Cryopreservation of human fetal organs
Cryopreservation of human fetal organs
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人类胎儿器官的冷冻保存
DOI：
10.1007/bf00318342
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发表时间：
2004
期刊：
影响因子：
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通讯作者：
G. Haselbacher
中科院分区：
文献类型：
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作者：
P. Groscurth;M. Erni;M. Balzer;H. Peter;G. Haselbacher
采用透射电镜（TEM）、扫描电镜（SEM）、体外培养和异种移植等方法，研究了低温保存对人胚胎不同器官形态、活力和分化能力的影响。为此目的，通过三步冷却程序冷冻来自10个人胎儿（9至14周龄）的肺、肾、小肠、甲状腺、脑、肝和脾的碎片。在3至12个月后，将样本快速解冻并进行TEM和/或体外培养和/或移植到裸鼠中。对冷冻的肺、肾和肠的TEM研究显示，大体上超微结构良好，而肝和脾碎片出现高度坏死。取三个胎儿的冷冻肠和肺标本，用于单层培养的建立。胰蛋白酶消化后，上皮细胞和间充质细胞在塑料瓶底部形成连续层。在进一步的传代过程中，上皮细胞的数量减少，导致纯成纤维细胞培养物的形成。来自一个胎儿的冷冻脑组织也被成功培养，在玻璃盖片表面形成不同大小的细胞簇和纤维束。皮下植入裸鼠后，绝大多数来自肺、肾、肠和甲状腺的碎片在受体体内生长。通过TEM或放射自显影研究，异种移植物的生长伴随着细胞分化的持续（肾、肠）或甚至增加（肺、甲状腺）。然而，移植的肝和脾碎片在裸鼠皮下组织中有规律地退化形成固体瘢痕。综上所述，三步冷却程序是保存不同人胎器官的有效方法，可以保持组织的形态、活力和分化能力。解冻的标本可用于几项实验，包括体内和体外研究。
The effects of low temperature preservation on morphology, viability and differentiation capacity of different human fetal organs were studied using transmission (TEM) and scanning (SEM) electron microscopy, in vitro cultivation as well as xenogeneic transplantation. For this purpose fragments of lung, kidney, small intestine, thyroid, brain, liver and spleen from 10 human fetuses (aged 9 to 14 weeks of gestation) were frozen by a three step cooling procedure. After 3 to 12 months the specimens were thawed rapidly and processed for TEM and/or in vitro cultivation and/or transplantation into nude mice. TEM studies on frozen lung, kidney and intestine revealed generally a well preservedultrastructurewhereas liver and spleen fragments appeared highly necrotic. From three fetuses, frozen intestine and lung specimens were used for the establishment ofmonolayer cultures. Following trypsinization, both epithelial and mesenchymal cells formed a continous layer on the bottom of plastic bottles. During further subpassages the number of epithelial cells decreased resulting in the formation of pure fibroblast cultures. Frozen brain tissue from one fetus was also successfully cultivated forming cell clusters and fiber bundles of variable size at the surface of glass cover slips. Following subcutaneous implantation intonude mice, the vast majority of fragments from lung, kidney, intestine and thyroid was found to growth in the recipients. The growth of xenografts was accompanied by persistence (kidney, intestine) or even increase (lung, thyroid) in cellular differentiation studied by TEM or autoradiography. Transplanted liver and spleen fragments, however, regularly regressed forming solid scars in the subcutaneous tissue of nude mice.In summary, the three-step cooling procedure is an effective method for storage of different human fetal organs preserving morphology, viability and differentiation capacity of the tissues. Thawed specimens may be used for several experiments including in vivo and in vitro studies.