A molecular biologic study on delayed type cell injury of the airway mucosa.

用白三烯D_4(LTD_4)和血小板活化因子(PAF)分别诱导培养的人鼻上皮细胞诱导型一氧化氮合酶m RNA(INOS M RNA)、黄嘌呤氧化酶m RNA(XO M RNA)和环氧合酶-2 m RNA(COX-2 m RNA)，20h后提取总RNA。用实时荧光定量聚合酶链式反应(Real Time-PCR)检测每条基因的表达情况。结果：(1)与PBS相比，LTD_4诱导的iNOS、XO和COX-2mRNA的表达水平相同。(2)与PBS相比，PAF诱导的iNOS基因表达水平相同，而XO和COX-2基因表达水平较高。(3)LTD_4可增加细胞内iNOS蛋白的浓度，而PAF则略有增加，提示LTD_4可能通过iNOS参与NO的过量产生，PAF可能通过XO产生超氧阴离子。这些脂质介质在SU…中产生炎症细胞较多，如嗜酸性粒细胞、中性粒细胞和巨噬细胞；尤其是PAF在嗜酸性粒细胞中大量产生。因此，上皮细胞也可能与过敏性炎症过程中NO和超氧化物的过度产生有关。这些结果表明，LTD_4和PAF从炎症细胞如嗜酸性粒细胞等释放出来，其他白细胞参与了与XO的其他促进因子相关的超氧化物的产生。众所周知，炎症细胞会产生超氧化物，但其机制尚未阐明。LTB_4还具有促进白细胞产生超氧化物的能力。因此，在过量生成NO和超氧化物的情况下，这些分子相互反应生成具有强烈伤害活性的过氧亚硝酸盐，并攻击酪氨酸，留下微量的硝基酪氨酸。我们先前的研究显示，过敏性鼻炎患者的鼻腔杠杆液中含有硝基酪氨酸。在进一步的研究中，主要细胞因子在诱导人鼻腔上皮细胞iNOS、XO和COX-2mRNA表达中的作用有待进一步研究。较少

Induction of inducible nitric oxide synthase m RNA (iNOS mRNA), xanthine oxidase mRNA (XO mRNA) and cylooxygenase-2 mRNA (COX-2 mRNA) in cultured human nasal epithelial cells were studied by exposing to 10^<-8>M leukotrien D_4(LTD_4) and 10^<-8>M platelet activating factor (PAF), respectively and 20 hours later total RNA was extracted. Each mRNA was investigated by real time -PCR. The concentration of the iNOS protein in the cell solution was also measured in a duplicate manner.Results : (1)Comparing PBS, LTD_4 showed the same level induction of iNOS mRNA, XO mRNA and COX-2 mRNA. (2)Comparing PBS, PAF showed the same level induction of iNOS mRNA although PAF did higher level of induction of both XO mRNA and COX-2 mRNA. (3)LTD_4 augmented the concentration of the iNOS protein in the cell solution, while PAF did slightly.These results suggest that LTD_4 may take part in excessive production of NO via iNOS, and PAF may do superoxide production via XO. These lipid mediators generated in su … More ch inflamed cells as eosinophils, neutrophils and macrophages ; especially PAF is generated in eosinophils in greater amount. Therefore, epithelial cells may also relate with excess production of NO and superoxide in allergic inflammation process.These results suggest that LTD_4 and PAF released from inflamed cells such as eosinophils, other leukocytes takes part in superoxide production associated with other promoting factors of XO. It has been well known that inflamed cells produce superoxide, however, its mechanism has not been elucidated. LTB_4 has also the capacity of promoting superoxide generation in leukocytes. Therefore, under excessive generation of NO and superoxide generation, these molecules react each other generating peroxynitrite, which has strong injurious activity and attack tyrosine leaving a trace, nitrotyrosine. Our previous study showed nitrotyrosine in nasal leverage fluids of allergic patients with rhinitis. In the further study the effect of primary cytokines on induction of iNOS mRNA, XO mRNA and COX-2 mRNA in cultured human nasal epithelial cells should be elucidated. Less