Loftin C D, Eling T E
Laboratory of Molecular Biophysics, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
Arch Biochem Biophys. 1996 Jun 15;330(2):419-29. doi: 10.1006/abbi.1996.0271.
BALB/c mouse keratinocytes (BALB/MK) are nontumorigenic epithelial cells which are dependent on mouse epidermal growth factor (EGF) for maintaining proliferation in culture. In BALB/MK the oxygenation of both arachidonic acid and linoleic acid was dependent on EGF. EGF stimulated the formation of prostaglandin E2 and prostaglandin F2 alpha from arachidonic acid and 9- and 13-hydroxyoctadecadienoic acid (HODE) from linoleic acid. Analysis of the linoleic acid metabolites determined the ratio of 9-HODE to 13-HODE was approximately 6 to 4, and the 9-HODE was the (R) enantiomer, consistent with metabolism by prostaglandin G/H synthase (PGHS). The formation of these linoleic acid metabolites was sensitive to indomethacin, a PGHS inhibitor. EGF induced the expression of PGHS-2 mRNA after 30 min, which peaked after 1 h, and remained expressed for at least 24 h after the addition of EGF. A less significant increase in the expression of PGHS-1 mRNA occurred 4 h after EGF stimulation. Immunoblot analysis did not detect expression of PGHS-1 protein. However, PGHS-2 protein expression was increased 2 h after EGF exposure and was dependent on EGF. PGHS-2 protein was not transiently expressed as reported with other cell types, but was continually expressed in proliferating cells maintained with EGF at a subconfluent density. Indomethacin significantly attenuated EGF-dependent mitogenesis and cell proliferation. These results suggest that PGHS-2 activity contributes to the proliferative response of BALB/MK to EGF.
BALB/c小鼠角质形成细胞（BALB/MK）是无致瘤性的上皮细胞，在培养中依赖小鼠表皮生长因子（EGF）来维持增殖。在BALB/MK中，花生四烯酸和亚油酸的氧化均依赖于EGF。EGF刺激花生四烯酸生成前列腺素E2和前列腺素F2α，以及亚油酸生成9-和13-羟基十八碳二烯酸（HODE）。对亚油酸代谢产物的分析确定9-HODE与13-HODE的比例约为6比4，且9-HODE为（R）对映体，这与前列腺素G/H合酶（PGHS）的代谢一致。这些亚油酸代谢产物的形成对PGHS抑制剂吲哚美辛敏感。EGF在30分钟后诱导PGHS-2 mRNA的表达，1小时后达到峰值，并在添加EGF后至少24小时持续表达。EGF刺激4小时后，PGHS-1 mRNA的表达有较不显著的增加。免疫印迹分析未检测到PGHS-1蛋白的表达。然而，EGF暴露2小时后PGHS-2蛋白表达增加，且依赖于EGF。PGHS-2蛋白不像其他细胞类型那样短暂表达，而是在以亚汇合密度用EGF维持的增殖细胞中持续表达。吲哚美辛显著减弱了EGF依赖性的有丝分裂和细胞增殖。这些结果表明PGHS-2活性有助于BALB/MK对EGF的增殖反应。
