Oxidative stress-induced inhibition of adrenal steroidogenesis requires participation of p38 mitogen-activated protein kinase signaling pathway.
Previous studies from this laboratory identified excessive oxidative stress as an important mediator of age-related decline in steroid hormone production. Here, we investigated whether oxidative stress exerts its antisteroidogenic action through modulation of oxidant-sensitive mitogen-activated protein kinase (MAPK) signaling pathways. To accomplish these studies, we employed a highly responsive mouse adrenocortical cell line, Y1-BS1 cells that secrete large quantities of steroids when stimulated with lipoprotein plus hormone. Treatment of these cells with superoxide, H(2)O(2) or 4-hydroxy-2-nonenal (HNE) significantly inhibited steroid production and increased phosphorylation and activation of p38 MAPK. None of the treatments altered the phosphorylation of either extracellular signal-regulated kinases or c-Jun N-terminal kinases (JNKs). Pretreatment of Y1-BS1 cells with MnTMPyP, a cell-permeable superoxide-dismutase/catalase mimetic reactive oxygen species (ROS scavenger), completely prevented the superoxide- and H(2)O(2)-mediated inhibition of steroid production. Likewise, antioxidant N-acetylcysteine completely blocked the HNE-induced loss of steroidogenic response. Incubation of Y1-BS1 cells with either MnTMPyP or NAC also upregulated Bt(2)cAMP and Bt(2)cAMP+hHDL(3)-stimulated steroid synthesis, indicating that endogenously produced ROS can inhibit steroidogenesis. Inhibition of p38 MAPK with SB203580 or SB202190 upregulated the basal steroid production and also prevented the oxidant-mediated inhibition of steroid production. mRNA measurements by qPCR indicated that Y1-BS1 adrenal cells predominantly express p38 MAPKalpha isoform, along with relatively low-level expression of p38 MAPKgamma. By contrast, little or no expression was detected for p38 MAPKbeta and p38 MAPKdelta isoforms in these cells. Transfection of Y1-BS1 cells with either caMKK3 or caMMK6 construct, the upstream p38 MAPK activators, decreased steroidogenesis, whereas transfection with dnMKK3 or dnMKK6 plasmid DNA increased steroidogenesis. Similarly, transfection of cells with a dnp38 MAPKalpha or dnp38 MAPKbeta construct also increased steroid hormone production; however, the effect was less pronounced after expression of either dnp38 MAPKgamma or dnp38 MAPKdelta construct. These results indicate that activated p38 MAPK mediates oxidant (excessive oxidative stress)-induced inhibition of adrenal steroidogenesis.
Abidi
Parveen
P
Department of Veterans Affairs Palo Alto Health Care System, Geriatric Research, Education and Clinical Center, 3801 Miranda Avenue, Palo Alto, California 94304, USA.
Zhang
Haiyan
H
Zaidi
Syed M
SM
Shen
Wen-Jun
WJ
Leers-Sucheta
Susan
S
Cortez
Yuan
Y
Han
Jiahuai
J
Azhar
Salman
S
D004092:20-alpha-Dihydroprogesterone / Q000096:biosynthesis*; D000311:Adrenal Glands / Q000378:metabolism*; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology; D000818:Animals; D045744:Cell Line, Tumor; D003563:Cyclic CMP / Q000031:analogs & derivatives / Q000494:pharmacology; D048371:MAP Kinase Kinase 3 / Q000502:physiology; D048669:MAP Kinase Kinase 6 / Q000502:physiology; D020935:MAP Kinase Signaling System; D051379:Mice; D018384:Oxidative Stress; D010750:Phosphoproteins / Q000235:genetics; D010766:Phosphorylation; D013481:Superoxides / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D000098682:Steroidogenic Acute Regulatory Protein
D010750:Phosphoproteins; D000098682:Steroidogenic Acute Regulatory Protein; D013481:Superoxides; D004092:20-alpha-Dihydroprogesterone; D003563:Cyclic CMP; C053441:dibutyryl cyclic-3',5'-cytidine monophosphate; D000324:Adrenocorticotropic Hormone; D048051:p38 Mitogen-Activated Protein Kinases; D048371:MAP Kinase Kinase 3; D048669:MAP Kinase Kinase 6; C482531:Map2k3 protein, mouse; C482567:Map2k6 protein, mouse