Pulmonary Surfactant In Treatment Of HCL-induced Early Stage Acute Lung Injury In Rabbit

Gastric acid inhalation is one of the common clinical critical complications in emergency patients, and it is also an important cause of acute lung injury / acute respiratory distress syndrome (ALI / ARDS).ALI is considered as the early stage of ARDS, so ALI model is very meaningful for the study of the treatment of ARDS. A biphasic injury pattern is observed with injury peaks at 1 hr and 4 hr. Those studies suggest that gastric acid inhalation results in a biphasic acute injury .We hypothesize that the first phase results from a direct physiochemical process.The second phase, occurring 2–3 hr later, is mediated by neutrophils and is consistent with an acute inflammatory response. There have been many case reports and uncontrolled studies evaluating exogenous pulmonary surfactant in the first phase patients with ALI / ARDS. They all agreed that pulmonary surfactant could improve pulmonary ventilation function and mechanical properties and all studied objects have a good prognosis. This article will discuss the efficacy of pulmonary surfactant treatment in the second phase of ALI / ARDS induced by hydrochloric acid inhalation and provide a new thinking for clinical treatment.Objective To establish an animal model of ALI and observe the changes of arterial blood-gas on every scheduled time,pulmonary tissue permeability,pulmonary tissue pathology after pulmonary surfactant treatment in the second phase of hydrochloric acid inhalation induced-ALI/ ARDS. Pulmonary tissue permeability includes wet and dry lung ratio,the number of white blood cell and total protein content in bronchoalveolar lavage fluid. Material and methodsStudy groups:Eighteen New Zealand white male rabbits, weighing 1.7 ~ 2.3 kg, were from Forth Military Medical University. Animals were randomly divided into three groups: healthy control group (n=6),model group (n=6),and treatment group (n=6).1. Establish the model of lung injury after hydrochloric acid inhalationRabbits were anesthetized with Sumianxin(0.2ml/ kg)by intramuscular injection. Rabbits were maintained in the supine position during experiments. A heparin-filled catheter was inserted into the right carotid artery for withdrawal of references blood flow samples. Trachea was isolated, and a 10 ml syringe needle was inserted and fixed to a position approximately 1 cm above the carina for the inhalation of dilute hydrochloric acid and the pulmonary surfactant delivery. Following a 15 min stable baseline period, hydrochloric acid (pH=1.5,0.6×2 ml/ kg ) was instilled slowly through syringe needle at the right side of body, the head raised by a 45-degree angle, then 2 ml air was pushed into lung slowly. 5 minutes later, hydrochloric acid (pH=1.5,0.4×2 ml/ kg) was instilled slowly through syringe needle at the left side of body. Arterial blood gas analysis showed PaO2/ FiO2 < 300mmHg, the models are reckoned to have been successfully established.3. MethodTreatment group: pulmonary surfactant (80 mg / ml, 120 mg / kg ) was given to intervene through syringe needle as the same manner as hydrochloric acid inhalation; control group and model group: the same volume of normal saline was instilled slowly at the appropriate time. Arterial blood gas analysis were performed at 30 min before and 4h after hydrochloric acid inhalation and 30 min, 2h, 4h, 6h after pulmonary surfactant was given. Trachea was cut on the position approximately 0.3cm above the carina after rabbit was sacrificed. Then a tracheal intubation was inserted and fixed. The left main bronchus was ligated with surgical line. The right lung was lavaged with 20 ml sterile saline. This action was repeated 3 times. Gathered bronchoalveolar lavage fluid was filtered and centrifuged with the speed of 1000r / min for 10 min at 4℃. The supernatant was stored at - 80℃for evaluating total protein content. The remained sediment was suspended in 1ml normal saline. A drop of liquid was stained with violaceum methylium, and total white blood cell were counted under the microscope. The inferior lobe of left lung was removed and dried to a constant weight (80℃for 48h). The dried lung was weighed and the lung dry/wet radio was then calculated. The superior lobe of left lung was fixed in 10% formalin, and then embedded in paraffin wax, sliced, and stained with hematoxylin-eosin.Results1. Arterial blood gas analysisBefore the start of hydrochloric acid inhalation, the differences of PaO2 and PaCO2 of all animals were not significant. After hydrochloric acid inhalation, PaO2 decreased significantly and PaCO2 increased significantly in model group and treatment group. Compared with model group, treatment groups'value of PaO2 and PaCO2 at different time points improved significantly after treatment with pulmonary surfactant, but was still unable to return to the level of healthy control group.2. Pulmonary tissue permeabilityWet and dry lung ratio,the number of white blood cell and total protein content in bronchoalveolar lavage fluid in the treatment group were significantly lower than model group, but still unable to return to the level of healthy control group.3. Gross appearance of lung tissueControl group was considered as normal lung tissue. Compared with the control group, we can see kermesinus lamellar and (or) hemorrhage that are mainly located at the lower lung and extrude many bloody edema from cross-section. Pulmonary edema is very severe. Treatment group showed pink, uniformity in the surface of lung and no significant edema, having a small amount of exudates in part of section.4. Common light microscopeThe lung tissue structure of control group was seen as normal structure. Model group of lung tissue sections stained by hematoxylin and eosin presented alveolus and interstitial edema at different degree, with alveolar septum and interlobular septum broken, and we can see scatter hemorrhage. Blood capillary of alveolar wall engorgement and pulmonary hyaline membrane are formed, and the infiltration of inflammatory cells in alveolar spaces. In the treatment group, these changes were less obvious, and the amplitude decreased.ConclusionAt the lung injury secondary stage induced by hydrochloric acid inhalation, we can see significant improvements in arterial blood gas analysis, if we give exogenous pulmonary surfactant to intervene. Wet and dry lung ratio,the number of white blood cell and total protein content in bronchoalveolar lavage fluid all descent. Pathological structure of the lung tissue is improved significantly ,but they are still unable to return to the level of the control group.