Evidence that a leukocyte type of 12-lipoxygenase is expressed and regulated by angiotensin II in human adrenal glomerulosa cells.
The 12-lipoxygenase (LO) pathway of arachidonic acid plays an important role in angiotensin II (AII)-mediated aldosterone synthesis. Several distinct isoforms of 12-LO have been cloned. However, in humans only the platelet form of 12-LO has been reported to be present. Western immunoblotting analysis in cultured human adrenal glomerulosa cells using polyclonal antibodies to porcine leukocyte 12-LO enzyme or peptide showed a specific 72-kilodalton band, which is identical to the molecular size of the porcine leukocyte form of 12-LO. In addition, AII (10(-7)) increased the intensity of the 72-kilodalton band nearly 2-fold over basal. In situ hybridization analysis indicated a strong positive reaction with the porcine leukocyte type of 12-LO antisense riboprobe in the zona glomerulosa of the adrenal cortex. The 12-LO probe also recognized a near 4-kilobase messenger RNA (mRNA) from human glomerulosa cells in Northern blots. Since the leukocyte type of 12-LO is highly homologous to human 15-LO, a reverse transcriptase and polymerase chain reaction was used to analyze the specific type of 12-LO mRNA in human cells. The mRNA for a porcine leukocyte type of 12-LO was detected in human adrenal glomerulosa cells, and the level of 12-LO transcripts was increased approximately 60-fold by AII (10(-7) M). The leukocyte type of 12-LO also was detected in human monocyte-like U937 cells, but not in IM-9 lymphocytes or human erythroleukemia cells. These results suggest that human adrenal glomerulosa cells and human monocyte-like U937 cells express a 12-LO which has immunological and molecular biological characteristics similar to the porcine leukocyte 12-LO.
Gu
J L
JL
Department of Diabetes, Endocrinology, and Metabolism, City of Hope National Medical Center, Duarte, California 91010.
Natarajan
R
R
Ben-Ezra
J
J
Valente
G
G
Scott
S
S
Yoshimoto
T
T
Yamamoto
S
S
Rossi
J J
JJ
Nadler
J L
JL
D000804:Angiotensin II / Q000502:physiology*; D001092:Arachidonate 12-Lipoxygenase / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D015152:Blotting, Northern; D002478:Cells, Cultured; D006801:Humans; D017403:In Situ Hybridization; D007962:Leukocytes / Q000201:enzymology*; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes / Q000235:genetics; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D014158:Transcription, Genetic; D015384:Zona Glomerulosa / Q000166:cytology / Q000378:metabolism*
D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D000804:Angiotensin II; D001092:Arachidonate 12-Lipoxygenase