摘要
目的利用小型猪多个系统组织结构和功能接近于人类的特性,建立模拟失重动物模型,观察其病理生理学变化,为航空航天失重环境研究提供新方法。方法选取9头普通级小型猪,随机分为实验组(n=7)和对照组(n=2)。实验组小型猪使用定制金属笼固定,帆布吊带悬挂使其后肢离地去负荷,身体与地面呈-20°角,模拟失重30 d(每天24 h)。通过采集不同时间点各组小型猪的体重、血容量、血生化指标等数据,对其基础体征的变化进行统计分析。实验结束后,对小型猪实施安乐死并解剖取材,对心血管、骨骼、骨骼肌等各系统组织脏器进行HE、Masson染色后的组织病理学观察,并对各动脉血管厚度、骨骼肌肌纤维直径进行统计分析。同时,采用蛋白质免疫印迹法检测骨骼肌肌肉萎缩蛋白包括肌环指蛋白1(muscle-specific RING finger protein 1,MuRf-1)和肌萎缩素1(muscle atrophy F-box,MAFbx,又称Atrogin-1)的表达量,并采用免疫组织化学染色法检测脑内星形胶质细胞激活相关蛋白即胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)的表达量,以反映各系统的病理生理学功能变化。结果后肢去负荷造模后,实验组小型猪体重显著下降(P<0.001),血容量也显著下降(P<0.01)。实验期间,实验组小型猪的血红蛋白、红细胞比容和红细胞计数水平显著降低(P<0.05),随后逐渐恢复;丙氨酸氨基转移酶和γ-谷氨酰转移酶表达水平呈现先下降(P<0.05)后回升的趋势,白蛋白水平显著下降(P<0.001),球蛋白水平显著上升(P<0.01),肌酐水平显著下降(P<0.05)。实验组小型猪腓肠肌的平均肌纤维直径显著缩短(P<0.05),肌纤维直径分布左移,小直径肌纤维增多;同时腓肠肌、椎旁肌肉Atrogin-1的表达水平显著增加(P<0.05)。这些变化与航天失重对人和动物的影响基本一致。此外,实验组小型猪的脑皮质顶叶、额叶和海马区域部分神经元变性,小脑区域浦肯野细胞数量轻度减少,而海马区的GFAP阳性信号显著增强(P<0.05)。结论-20°角后肢去负荷30 d的小型猪可作为一种新的模拟失重动物模型,用于航空航天领域相关研究。
Objective To establish a weightlessness simulation animal model using miniature pigs,leveraging the characteristic of multiple systems’tissue structures and functions similar to those of humans,and to observe pathophysiological changes,providing a new method for aerospace research.Methods Nine standard-grade miniature pigs were selected and randomly divided into an experimental group(n=7)and a control group(n=2).The experimental group was fixed using customized metal cages,with canvas slings suspending their hind limbs off the ground,and the body positioned at a-20°angle relative to the ground to simulate unloading for 30 days(24 hours a day).Data on body weight,blood volume,and blood biochemistry indicators were collected at different time points for statistical analysis of basic physiological changes.After the experiment,the miniature pigs were euthanized and tissue samples were collected for histopathological observation of the cardiovascular,skeletal and muscle systems HE and Masson staining.Statistical analysis was also conducted on the thickness of arterial vessels and the diameter of skeletal muscle fibers.Additionally,western blotting was employed to detect the expression levels of skeletal muscle atrophy-related proteins,including muscle-specific RING finger protein 1(MuRf-1)and muscle atrophy F-box(MAFbx,as known as Atrogin-1),while immunohistochemistry was used to detect the expression of glial fibrillary acidic protein(GFAP),an indicator of astrocyte activation in the brain,reflecting the pathophysiological functional changes across systems.Results After hindlimb unloading,the experimental group showed significant decreases in body weight(P<0.001)and blood volume(P<0.01).During the experiment,hemoglobin,hematocrit,and red blood cell count levels significantly decreased(P<0.05)but gradually recovered.The expression levels of alanine aminotransferase andγ-glutamyltransferase initially decreased(P<0.05)before rebounding,while albumin significantly decreased(P<0.001)and globulin significantly increased(P<0.01).Creatinine significantly decreased(P<0.05).The average diameter of gastrocnemius muscle fibers in the experimental group significantly shortened(P<0.05),with a leftward shift in the distribution of muscle fiber diameters and an increase in small-diameter muscle fibers.Simultaneously,Atrogin-1 expression in the gastrocnemius and paravertebral muscles significantly increased(P<0.05).These changes are generally consistent with the effects of weightlessness on humans and animals in space.Furthermore,degenerative changes were observed in some neurons of the cortical parietal lobe,frontal lobe,and hippocampal regions of the experimental group,with a slight reduction in the number of Purkinje cells in the cerebellar region,and a significant enhancement of GFAP-positive signals in the hippocampal area(P<0.05).Conclusion Miniature pigs subjected to a-20°angle hind limb unloading for 30 days maybe serve as a new animal model for simulating weightlessness,applicable to related aerospace research.
作者
涂颖欣
纪依澜
王菲
杨东明
王冬冬
孙芷馨
戴悦欣
王言吉
阚广捍
吴斌
赵德明
杨利峰
TU Yingxin;JI Yilan;WANG Fei;YANG Dongming;WANG Dongdong;SUN Zhixin;DAI Yuexin;WANG Yanji;KAN Guanghan;WU Bin;ZHAO Deming;YANG Lifeng(College of Veterinary Medicine,China Agricultural University,Beijing 100193,China;China Astronaut Research and Training Center,Beijing 100094,China)
出处
《实验动物与比较医学》
CAS
2024年第5期475-486,共12页
Laboratory Animal and Comparative Medicine
基金
国家重点研发计划“实验动物新品种、新品系开发与评价”(2022YFF0710501)
航天医学基础与应用国家重点实验室研究基金资助自主研究项目“模拟失重和压力变化复合因素对小型巴马猪咽鼓管功能变化规律研究”(SMFA22B05)。
关键词
模拟失重
小型猪
动物模型
后肢去负荷
Simulated weightlessness
Miniature pig
Animal model
Hindlimb unloading
