Wang Ya-ting, Chen Sheng-li, Xu Shu-yun
Department of Peadiatrics, The First Affiliated Hospital, Anhui Medical University, Hefei, 230022, China.
Zhonghua Er Ke Za Zhi. 2004 Feb;42(2):94-7.
Nuclear factor-kappa B (NF-kappaB) is a critical transcription factor governing the expression of many cytokines that are involved in the pathogenesis of inflammatory diseases, such as asthma and rheumatoid arthritis. Melatonin (MT), a relatively safe and potent antioxidant which has shown efficacy in several chronic inflammatory models, may inhibit the expression of NF-kappaB and therefore might have a therapeutic use in asthma. This study aimed at observing the effect of MT on the expression of NF-kappaB and airway inflammation in a rat model of bronchial asthma.
Twenty-four male Sprague-Dawley (SD) rats weighing 120 g to 170 g were randomly divided into three experimental groups (8 in each): (1) Asthmatic group: Rats were immunized on day 1 by intraperitoneal injection of 100 mg ovalbumin (OVA) in 1 ml of saline with 100 mg of alu minum hydroxide. From day 15 the animals were challenged with aerosolized OVA (1% in saline) for 20 minutes per day for 7 consecutive days. (2) MT group: OVA-sensitized rats were injected intraperitoneally with 10 mg/kg MT 30 minutes before each OVA challenge. (3) CONTROL GROUP: OVA for inhalation and MT for intraperitoneal injection was replaced with normal saline (NS). Airway responsiveness to aerosolized acetylcholine of 24 rats was detected six hours after the last challenge. Then the rats were lavaged and total and differentiated leukocytes counts in bronchoalveolar lavage fluid (BALF) were performed after staining with Wright-Giemsa staining. At the same time, levels of nitric oxide (NO) in BALF, inducible nitric oxide synthesis (iNOS) and constitute nitric oxide synthesis (cNOS) in the lung tissues were assessed with the use of nitrate reductase and chemical colorimetry, respectively. The expression of NF-kappaB in the lung tissues was observed by means of immunohistochemical staining.
(1) After OVA challenge, there was a significant decrease in airway responsiveness, lymphocytes and eosinophils in BALF in MT group compared with asthmatic group (P < 0.01 respectively); (2) There was a significant decrease in amounts of NO(2)(-)/NO(3)(-) in the BALF and levels of iNOS in the lung tissues in MT group comparing with asthmatic group (P < 0.01 respectively); and the levels of iNOS in the lung tissues was correlated positively with NO(2)(-)/NO(3)(-) in the BALF (P < 0.01), but there were no significant differences in activity of cNOS in any of the groups analyzed. (3) There was a significant increase in expression of NF-kappaB in lung tissues in asthmatic group compared with the other groups (P < 0.01), and so was in MT group compared with control group (P < 0.05).
MT could partially inhibit the expression of NF-kappaB and down-regulate the activity of iNOS in lung tissue, decrease the production of NO in BALF. These data suggest that the inhibitory effect of MT probably play a role in decreasing airway hyperresponsiveness and airway inflammation of asthmatic rats model.
核因子-κB（NF-κB）是一种关键的转录因子，可调控许多细胞因子的表达，这些细胞因子参与炎症性疾病如哮喘和类风湿关节炎的发病机制。褪黑素（MT）是一种相对安全且有效的抗氧化剂，已在多种慢性炎症模型中显示出疗效，可能抑制NF-κB的表达，因此或许可用于哮喘的治疗。本研究旨在观察MT对支气管哮喘大鼠模型中NF-κB表达及气道炎症的影响。
将24只体重120至170克的雄性Sprague-Dawley（SD）大鼠随机分为三个实验组（每组8只）：（1）哮喘组：第1天腹腔注射1毫升含100毫克卵清蛋白（OVA）和100毫克氢氧化铝的生理盐水进行免疫。从第15天起，动物每天用雾化OVA（1%生理盐水溶液）激发20分钟，连续7天。（2）MT组：OVA致敏大鼠在每次OVA激发前30分钟腹腔注射10毫克/千克MT。（3）对照组：雾化吸入的OVA和腹腔注射的MT均用生理盐水（NS）替代。最后一次激发6小时后，检测24只大鼠对雾化乙酰胆碱的气道反应性。然后对大鼠进行灌洗，用瑞氏-吉姆萨染色后进行支气管肺泡灌洗液（BALF）中总白细胞和分化白细胞计数。同时，分别用硝酸还原酶法和化学比色法评估BALF中一氧化氮（NO）水平、肺组织中诱导型一氧化氮合酶（iNOS）和组成型一氧化氮合酶（cNOS）活性。通过免疫组织化学染色观察肺组织中NF-κB的表达。
（1）OVA激发后，与哮喘组相比，MT组气道反应性、BALF中淋巴细胞和嗜酸性粒细胞显著降低（P均<0.01）；（2）与哮喘组相比，MT组BALF中NO₂⁻/NO₃⁻量及肺组织中iNOS水平显著降低（P均<0.01）；肺组织中iNOS水平与BALF中NO₂⁻/NO₃⁻呈正相关（P<0.01），但各分析组中cNOS活性无显著差异。（3）与其他组相比，哮喘组肺组织中NF-κB表达显著增加（P<0.01），MT组与对照组相比也显著增加（P<0.05）。
MT可部分抑制肺组织中NF-κB的表达，下调iNOS活性，减少BALF中NO的产生。这些数据表明MT的抑制作用可能在降低哮喘大鼠模型的气道高反应性和气道炎症中发挥作用。
