犬诱导多能干细胞的研究进展与应用前景

doi:10.16431/j.cnki.1671-7236.2025.08.004

摘要/Abstract

摘要： 诱导多能干细胞(induced pluripotent stem cells，iPSCs)是通过将外源转录因子诱导进入体细胞内而表达内源性多能性网络的一种干细胞。iPSCs在形态、功能、分子层面的表达上与胚胎干细胞(embryonic stem cells，ESCs)高度相似，具有广泛的自我更新和分化能力；与ESCs相比，iPSCs具有相对易获得、免疫排斥风险低、伦理问题少等优势，可为疾病建模、药物筛选、再生医学等领域提供新方法，因此，iPSCs具有重要研究价值及广阔的市场价值。遵循“One Health”的倡议，犬诱导多能干细胞(canine induced pluripotent stem cells，ciPSCs)的出现不仅为犬临床疾病治疗开辟全新的干细胞疗法，同时犬作为人类疾病的最佳转化模型之一，也可促进人类再生医学的发展。值得注意的是，ciPSCs仍缺少规范且高效率的生产方案，ciPSCs转基因完全沉默失败或再激活的问题普遍存在，安全性、成本、可行性等因素制约着临床广泛应用ciPSCs。文章综述了ciPSCs重编程方案，探讨了ciPSCs构建所面临的挑战性和可能性，展望了ciPSCs在兽医学上的广阔应用前景，可为进一步完善ciPSCs的建立提供参考。

关键词: 犬诱导多能干细胞(ciPSCs); 重编程; 疾病治疗

Abstract: Induced pluripotent stem cells (iPSCs) are a type of stem cells generated by introducing exogenous transcription factors into somatic cells,thereby activating the endogenous pluripotency network.iPSCs exhibit high similarity to embryonic stem cells (ESCs) in terms of morphology,functionality,and molecular expression,demonstrating extensive self-renewal and differentiation capabilities.Compared with ESCs,iPSCs offer advantages such as relative ease of acquisition,lower risk of immune rejection,and fewer ethical concerns,providing new approaches for disease modeling,drug screening,regenerative medicine,and other fields.Therefore,iPSCs hold significant research value and broad market potential.In alignment with the "One Health" initiative,the emergence of canine induced pluripotent stem cells (ciPSCs) not only opens up novel stem cell-based therapies for canine clinical diseases but also promotes advancements in human regenerative medicine,given that dogs serve as one of the best translational models for human diseases.However,it is noteworthy that ciPSCs still lack standardized and highly efficient production protocols,and issues such as incomplete transgene silencing or reactivation are prevalent.Factors such as safety,cost,and feasibility continue to limit the widespread clinical application of ciPSCs.This review summarizes the reprogramming strategies for ciPSCs,discusses the challenges and possibilities in their generation,and explores the promising applications of ciPSCs in veterinary medicine,aiming to provide insights for further refining the establishment of ciPSCs.

Key words: canine induced pluripotent stem cells(ciPSCs); reprogramming; diseases treatment

中图分类号:

S852.21

高佳璇, 高晨, 李雨珏, 钟友刚. 犬诱导多能干细胞的研究进展与应用前景[J]. 中国畜牧兽医, 2025, 52(8): 3540-3550.

GAO Jiaxuan, GAO Chen, LI Yujue, ZHONG Yougang. Research Progress and Application Prospect of Canine Induced Pluripotent Stem Cells[J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(8): 3540-3550.

参考文献

[1] TAKAHASHI K,YAMANAKA S.Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors[J].Cell,2006,126(4):663-676.
[2] CONRAD J V,MEYER S,RAMESH P S,et al.Efficient derivation of transgene-free porcine induced pluripotent stem cells enables in vitro modeling of species-specific developmental timing[J].Stem Cell Reports,2023,18(12):2328-2343.
[3] TSUKAMOTO M,KIMURA K,YOSHIDA T,et al.Generation of canine induced pluripotent stem cells under feeder-free conditions using Sendai virus vector encoding six canine reprogramming factors[J].Stem Cell Reports,2024,19(1):141-157.
[4] SU Y,WANG L,FAN Z,et al.Establishment of bovine-induced pluripotent stem cells[J].International Journal of Molecular Sciences,2021,22(19):10489.
[5] LIU M,ZHAO L,WANG Z,et al.Generation of sheep induced pluripotent stem cells with defined dox-inducible transcription factors via piggybac transposition[J].Frontiers in Cell and Developmental Biology,2021,9:785055.
[6] FALK J,DONADEU F X.Equine induced pluripotent stem cell culture[J].Methods in Molecular Biology,2024,2749:175-184.
[7] LIU Y,ZHANG S,ZOU G,et al.Generation and characterization of giant panda induced pluripotent stem cells[J].Science Advances,2024,10(38):eadn7724.
[8] KATAYAMA M,FUKUDA T,KANEKO T,et al.Induced pluripotent stem cells of endangered avian species[J].Communications Biology,2022,5(1):1049.
[9] DÉJOSEZ M,MARIN A,HUGHES G M,et al.Bat pluripotent stem cells reveal unusual entanglement between host and viruses[J].Cell,2023,186(5):957-974.e28.
[10] KANEGI R,HATOYA S,KIMURA K,et al.Generation,characterization,and differentiation of induced pluripotent stem-like cells in the domestic cat[J].Journal of Reproduction and Development,2023,69(6):317-327.
[11] ARZI B,WEBB T L,KOCH T G,et al.Cell therapy in veterinary medicine as a proof-of-concept for human therapies:Perspectives from the North American veterinary regenerative medicine association[J].Frontiers in Veterinary Science,2021,8:779109.
[12] PARKER H G,SHEARIN A L,OSTRANDER E A.Man’s best friend becomes biology’s best in show:Genome analyses in the domestic dog[J].Annual Review of Genetics,2010,44:309-336.
[13] SABA E,SANDHU M A,PELAGALLI A.Canine mesenchymal stromal cell exosomes:State-of-the-art characterization,functional analysis and applications in various diseases[J].Veterinary Sciences 2024,11(5):187.
[14] JEUNG S,AN J H,KIM S S,et al.Safety and efficacy of canine gonadal tissue-derived mesenchymal stem cells for early myxomatous mitral valve disease[J].Frontiers in Veterinary Science,2024,11:1404607.
[15] EZASHI T,YUAN Y,ROBERTS R M.Pluripotent stem cells from domesticated mammals[J].Annual Review of Animal Biosciences,2016,4:223-253.
[16] SHIMADA H,NAKADA A,HASHIMOTO Y,et al.Generation of canine induced pluripotent stem cells by retroviral transduction and chemical inhibitors[J].Molecular Reproduction and Development,2010,77(1):2.
[17] YOSHIMATSU S,MURAKAMI R,NAKAJIMA M,et al.Establishment of an induced pluripotent stem cell line from a female domestic ferret (Mustela putorius furo) with an X chromosome instability[J].Stem Cell Research,2021,53:102385.
[18] LUO J,SUHR S T,CHANG E A,et al.Generation of leukemia inhibitory factor and basic fibroblast growth factor-dependent induced pluripotent stem cells from canine adult somatic cells[J].Stem Cells and Development,2011,20(10):1669-1678.
[19] BAIRD A,BARSBY T,GUEST D J.Derivation of canine induced pluripotent stem cells[J].Reproduction in Domestic Animals,2015,50(4):669-676.
[20] KIMURA K,TSUKAMOTO M,TANAKA M,et al.Efficient reprogramming of canine peripheral blood mononuclear cells into induced pluripotent stem cells[J].Stem Cells and Development,2021,30(2):79-90.
[21] BARRACHINA L,ARSHAGHI T E,O’BRIEN A,et al.Induced pluripotent stem cells in companion animals:How can we move the field forward?[J].Frontiers in Veterinary Science,2023,10:1176772.
[22] YAGI M,HORNG J E,HOCHEDLINGER K.Manipulating cell fate through reprogramming:Approaches and applications[J].Development,2024,151(19):dev203090.
[23] SPINELLI V,GUILLOT P V,DE COPPI P.Induced pluripotent stem (IPS) cells from human fetal stem cells (HFSCS)[J].Organogenesis,2013,9(2):101-110.
[24] STRÄSSLER E T,AALTO-SETÄLÄ K,KIAMEHR M,et al.Age is relative-impact of donor age on induced pluripotent stem cell-derived cell functionality[J].Frontiers in Cardiovascular Medicine,2018,5:4.
[25] HORTON C,DAVIES T J,LAHIRI P,et al.Induced pluripotent stem cells reprogrammed from primary dendritic cells provide an abundant source of immunostimulatory dendritic cells for use in immunotherapy[J].Stem Cells,2020,38(1):67-79.
[26] QUATTROCELLI M,SWINNEN M,GIACOMAZZI G,et al.Mesodermal ipsc-derived progenitor cells functionally regenerate cardiac and skeletal muscle[J].Journal of Clinical Investigation,2015,125(12):4463-4482.
[27] NISHIMURA T,HATOYA S,KANEGI R,et al.Feeder-independent canine induced pluripotent stem cells maintained under serum-free conditions[J].Molecular Reproduction and Development,2017,84(4):329-339.
[28] KIMURA K,NAGAKURA H,TSUKAMOTO M,et al.Canine induced pluripotent stem cells can be successfully maintained in weekend-free culture systems[J].Journal of Veterinary Medical Science,2024,86(3):247-257.
[29] TSUKAMOTO M,NISHIMURA T,YODOE K,et al.Generation of footprint-free canine induced pluripotent stem cells using auto-erasable Sendai virus vector[J].Stem Cells and Development,2018,27(22):1577-1586.
[30] ERVIN E H,FRENCH R,CHANG C H,et al.Inside the stemness engine:Mechanistic links between deregulated transcription factors and stemness in cancer[J].Seminars in Cancer Biology,2022,87:48-83.
[31] MADDEN S K,DE ARAUJO A D,GERHARDT M,et al.Taking the MYC out of cancer:Toward therapeutic strategies to directly inhibit c-MYC[J].Molecular Cancer,2021,20(1):3.
[32] KUNITOMI A,HIROHATA R,ARREOLA V,et al.Improved Sendai viral system for reprogramming to naive pluripotency[J].Cell Reports Methods,2022,2(11):100317.
[33] YOSHIMATSU S,NAKAJIMA M,IGUCHI A,et al.Non-viral induction of transgene-free IPSCS from somatic fibroblasts of multiple mammalian species[J].Stem Cell Reports,2021,16(4):754-770.
[34] RONIMA K R,DEY C,THUMMER R P.An insight into the role of Glis1 in embryonic development,IPSC generation,and cancer[J].Advances in Experimental Medicine and Biology,2024,1470:97-113.
[35] LIU Z,CAI Y,WANG Y,et al.Cloning of macaque monkeys by somatic cell nuclear transfer[J].Cell,2018,172(4):881-887.e7.
[36] WHITWORTH D J,OVCHINNIKOV D A,WOLVETANG E J.Generation and characterization of LIF-dependent canine induced pluripotent stem cells from adult dermal fibroblasts[J].Stem Cells and Development,2012,21(12):2288-2297.
[37] GHAEDI M,NIKLASON L E.Human pluripotent stem cells (IPSC) generation,culture,and differentiation to lung progenitor cells[J].Methods in Molecular Biology,2019,1576:55-92.
[38] MISHRA S,KACIN E,STAMATIADIS P,et al.The role of the reprogramming method and pluripotency state in gamete differentiation from patient-specific human pluripotent stem cells[J].Molecular Human Reproduction,2018,24(4):173-184.
[39] YE H,WANG Q.Efficient generation of non-integration and feeder-free induced pluripotent stem cells from human peripheral blood cells by Sendai virus[J].Cellular Physiology and Biochemistry,2018,50(4):1318-1331.
[40] CHOW L,JOHNSON V,REGAN D,et al.Safety and immune regulatory properties of canine induced pluripotent stem cell-derived mesenchymal stem cells[J].Stem Cell Research,2017,25:221-232.
[41] TSUKAMOTO M,KIMURA K,TANAKA M,et al.Generation of footprint-free canine induced pluripotent stem cells from peripheral blood mononuclear cells using Sendai virus vector[J].Molecular Reproduction and Development,2020,87(6):663-665.
[42] KIM M,HWANG S U,YOON J D,et al.Optimized approaches for the induction of putative canine induced pluripotent stem cells from old fibroblasts using synthetic RNAs[J].Animals (Basel),2020,10(10):1848.
[43] MOSHREF M,QUESTA M,LOPEZ-CERVANTES V,et al.Panobinostat effectively increases histone acetylation and alters chromatin accessibility landscape in canine embryonic fibroblasts but does not enhance cellular reprogramming[J].Frontiers in Veterinary Science,2021,8:716570.
[44] LUO J,CIBELLI J B.Conserved role of bfgf and a divergent role of LIF for pluripotency maintenance and survival in canine pluripotent stem cells[J].Stem Cells and Development,2016,25(21):1670-1680.
[45] MENON D V,BHASKAR S,SHESHADRI P,et al.Positioning canine induced pluripotent stem cells (IPSCS) in the reprogramming landscape of naïve or primed state in comparison to mouse and human ipscs[J].Life Sciences,2021,264:118701.
[46] SMITH L A,HIDALGO AGUILAR A,OWENS D D G,et al.Using advanced cell culture techniques to differentiate pluripotent stem cells and recreate tissue structures representative of teratoma xenografts[J].Frontiers in Cell and Developmental Biology,2021,9:667246.
[47] GONÇALVES N J N,BRESSAN F F,ROBALLO K C S,et al.Generation of LIF-independent induced pluripotent stem cells from canine fetal fibroblasts[J].Theriogenology,2017,92:75-82.
[48] MENZOROV A G.Pluripotent stem cells of order carnivora:Technical perspective[J].International Journal of Molecular Sciences,2023,24(4):3905.
[49] HOU P,LI Y,ZHANG X,et al.Pluripotent stem cells induced from mouse somatic cells by small-molecule compounds[J].Science,2013,341(6146):651-654.
[50] GUAN J,WANG G,WANG J,et al.Chemical reprogramming of human somatic cells to pluripotent stem cells[J].Nature,2022,605(7909):325-331.
[51] LIUYANG S,WANG G,WANG Y,et al.Highly efficient and rapid generation of human pluripotent stem cells by chemical reprogramming[J].Cell Stem Cell,2023,30(4):450-459.e9.
[52] KIMURA K,TSUKAMOTO M,YOSHIDA T,et al.Canine induced pluripotent stem cell maintenance under feeder-free and chemically-defined conditions[J].Molecular Reproduction and Development,2021,88(6):395-404.
[53] CHANDRASEKARAN A,THOMSEN B B,AGERHOLM J S,et al.Neural derivates of canine induced pluripotent stem cells-like cells from a mild cognitive impairment dog[J].Frontiers in Veterinary Science,2021,8:725386.
[54] GAAR-HUMPHREYS K R,SPANJERSBERG T C F,SANTARELLI G,et al.Genetic basis of dilated cardiomyopathy in dogs and its potential as a bidirectional model[J].Animals (Basel),2022,12(13):1679.
[55] NISKANEN J E,OHLSSON Å,LJUNGVALL I,et al.Identification of novel genetic risk factors of dilated cardiomyopathy:From canine to human[J].Genome Medicine,2023,15(1):73.
[56] CHOI H,KIM H J,YANG J,et al.Acetylation changes tau interactome to degrade tau in Alzheimer’s disease animal and organoid models[J].Aging Cell,2020,19(1):e13081.
[57] YIN X,LI Q,SHU Y,et al.Exploiting urine-derived induced pluripotent stem cells for advancing precision medicine in cell therapy,disease modeling,and drug testing[J].Journal of Biomedical Science,2024,31(1):47.
[58] MONDAL T,DAS K,SINGH P,et al.Thin films of functionalized carbon nanotubes support long-term maintenance and cardio-neuronal differentiation of canine induced pluripotent stem cells[J].Nanomedicine:Nanotechnology,Biology,and Medicine,2022,40:102487.
[59] CHOW L,MCGRATH S,DE ARRUDA SALDANHA C,et al.Generation of neural progenitor cells from canine induced pluripotent stem cells and preliminary safety test in dogs with spontaneous spinal cord injuries[J].Frontiers in Veterinary Science,2020,7:575938.
[60] NATARAJAN M,SINGH P,MONDAL T,et al.In vitro propagation and cardiac differentiation of canine induced pluripotent stem cells on carbon nanotube substrates[J].Tissue Cell,2021,71:101571.
[61] LEE A S,XU D,PLEWS J R,et al.Preclinical derivation and imaging of autologously transplanted canine induced pluripotent stem cells[J].Journal of Biological Chemistry,2011,286(37):32697-32704.
[62] NISHIMURA T,HATOYA S,KANEGI R,et al.Generation of functional platelets from canine induced pluripotent stem cells[J].Stem Cells and Development,2013,22(14):2026-2035.
[63] WHITWORTH D J,FRITH J E,FRITH T J R,et al.Derivation of mesenchymal stromal cells from canine induced pluripotent stem cells by inhibition of the TGFβ/activin signaling pathway[J].Stem Cells and Development,2014,23(24):3021-3033.
[64] WANG R,ZHU T,HOU B,et al.An IPSC-derived exosome-pulsed dendritic cell vaccine boosts antitumor immunity in melanoma[J].Molecular Therapy,2023,31(8):2376-2390.
[65] ARYA S B,COLLIE S P,PARENT C A.The ins-and-outs of exosome biogenesis,secretion,and internalization[J].Trends in Cell Biology,2024,34(2):90-108.
[66] LEE S,HAN J,YANG J,et al.Exosomes from human IPSC-derived retinal organoids enhance corneal epithelial wound healing[J].International Journal of Molecular Sciences,2024,25(16):8925.
[67] WANG S,HOU Y,LI X,et al.Comparison of exosomes derived from induced pluripotent stem cells and mesenchymal stem cells as therapeutic nanoparticles for treatment of corneal epithelial defects[J].Aging,2020,12(19):19546-19562.
[68] LEVY D,ABADCHI S N,SHABABI N,et al.Induced pluripotent stem cell-derived extracellular vesicles promote wound repair in a diabetic mouse model via an anti-inflammatory immunomodulatory mechanism[J].Advanced Healthcare Materials,2023,12(26):e2300879.
[69] DANE D M,CAO K,ZHANG Y A,et al.Inhalational delivery of induced pluripotent stem cell secretome improves postpneumonectomy lung structure and function[J].Journal of Applied Physiology,2020,129(5):1051-1061.