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中华移植杂志(电子版)

中华移植杂志(电子版) ›› 2020, Vol. 14 ›› Issue (04) : 229 -233. doi: 10.3877/cma.j.issn.1674-3903.2020.04.007

所属专题： 文献；

论著

大鼠卵巢体外低温机械灌注模型的建立
张诗辰1, 姚翰林1, 杨志坚1, 叶啟发2, 王彦峰1,†()   
  1. 1. 430071　武汉大学中南医院，武汉大学肝胆疾病研究院，武汉大学移植医学中心，移植医学技术湖北省重点实验室
    2. 430071　武汉大学中南医院，武汉大学肝胆疾病研究院，武汉大学移植医学中心，移植医学技术湖北省重点实验室；410013　长沙，中南大学湘雅三医院，卫生部移植医学工程技术研究中心
  • 收稿日期:2020-03-06 出版日期:2020-08-25
  • 通信作者: 王彦峰
  • 基金资助:
    国家自然科学基金面上项目(81570079)

The establishment of hypothermic mechanical perfusion model of rat ovary in vitro

Shichen Zhang1, Hanlin Yao1, Zhijian Yang1, Qifa Ye2, Yanfeng Wang1,†()   

  1. 1. Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Hubei Key Laboratory of Medical Technology on Transplantation, Wuhan, 430071, China
    2. Zhongnan Hospital of Wuhan University, Institute of Hepatobiliary Diseases of Wuhan University, Transplant Center of Wuhan University, Hubei Key Laboratory of Medical Technology on Transplantation, Wuhan, 430071, China; The 3rd Xiangya Hospital of Central South University, Research Center of National Health Ministry on Transplantation Medicine Engineering and Technology, Changsha 410013, China
  • Received:2020-03-06 Published:2020-08-25
  • Corresponding author: Yanfeng Wang
  • About author:
    Corresponding author: Wang Yanfeng, Email:
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引用本文：

张诗辰, 姚翰林, 杨志坚, 叶啟发, 王彦峰. 大鼠卵巢体外低温机械灌注模型的建立[J]. 中华移植杂志(电子版), 2020, 14(04): 229-233.

Shichen Zhang, Hanlin Yao, Zhijian Yang, Qifa Ye, Yanfeng Wang. The establishment of hypothermic mechanical perfusion model of rat ovary in vitro[J]. Chinese Journal of Transplantation(Electronic Edition), 2020, 14(04): 229-233.

目的

建立大鼠卵巢体外低温机械灌注(HMP)模型，并验证其有效性。

方法

26只雌性8～10周龄SD大鼠，供体组、受体组、正常对照组和卵巢摘除组各6只，另有2只大鼠HMP结束后留取卵巢标本以备检测。供体组和取材大鼠获取卵巢和肾脏后置于0～4 ℃林格氏液中保存2 h，再行HMP 2 h。供体组大鼠卵巢组织HMP结束后移植至受体组大鼠左肾包膜下，移植7 d后检测血清抗苗勒管激素(AMH)水平，同时获取移植卵巢进行组织病理检查以及Ki67和CD31免疫组织化学染色。采用单因素方差分析比较正常对照组、卵巢摘除组和受体组大鼠血清AMH水平，采用t检验比较正常对照组和受体组卵泡计数以及Ki67和CD31阳性细胞面积百分比。P<0.05为差异有统计学意义。

结果

HMP后的卵巢组织血管未见明显扩张，受体组大鼠移植7 d后的卵巢组织卵泡结构完整，成熟卵泡数为(7.0±0.6)个，低于正常对照组(26.3±3.4)个，差异有统计学意义(t＝5.633，P<0.05)。受体组血清AMH为(4.63±0.87)ng/mL，高于卵巢摘除组(2.55±0.16)ng/mL，低于正常对照组(7.39±1.04)ng/mL，但差异均无统计学意义(P均>0.05)。移植7 d后受体组大鼠卵巢组织Ki67和CD31阳性细胞面积百分比分别为(29.0±2.4)%和(12.3±1.0)%，正常对照组分别为(21.9±3.8)%和(12.6±1.2)%，差异均无统计学意义(t＝1.634和0.178，P均>0.05)。

结论

本研究构建的大鼠卵巢体外HMP模型，方法简单，可短时间维持卵巢的正常功能，为卵巢器官保存提供了一个新的方式。

关键词: 大鼠, 卵巢移植, 低温机械灌注, 卵巢
Objective

To establish a hypothermic mechanical perfusion (HMP) model of rat ovary in vitro and test its effectiveness.

Methods

26 female SD rats aged 8-10 weeks were divided into donor group, recipient group, normal group and ovariectomy group, with 6 rats in each group. Ovarian tissue of 2 rats was harvested for test after HMP. Ovaries and kidneys from donor group and HMP management only group were placed in Ringer′s solution at 0-4 ℃ for 2 h, and follwed by HMP for 2 h. Ovarian tissue from donor group was transplanted under the recipient′s left kidney capsule after HMP, and the serum anti-Mullerian hormone (AMH) in recipient rat was tested 7 days after transplantation, and the ovaries were harvested for histological analysis, and immunohistochemistry for CD31 and Ki67. ANOVA was used to compare the serum AMH levels in the normal group, the ovariectomy group and the recipient group. The t-test was used to compare the follicle counts and the percentages of Ki67 and CD31 positive cells in the normal group and the recipient group. P<0.05 was considered statistically significant.

Results

No obvious dilatation was observed in the blood vessels of ovarian tissue after HMP. The follicle structure of rats from recipient group was complete 7 days after transplantation, and the number of mature follicles was (7.0±0.6), which was lower than that of the normal group (26.3±3.4), the difference had statistical significance (t=5.633, P<0.05). The serum AMH of recipient group was (4.63±0.87) ng/mL, higher than that of the ovariectomy group [(2.55±0.16) ng/mL], and lower than that of the normal group [(7.39±1.04) ng/mL]. The percentage of Ki67 and CD31 positive cells in ovarian tissue from recipient group 7 days after transplantation were (29.0±2.4)% and (12.3±1.0)%, which had no significant difference with normal group [(21.9±3.8%) and (12.6±1.2%)] (t=1.634 and 0.178, P all>0.05).

Conclusions

The method to construct the model of HMP in vitro of rat ovary in this study is simple and can maintain the normal function of ovary for a short time, providing a new method for organ preservation of ovary.

Key words: Rats, Ovarian transplantation, Hypothermic mechanical perfusion, Ovary
图1 SD大鼠卵巢获取解剖示意图
图2 SD大鼠卵巢体外低温机械灌注前后大体观
图3 SD大鼠卵巢体外低温机械灌注前后和移植7 d后病理学检查结果(HE ×100)
图4 SD大鼠卵巢免疫组织化学染色结果(×200)
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