Zhun Yun-Fu, Dai Ai-Guo, Hu Rui-Cheng, Jian Yong-Lian
Department of Respiratory Medicine, Hunan Institute of Gerontology, Hunan Province Geriatric Hospital, Changsha 410001, China.
Zhongguo Ying Yong Sheng Li Xue Za Zhi. 2006 Nov;22(4):492-6.
To investigate the effects of Nrf2 (Nuclear-E2 related factor) on gamma-glutamylcysteine synthase (gamma-GCS) in lung of guinea pigs with bronchial asthma.
20 adult male guinea pigs were randomly divided into two groups (n = 10): control group (C group) and asthmatic group (A group), asthmatic model was established by ovalbumin intraperitoneal and ovalbumin inhalation. The reactive oxygen piece (ROS), reduced glutathione (GSH), oxidant glutathione (GSSG) and total GSH in lung tissue were examined respectively. Inflammatory cell infiltration and index of remodeling of bronchiole were detected. In situ hybridization detected the gamma-GCS heavy subunit (gamma-GCS h) mRNA in lung tissue. Immunohistochemistry detected the expression of Nrf2 protein and gamma-GCS protein in lung tissue. RT-PCR measured the expression of Nrf2 mRNA in lung tissue. The activity of gamma-GCS was measured by coupled enzyme assay.
(1) The number of eosinophils and lymphocytes in bronchiole of A group were significantly higher than that of C group (P < 0.05), the remodeling of bronchiole in A group was definite. (2) ROS (U/mg pro), GSSG (micromol/g pro) and total GSH in lung tissue of A group were significantly higher than that of C group (P < 0.01). The GSH/GSSG in lung tissue of A group was much lower than that of C group (P < 0.01), GSH in lung tissue showed no difference between A group and C group. (3) Immunohistochemistry indicated that Nrf2 protein and gamma-GCS protein were more positively expressed in A group than that in C group (P < 0.01). In situ hybridization discovered that the expression of gamma-GCS-h mRNA in lung tissue of A group was more positive than that of C group. (4) RT-PCR showed that the expression of Nrf2 mRNA was no difference between A group and C group (P > 0.05). (5) The activity of gamma-GCS of A group was (28 +/- 8)U which was significantly higher than that of C group (9 +/- 2)U (P < 0.01). (6) Linear correlation analysis indicated that in lung tissue of guinea pig with asthma there existed strongly positive relationship among ROS, GSSG and the expression of Nrf2, gamma-GCS mRNA, gamma-GCS protein, the activity of gamma-GCS, there existed strongly negative relationship among GSH/GSSG and the expression of Nrf2, gamma-GCS mRNA, gamma-GCS protein, the activity of gamma-GCS.
There existed oxidative stress in lung of guinea pigs with bronchial asthma, which possibly positively regulated gamma-GCS via up regulating transcription factor Nrf2.
探讨核因子E2相关因子2（Nrf2）对支气管哮喘豚鼠肺组织中γ-谷氨酰半胱氨酸合成酶（γ-GCS）的影响。
将20只成年雄性豚鼠随机分为两组（n = 10）：对照组（C组）和哮喘组（A组），通过卵白蛋白腹腔注射和雾化吸入建立哮喘模型。分别检测肺组织中活性氧（ROS）、还原型谷胱甘肽（GSH）、氧化型谷胱甘肽（GSSG）及总GSH水平。检测炎症细胞浸润及细支气管重塑指数。采用原位杂交法检测肺组织中γ-GCS重链（γ-GCS h）mRNA表达。采用免疫组织化学法检测肺组织中Nrf2蛋白和γ-GCS蛋白表达。采用RT-PCR法检测肺组织中Nrf2 mRNA表达。采用偶联酶法检测γ-GCS活性。
（1）A组细支气管内嗜酸性粒细胞和淋巴细胞数量显著高于C组（P < 0.05），A组细支气管重塑明显。（2）A组肺组织中ROS（U/mg pro）、GSSG（μmol/g pro）及总GSH水平显著高于C组（P < 0.01）。A组肺组织中GSH/GSSG显著低于C组（P < 0.01），A组与C组肺组织中GSH水平差异无统计学意义。（3）免疫组织化学结果显示，A组Nrf2蛋白和γ-GCS蛋白阳性表达高于C组（P < 0.01）。原位杂交发现，A组肺组织中γ-GCS-h mRNA表达阳性率高于C组。（4）RT-PCR结果显示，A组与C组Nrf2 mRNA表达差异无统计学意义（P > 0.05）。（5）A组γ-GCS活性为（28±8）U，显著高于C组（9±2）U（P < 0.01）。（6）直线相关分析表明，哮喘豚鼠肺组织中ROS、GSSG与Nrf2、γ-GCS mRNA、γ-GCS蛋白表达及γ-GCS活性呈显著正相关，GSH/GSSG与Nrf2、γ-GCS mRNA、γ-GCS蛋白表达及γ-GCS活性呈显著负相关。
支气管哮喘豚鼠肺组织存在氧化应激，可能通过上调转录因子Nrf2对γ-GCS进行正向调控。
