过氧亚硝酸根联合碱性成纤维细胞生长因子对人晶状体上皮细胞细胞外信号调节激酶（ERK）磷酸化水平的影响

目的　探讨过氧亚硝酸根（ｐｅｒｏｘｙｎｉｔｒｉｔｅ，ＯＮＯＯ－）和重组人碱性成纤维细胞生长因子（ｒｅｃｏｍｂｉｎａｎｔｈｕｍａｎｂａｓｉｃｆｉｂｒｏｂｌａｓｔｇｒｏｗｔｈｆａｃｔｏｒ，ｂＦＧＦ）共同作用对人晶状体上皮细胞细胞外信号调节激酶（ＥＲＫ）磷酸化水平的影响。初步探讨ＯＮＯＯ－导致白内障发生的可能机制。方法　将培养的ＨＬＥＣ细胞分为４组，对照组（未经ＯＮＯＯ－和ｂＦＧＦ处理）、ｂＦＧＦ单独处理组、ｂＦＧＦ＋ＯＮＯＯ－处理组［ｂＦＧＦ预处理１ｈ后加入不同浓度ＯＮＯＯ－（５０μｍｏｌ·Ｌ－１、１００μｍｏｌ·Ｌ－１、２００μｍｏｌ·Ｌ－１）处理１ｈ］和ＯＮＯＯ－ ＋ｂＦＧＦ处理组［不同浓度ＯＮＯＯ－（５０μｍｏｌ·Ｌ－１、１００μｍｏｌ·Ｌ－１、２００μｍｏｌ·Ｌ－１）预处理１ｈ后加入ｂＦＧＦ处理１ｈ］。检测各组细胞内ＥＲＫ磷酸化水平的变化。结果　ｂＦＧＦ单独处理组和ｂＦＧＦ ＋ＯＮＯＯ－（５０μｍｏｌ·Ｌ－１、１００μｍｏｌ·Ｌ－１、２００μｍｏｌ·Ｌ－１）处理组中磷酸化ＥＲＫ相对表达值分别为５．１７±０．３５、４．４２±０．１２、３．９１±０．１５和０．４９±０．０８，与对照组（１．００±０．０５）相比差异有统计学意义（Ｆ＝４０２．８３，Ｐ＜０．００１），结果显示ＯＮＯＯ－可以抑制由ｂＦＧＦ诱导的ＥＲＫ磷酸化水平的增加（Ｆ＝３１０．３４，Ｐ＜０．０５）。ＯＮＯＯ－（５０μｍｏｌ·Ｌ－１、１００μｍｏｌ·Ｌ－１、２００μｍｏｌ·Ｌ－１）＋ｂＦＧＦ处理组中磷酸化ＥＲＫ相对表达值分别为３．３６±０．０４、３．３２±０．０６和２．９２±０．０８，与对照组（１．００±０．０２）相比，所有处理组的ＥＲＫ磷酸化水平均显著增加（Ｆ＝５１８．９４，Ｐ＜０．００１）；５０μｍｏｌ·Ｌ－１和１００μｍｏｌ·Ｌ－１ＯＮＯＯ－处理细胞后经ｂＦＧＦ处理，ＥＲＫ磷酸化水平较ｂＦＧＦ单独处理组（３．０４±０．０５）显著增加（Ｆ＝３５．５３，Ｐ＜０．０５），而２００μｍｏｌ·Ｌ－１ＯＮＯＯ－处理组与ｂＦＧＦ单独处理组相比差异无统计学意义（Ｐ＞０．０５）。结论　ＯＮＯＯ－诱导的白内障的发生可能与ＥＲＫ磷酸化的紊乱有关。

Effects of peroxynitrite and basic fibroblast growth factor on level of ERK phosphorylation in human lens epithelial cells

Objective To discuss the effects of peroxynitrite (ONOO - ) and recombinant human basic fibroblast growth factor ( bFGF) on the level of ERK phosphorylation ( pERK) in human lens epithelial cells ( HLEC) . and investigate the mechanisms of ONOO - on the development of cataract. Methods HLEC cell line SRA01/04 cells were cultured in DMEM medium and treated with ONOO - and bFGF. Cells were divided into four groups : Control group ( treated without bFGF and ONOO - ) ; bFGF treatment alone group ;bFGF + ONOO - treatment group HLEC cells were treated with bFGF for I hour and then cultured with different concentrations ( 50 ymol . L ’1 ,100 Vmol . L-l , 200 Vmol . L-’) of ONOO - for I hour] ;ONOO - + bFGF treatment group HLEC cells were treated with different concentrations (50 ymol . 1-1 ,100 pdmol . 1-1 ,200 pmol . L") of ONOO- for I hour and then cultured with bFGF for I hour). The levels of pERK were assayed by Western blot. Results The levels of pERK in bFGF treatment alone group and bFGF + ONOO - treatment group (50 Vmol . L’l , 100 ymol . 1’1 .200 ymol . L -l ONOO - ) were 5. 17 +0. 35 ,4. 42 + 0. 12 .3. 91 +0. 15 and 0. 49 +0. 08 , respectively,the difference of pERK was statistically significant compared with the control group ( 1. 00 + 0. 05 ) ( F = 402. 83 ,P < 0. 001 ) . The results showed that HQ could inhibit the increase of pERK induced by bFGF (F = 310. 34 .P < 0. 05 ) . The levels of pERK in ONOO - (50 Vmol . 1-1 ,100 ymol . 1-1 ,200 Vmol . L-’) + bFGF treatment group were 3. 36 + 0. 04 , 3. 32 + 0. 06 and 2. 92 + 0. 08 , respectively , the difference of pERK was statistically significant compared with the control group ( 1. 00 + 0. 02 ) ( F = 518. 94 , P < 0. 001) . and showed that bFGF could increase the levels of pERK induced by low concentrations of ONOO - (50 Vmol . L-I , 100 ymol . L’l ) .which was higher than that in the bFGF treatment alone group ( F = 35. 53 , P < 0. 01 ) . but there was no statistical difference between 200 ymol . L -l ONOO - treatment group and bFGF treatment alone group (P > 0. 05 ) . Conclusion ONOO - induced cataract may be associated with the disorder of ERK phosphorylation.