The Effects Of Erigeron Breviscapine On Ischemia Reperfusion Injury Of Rat Lung

Background and Objective Ischemia reperfusion injury of the lung(LIRI)is still a challenge for the thoracic surgeon, which occurs in many clinical conditions, such as cardiopulmonary bypass, lung embolism, shock etc, especially during the lung transplantation. Those may lead to acute lung injury or even multiple organ dysfunction syndrome(MODS). Ischemia-reperfusion induced lung injury contributes to a significant cause of morbidity and mortality after the lung transplantation, so it is necessary to better understand the mechanisms of lung ischemia reperfusion injury and investigate more reliable strategies to reduce the incidence of the injury. This experiment was carried out, through establishing an in situ hilar occlusion and in vivo rat lung ischemia reperfusion injury model. The wet-to-dry weight ratio(W/D), myeloperoxidase(MPO)of lung tissue and the content of nuclear NF-κB p65 were measured. The histopathological and ultrastructural changes of the lung tissue were observed, to explore the effects of erigeron breviscapine on ischemia reperfusion injury of rat lung in vivo.Materials and Methods Thirty-two male Sprague-Dawley rats were anesthetized with an intraperitoneal injection of sodium pentobarbital(30mg/kg). The animals were shaved, endotracheal intubated with a 14 gauge tube, and ventilated with a rodent ventilator at a respiratory rate of 60 breaths/min, a 1:1.5 of I/E ratio and a mean peak pressure of 0.02 MPa. All animals received 0.04 mg/kg of intramuscular atropine after being anesthetized. The chest was opened via a left thoracotomy through the fifth intercostal space. After the left pulmonary hilum including neural, vascular, lymphatic, and connective tissue was stripped, a ligature was placed through it, and the ends of the tie were threaded through a small plastic tube to form a snare for reversible left pulmonary hilar occlusion. Heparinization was maintained during the experimental period with a bolus injection of 100U/kg sodium heparin via the dorsal penile vein.All animals were randomized into four groups with 8 animals in each group.①sham operation group(Ⅰ),②IR group(Ⅱ),③erigeron 25 mg/kg group(Ⅲ),④erigeron 50 mg/kg group(Ⅳ). IR injury consisted of 45 minutes of lung cross-clamping followed by 2 hours of reperfusion, while the sham operation animals had a thoracotomy and pulmonary hilus cleating and stripping only. Erigeron was given with an intraperitoneal injection qd×3 before operation. The sham operation and IR groups received N.S. 10ml/kg according to the same protocol.At the end of the experiment, all operated rats were killed by blood letting through carotid artery. The left lung tissue were retrieved, and then saved in a -70℃refrigeratory for determination. The W/D and MPO of lung tissue, the content of nuclear NF-κB p65 were detected by immunohistochemical stain and Western blot technically. The histopathological and ultrastructural changes of the lung tissue were observed under the optical and electronic microscopy.Results(1) The ratio of W/D and MPO activity in lung tissue. After reperfusion 2 hours, the W/D and MPO increased markedly in IR group and had significantly change compared with groupⅠ(p＜0.01). Treatment with erigeron alleviated the increasd of W/D and MPO. Moreover, W/D and MPO in groupⅣwere lower than groupⅢ(p＜0.01).(2) The level of NF-κB in lung tissue.Immunohistochemical stain analysis of lung NF-κB. The expression of NF-κB p65 in sham operation group showed light brown immunostaining in cytoplasm and little staining in the nuclei. The significant positive expressions of NF-κB p65 as strong brown staining in cytoplasm and nuclei were observed in IR group. Compared to IR group, the positive rates of NF-κB p65 expression decreased obviously in erigeron groups.Western blot assessment of lung NF-κB. Western blot showed weak NF-κB p65 positive signals in the lungs of sham-operated animals. In contrast, significant increase of NF-κB p65 protein expression was found in IR group(p＜0.01). In the rats treated with erigeron 25 mg/kg or 50mg/kg, the signals weakened evidently compared with IR group(p＜0.01). Furthermore, compared to low dose erigeron group, the signals of high dose one weaken distinctly(p＜0.01).(3) Histological evaluation. In the process of ischemia-reperfusion, the lung injury was aggravating progressively in the IR group. Marked pulmonary capillary congestion, edema in pulmonary stroma, neutrophil infiltration, inflammatory exudation, thickening of alveolar wall and intra-alveolar hemorrhage could be observed. These changes in morphology were less pronounced in rats that had been pretreated with erigeron. And that high dose one largely attenuated these pathological alterations associated with reperfusion.(4) Ultrastructural examination. The effect of ischemia-reperfusion on the lung, as demonstrated by electron microscopy, revealed massive alterations of pneumocyte morphology over baseline samples as indicated by cytoplasmic vacuolation, nuclear membrane deformity, the swelling of both mitochondria and endoplasmic reticulum and contraction of endothelial cells(EC). In contrast, erigeron treatment resulted in a marked attenuation of these morphological alterations.Conclusions (1) The above results demonstrated that erigeron breviscapine can attenuate the lung ischemia-reperfusion injury in rats. And the protective change has dose-depedent effect. (2) Erigeron may be restrain NF-κB activation in the pulmonary cells, and then modulate the secretion of inflammatory mediators in transcription degree that reduce inflammatory response. (3) Erigeron can inhibit neutrophil aggregation and activation in the lung to protect it against ischemia reperfusion injury.