孤雌胚胎干细胞更易向神经元样细胞分化

doi:10.3969/j.issn.1673-5374.2013.04.001

中国神经再生研究(英文版) ›› 2013, Vol. 8 ›› Issue (4): 293-300.doi: 10.3969/j.issn.1673-5374.2013.04.001

• 原著:神经损伤修复保护与再生 • 下一篇

孤雌胚胎干细胞更易向神经元样细胞分化

收稿日期:2012-10-13 修回日期:2012-12-10 出版日期:2013-02-05 发布日期:2013-02-05

Differentiation of neuron-like cells from mouse parthenogenetic embryonic stem cells

Xingrong Yan¹, Yanhong Yang², Wei Liu¹, Wenxin Geng¹, Huichong Du¹, Jihong Cui¹, Xin Xie¹, Jinlian Hua³, Shumin Yu⁴, Liwen Li¹, Fulin Chen¹

1 College of Life Sciences, Northwest University, Xi’an 710069, Shaanxi Province, China
2 Department of Obstetrics & Gynecology, Tangdu Hospital, the Fourth Military Medical University of Chinese PLA, Xi’an 710038, Shaanxi Province, China

3 College of Veterinary Medicine, Northwest A&F University, Yangling 712100, Shaanxi Province, China
4 College of Veterinary Medicine, Sichuan Agricultural University, Yaan 625001, Sichuan Province, China

Received:2012-10-13 Revised:2012-12-10 Online:2013-02-05 Published:2013-02-05
Contact: Xin Xie, Ph.D., Lecturer, College of Life Sciences, Northwest University, Xi’an 710069, Shaanxi Province, China, xiexhd@126.com. Fulin Chen, Ph.D., Professor, College of Life Sciences, Northwest University, Xi’an 710069, Shaanxi Province, China, tengda111111@ 163.com
About author:Xingrong Yan☆, Ph.D., Master’s supervisor. Xingrong Yan and Yanhong Yang contribute equally to this work
Supported by:
This work was supported by the National Natural Science Foundation of China, No. 30900155 and 81070496; the Research Foundation of Education Bureau of Shaanxi Province, China, No. 09JK785; Foundation of Interdisciplinary for Postgraduates from Northwest University, No. 08YJC22 and the Key Laboratory Funding of Northwestern University, Shaanxi Province in China

摘要/Abstract

摘要：

孤雌胚胎干细胞具有与受精胚来源的胚胎干细胞相似的分化潜能，故实验对来源于C3H小鼠的孤雌胚胎干细胞和受精胚来源的胚胎干细胞向神经细胞分化的潜力进行了比较。在诱导分化前分析细胞核型，发现孤雌胚胎干细胞和受精胚来源的胚胎干细胞的核型保持正常，性染色体为XX。实时定量PCR及免疫细胞化学染色检测发现细胞高表达多能性蛋白Oct4 mRNA及蛋白，说明细胞具有多向分化潜能。应用维甲酸诱导2种干细胞形成拟胚体，再将拟胚体离散为单细胞，应用N2培养基诱导分化9 d。免疫荧光细胞化学染色显示，经诱导的孤雌胚胎干细胞和受精胚来源的胚胎干细胞均表达神经细胞标志物Nestin、β-Ⅲ微管蛋白及髓鞘碱性蛋白；实时定量PCR结果显示，经诱导的孤雌胚胎干细胞和受精胚来源的胚胎干细胞表达神经发生相关基因Sox-1, Nestin, GABA, Pax6, Zic5, Pitx1，而多能性基因Oct4的表达显著降低，且孤雌胚胎干细胞中Nestin和Pax6基因的表达明显高于受精胚来源的胚胎干细胞。可见在基因水平上，孤雌胚胎干细胞比受精胚来源的胚胎干细胞具有更强的向神经细胞分化的潜力。

关键词: 神经再生, 干细胞, 孤雌发生, 孤雌胚胎干细胞, 胚胎干细胞, 神经细胞, 核型, Oct4, 分化, 拟胚体, 小鼠, 基金资助文章, 图片文章

Abstract:

Parthenogenetic embryonic stem cells have pluripotent differentiation potentials, akin to fertilized embryo-derived embryonic stem cells. The aim of this study was to compare the neuronal differentiation potential of parthenogenetic and fertilized embryo-derived embryonic stem cells. Before differentiation, karyotype analysis was performed, with normal karyotypes detected in both parthenogenetic and fertilized embryo-derived embryonic stem cells. Sex chromosomes were identified as XX. Immunocytochemistry and quantitative real-time PCR detected high expression of the pluripotent gene, Oct4, at both the mRNA and protein levels, indicating pluripotent differentiation potential of the two embryonic stem cell subtypes. Embryonic stem cells were induced with retinoic acid to form embryoid bodies, and then dispersed into single cells. Single cells were differentiated in N2 differentiation medium for 9 days. Immunocytochemistry showed parthenogenetic and fertilized embryo-derived embryonic stem cells both express the neuronal cell markers nestin, βIII-tubulin and myelin basic protein. Quantitative real-time PCR found expression of neurogenesis related genes (Sox-1, Nestin, GABA, Pax6, Zic5 and Pitx1) in both types of embryonic stem cells, and Oct4 expression was significantly decreased. Nestin and Pax6 expression in parthenogenetic embryonic stem cells was significantly higher than that in fertilized embryo-derived embryonic stem cells. Thus, our experimental findings indicate that parthenogenetic embryonic stem cells have stronger neuronal differentiation potential than fertilized embryo-derived embryonic stem cells.

Key words: neural regeneration, stem cells, parthenogenesis, parthenogenetic embryonic stem cells, embryonic stem cells, neuronal cells, karyotypes, Oct4, differentiation, embryoid body, mice, grants-supported paper, photographs-containing paper, neuroregeneration

. 孤雌胚胎干细胞更易向神经元样细胞分化[J]. 中国神经再生研究(英文版), 2013, 8(4): 293-300.

Xingrong Yan, Yanhong Yang, Wei Liu, Wenxin Geng, Huichong Du, Jihong Cui, Xin Xie, Jinlian Hua, Shumin Yu, Liwen Li, Fulin Chen. Differentiation of neuron-like cells from mouse parthenogenetic embryonic stem cells[J]. Neural Regeneration Research, 2013, 8(4): 293-300.

参考文献

[1] Alvarez CV, Garcia-Lavandeira M, Garcia-Rendueles ME, et al. Defining stem cell types: understanding the therapeutic potential of ESCs, ASCs, and iPS cells. J Mol Endocrinol. 2012;49(2):R89-111.

[2] Sills ES, Takeuchi T, Tanaka N, et al. Identification and isolation of embryonic stem cells in reproductive endocrinology: theoretical protocols for conservation of human embryos derived from in vitro fertilization. Theor Biol Med Model. 2005;2(25):1-8.

[3] Lin G, OuYang Q, Zhou X, et al. A highly homozygous and parthenogenetic human embryonic stem cell line derived from a one-pronuclear oocyte following in vitro fertilization procedure. Cell Res. 2007;17(12):999-1007.

[4] Okamoto S, Takahashi M. Induction of Retinal Pigment Epithelial Cells from Monkey Invest. Invest Ophthalmol Vis Sci. 2011;52(12):8785-8790.

[5] Jiang J, Ding G, Lin J, et al. Different developmental potential of pluripotent stem cells generated by different reprogramming strategies. J Mol Cell Biol. 2011;3(3): 197-199.

[6] Yang X, Smith SL, Tian XC, et al. Nuclear reprogramming of cloned embryos and its implications for therapeutic cloning. Nat Genet. 2007;39(3):295-302.

[7] Rideout WM 3rd, Hochedlinger K, Kyba M, et al. Correction of a genetic defect by nuclear transplantation and combined cell and gene therapy. Cell. 2002;109(1): 17-27.

[8] Robertson JA. Embryo stem cell research: ten years of controversy. J Law Med Ethics. 2010;38(2):191-203.

[9] Takahashi K, Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell. 2006;126(4):663-676.

[10] Zou J, Mali P, Huang X, et al. Site-specific gene correction of a point mutation in human iPS cells derived from an adult patient with sickle cell disease. Blood. 2011;118(17): 4599-4608.

[11] Alipio Z, Liao W, Roemer EJ, et al. Reversal of hyperglycemia in diabetic mouse models using induced-pluripotent stem (iPS)-derived pancreatic β-like cells. Proc Natl Acad Sci U S A. 2010;107(30): 13426-13431.

[12] Mou CL, Chen LY. Battle for pluripotency: Derivation of induced pluripotent stem cells. Recent Pat Regen Med. 2011;1:123-130.

[13] Zeitouni S, Krause U, Clough BH, et al. Human mesenchymal stem cell-derived matrices for enhanced osteoregeneration. Sci Transl Med. 2012;4(132): 132-155.

[14] Van Keymeulen A, Blanpain C. Tracing epithelial stem cells during development, homeostasis, and repair. J Cell Biol. 2012;197(5):575-584.

[15] Rode I, Boehm T. Regenerative capacity of adult cortical thymic epithelial cells. Proc Natl Acad Sci U S A. 2012; 109(9):3463-3468.

[16] Kaufman MH, Robertson EJ, Handyside AH, et al. Establishment of pluripotential cell lines from haploid mouse embryos. J Embryol Exp Morphol. 1983;73: 249-261.

[17] Kim K, Lerou P, Yabuuchi A, et al. Histocompatible embryonic stem cells by parthenogenesis. Science. 2007;315(5811):482-486.

[18] Revazova ES, Turovets NA, Kochetkova OD, et al. Patient-specific stem cell lines derived from human parthenogenetic blastocysts. Cloning Stem Cells. 2007; 9:432-449.

[19] Bibel M, Richter J, Schrenk K, et al. Differentiation of mouse embryonic stem cells into a defined neuronal lineage. Nat Neurosci. 2004;7(9):1003-1009.

[20] Stavridis MP, Smith AG. Neural differentiation of mouse embryonic stem cells. Biochem Soc Trans. 2003;31(Pt 1): 45-49.

[21] Yu SM, Yan XR, Chen DM, et al. Isolation and characterization of parthenogenetic embryonic stem (pES) cells containing genetic background of the Kunming mouse strain. Asian-Aust J Anim Sci. 2011;24(1):37-44.

[22] Yan X, Yu S, Lei A, et al. The four reprogramming factors and embryonic development in mice. Cell Reprogram. 2010;12(5):565-570.

[23] Gong SP, Kim H, Lee EJ, et al. Change in gene expression of mouse embryonic stem cells derived from parthenogenetic activation. Hum Reprod. 2009;24(4): 805-814.

[24] Jäderstad J, Jäderstad LM, Li J, et al. Communication via gap junctions underlies early functional and beneficial interactions between grafted neural stem cells and the host. Proc Natl Acad Sci U S A. 2010;107(11):5184-5189.

[25] Cao Q, He Q, Wang Y, et al. Transplantation of ciliary neurotrophic factor-expressing adult oligodendrocyte precursor cells promotes remyelination and functional recovery after spinal cord injury. J Neurosci. 2010;30(8): 2989-3001.

[26] Zhou J, Tian GP, Wang J, et al. In vitro differentiation of adipose-derived stem cells and bone marrow-derived stromal stem cells into neuronal-like cells. Neural Regen Res. 2011;6(19):1467-1472.

[27] Pevny LH, Sockanathan S, Placzek M, et al. A role for SOX1 in neural determination. Development. 1998;125: 1967-1978.

[28] Park D, Xiang AP, Mao FF, et al. Nestin is required for the proper self-renewal of neural stem cells. Stem Cells. 2101;28(12):2162-2171.

[29] Jia H, Tao H, Feng R, et al. Pax6 regulates the epidermal growth factor-responsive neural stem cells of the subventricular zone. Neuroreport. 2011;22(9):448-452.

[30] Corti S, Nizzardo M, Nardini M, et al. Embryonic stem cell-derived neural stem cells improve spinal muscular atrophy phenotype in mice. Brain. 2010;133(Pt 2): 465-481.

[31] Elkouris M, Balaskas N, Poulou M, et al. Sox1 maintains the undifferentiated state of cortical neural progenitor cells via the suppression of Prox1-mediated cell cycle exit and neurogenesis. Stem Cells. 2011;29(1):89-98.

[32] Jin Z, Liu L, Bian W, et al. Different transcription factors regulate nestin gene expression during P19 cell neural differentiation and central nervous system development. J Biol Chem. 2009;284(12):8160-8173.

[33] Suzuki S, Namiki J, Shibata S, et al. The neural stem/progenitor cell marker nestin is expressed in proliferative endothelial cells, but not in mature vasculature. J Histochem Cytochem. 2010;58(8):721-730.

[34] Zhang X, Huang CT, Chen J, et al. Pax6 is a human neuroectoderm cell fate determinant. Cell Stem Cell. 2010;7(1):90-100.

[35] Gao J, Wang J, Wang Y, et al. Regulation of Pax6 by CTCF during induction of mouse ES cell differentiation. PLoS One. 2011;6(6):e20954.

[36] Sansom SN, Griffiths DS, Faedo A, et al. The level of the transcription factor pax6 is essential for controlling the balance between neural stem cell self-renewal and neurogenesis. PLoS Genet. 2009;5(6):e1000511.

[37] Tan KR, Yvon C, Turiault M, et al. GABA neurons of the VTA drive conditioned place aversion. Neuron. 2012; 73(6):1173-1183.

[38] Inoue T, Hatayama M, Tohmonda T, et al. Mouse Zic5 deficiency results in neural tube defects and hypoplasia of cephalic neural crest derivatives. Dev Biol. 2004;270(1): 146-162.

[39] National Research Council (US) Committee for the Update of the Guide for the Care and Use of Laboratory Animals. Guide for the Care and Use of Laboratory Animals. 8th edition. Washington (DC): National Academies Press (US). 2011.

[40] Luo C, Zuñiga J, Edison E, et al. Superovulation strategies for 6 commonly used mouse strains. J Am Assoc Lab Anim Sci. 2011;50(4):471-478.

[41] González S, Ibáñez E, Santaló J. Establishment of mouse embryonic stem cells from isolated blastomeres and whole embryos using three derivation methods. J Assist Reprod Genet. 2010;27(12):671-682.

[42] Nagy A, Gertsenstein M, Vintersten K, et al. Karyotyping Mouse Cells. CSH Protoc. 2008;3(5):1-3.

[43] Cook MS, Munger SC, Nadeau JH, et al. Regulation of male germ cell cycle arrest and differentiation by DND1 is modulated by genetic background. Development. 2011; 138(1):23-32.

引言

材料方法

Design

A controlled observational study examining cell cytology.

Time and setting

Experiments were performed from March 2011 to June 2012 in the Shaanxi Provincial Key Laboratory of Biotechnology, Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, China.

Materials

Female (4–6 weeks old) and male (6–8 weeks old) Kunming mice were purchased from the Experimental Animal Center at the Fourth Military Medical University of Chinese PLA, China (license No. SCXK (Military) 2002-005). C3H female mice (4 weeks old) were purchased from Vital River Laboratory (license No. SCXK (Beijing) 2007-0001) in China. Animal treatments were performed in accordance with the Guidance Suggestions for the Care and Use of Laboratory Animals^[39].

Methods

Isolation of parthenogenetic and fertilized embryo-derived embryonic stem cells

Female Kunming mice were intraperitoneally injected with 10 IU of pregnant mare serum gonadotropin (Sigma, St. Louis, MO, USA), and at 48-hour intervals intraperitoneally injected with 10 IU of human chorionic gonadotropin (Sigma). Immediately following human chorionic gonadotropin injection, female mice were mated (1:1) to male mice of proven fertility and the same strain^[40]. Metaphase II oocytes were collected from oviducts of C3H female mice that were superovulated and artificially activated by chemical methods^[22]. Blastocysts were assembled by culturing parthenogenetic embryos in vitro. Fertilized blastocysts were derived from natural matings of C3H female mice at 3.5 days post-coitum. The zona pellucida of parthenogenetic and fertilized blastocysts were removed using acid Tyrode’s solution (Sigma), and the zona-free blastocyst was cultured on a feeder layer of mouse embryo fibroblasts treated with mitomycin C (Sigma) in gelatin-coated 4-well multi-dishes. Cells were cultured in knock-out Dulbecco’s minimal essential medium (Gibco, Carlsbad, CA, USA), supplemented with 0.1 mM β-mercaptoethanol, 1% (v/v) non-essential amino acids, 2 mM L-glutamine, 1% (v/v) penicillin and streptomycin, and 2 000 U/mL mouse leukemia inhibitory factor (Chemicon, Temecula, CA, USA). After 3 days culture, colonies derived from the inner cell mass were dissociated with accutase (Gibco) and subcultured on gelatin-coated 4-well multi-dishes in the presence of mouse embryo fibroblast feeder cells. When cultured embryonic stem cells had obvious colonies, subcultures were conducted at 3–4 day intervals with the same protocols, as previously reported^[41]. Both embryonic stem cell types were demonstrated to be pluripotent by embryoid body formation, differentiation of three germ layers and expression of pluripotent genes (data not shown).

Karyotype analysis of parthenogenetic and fertilized embryo-derived embryonic stem cells

Karyotype analysis of parthenogenetic and fertilized embryo-derived embryonic stem cells was performed as described previously^[42]. Briefly, there were at least three passages in feeder-free conditions, aimed to remove cells derived from mouse embryo fibroblasts. Parthenogenetic and fertilized embryo-derived embryonic stem cells were incubated in colchicine (10 μg/mL; Sigma) for 2 hours. Subsequently, cells were dissembled into single-cell suspensions, followed by incubation for 15 minutes in 0.075 mM KCl. Cells were fixed in a methanol/acetic acid mixture at room temperature, at least 3 times (the last time, temperature of fix solution is at –20°C), then spread over slides and stained with Giemsa solution (Sigma). Karyotypes were observed and imaged using a Digital Microscope (Leica, Heidelberg, Germany).

Embryoid body formation and differentiation into neuronal lineages

For embryoid body formation, 3 × 10⁶ parthenogenetic and fertilized embryo-derived embryonic stem cells were plated on non-adherent bacterial dishes in embryoid body medium, without leukemia inhibitory factor. The medium was changed every 2 days, and 5 µM retinoic acid (Sigma) was added after 4 days, with continued culture for 8 days. Embryoid bodies were dissociated using accutase and cells (1 × 10⁴/mL) plated on polyornithine-coated plates in N2 medium (Invitrogen, Carlsbad, CA, USA). The medium was changed every 2 days, as in previous reports^[20].

Immunocytochemistry

For immunofluorescence, staining was carried out as reported previously^[43]. Briefly, single cells were plated on coverslips and neuronal differentiation was continued for 9 days. Cells were fixed in 4% paraformaldehyde for 3 hours, rinsed with PBS at least three times, blocked in 10% goat serum with 0.2% Triton X-100 for 1 hour, and then washed, at least three times, with PBS and Triton X-100 (0.2%). Cells were incubated at 4°C overnight with primary monoclonal rabbit anti-mouse antibodies (all 1:100; Chemicon, Santa Cruz, CA, USA): βIII-tubulin, nestin and myelin basic protein. After at least five PBS washes, FITC-conjugated secondary antibody (goat anti-rabbit IgG; 1:150; Chemicon) was incubated at 37°C for 1 hour. 4’,6-diamidino-2-phenylindole (1 μg/mL; Chemicon) was used to stain the cell nucleus.

For immunocytochemistry, fertilized embryo-derived and parthenogenetic embryonic stem cells were cultured on coverslips with a feeder cell layer. Immunocytochemistry was performed according to the manufacturer’s instructions, SP-9000 General Immunocytochemical Kit (Chemicon). Briefly, cells were blocked with 10% goat serum with 0.2% Triton X-100, then incubated at 4°C overnight with primary rabbit anti-mouse Oct4 monoclonal antibody (1:100; Chemicon). Finally, cells were exposed to horseradish peroxidase-conjugated secondary antibodies (goat anti-rabbit IgG) with positive 3,3-diaminobenzidine expression detected as red- or yellow-brown^[21].

Quantitative real-time PCR detection

Total RNA was extracted from fertilized embryo-derived and parthenogenetic embryonic stem cells, and differentiating neuronal cells as described previously^[22]. Reverse transcription was performed using a reverse transcription kit (Takara, Dalian, China) according to the manufacturer’s instructions. For mRNA expression, PCR amplification of genes was performed using EXTaq polymerase (Takara), with a program of 94°C for 5 minutes, 35 cycles of 94°C for 30 seconds, 52–59°C for 30 seconds, 72°C for 30 seconds, and final extension at 72°C for 10 minutes. Real-time reverse-transcription PCR was performed on a Bio-Rad CFX96 Sequence Detection System (Bio-Rad, Hercules, CA, USA) using the SYBR Green PCR Mix (Takara). PCR reactions contained 10 μL SYBR Green PCR Master Mix, 1 μL forward and reverse primers (10 mM), 8.5 μL water and 0.5 μL cDNA template, in a total 20 μL volume. Sox-1, nestin, GABA, Pax6, Zic5, Pitx1 and Oct4 were detected by quantitative real-time PCR. Primer sequences are shown in Table 1. Results were normalized to β-actin levels. Experiments were repeated three times.

讨论

文章快阅

延伸阅读

1. 内容介绍:

孤雌胚胎干细胞是未来具有重大应用潜力的细胞原因有以下几点：1，与胚胎干细胞一样孤雌胚胎干细胞具有分化的多能性，在体外能长期传代，并维持其多能性；2，孤雌胚胎干细胞来源与未受精的卵母细胞，分离过程中没有破坏能发育为个体的胚胎，因此没有伦理问题的限制。在孤雌胚胎干细胞应用以前还需大量的基础理论研究；3，孤雌胚胎干细胞来源于自体，可避免免疫排斥的限制。本研究来源的ESCs和pESCs均来源于C3H的小鼠，同时性染色体均为XX，因此具有较强的可比性。细胞特性方面进行了检测，核型分析发现80%以上的细胞核型保持正常。多能性基因之一，Oct4在两同品系来源的ESCs和pESCs之间的表达水平没有显著差异，在向神经细胞分化的第9 d表达显著降低。神经发生相关的基因和蛋白在分化第9 d已经表达，如神经早期发生关键蛋白Nestin，神经细胞特异性蛋白β-Ⅲ Tubilin和MBP。神经发生有关的基因也进行了检测，这些基因包括Sox-1, Nestin, Gaba, Pax6, Zic, pitx1，发现Nestin和pax6在pESCs来源的分化细胞中显著升高，说明在基因水平上pESCs具有向神经细胞发育的潜力。

2. 实验设计及文章构思：

（1）分离获得相同品系来源的ESCs和pESCs系。

（2）以其中一个基因为例说明两者多能性基因表达没有差异，同时在分化后同时显著降低，分化后两种细胞之间oct4的表达没有显著差异。

（3）检测细胞核型，为后续研究的细胞提供保障。

（4）神经相关蛋白（早期和后期）的表达，在细胞水平上观察表达程度和变化。

（5）神经发生相关基因的表达，检测这些基因的表达变化，确定细胞向神经分化的潜力。

3. 文章学术水平与国内外同类研究的比较：

在pubmed中输入“parthenogenetic embryonic stem cell”和“neuron”共出现12篇文献，这些研究报道了孤雌胚胎干细胞向神经细胞的分化研究并取得许多重要的结果。本研究与以前报道不同的是，ESCs和pESC来源于相同的小鼠品系，比较两种细胞在早期向神经谱系的分化潜力。结果发现pESCs具有向神经分化较强的潜力，为我们用这株细胞向神经细胞后续的比较研究提供基础。

孤雌胚胎干细胞更易向神经元样细胞分化

Differentiation of neuron-like cells from mouse parthenogenetic embryonic stem cells

PDF

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

引言

材料方法

讨论

文章快阅

延伸阅读

编辑推荐

Metrics

本文评价

[1]	. 牙髓干细胞移植对血管性痴呆模型大鼠的治疗作用[J]. 中国神经再生研究(英文版), 2021, 16(8): 1645-1651.
[2]	. 神经基质膜包裹周围神经损伤的安全和有效性：一项随机单盲多中心临床试验[J]. 中国神经再生研究(英文版), 2021, 16(8): 1652-1659.
[3]	. 硫化氢可增强帕金森病小鼠模型的成年神经发生[J]. 中国神经再生研究(英文版), 2021, 16(7): 1353-1358.
[4]	. 复合神经干细胞的胶原/硫酸肝素多孔支架可改善创伤性脑损伤大鼠的神经功能[J]. 中国神经再生研究(英文版), 2021, 16(6): 1068-1077.
[5]	. 牙髓干细胞移植治疗阿尔茨海默病的潜力[J]. 中国神经再生研究(英文版), 2021, 16(5): 893-898.
[6]	. 人源多能干细胞可诱导分化为Mu和Kappa阿片受体的神经元[J]. 中国神经再生研究(英文版), 2021, 16(4): 653-658.
[7]	. 下调窖蛋白1表达可促进人脂肪间充质干细胞向多巴胺能神经元样分化[J]. 中国神经再生研究(英文版), 2021, 16(4): 714-720.
[8]	. 硬膜外电刺激结合离体三基因疗法治疗脊髓损伤[J]. 中国神经再生研究(英文版), 2021, 16(3): 550-560.
[9]	. miR-132/甲基CpG结合蛋白2调节神经干细胞分化中的作用[J]. 中国神经再生研究(英文版), 2021, 16(2): 345-349.
[10]	. 抑制γ-氨基丁酸A型受体亚型GABAA-ρ诱导斑马鱼的视网膜再生[J]. 中国神经再生研究(英文版), 2021, 16(2): 367-374.
[11]	. 干细胞治疗糖尿病视网膜病变的研究热点：文献可视化分析[J]. 中国神经再生研究(英文版), 2021, 16(1): 172-178.
[12]	. 外生内皮细胞形成功能性脑屏障并恢复其完整性[J]. 中国神经再生研究(英文版), 2020, 15(6): 1071-1078.
[13]	. CXCR7中和抗体对脑缺血慢性期海马齿状回神经再生认知功能的影响[J]. 中国神经再生研究(英文版), 2020, 15(6): 1079-1085.
[14]	. 脊髓损伤模型小鼠MicroRNA-21-5p的比较蛋白质组学变化及在不同途径中的可能作用[J]. 中国神经再生研究(英文版), 2020, 15(6): 1102-1110.
[15]	. miR-124在调节视黄酸诱导N2a细胞分化中的作用[J]. 中国神经再生研究(英文版), 2020, 15(6): 1133-1139.