原青花素对大鼠急性肺损伤伴肾功能损害的拮抗作用

Effects of grasp seed procyanidins(原青花素) on acute lung injury and renal function damage in rats

目的:探讨原青花素(GSP)对脂多糖(LPS)诱导大鼠急性肺损伤(ALI)伴肾功能损害的保护作用及其分子机制。方法:采用尾静脉注射LPS制备SD大鼠ALI模型;同时腹腔注射GSP 50 mg/kg进行干预。给药后6 h取静脉血,并制备肺、肾皮质组织匀浆;检测血中肌酐(SCr)、尿素氮(BUN)、乳酸(Lac)及一氧化氮(NO)含量;用酶联免疫吸附法(ELISA)测定血清、肺和肾组织肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)、单核细胞趋化蛋白-1(MCP-1)及IL-6水平。观察肺组织病理学变化、肺血管通透性变化及肺组织湿/干重(W/D)比值,检测肺和肾组织丙二醛(MDA)含量,Na+-K+-ATP酶、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)及髓过氧化物酶(MPO)活性。用蛋白质免疫印迹法(Western blotting)检测肺组织丝裂原活化蛋白激酶(MAPKs)变化;用电泳迁移率实验(EMSA)检测肺组织核转录因子-κB(NF-κB)的DNA结合活性。结果:1LPS大鼠肺间质弥漫性出血,肺泡腔内可见大量粒细胞聚集、浸润,并可见弥漫性肺泡间隔增厚,GSP干预可明显减轻上述病理表现。2LPS大鼠肺组织W/D和肺血管通透性显著增高(P<0.05或P<0.01);而GSP干预可显著降低上述指标(P均<0.01)。3LPS组血清及肺组织TNF-α、IL-1β、MCP-1和IL-6、MDA和MPO水平均明显高于对照组,而Na+-K+-ATP酶、SOD及GSH-Px活性均明显低于对照组(P<0.05或P<0.01)。LPS大鼠肺组织MAPKs磷酸化水平及NF-κB的DNA结合活性也显著升高,GSP干预后上述指标明显减轻。结论:GSP通过阻断MAPKs的激活及抑制NF-κB的DNA结合活性,降低肺组织的炎症反应,减轻肺组织的损害,对LPS所致大鼠ALI伴肾功能损伤具有显著的保护作用。

Objective： To investigate the effects of grasp seed procyanidins （GSP, 原青花素） on lipopolysaccharide （LPS）-induced acute lung injury （ALI） in rats with renal function damage and the related possible molecular mechanisms. Methods： The homogenates of lung and kidney were prepared and venous blood were collected at 6 hours after injection of LPS and medicine. The changes of contents of creatinine （Cr）, blood urea nitrogen （BUN）, lactic acid （Lac） and nitric oxide （NO） in the blood were measured. The enzyme linked immunosorbent assay （ELISA） was used to measure the levels of tumor necrosis factor -α （TNF-α）, interleukin - 1β （IL - 1β）, monocyte chemoattractant protein - 1 （MCP - 1） and IL - 6 in the serum, lung and renal cortex tissue homogenate in various groups. The histopathological changes of lung tissues were observed. The pulmonary vascular permeability and the lung wet/dry （W/D） weight ratio were determined; the malonaldehyde （MDA） content, Na^＋- K^＋ - ATPase, superoxide dismutase （SOD）, myeloperoxidase （MPO） and glutathion peroxidase （GSH- Px） activities in lung and renal tissues were also determined. Changes of mitogen activated protein kinase （MAPKs） were detected by Western blotting, and the combination activity of nuclear factor - κB （NF -κB） to DNA was detected by electrophoretic mobility shift assay （EMSA） in lung tissues. Results： ①Compared with the normal rats in control group, the lungs of the rats in LPS treatment group and GSP group had significant hyperemia and spotted hemorrhage. The inflammatory granulocyte infiltration, diffuse alveolar septum thickening and spotted hemorrhage were observed in the pathological examinations, while in LPS plus GSP group the above mentioned pathological changes were milder. ②Compared with control group, the lung W/D and pulmonary vascular permeability were much higher in the LPS treatment groups （P〈0.05 or P0.01）, while they were significantly less in the groups treated by GSP （all P〈0.01）. ③MPO activity and MDA content, the levels of TNF -α, IL - 1β, MCP -1, IL -6 in the serum and lung tissue were higher, while activities of Na＋^- K＋^- ATPase, SOD, as well as GSH - Px were significantly less in LPS treatment group than those in the control group （P〈0.05 or P〈 0. 01 ）. Levels of phosphorylation in MAPKs and combination of NF -κB to DNA activity were elevated markedly in the LPS treatment group. However, in the group treated by GSP, all the above -mentioned measurements were decreased significantly. Conclusion： GSP may decrease inflammatory reaction of lung and kidney and further decrease lung damage by interdicting MAPK activity, inhibiting the combination activity of NF-κB to DNA and renal function damage induced by LPS in rats, all indicating protection of GSP against ALl.