Dexamethasone (DEX) increased mucosal glutaminase activity by 45%, with maximal response at 12 hr. This increase was dose-dependent and was significant at doses of 1 and 10 microM. The dexamethasone-mediated increase in glutaminase activity was associated with a 40% increase in glutaminase mRNA. The DEX-induced increase in glutaminase activity was inhibited by actinomycin D and cycloheximide, indicating the requirement for de novo RNA and protein synthesis. Glucocorticoids stimulate glutamine metabolism in these human enterocytic cells by increasing the activity of glutaminase, a response that is preceded by an increase in gene transcription. This glucocorticoid-mediated increase in glutaminase activity may be a mechanism by which gut glutamine metabolism is maintained during critical illness when blood glutamine levels are diminished and food intake is often interrupted.
Sarantos
P
P
Department of Surgery, University of Florida College of Medicine, Gainesville 32611.
Abouhamze
Z
Z
Copeland
E M
EM
Souba
W W
WW
D001345:Autoradiography; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology; D015870:Gene Expression; D005938:Glucocorticoids / Q000494:pharmacology*; D005972:Glutaminase / Q000235:genetics*; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology / Q000201:enzymology*; D012333:RNA, Messenger / Q000378:metabolism; D014407:Tumor Cells, Cultured
D005938:Glucocorticoids; D012333:RNA, Messenger; D003609:Dactinomycin; D003907:Dexamethasone; D003513:Cycloheximide; D005972:Glutaminase