【研究背景】

慢性阻塞性肺疾病（Chronic Obstructive Pulmonary Disease，COPD）是一种以持续性呼吸道症状和气流受限为特征的可以预防和治疗的呼吸系统慢性疾病。COPD患病率高、死亡率高，我国总患病人数已达1亿，是仅次于高血压和糖尿病的第3大慢性疾病，目前临床上仍缺乏有效的治疗方法和药物，常用药物有糖皮质激素、支气管扩张剂等，但往往只能在一定程度上缓解症状，无法阻止肺功能进行性下降，效果甚微，患者的生活质量和劳动能力受到限制，严重影响着国民健康和社会经济发展，迫切需要探寻新药物。

阿胶是我国一味传统中药，药用历史悠久，临床上常用于妇科疾病和血液系统疾病的治疗，典型的药理作用有补血造血、滋阴润肺、抗氧化、抗肿瘤、抗衰老、抗休克、抗疲劳、加快钙吸收、改善睡眠质量、增强免疫力、安胎保胎和美容养颜等，补血造血功能已广为流传和证实，但从动物模型和细胞水平等层面上对其滋阴润肺、抗炎、耐缺氧等药理作用的机制报道很少见，对于COPD的治疗尚无系统的循证医学证据，其药理机制也尚未完全阐明。

【研究目的】

以SD大鼠为研究对象，通过烟熏法构建COPD大鼠模型，观察阿胶灌胃给药对COPD大鼠的疗效，为COPD临床治疗提供新的药物，也为阿胶在COPD中的应用提供实验依据。

【研究方法】

将25只9~10周龄健康SPF级雄性SD大鼠随机分为5组：对照组（control）、模型组（COPD）、阿胶低剂量组（low-dose CCA）、阿胶中剂量组（medium-dose CCA）和阿胶高剂量组（high-dose CCA），每组5只。采用单纯卷烟烟雾暴露的方法构建COPD大鼠模型：除对照组外，其余4组大鼠均放置于暴露装置中，进行卷烟烟雾被动吸入，对照组大鼠同时置于清洁空气中进行伪暴露，总暴露时间是336天。烟雾暴露结束次日起开始阿胶干预处理：将阿胶粉末溶解、震荡、混匀配制成阿胶溶液；灌胃给予1 g/kg、2 g/kg和4 g/kg三种不同浓度的低、中、高剂量阿胶溶液，对照组和模型组大鼠同时灌胃等量生理盐水，每次现配现用，每日1次，连续灌胃28天；造模期间和阿胶灌胃期间观察各组大鼠一般情况并称重。灌胃结束次日采用动物肺功能仪活体检测各组大鼠的肺功能，麻醉后经右心室采集血液处死，收集左心、右心、肺脏、肝脏、脾脏、肾脏、肾上腺、胸腺、睾丸、脑组织标本，称量湿重，计算脏器系数和脑脏比。肺组织常规制作石蜡切片，行苏木素-伊红（HE）染色镜下观察病理形态改变。检测各组大鼠动脉血清电解质Na⁺、K⁺、Cl^-和Ca²⁺水平。检测各组大鼠血脂总胆固醇（TCHO）和甘油三酯（TG）水平。检测各组大鼠血液红细胞系统参数，包括红细胞计数（RBC）、血红蛋白含量（Hb）、红细胞分布宽度（RDW）、红细胞压积（HCT）、平均红细胞体积（MCV）、平均红细胞血红蛋白含量（MCH）、平均红细胞血红蛋白浓度（MCHC）。检测各组大鼠血小板计数（PLT）和血小板平均体积（MPV）两项血小板参数。采用Graph Pad Prism 8.0统计软件分析数据和绘图，进行各指标在5组间的差异比较，以P < 0.05为差异具有统计学意义。

【研究结果】

对照组大鼠一般状况良好，体重增长明显，肺功能各项指标正常，肺组织外观正常，肺泡结构完整，无肺气肿改变，无炎性浸润。各脏器体积正常，无扩大或缩小，质量均正常，Na⁺、K⁺、Cl^-、Ca²⁺、TCHO、TG、RBC、Hb、RDW、HCT、MCV、MCH、MCHC）、PLT、MPV水平均处于正常值范围。

和对照组大鼠比较，模型组大鼠随卷烟烟雾暴露时间的延长，逐渐出现咳嗽、喷嚏等呼吸道症状，进食量减少，精神差，体重增长缓慢，肺功能指标TV、EV、Te、RT、EEP和Penh均显著增加（P < 0.05），而PEF、PIF、MV、EF₅₀和f均显著降低（P < 0.05），肺组织HE染色结果显示出现肺气肿和炎性病理性改变，肺组织病变加重；左心质量、右心质量和肝脏质量明显增加（P < 0.05），肾脏质量、肾脏指数明显减小（P < 0.05），其余脏器质量及其脏器系数均无显著性差异（P > 0.05），左心、右心和肝脏的脏脑比系数均显著增大（P < 0.05），肾脏的脏脑比系数显著减小（P < 0.05），其余脏器的脏脑比系数无显著性差异（P > 0.05）；（P > 0.05），TCHO、TG、RBC、Hb、HCT、MCV、MCH、MCHC、PLT和MPV明显增多（P < 0.05），Na⁺、K⁺、Cl^-、Ca²⁺和RDW水平无显著变化（P > 0.05）。

与模型组大鼠相比，阿胶低、中和高剂量组大鼠灌胃期间咳嗽、喷嚏减少，摄食量增加，精神状况好转，体重增长加快。阿胶低剂量组大鼠各脏器质量无显著性变化（P > 0.05），肾脏指数明显增大（P < 0.05），各脏器脏脑比系数均无显著性差异（P > 0.05），TCHO、Hb、HCT明显减少（P < 0.05），Na⁺、K⁺、Cl^-、Ca²⁺、TG、RBC、MCV、MCH、MCHC、RDW、PLT和MPV无显著性变化（P > 0.05）；阿胶中剂量组大鼠肺功能指标Penh明显降低（P < 0.05），各脏器质量均无显著性变化（P > 0.05），肾脏指数明显增大（P < 0.05），各脏器脏脑比系数均无显著性差异（P > 0.05），TCHO、HCT、MCV明显减少（P < 0.05），Na⁺、K⁺、Cl^-、Ca²⁺、TG、RBC、Hb、MCH、MCHC、RDW、PLT和MPV均无显著性变化（P > 0.05）；阿胶高剂量组Te、RT、Penh显著下降（P < 0.05），MV、PIF、EF₅₀显著增加（P < 0.05），肺部炎性细胞浸润减少，肺泡扩张减轻，肺大泡减少，肺气肿程度减轻，肺组织病理损伤明显缓解，左心质量明显减少（P < 0.05），其余脏器质量均无显著性变化（P > 0.05），肾脏指数明显增大（P < 0.05），其余脏器系数均无显著差异（P > 0.05），各脏器脏脑比系数均无显著性差异（P > 0.05），TCHO、TG、Hb、HCT、MCV、MCH、MCHC、PLT和MPV明显减少（P < 0.05），Na⁺、K⁺、Cl^-、Ca²⁺、RBC、RDW无显著性变化（P > 0.05）。

【研究结论】

1.阿胶可以改善COPD大鼠体重的减轻，缓解卷烟烟雾暴露对大鼠生长的抑制作用。

2.高剂量阿胶可以缓解COPD大鼠缺氧状态，改善肺功能，减轻肺组织炎性反应，降低高血脂水平，还可能减轻卷烟烟雾暴露对心脏、肝脏和肾脏造成的损伤。

3.不同剂量阿胶对COPD大鼠血清电解质水平无显著影响。

Background

Chronic obstructive pulmonary disease (COPD) is a preventable and treatable chronic disease of the respiratory system characterized by persistent respiratory symptoms and restricted airflow. The prevalence of COPD is high and the mortality rate is high. The total number of COPD patients in China has reached 100 million, which is the third largest chronic disease after hypertension and diabetes. At present, there is still a lack of effective therapeutic methods and drugs in clinical practice, commonly used drugs include corticosteroids, bronchodilators, etc. However, the symptoms can only be alleviated to a certain extent, and the progressive decline of lung function can’t be prevented, with little effect. The quality of life and working ability of patients are limited, which seriously affects the national health and social and economic development. Therefore, it’s urgent to explore new drugs.

Ejiao (Latin: Colla Corii Asini) is a traditional Chinese medicine with a long history of medicinal use. Clinically, it is often used in the treatment of gynecological diseases and hematological diseases. Typical pharmacological effects include blood tonifying hematopoiesis, ziyin runfei, anti-oxidation, anti-tumor, anti-aging, anti-shock, anti-fatigue, accelerating calcium absorption, improving sleep quality, enhancing immunity, fetal safety and beautify the appearance, etc. Blood tonifying and hematopoietic function has been widely spread and confirmed, but there are few reports on the mechanism of its pharmacological effects on zi yin run fei, anti-inflammation and anti-hypoxia at the animal model and cell level. There is no systematic evidence-based medicine for the treatment of COPD, and it’s pharmacological mechanism has not been fully clarified.

Objective

Taking SD rats as the research object, and COPD rat model was established by smoke fumigation method to observe the efficacy of oral administration of Colla Corii Asini on COPD rats, to provide a new drug for clinical treatment of COPD, and also provide experimental basis for the application of Colla Corii Asini in COPD.

Methods

25 healthy SPF male SD rats aged 9~10 weeks were randomly divided into 5 groups: namely the control group, model group (COPD), low-dose CCA group, medium-dose CCA group and high-dose CCA group, with 5 rats in each group. COPD rats model was established by cigarette smoke exposure alone: except the control group, the other 4 groups of rats were placed in the exposure device for passive inhalation of cigarette smoke, while the control group rats were placed in clean air for pseudo-exposure at the same time, and the total exposure time was 336 days. From the next day after modeling, the intervention treatment of CCA was started: the CCA powder was dissolved, shaken and mixed into prepare gelatin solution; three different concentrations of low, medium and high doses of CCA solution were given by gavage: 1 g/kg, 2 g/kg and 4 g/kg. The rats in control group and model group were gavaged with the same amount of physiological saline at the same time, once a day, for 28 days. The general conditions of rats in each group were observed and weighed during the modeling period and the intragastric administration of CCA. The next day after intragastric administration, the lung function of rats in each group was detected by animal lung function apparatus. After anesthesia, blood was collected through the right ventricle and sacrificed. Tissue specimens of the left heart, right heart, lung, liver, spleen, kidney, adrenal gland, thymus, testis and brain were collected, weighing wet weight, calculating viscera coefficient and brain-viscera ratio. Paraffin sections of lung tissue were routinely made, and the pathological changes were observed under microscope by hematoxylin-eosin (HE) staining. The levels of electrolyte Na⁺, K⁺, Cl^- and Ca²⁺ in arterial serum of rats in each group were detected. The levels of total cholesterol (TCHO) and triglyceride (TG) were determined in each group. The RBC system parameters of rats in each group was detected, including red blood cell count (RBC), hemoglobin content (Hb), red blood cell distribution width (RDW), hematocrit (HCT), mean red blood cell volume (MCV), mean red blood cell hemoglobin content (MCH), and mean red blood cell hemoglobin concentration (MCHC). Platelet count (PLT) and mean platelet volume (MPV) were measured. Graph Pad Prism 8.0 statistical software was used to analyze the data and drawings, and the differences of each index among the 5 groups were compared P < 0.05 indicates that the differences is statistically significant.

Results

The rats in the control group were generally in good condition, with obvious weight gain, normal indicators of lung function, normal appearance of lung tissue, complete alveolar structure, no emphysema changes, and no inflammatory infiltration. The volume of each organ was normal without enlargement or contraction, and the mass was normal. The levels of Na⁺, K⁺, Cl^-, Ca²⁺, TCHO, TG, RBC, Hb, RDW, HCT, MCV, MCH, MCHC, PLT and MPV were all in the normal range.

Compared with the control group, with the prolongation of cigarette smoke exposure time, the model group rats gradually development respiratory symptoms, such as cough and sneeze, decreased food intake, poor mental performance, slow weight gain, and significantly increased pulmonary function indexes, such as TV, EV, TE, RT, EEP and Penh ( P < 0.05), and PEF, PIF, MV, EF₅₀ and F were significantly decreased ( P < 0.05), lung tissue HE staining showed emphysema and inflammatory pathological changes, lung tissue lesions were aggravated, Left heart mass, right heart mass and liver mass were significantly increased ( P < 0.05), renal mass and renal index decreased significantly ( P < 0.05), and there was no significant difference in the mass and coefficient of other organs ( P > 0.05), the ratio coefficitnts of left heart , right heart and liver were significantly increased ( P < 0.05), the ratio of kidney to brain decreased significantly ( P < 0.05), and there was no significant differences in the ratio coefficients of the other organs ( P > 0.05). There were no significant changes in serum Na⁺, K⁺, Cl^-, Ca²⁺ levels ( P > 0.05), THCO, TG, RBC, Hb, HCT, MCV, MCH, MCHC, PLT and MPV were significantly increased ( P < 0.05), there was no significant difference in RDW ( P > 0.05).

Compared with the model group, cough and sneeze decreased, food intake increased, mental state improved and weight gain accelerated in the low-dose, medium-dose and high-dose CCA groups during intragastric administration. There was no significant change in the weight of each organ in the low-dose CCA group ( P > 0.05 ), the renal index increased significantly ( P < 0.05), and there was no significant difference in the ratio of viscera to brain among all organs ( P > 0.05), THCO, Hb and HCT were significantly decreased ( P < 0.05), there were no significant changes in Na⁺, K⁺, Cl^-, Ca²⁺, TG, RBC, MCV, MCH, MCHC, RDW, PLT and MPV ( P > 0.05 ); The lung function index Penh of rats in the medium-dose CCA group was significantly decreased ( P < 0.05), there was no significant changes in the mass of each organ ( P > 0.05), the renal index increased significantly ( P > 0.05), and there was no significant difference in the ratio of viscera to brain among all organs ( P > 0.05), TCHO, HCT and MCV were significantly decreased ( P < 0.05), there were no significant changes in Na⁺, K⁺, Cl^-, Ca²⁺, TG, RBC, Hb, MCH, MMCHC, RDW, PLT and MPV ( P > 0.05); The Te, RT and Penh of high-dose CCA group were significantly decreased ( P < 0.05), MV, PIF and EF₅₀ were significantly increased ( P < 0.05), lung inflammatory cell infiltration was reduced, alveolar dilatation was reduced, lung bullae was reduced, emphysema was reduced, pathological injury of lung tissue was significantly relieved, and left heart mass was significantly reduced ( P < 0.05),the mass of other organs had no significant change ( P > 0.05), the renal index increased significantly ( P < 0.05), the coefficients of other prgans has no significant difference ( P > 0.05), and there was no significant difference in the ratio of viscera to brain among all organs ( P > 0.05), TCHO, TG, Hb, HCT, MCV, MCH, MCHC, PLT and MPV were aignificantly decreased ( P < 0.05), there were no significant changes in Na⁺, K⁺, Cl^-, Ca²⁺, RBC and RDW ( P > 0.05).

Conclusion

1. Colla Corii Asini can improve the weight loss of COPD rats and alleviate the inhibitory effect of cigarette smoke exposure on the growth of rats.

2. High-dose Colla Corii Asini can alleviate hypoxia, improve lung function, reduce the inflammatory response of lung tissue, and reduce hyperlipidemia in COPD rats. It may also reduce the damage to the heart, liver and kidney caused by cigarette smoke exposure.

3. Different doses of Colla Corii Asini had no significant effect on serum electrolyte level of COPD rats.