Hill E M, Eling T, Nettesheim P
Laboratories of Molecular Carcinogenesis and Pulmonary Pathobiology, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, USA.
Am J Respir Cell Mol Biol. 1998 May;18(5):662-9. doi: 10.1165/ajrcmb.18.5.2985.
The purpose of our studies was to examine differentiation-dependent expression of 15-lipoxygenase (15-LO) and prostaglandin H synthase (PGHS) isoforms in cultured normal human tracheobronchial epithelial cells. In the presence of retinoic acid (RA) the cultures differentiated into a mucociliary epithelium. When cultured in RA-depleted media, the cultures differentiated into a squamous epithelium. In the absence of RA the cultures did not express 15-LO or either of the PGHS isoforms. The PGHS-1 isoform was not expressed in RA-sufficient cultures, but both PGHS-2 messenger RNA (mRNA) and protein were strongly expressed, and prostaglandin E2 (PGE2) was produced during the predifferentiation phase. No PGHS-2 expression or PGE2 could be detected in fully differentiated mucociliary cultures. 15-LO showed the opposite expression pattern: neither mRNA nor protein were detected during the predifferentiation stage, but both were strongly expressed once mucous differentiation had occurred. Cytosolic phospholipase A2 protein was expressed throughout all stages of growth and differentiation. The cultures generated no 15-LO metabolites when incubated with 10 microM to 50 microM arachidonic acid (AA) and stimulated with ionophore. However, lysates prepared from such cultures generated 15-hydroxyeicosatetraenoic acid (15-HETE) and 12-HETE from AA, indicating that the cells contained active enzyme. When cultures expressing 15-LO protein were incubated with 10 microM linoleic acid (LA) instead of AA, and were stimulated with ionophore, they generated 13-hydroxy-9,11-octadecadienoic acid. LA rather than AA appeared to be the preferred substrate for the 15-LO enzyme. Our studies indicated that the expression of 15-LO and PGHS-2 is differentiation dependent in airway epithelial cells.
我们研究的目的是检测培养的正常人气管支气管上皮细胞中15-脂氧合酶（15-LO）和前列腺素H合成酶（PGHS）亚型的分化依赖性表达。在视黄酸（RA）存在的情况下，培养物分化为黏液纤毛上皮。当在缺乏RA的培养基中培养时，培养物分化为鳞状上皮。在没有RA的情况下，培养物不表达15-LO或任何一种PGHS亚型。PGHS-1亚型在RA充足的培养物中不表达，但PGHS-2信使核糖核酸（mRNA）和蛋白均强烈表达，并且在预分化阶段产生前列腺素E2（PGE2）。在完全分化的黏液纤毛培养物中未检测到PGHS-2表达或PGE2。15-LO表现出相反的表达模式：在预分化阶段未检测到mRNA和蛋白，但一旦发生黏液分化，两者均强烈表达。胞质磷脂酶A2蛋白在生长和分化的所有阶段均有表达。当用10微摩尔/升至50微摩尔/升花生四烯酸（AA）孵育并用电离子载体刺激时，培养物不产生15-LO代谢产物。然而，从这些培养物制备的裂解物能从AA产生15-羟基二十碳四烯酸（15-HETE）和12-HETE，表明细胞含有活性酶。当将表达15-LO蛋白的培养物用10微摩尔/升亚油酸（LA）而非AA孵育，并用电离子载体刺激时，它们产生13-羟基-9,11-十八碳二烯酸。LA而非AA似乎是15-LO酶的首选底物。我们的研究表明，15-LO和PGHS-2的表达在气道上皮细胞中是分化依赖性的。
