濒危基因编辑小鼠品系拯救技术及其应用探讨

doi:10.12300/j.issn.1674-5817.2023.080

摘要/Abstract

摘要：

基因编辑小鼠模型优势显著，是人类基因功能研究、疾病机制探索和新药研发等理想的实验动物。基因编辑小鼠因自身生育力低下、年老、疾病等因素导致的品系断代会对科学研究造成不可挽回的损失。相关从业人员需针对不同情况采用相应的技术手段进行品系拯救，同时要综合考虑技术选择和成本控制，尽可能降低小鼠品系拯救的实验成本。首先，当仅存的处于生育年龄的基因编辑小鼠突然死亡时，对死亡雄鼠可以通过体外受精（in vitro fertilization，IVF）、单精子卵胞质显微注射（intracytoplasmic sperm injection，ICSI），对雌鼠可通过卵巢移植等技术进行品系拯救。其次，由于年老或疾病因素具有断代风险的小鼠，可以通过IVF-胚胎移植（embryo transfer，ET）、单侧附睾尾活体辅助生殖等技术进行品系拯救。再者，生命状态差的性成熟前小鼠，可以采用圆形精子注射（round spermatid injection，ROSI）或卵巢移植等技术进行品系拯救。本文对常用濒危小鼠品系拯救技术及其应用进行了探讨，以期为相关从业人员更好地做好基因编辑小鼠品系维护提供参考。

关键词: 基因编辑, 濒危小鼠, 品系拯救, 辅助生殖技术

Abstract:

Gene-edited mice are the most ideal laboratory animals for studying human gene functions, exploring disease mechanisms, and developing new drugs. Strain resulting from low fertility, aging, illness, etc. can cause irreversible losses to scientific research, so strain rescues of genetically engineering mice require different measures accordingly. Meanwhile, cost control is another key point when a specific technology is applied. First of all, when the only remaining gene-edited mouse in reproductive age suddenly dies, the dead male mouse can be rescued by in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI), while the female mouse can be saved by ovarian transplantation, etc. Secondly, due to aging or diseases, mice can be saved through IVF-embryo transfer (ET) and unilateral epididymal tail assisted reproduction. Thirdly, round sperm injection (ROSI) and ovarian transplantation can be used to save endangered mice before sexual maturity with poor life status. This paper reviews rescue techniques of common endangered mice and their applications, which provides a reference for relevant practitioners to better maintain gene-edited mouse strains.

Key words: Gene-edited, Endangered mouse, Strain rescue, Assisted reproductive technology

中图分类号:

Q95-33

聂永强, 王朝霞. 濒危基因编辑小鼠品系拯救技术及其应用探讨[J]. 实验动物与比较医学, 2023, 43(6): 636-640.

Yongqiang NIE, Zhaoxia WANG. Rescue Technology and Its Application of Endangered Gene-Edited Mice[J]. Laboratory Animal and Comparative Medicine, 2023, 43(6): 636-640.

参考文献 32

1	LIU G W, LIN Q P, JIN S, et al. The CRISPR-Cas toolbox and gene editing technologies[J]. Mol Cell, 2022, 82(2):333-347. DOI: 10.1016/j.molcel.2021.12.002 .
2	KNOTT G J, DOUDNA J A. CRISPR-Cas guides the future of genetic engineering[J]. Science, 2018, 361(6405):866-869. DOI: 10.1126/science.aat5011 .
3	洪胜辉. 利用卵巢移植技术挽救濒危转基因小鼠方法的建立[J]. 畜牧兽医科技信息, 2017(8): 21-22. DOI: 10.3969/J.ISSN.1671-6027.2017.08.012 .
	HONG S H. Establishment of a method to save endangered transgenic mice by ovarian transplantation[J]. Chin J Animal Husb Vet Med, 2017(8): 21-22. DOI: 10.3969/J.ISSN.1671-6027.2017.08.012 .
4	TAFT R. In vitro fertilization in mice[J]. Cold Spring Harb Protoc, 2017, 2017(11): pdb.prot094508. DOI: 10.1101/pdb.prot094508 .
5	GRIESINGER G, LARSSON P. Conventional outcome reporting per IVF cycle/embryo transfer may systematically underestimate chances of success for women undergoing ART: relevant biases in registries, epidemiological studies, and guidelines[J]. Hum Reprod Open, 2023, 2023(2): hoad018. DOI: 10.1093/hropen/hoad018 .
6	O'NEILL C L, CHOW S, ROSENWAKS Z, et al. Development of ICSI[J]. Reproduction, 2018, 156(1): F51-F58. DOI: 10.1530/REP-18-0011 .
7	HADDAD M, STEWART J, XIE P, et al. Thoughts on the popularity of ICSI[J]. J Assist Reprod Genet, 2021, 38(1):101-123. DOI: 10.1007/s10815-020-01987-0 .
8	MIZUNO K, HOSHI K, HUANG T. Fertilization and embryo development in a mouse ICSI model using human and mouse sperm after immobilization in polyvinylpyrrolidone[J]. Hum Reprod, 2002, 17(9):2350-2355. DOI: 10.1093/humrep/17.9.2350 .
9	CHINÈ A, RESCHINI M, FORNELLI G, et al. Low ovarian reserve and risk of miscarriage in pregnancies derived from assisted reproductive technology[J]. Hum Reprod Open, 2023, 2023(3): hoad026. DOI: 10.1093/hropen/hoad026 .
10	HIRAOKA K, ISUGE M, KAMADA Y, et al. Piezo-ICSI for human oocytes[J]. J Vis Exp, 2021(170): e60224. DOI: 10.3791/60224 .
11	CADDY M, POPKISS S, WESTON G, et al. PIEZO-ICSI increases fertilization rates compared with conventional ICSI in patients with poor prognosis[J]. J Assist Reprod Genet, 2023, 40(2):389-398. DOI: 10.1007/s10815-022-02701-y .
12	SALGADO R M, BROM-DE-LUNA J G, RESENDE H L, et al. Lower blastocyst quality after conventional vs. Piezo ICSI in the horse reflects delayed sperm component remodeling and oocyte activation[J]. J Assist Reprod Genet, 2018, 35(5):825-840. DOI: 10.1007/s10815-018-1174-9 .
13	BEHRINGER R. Mouse ovary transplantation[J]. Cold Spring Harb Protoc, 2017, 2017(3): pdb.prot094458. DOI: 10.1101/pdb.prot094458 .
14	左琴, 王劲松, 范涛, 等. 遗传工程小鼠冻存卵巢异体原位移植比较研究[J]. 实验动物科学, 2020, 37(3): 65-68, 72. DOI: 10.3969/j.issn.1006-6179.2020.03.011 .
	ZUO Q, WANG J S, FAN T, et al. Study for cryopreservation and orthotopic transplantation of genetically engineering mice ovary[J]. Lab Anim Sci, 2020, 37(3): 2020, 37(3): 65-68, 72. DOI: 10.3969/j.issn.1006-6179.2020.03.011 .
15	DAY J R, DAVID A, BARBOSA M G M, et al. Encapsulation of ovarian allograft precludes immune rejection and promotes restoration of endocrine function in immune-competent ovariectomized mice[J]. Sci Rep, 2019, 9(1):16614. DOI: 10.1038/s41598-019-53075-8 .
16	YANG J X, MEDVEDEV S, YU J Y, et al. Deletion of the DNA/RNA-binding protein MSY2 leads to post-meiotic arrest[J]. Mol Cell Endocrinol, 2006, 250(1-2):20-24. DOI: 10.1016/j.mce.2005.12.019 .
17	HUANG C Z, ZHAO S M, YANG Y J, et al. TP63 gain-of-function mutations cause premature ovarian insufficiency by inducing oocyte apoptosis[J]. J Clin Invest, 2023, 133(5): e162315. DOI: 10.1172/JCI162315 .
18	OYAMA Y, MIYATA H, SHIMADA K, et al. TULP2 deletion mice exhibit abnormal outer dense fiber structure and male infertility[J]. Reprod Med Biol, 2022, 21(1): e12467. DOI: 10.1002/rmb2.12467 .
19	LU S, GU Y Y, WU Y F, et al. Bi-allelic variants in human WDR63 cause male infertility via abnormal inner dynein arms assembly[J]. Cell Discov, 2021, 7(1):110. DOI: 10.1038/s41421-021-00327-5 .
20	LI M W, KINCHEN K L, VALLELUNGA J M, et al. Safety, efficacy and efficiency of laser-assisted IVF in subfertile mutant mouse strains[J]. Reproduction, 2013, 145(3):245-254. DOI: 10.1530/REP-12-0477 .
21	FANG Q Y, JIANG X H, BAI S, et al. Safety of early cumulus cell removal combined with early rescue ICSI in the prevention of fertilization failure[J]. Reprod Biomed Online, 2023: S1472-6483(23)00216. DOI: 10.1016/j.rbmo.2023.04.005 .
22	洪胜辉, 卫振, 刘平, 等. 高校外来基因工程小鼠质量控制技术分析与探讨[J]. 实验技术与管理, 2022, 39(7): 224-227, 237. DOI: 10.16791/j.cnki.sjg.2022.07.039 .
	HONG S H, WEI Z, LIU P, et al. Analysis and discussion on quality control technology of foreign genetically engineered mice in universities[J]. Exp Technol Manag, 2022, 39(7): 224-227, 237. DOI: 10.16791/j.cnki.sjg.2022.07.039 .
23	朱莲, 孔彭成, 王美珊, 等. 剖宫产与体外授精技术在小鼠生物净化中的应用[J]. 实验动物与比较医学, 2013, 33(3): 218-221. DOI: 10.3969/j.issn.1674-5817.2013.03.011 .
	ZHU L, KONG P C, WANG M S, et al. Application of Caesarean and in vitro fertilization for bio-cleansing in mice[J]. Lab Anim Comp Med, 2013, 33(3): 218-221. DOI: 10.3969/j.issn.1674-5817.2013.03.011 .
24	张曼, 彭丽娜, 高则发, 等. 基因工程小鼠剖宫产生物净化技术的研究[J]. 实验动物科学, 2021, 38(6): 2021, 38(6): 76-79, 84. DOI: 10.3969/j.issn.1006-6179.2021.06.015 .
	ZHANG M, PENG L N, GAO Z F, et al. Study on purification technology of genetically engineered mice cesarean section[J]. Lab Anim Sci, 2021, 38(6): 2021, 38(6): 76-79, 84. DOI: 10.3969/j.issn.1006-6179.2021.06.015 .
25	KIM H, BANG J, BAEK S H, et al. Eliminating murine norovirus, Helicobacter hepaticus, and intestinal protozoa by embryo transfer for an entire mouse barrier facility[J]. Exp Anim, 2022, 71(1):28-35. DOI: 10.1538/expanim.21-0026 .
26	TEKAYEV M, VURUSKAN A K. Clinical values and advances in round spermatid injection (ROSI)[J]. Reprod Biol, 2021, 21(3):100530. DOI: 10.1016/j.repbio.2021.100530 .
27	MIKI H, LEE J, INOUE K, et al. Microinsemination with first-wave round spermatids from immature male mice[J]. J Reprod Dev, 2004, 50(1):131-137. DOI: 10.1262/jrd.50.131 .
28	TAO Y. Oocyte activation during round spermatid injection: state of the art[J]. Reprod Biomed Online, 2022, 45(2):211-218. DOI: 10.1016/j.rbmo.2022.03.024 .
29	PAES D, SCHEPERS M, WILLEMS E, et al. Ablation of specific long PDE4D isoforms increases neurite elongation and conveys protection against amyloid-β pathology[J]. Cell Mol Life Sci, 2023, 80(7):178. DOI: 10.1007/s00018-023-04804-w .
30	OH J, PARK C, KIM S, et al. High levels of intracellular endotrophin in adipocytes mediate COPII vesicle supplies to autophagosome to impair autophagic flux and contribute to systemic insulin resistance in obesity[J]. Metabolism, 2023, 145:155629. DOI: 10.1016/j.metabol.2023.155629 .
31	VAN DER WEYDEN L, JONKERS J, ADAMS D J. The use of CRISPR/Cas9-based gene editing strategies to explore cancer gene function in mice[J]. Curr Opin Genet Dev, 2021, 66:57-62. DOI: 10.1016/j.gde.2020.12.005 .
32	KANEKO T, FUKUMOTO K, HARUGUCHI Y, et al. Fertilization of C57BL/6 mouse sperm collected from cauda epididymides after preservation or transportation at 4 degrees C using laser-microdissected oocytes[J]. Cryobiology, 2009, 59(1):59-62. DOI: 10.1016/j.cryobiol.2009.04.006 .

[1]	洪胜辉, 张旭亮, 王芊芊, 刘平, 刘迪文. 抑制素基因敲除小鼠模型的构建及表型初步分析[J]. 实验动物与比较医学, 2020, 40(4): 306-.
[2]	匡德宣, 王文广, 孙晓梅, 代解杰. 恶性疟原虫动物模型及基因编辑研究进展[J]. 实验动物与比较医学, 2019, 39(1): 65-71.
[3]	成大欣, 徐丽然, 于清清, 高守翠, 王晓靖, 刘懿, 刘恩岐, 赵四海. 一种简单的gRNA体外筛选方法[J]. 实验动物与比较医学, 2018, 38(2): 126-129.
[4]	王新蕾1，邵义祥1，王鹏2. 动物超数排卵技术研究进展[J]. , 2007, 27(4): 280-284.