积雪草粗提取物对脂多糖诱导小鼠急性肺损伤的治疗作用

发布时间：2018-10-08 06:34

【摘要】：急性肺损伤（Acute Lung Injury，ALI）是临床上比较常见的一种危重病症，它是过度的炎性反应在肺部的表现。因其发病急且严重，发病率和死亡率高，到目前为止仍无有效手段和特效药治疗。当机体发生急性肺损伤时,会出现诸多的病理变化，其中最为显著地是：肺内过度失控的炎性反应，肺泡表面活性物质的降低、肺水肿和肺膨胀不全，呼吸肌疲劳导致通气不足而产生的二氧化碳过多蓄积等症状。如果急性肺损伤得不到及时的治疗，随着病程的发展，其有可能会发展成为急性呼吸窘迫综合征，而使疾病无法控制，最终会导致死亡。据报道，急性肺损伤的发病率逐年升高,其死亡率甚至高达40%~60%。脂多糖（Lipopolysaccharide，LPS）是炎症反应经典的诱导剂，作为革兰氏阴性菌细胞壁上的主要成分，是细菌治病的重要毒素，也是引起急性肺损伤的重要原因之一。当机体受到革兰氏阴性菌感染时，LPS会与细胞膜上的受体结合激活胞内一系列的信号传导途径，致使上皮细胞、内皮细胞、单核/巨噬细胞等合成和释放大量的内源性生物活性因子如炎症细胞因子、趋化因子、生长因子等，这些生物活性因子可以通过血液循环到达全身各处，引起全身性的炎症反应，由此会引起中毒性休克、全身炎症反应综合症和多器官功能障碍综合症等恶性疾病。积雪草(Centella Asiatsca)作为一种传统中药材，用药历史悠久,最早记载于《神农本草经》，性寒，味苦、辛，，具有清热利湿、解毒消肿之功效，近代研究发现积雪草具有抗氧化，抵抗炎症，抑制肿瘤细胞生长等作用。本研究通过LPS诱导RAW264.7细胞的炎症反应和Balb/c小鼠的急性肺损伤模型，从体外和体内两个方面说明了积雪草粗提取物对炎症反应有比较明显的抑制作用，从而发现了其对于小鼠急性肺损伤的各种病变有显著地缓解作用，一定程度上起到保护小鼠肺组织的作用，初步探讨了积雪草对于LPS引发的炎症反应会产生抑制作用的分子机制。实验方法：利用LPS滴鼻建立小鼠急性肺损伤模型，并且利用酶联免疫吸附试验（Enzyme-linkedimmunosorbent assay，ELISA）检测了肿瘤坏死因子-α（tumornecrosis factor-α，TNF-α）和白细胞介素-6（IL-6）在细胞上清和小鼠肺泡灌洗液中的含量。同时，免疫蛋白印迹（Western blot）实验检测NF-κB信号通路的活化，我们还观察了小鼠肺组织病理变化，测定了肺组织中髓过氧化物酶(MPO)含量及肺湿/干重比(W/D)。实验结果：在LPS诱导的炎症反应中，积雪草粗提取物能显著的减少炎性细胞因子TNF-α和IL-6的表达水平，并且呈现一定剂量依赖性，高剂量效果显著。另外，通过观察小鼠肺部病理组织变化，测定小鼠肺组织内髓过氧化物酶（Myeloperoxidase，MPO）以及干湿重比等指标，综合的说明了积雪草粗提取物对于小鼠急性肺损伤的诸多病变有显著地缓解，表现出了一定程度的治疗作用。并且Western blot实验结果说明了积雪草粗提取物显著的减弱了NF-κB信号通路的活化，为其机制的研究奠定基础。实验结论：基于以上的研究，我们证明了积雪草具有体内外的抗炎效果，并且对于小鼠急性肺损伤病变的缓解效果显著，为该药的广泛使用提供了理论基础和实验依据。
[Abstract]:Acute lung injury (ALI) is a very common serious condition in clinic, which is an excessive inflammatory response in the lungs. Due to its acute and serious morbidity and mortality, there has been no effective means and therapeutic treatment so far. In the case of acute lung injury in the body, many pathological changes occur, most notably: inflammatory response of over-control in the lungs, decreased alveolar surfactant, pulmonary edema, and incomplete lung expansion, Respiratory muscle fatigue leads to excessive accumulation of carbon dioxide and other symptoms due to insufficient ventilation. If acute lung injury is not treated in a timely manner, as the course progresses, it may develop into acute respiratory distress syndrome, which will not control the disease and ultimately lead to death. It was reported that the incidence of acute lung injury increased year by year, and the mortality rate was even 40% ~ 60%. Lipopolysaccharide (LPS) is a classical inducer of inflammatory response. As the main component of the cell wall of gram-negative bacteria, it is one of the most important causes of acute lung injury. When the body is infected by Gram-negative bacteria, the LPS binds to the receptors on the cell membrane to activate a series of signaling pathways in the cell, such that epithelial cells, endothelial cells, monocytes/ macrophages, and the like are synthesized and released a large number of endogenous bioactive factors such as inflammatory cytokines, Chemokine, growth factor, etc., these biological activity factors can reach the whole body through the blood circulation, cause systemic inflammatory reaction, thus can cause toxic shock, systemic inflammatory response syndrome and multiple organ dysfunction syndrome and other malignant diseases. Centella Asiana sca, as a traditional Chinese medicinal material, has a long history, which is first recorded in Shennong Cao Jing>, cold, bitter and pungent, and has the effects of clearing away heat, promoting diuresis, removing toxic materials and detumescence, and modern research finds that the traditional Chinese medicine has antioxidant and resistance. Inflammation, inhibition of tumor cell growth, etc. In this study, the inflammatory response of RAW264.7 cells was induced by LPS and the model of acute lung injury in BALB/ c mice. It has been found that it has a significant effect on various pathological changes of acute lung injury in mice, which plays a role in protecting the lung tissues of mice, and preliminarily probes into the molecules which can inhibit the inflammatory response induced by LPS. Methods: The acute lung injury model of mice was established by LPS nasal drops, and tumor necrosis factor-7721 (TNF-hIL) and interleukin-6 (IL-6) were detected by enzyme-linked immunosorbent assay (ELISA). The activity of NF-Sepharose B signaling pathway was detected by Western blot. The pathological changes of lung tissue were also observed. The content of MPO in lung tissue and the ratio of lung wet/ dry weight were determined. (W/ D). The results showed that in the LPS-induced inflammatory response, the crude extract could significantly reduce the expression level of IL-6 in inflammatory cytokines and present a dose-dependent dose. In addition, by observing the changes of lung pathological tissues of mice, the indexes such as myelopoxidase (MPO) and dry-wet weight ratio in lung tissues of mice were measured. There was a significant response to the lesion, showing a The results of Western blot showed that the crude extract significantly attenuated the activation of NF-Sepharose B signaling pathway, Conclusion: Based on the above researches, we have demonstrated that the anti-inflammatory effect in mice has an external anti-inflammatory effect, and has a significant effect on the acute lung injury in mice, and it is widely used in the study.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S853.7

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