The Study Of 17β-estradiol Admimistration Attenmates Seawater Aspiration-induced Pulmensry Edrna In Rats

Worldwide, some half a million people die each year from drowning. The sites of drowning are varied, with the majority occurring in freshwater. But, the pulmonary injuries are induced by seawater drowning, such as lung edema, hypoxemia and inflammatory reaction, are more serious compared to that induced by freshwater drowning. As all known, seawater is hyperosmolar fluid. The NaCl concentration is 3%-3.5%, about 3 fold of the physiologic saline, water aspiration can induce acute lung injury (ALI) and its more severe form, the acute respiratory distress syndrome (ARDS).In addition, pulmonary edema is a common feature in early seawater aspiration-induced ALI and careful attention must be taken to fluid status. Aquaporins (AQPs), a family of trans-membrane water channel proteins, play a major role in trans-cellular and trans-epithelial water movement. Furthermore, estrogen plays a major role in the lung under both physiological and pathophysiological conditions in animals. Previous results have shown that 17β-estradiol (E2) was effective for the treatment of pulmonary diseases, However, as for whether E2 plays an very important role in ALI/ARDS induced by seawater drowning, there is also very little evidence to demonstrate. In the present study, we explored the effect of E2 on PE-SWD both in vivo and in vitro, and examine the effects of E2 on Aquaporin 1 (AQP1) and Aquaporin 5 (AQP5) in PE-SWD and its underlying mechanisms.Objective:To investigate whether E2 treatment attenuates seawater aspiration-induced acute pulmonary edema, and examines the effects of E2 administration on AQP1 and AQP5 in seawater aspiration-induced acute pulmonary edema and its underlying mechanisms by extracellular-signalregulated protein kinase (ERK) signaling pathway.Materials and methods:1. Animal experiment:Male Sprague-Dawley rats were randomly divided into four groups:normal group (NG),17β-estradiol (E2) group, seawater aspiration (SW) group, and 17β-estradiol treatment (SW+E2) group.1.1 Assess the acute lung injury in rats:Rats were killed by decapitation at the indicated time points. The changes of PaO2and pH (30min, 1h,2h,4h,8h after seawater aspiration), pulmonary vascular permeability (PMVP), Wet/Dry (W/D) weight ratios and pulmonary histological changes were investigated to assess the degree of lung injury. And then the rats consistent with the degree of acute lung injury had been taken to the following experiment.1.2 AQP1 and AQP5 protein and mRNA expression:The lungs were harvested 4h after seawater-aspiration for Western-blotting, immunohistochemistry and RT-RCR studies.2. Cell experiment:The A549 cells were randomly divided into five groups: normal group (NG),17β-estradiol (E2) group, seawater aspiration (SW) group, 17β-estradiol treatment (SW+E2) group and ERK inhibitor (SW+E2+U0126) group. To detected AQP1, AQP5 subunit and ERK protein expression, cell protein was harvested 4h after seawater administration for Western blotting.Results1. Results of animal experiment1.1 After seawater aspiration the PaO2 and pH were consistently decreased in SW and SG+E2 group, and then increased slowly but still lower than normal group. In SW+E2 group, the PaO2 and pH were slight for better compared to seawater aspirated rats. Treatment with E2, PaO2 was significantly increased compared with that of seawater group from 2 hours, and increased to peak at 4h post seawater instillation, so we choose 4h time point for the next studies.1.2 Seawater aspirated rats followed much higher PMVP and W/D ratios, which were reduced by estrogen administration.1.3 Seawater aspiration induced prominent lesions, as well as focal hemorrhage, distortion, and alveolar thickening. However, estrogen treatment significantly attenuated the lung injury induced by seawater, and even some parts of the lungs exhibited normal histological features.1.4 The western-blot data showed that compared with SW group, the expressions of AQP1 and AQP5 were lower in SW+E2 group. The immunohistochemistry staining further confirmed the above results from western blotting data. The RT-PCR data showed that compared with SW group, the expressions of AQP1 and AQP5 mRNA were lower in SW+E2 group. Estrogen treatment inhibits the upregulation of AQP1 and AQP5 protein and mRNA expression induced by seawater aspiration.2. Results of cell experiment2.1 Seawater induced the decrease in p-ERK expression. However, E2 treatment increased p-ERK following seawater incubation, which manifested that E2 can activate p-ERK in A549 cells after seawater administration. 2.2 The cell experiment indicted that the change of AQP1,AQP5 protein expression in A549 cells were similar to the lung tissues. Compared with SW group, the expressions of AQP1 and AQP5 were lower in SW+E2 group; Moreover, we examine the mechanism of E2 protective effect on AQP1 and AQP5 in seawater treated cells. Pretreated with ERK inhibitor U-0126, AQP1 and AQP5 protein expression were decreased compared with E2 treatment following seawater incubation.Conclusion:1. Our results showed 17β-estradiol treatment could alleviate the PE-SWD which was characterized by the improvement of blood gas index, pulmonary edema and histopathologic changes.2.17β-estradiol treatment could alleviate seawater aspiration-induced ALI in rat lung, and suppress the seawater-induced AQP1 and AQP5, results in an attenuation of pulmonary edema.3. After seawater administration,17β-estradiol activated the ERK in order to regulate the increased AQP1 and AQP5 in A549 cells.