FREE SHIPPING over $40
OVER 10000 FIVE-STAR REVIEWS
5% OFF ALL SUBSCRIPTIONS
100% HAPPINESS GUARANTEE
Analyze Diet
0
Home / Equine Research Bank / Animals (Basel) / Edition of Prostaglandin E2 Receptors EP2 and EP4 by CRISPR/...
Animals : an open access journal from MDPI2020; 10(6); 1078; doi: 10.3390/ani10061078

Edition of Prostaglandin E2 Receptors EP2 and EP4 by CRISPR/Cas9 Technology in Equine Adipose Mesenchymal Stem Cells.

Abstract: In mesenchymal stem cells (MSCs), it has been reported that prostaglandin E2 (PGE2) stimulation of EP2 and EP4 receptors triggers processes such as migration, self-renewal, survival, and proliferation, and their activation is involved in homing. The aim of this work was to establish a genetically modified adipose (aMSC) model in which receptor genes EP2 and EP4 were edited separately using the CRISPR/Cas9 system. After edition, the genes were evaluated as to if the expression of MSC surface markers was affected, as well as the migration capacity in vitro of the generated cells. Adipose MSCs were obtained from Chilean breed horses and cultured in DMEM High Glucose with 10% fetal bovine serum (FBS). sgRNA were cloned into a linearized LentiCRISPRv2GFP vector and transfected into HEK293FT cells for producing viral particles that were used to transduce aMSCs. GFP-expressing cells were separated by sorting to obtain individual clones. Genomic DNA was amplified, and the site-directed mutation frequency was assessed by T7E1, followed by Sanger sequencing. We selected 11 clones of EP2 and 10 clones of EP4, and by Sanger sequencing we confirmed 1 clone knock-out to aMSC/EP2 and one heterozygous mutant clone of aMSC/EP4. Both edited cells had decreased expression of EP2 and EP4 receptors when compared to the wild type, and the edition of EP2 and EP4 did not affect the expression of MSC surface markers, showing the same pattern in filling the scratch. We can conclude that the edition of these receptors in aMSCs does not affect their surface marker phenotype and migration ability when compared to wild-type cells.
Get Full Text
Publication Date: 2020-06-23 PubMed ID: 32585798PubMed Central: PMC7341266DOI: 10.3390/ani10061078Google Scholar: Lookup
The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
LinkedInCopy
  • Journal Article

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research used CRISPR/Cas9 gene editing technology to modify Prostaglandin E2 (PGE2) receptors EP2 and EP4 on Mesenchymal stem cells (MSCs) from horse adipose tissue, and found that this modification did not affect the cells’ surface markers or ability to migrate.

Introduction and Objective

  • The research focuses on the modification of Prostaglandin E2 (PGE2) receptors EP2 and EP4 on Mesenchymal stem cells (MSCs), specifically those extracted from equine adipose (aMSCs).
  • PGE2 stimulation of EP2 and EP4 receptors has been known to trigger processes in MSCs such as migration, self-renewal, survival, and proliferation and their activation is involved in a process called homing.
  • The primary aim of the study was to create a genetically modified model of equine adipose by editing the receptor genes EP2 and EP4 separately using the CRISPR/Cas9 gene editing system.

Methodology

  • Adipose MSCs were harvested from horses of Chilean breed and cultured in a specific type of culture medium.
  • sgRNA, a component required for the CRISPR process, was cloned into a certain type of vector and subsequently, used to create viral particles through transfection into cell types known as HEK293FT.
  • The viral particles were then used to transduce the equine aMSCs, and the cells expressing Green Fluorescent Protein (GFP) were sorted out to establish individual clones.
  • Genomic DNA from these clones was amplified and assessed for mutation frequency through a process called T7E1 enzyme assay.
  • The mutations were then confirmed through a sequencing process known as Sanger sequencing.

Results and Conclusion

  • The results confirmed one clone knock-out for aMSC/EP2 and one heterozygous mutant clone for aMSC/EP4 through Sanger sequencing.
  • It was found that the edited cells demonstrated decreased expression of EP2 and EP4 receptors when compared to unedited cells (wild type).
  • Importantly, the modifications to the EP2 and EP4 receptors did not influence the expression of MSC surface markers nor did it affect the cells’ migration capacity.
  • The results of this study indeed suggest that modifying EP2 and EP4 receptors in equine aMSCs does not alter the cells’ phenotype or impede their migration ability when compared to unedited or wild-type cells.

Cite This Article

APA
Mançanares ACF, Cabezas J, Manríquez J, de Oliveira VC, Wong Alvaro YS, Rojas D, Navarrete Aguirre F, Rodriguez-Alvarez L, Castro FO. (2020). Edition of Prostaglandin E2 Receptors EP2 and EP4 by CRISPR/Cas9 Technology in Equine Adipose Mesenchymal Stem Cells. Animals (Basel), 10(6), 1078. https://doi.org/10.3390/ani10061078

Publication

Animals : an open access journal from MDPI
ISSN: 2076-2615
NlmUniqueID: 101635614
Country: Switzerland
Language: English
Volume: 10
Issue: 6
PII: 1078

Researcher Affiliations

Mançanares, Ana Carolina Furlanetto
  • Department of Animal Science, Faculty of Veterinary Science, Universidad de Concepción, Campus Chillan, Chillán 3780000, Chile.
Cabezas, Joel
  • Department of Animal Science, Faculty of Veterinary Science, Universidad de Concepción, Campus Chillan, Chillán 3780000, Chile.
Manríquez, José
  • Department of Animal Science, Faculty of Veterinary Science, Universidad de Concepción, Campus Chillan, Chillán 3780000, Chile.
de Oliveira, Vanessa Cristina
  • Department of Veterinary Medicine, Faculty of Animal Science and Food Engineering, University of São Paulo, Pirassununga, São Paulo 13630-000, Brazil.
Wong Alvaro, Yat Sen
  • Department of Animal Science, Faculty of Veterinary Science, Universidad de Concepción, Campus Chillan, Chillán 3780000, Chile.
Rojas, Daniela
  • Department of Animal Pathology, Faculty of Veterinary Sciences, Universidad de Concepción, Campus Chillan, Chillán 3780000, Chile.
Navarrete Aguirre, Felipe
  • Department of Animal Science, Faculty of Veterinary Science, Universidad de Concepción, Campus Chillan, Chillán 3780000, Chile.
Rodriguez-Alvarez, Lleretny
  • Department of Animal Science, Faculty of Veterinary Science, Universidad de Concepción, Campus Chillan, Chillán 3780000, Chile.
Castro, Fidel Ovidio
  • Department of Animal Science, Faculty of Veterinary Science, Universidad de Concepción, Campus Chillan, Chillán 3780000, Chile.

Grant Funding

  • Postdoctoral Fellowship 3170390 / Fondo Nacional de Desarrollo Cientu00edfico y Tecnolu00f3gico

Conflict of Interest Statement

None of the authors have any conflict of interest to declare.

References

This article includes 66 references
  1. Lee BC, Kim HS, Shin TH, Kang I, Lee JY, Kim JJ, Kang HK, Seo Y, Lee S, Yu KR, Choi SW, Kang KS. PGE2 maintains self-renewal of human adult stem cells via EP2-mediated autocrine signaling and its production is regulated by cell-to-cell contact.. Sci Rep 2016 May 27;6:26298.
    doi: 10.1038/srep26298pmc: PMC4882486pubmed: 27230257google scholar: lookup
  2. Sugimoto Y, Narumiya S. Prostaglandin E receptors.. J Biol Chem 2007 Apr 20;282(16):11613-7.
    doi: 10.1074/jbc.R600038200pubmed: 17329241google scholar: lookup
  3. Sheldon IM, Price SB, Cronin J, Gilbert RO, Gadsby JE. Mechanisms of infertility associated with clinical and subclinical endometritis in high producing dairy cattle.. Reprod Domest Anim 2009 Sep;44 Suppl 3:1-9.
    doi: 10.1111/j.1439-0531.2009.01465.xpubmed: 19660075google scholar: lookup
  4. Regan JW. EP2 and EP4 prostanoid receptor signaling.. Life Sci 2003 Dec 5;74(2-3):143-53.
    doi: 10.1016/j.lfs.2003.09.031pubmed: 14607241google scholar: lookup
  5. Smith WL, DeWitt DL, Garavito RM. Cyclooxygenases: structural, cellular, and molecular biology.. Annu Rev Biochem 2000;69:145-82.
    doi: 10.1146/annurev.biochem.69.1.145pubmed: 10966456google scholar: lookup
  6. Vukicevic S, Simic P, Borovecki F, Grgurevic L, Rogic D, Orlic I, Grasser WA, Thompson DD, Paralkar VM. Role of EP2 and EP4 receptor-selective agonists of prostaglandin E(2) in acute and chronic kidney failure.. Kidney Int 2006 Sep;70(6):1099-106.
    doi: 10.1038/sj.ki.5001715pubmed: 16871242google scholar: lookup
  7. Ushikubi F, Sugimoto Y, Ichikawa A, Narumiya S. Roles of prostanoids revealed from studies using mice lacking specific prostanoid receptors.. Jpn J Pharmacol 2000 Aug;83(4):279-85.
    doi: 10.1254/jjp.83.279pubmed: 11001172google scholar: lookup
  8. Wånggren K, Lalitkumar PG, Stavreus-Evers A, Ståbi B, Gemzell-Danielsson K. Prostaglandin E2 and F2alpha receptors in the human Fallopian tube before and after mifepristone treatment.. Mol Hum Reprod 2006 Sep;12(9):577-85.
    doi: 10.1093/molehr/gal058pubmed: 16820403google scholar: lookup
  9. Breyer RM, Bagdassarian CK, Myers SA, Breyer MD. Prostanoid receptors: subtypes and signaling.. Annu Rev Pharmacol Toxicol 2001;41:661-90.
    doi: 10.1146/annurev.pharmtox.41.1.661pubmed: 11264472google scholar: lookup
  10. Wang Y, Lai S, Tang J, Feng C, Liu F, Su C, Zou W, Chen H, Xu D. Prostaglandin E2 promotes human CD34+ cells homing through EP2 and EP4 in vitro.. Mol Med Rep 2017 Jul;16(1):639-646.
    doi: 10.3892/mmr.2017.6649pmc: PMC5482140pubmed: 28560401google scholar: lookup
  11. Samuelsson B, Morgenstern R, Jakobsson PJ. Membrane prostaglandin E synthase-1: a novel therapeutic target.. Pharmacol Rev 2007 Sep;59(3):207-24.
    doi: 10.1124/pr.59.3.1pubmed: 17878511google scholar: lookup
  12. Han J, Lu X, Zou L, Xu X, Qiu H. E-Prostanoid 2 Receptor Overexpression Promotes Mesenchymal Stem Cell Attenuated Lung Injury.. Hum Gene Ther 2016 Aug;27(8):621-30.
    doi: 10.1089/hum.2016.003pubmed: 27158855google scholar: lookup
  13. Lu X, Han J, Xu X, Xu J, Liu L, Huang Y, Yang Y, Qiu H. PGE2 Promotes the Migration of Mesenchymal Stem Cells through the Activation of FAK and ERK1/2 Pathway.. Stem Cells Int 2017;2017:8178643.
    doi: 10.1155/2017/8178643pmc: PMC5504996pubmed: 28740516google scholar: lookup
  14. Hoggatt J, Singh P, Sampath J, Pelus LM. Prostaglandin E2 enhances hematopoietic stem cell homing, survival, and proliferation.. Blood 2009 May 28;113(22):5444-55.
    doi: 10.1182/blood-2009-01-201335pmc: PMC2689046pubmed: 19324903google scholar: lookup
  15. Carrade DD, Lame MW, Kent MS, Clark KC, Walker NJ, Borjesson DL. Comparative Analysis of the Immunomodulatory Properties of Equine Adult-Derived Mesenchymal Stem Cells().. Cell Med 2012;4(1):1-11.
    doi: 10.3727/215517912X647217pmc: PMC3495591pubmed: 23152950google scholar: lookup
  16. Duffy MM, Pindjakova J, Hanley SA, McCarthy C, Weidhofer GA, Sweeney EM, English K, Shaw G, Murphy JM, Barry FP, Mahon BP, Belton O, Ceredig R, Griffin MD. Mesenchymal stem cell inhibition of T-helper 17 cell- differentiation is triggered by cell-cell contact and mediated by prostaglandin E2 via the EP4 receptor.. Eur J Immunol 2011 Oct;41(10):2840-51.
    doi: 10.1002/eji.201141499pubmed: 21710489google scholar: lookup
  17. Matysiak M, Orlowski W, Fortak-Michalska M, Jurewicz A, Selmaj K. Immunoregulatory function of bone marrow mesenchymal stem cells in EAE depends on their differentiation state and secretion of PGE2.. J Neuroimmunol 2011 Apr;233(1-2):106-11.
    doi: 10.1016/j.jneuroim.2010.12.004pubmed: 21354631google scholar: lookup
  18. Bolotin A, Quinquis B, Sorokin A, Ehrlich SD. Clustered regularly interspaced short palindrome repeats (CRISPRs) have spacers of extrachromosomal origin.. Microbiology (Reading) 2005 Aug;151(Pt 8):2551-2561.
    doi: 10.1099/mic.0.28048-0pubmed: 16079334google scholar: lookup
  19. Mojica FJ, Díez-Villaseñor C, García-Martínez J, Soria E. Intervening sequences of regularly spaced prokaryotic repeats derive from foreign genetic elements.. J Mol Evol 2005 Feb;60(2):174-82.
    doi: 10.1007/s00239-004-0046-3pubmed: 15791728google scholar: lookup
  20. Pourcel C, Salvignol G, Vergnaud G. CRISPR elements in Yersinia pestis acquire new repeats by preferential uptake of bacteriophage DNA, and provide additional tools for evolutionary studies.. Microbiology (Reading) 2005 Mar;151(Pt 3):653-663.
    doi: 10.1099/mic.0.27437-0pubmed: 15758212google scholar: lookup
  21. Niu J, Zhang B, Chen H. Applications of TALENs and CRISPR/Cas9 in human cells and their potentials for gene therapy.. Mol Biotechnol 2014 Aug;56(8):681-8.
    doi: 10.1007/s12033-014-9771-zpubmed: 24870618google scholar: lookup
  22. Shen Y, Zhang J, Yu T, Qi C. Generation of PTEN knockout bone marrow mesenchymal stem cell lines by CRISPR/Cas9-mediated genome editing.. Cytotechnology 2018 Apr;70(2):783-791.
    doi: 10.1007/s10616-017-0183-3pmc: PMC5851970pubmed: 29387984google scholar: lookup
  23. Bhat SA, Malik AA, Ahmad SM, Shah RA, Ganai NA, Shafi SS, Shabir N. Advances in genome editing for improved animal breeding: A review.. Vet World 2017 Nov;10(11):1361-1366.
    doi: 10.14202/vetworld.2017.1361-1366pmc: PMC5732344pubmed: 29263600google scholar: lookup
  24. Zarei A, Razban V, Hosseini SE, Tabei SMB. Creating cell and animal models of human disease by genome editing using CRISPR/Cas9.. J Gene Med 2019 Apr;21(4):e3082.
    doi: 10.1002/jgm.3082pubmed: 30786106google scholar: lookup
  25. Cabezas J, Rojas D, Navarrete F, Ortiz R, Rivera G, Saravia F, Rodriguez-Alvarez L, Castro FO. Equine mesenchymal stem cells derived from endometrial or adipose tissue share significant biological properties, but have distinctive pattern of surface markers and migration.. Theriogenology 2018 Jan 15;106:93-102.
    doi: 10.1016/j.theriogenology.2017.09.035pubmed: 29049924google scholar: lookup
  26. Castro FO, Torres A, Cabezas J, Rodríguez-Alvarez L. Combined use of platelet rich plasma and vitamin C positively affects differentiation in vitro to mesodermal lineage of adult adipose equine mesenchymal stem cells.. Res Vet Sci 2014 Feb;96(1):95-101.
    doi: 10.1016/j.rvsc.2013.12.005pubmed: 24377415google scholar: lookup
  27. Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F. Genome engineering using the CRISPR-Cas9 system.. Nat Protoc 2013 Nov;8(11):2281-2308.
    doi: 10.1038/nprot.2013.143pmc: PMC3969860pubmed: 24157548google scholar: lookup
  28. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method.. Methods 2001 Dec;25(4):402-8.
    doi: 10.1006/meth.2001.1262pubmed: 11846609google scholar: lookup
  29. Dominici M, Le Blanc K, Mueller I, Slaper-Cortenbach I, Marini F, Krause D, Deans R, Keating A, Prockop Dj, Horwitz E. Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement.. Cytotherapy 2006;8(4):315-7.
    doi: 10.1080/14653240600855905pubmed: 16923606google scholar: lookup
  30. Bundgaard L, Stensballe A, Elbæk KJ, Berg LC. Mapping of equine mesenchymal stromal cell surface proteomes for identification of specific markers using proteomics and gene expression analysis: an in vitro cross-sectional study.. Stem Cell Res Ther 2018 Oct 25;9(1):288.
    doi: 10.1186/s13287-018-1041-8pmc: PMC6202851pubmed: 30359315google scholar: lookup
  31. Connick P, Kolappan M, Crawley C, Webber DJ, Patani R, Michell AW, Du MQ, Luan SL, Altmann DR, Thompson AJ, Compston A, Scott MA, Miller DH, Chandran S. Autologous mesenchymal stem cells for the treatment of secondary progressive multiple sclerosis: an open-label phase 2a proof-of-concept study.. Lancet Neurol 2012 Feb;11(2):150-6.
    doi: 10.1016/S1474-4422(11)70305-2pmc: PMC3279697pubmed: 22236384google scholar: lookup
  32. Hmadcha A, Martin-Montalvo A, Gauthier BR, Soria B, Capilla-Gonzalez V. Therapeutic Potential of Mesenchymal Stem Cells for Cancer Therapy.. Front Bioeng Biotechnol 2020;8:43.
    doi: 10.3389/fbioe.2020.00043pmc: PMC7013101pubmed: 32117924google scholar: lookup
  33. Aggarwal S, Pittenger MF. Human mesenchymal stem cells modulate allogeneic immune cell responses.. Blood 2005 Feb 15;105(4):1815-22.
    doi: 10.1182/blood-2004-04-1559pubmed: 15494428google scholar: lookup
  34. Gao F, Chiu SM, Motan DA, Zhang Z, Chen L, Ji HL, Tse HF, Fu QL, Lian Q. Mesenchymal stem cells and immunomodulation: current status and future prospects.. Cell Death Dis 2016 Jan 21;7(1):e2062.
    doi: 10.1038/cddis.2015.327pmc: PMC4816164pubmed: 26794657google scholar: lookup
  35. Food U., Administration D.. Cellular products for joint surface repair. Cell. Tissue Gene Ther. Advis. Comm. 2005.
  36. Paris DB, Stout TA. Equine embryos and embryonic stem cells: defining reliable markers of pluripotency.. Theriogenology 2010 Sep 1;74(4):516-24.
    doi: 10.1016/j.theriogenology.2009.11.020pubmed: 20071015google scholar: lookup
  37. Mambelli LI, Winter GH, Kerkis A, Malschitzky E, Mattos RC, Kerkis I. A novel strategy of mesenchymal stem cells delivery in the uterus of mares with endometrosis.. Theriogenology 2013 Mar 15;79(5):744-50.
    doi: 10.1016/j.theriogenology.2012.11.030pubmed: 23270861google scholar: lookup
  38. Rink BE, Beyer T, French HM, Watson E, Aurich C, Donadeu FX. The Fate of Autologous Endometrial Mesenchymal Stromal Cells After Application in the Healthy Equine Uterus.. Stem Cells Dev 2018 Aug 1;27(15):1046-1052.
    doi: 10.1089/scd.2018.0056pmc: PMC6067096pubmed: 29790424google scholar: lookup
  39. Jang MW, Yun SP, Park JH, Ryu JM, Lee JH, Han HJ. Cooperation of Epac1/Rap1/Akt and PKA in prostaglandin E(2) -induced proliferation of human umbilical cord blood derived mesenchymal stem cells: involvement of c-Myc and VEGF expression.. J Cell Physiol 2012 Dec;227(12):3756-67.
    doi: 10.1002/jcp.24084pubmed: 22378492google scholar: lookup
  40. Kleiveland CR, Kassem M, Lea T. Human mesenchymal stem cell proliferation is regulated by PGE2 through differential activation of cAMP-dependent protein kinase isoforms.. Exp Cell Res 2008 May 1;314(8):1831-8.
    doi: 10.1016/j.yexcr.2008.02.004pubmed: 18395713google scholar: lookup
  41. Osma-Garcia IC, Punzón C, Fresno M, Díaz-Muñoz MD. Dose-dependent effects of prostaglandin E2 in macrophage adhesion and migration.. Eur J Immunol 2016 Mar;46(3):677-88.
    doi: 10.1002/eji.201545629pubmed: 26631603google scholar: lookup
  42. Concordet JP, Haeussler M. CRISPOR: intuitive guide selection for CRISPR/Cas9 genome editing experiments and screens.. Nucleic Acids Res 2018 Jul 2;46(W1):W242-W245.
    doi: 10.1093/nar/gky354pmc: PMC6030908pubmed: 29762716google scholar: lookup
  43. Petersen GF, Hilbert B, Trope G, Kalle W, Strappe P. Efficient transduction of equine adipose-derived mesenchymal stem cells by VSV-G pseudotyped lentiviral vectors.. Res Vet Sci 2014 Dec;97(3):616-22.
    doi: 10.1016/j.rvsc.2014.09.004pubmed: 25443656google scholar: lookup
  44. Kallifatidis G, Beckermann BM, Groth A, Schubert M, Apel A, Khamidjanov A, Ryschich E, Wenger T, Wagner W, Diehlmann A, Saffrich R, Krause U, Eckstein V, Mattern J, Chai M, Schütz G, Ho AD, Gebhard MM, Büchler MW, Friess H, Büchler P, Herr I. Improved lentiviral transduction of human mesenchymal stem cells for therapeutic intervention in pancreatic cancer.. Cancer Gene Ther 2008 Apr;15(4):231-40.
    doi: 10.1038/sj.cgt.7701097pubmed: 18202717google scholar: lookup
  45. Van Damme A, Thorrez L, Ma L, Vandenburgh H, Eyckmans J, Dell'Accio F, De Bari C, Luyten F, Lillicrap D, Collen D, VandenDriessche T, Chuah MK. Efficient lentiviral transduction and improved engraftment of human bone marrow mesenchymal cells.. Stem Cells 2006 Apr;24(4):896-907.
    doi: 10.1634/stemcells.2003-0106pubmed: 16339997google scholar: lookup
  46. Chen L, Qu J, Cheng T, Chen X, Xiang C. Menstrual blood-derived stem cells: toward therapeutic mechanisms, novel strategies, and future perspectives in the treatment of diseases.. Stem Cell Res Ther 2019 Dec 21;10(1):406.
    doi: 10.1186/s13287-019-1503-7pmc: PMC6925480pubmed: 31864423google scholar: lookup
  47. Goujon C, Jarrosson-Wuillème L, Bernaud J, Rigal D, Darlix JL, Cimarelli A. With a little help from a friend: increasing HIV transduction of monocyte-derived dendritic cells with virion-like particles of SIV(MAC).. Gene Ther 2006 Jun;13(12):991-4.
    doi: 10.1038/sj.gt.3302753pubmed: 16525481google scholar: lookup
  48. Lin P, Lin Y, Lennon DP, Correa D, Schluchter M, Caplan AI. Efficient lentiviral transduction of human mesenchymal stem cells that preserves proliferation and differentiation capabilities.. Stem Cells Transl Med 2012 Dec;1(12):886-97.
    doi: 10.5966/sctm.2012-0086pmc: PMC3659678pubmed: 23283550google scholar: lookup
  49. de Oliveira VC, Moreira GSA, Bressan FF, Gomes Mariano Junior C, Roballo KCS, Charpentier M, Concordet JP, Meirelles FV, Ambrósio CE. Edition of TFAM gene by CRISPR/Cas9 technology in bovine model.. PLoS One 2019;14(3):e0213376.
    doi: 10.1371/journal.pone.0213376pmc: PMC6405117pubmed: 30845180google scholar: lookup
  50. Sentmanat MF, Peters ST, Florian CP, Connelly JP, Pruett-Miller SM. A Survey of Validation Strategies for CRISPR-Cas9 Editing.. Sci Rep 2018 Jan 17;8(1):888.
    doi: 10.1038/s41598-018-19441-8pmc: PMC5772360pubmed: 29343825google scholar: lookup
  51. Jia C, Huai C, Ding J, Hu L, Su B, Chen H, Lu D. New applications of CRISPR/Cas9 system on mutant DNA detection.. Gene 2018 Jan 30;641:55-62.
    doi: 10.1016/j.gene.2017.10.023pubmed: 29031777google scholar: lookup
  52. Hsiau T., Conant D., Rossi N., Maures T., Waite K., Yang J., Joshi S., Kelso R., Holden K., Enzmann B.L.. Inference of CRISPR Edits from Sanger Trace Data. BioRxiv 2019.
    doi: 10.1101/251082google scholar: lookup
  53. Jin J, Xu Y, Huo L, Ma L, Scott AW, Pizzi MP, Li Y, Wang Y, Yao X, Song S, Ajani JA. An improved strategy for CRISPR/Cas9 gene knockout and subsequent wildtype and mutant gene rescue.. PLoS One 2020;15(2):e0228910.
    doi: 10.1371/journal.pone.0228910pmc: PMC7018052pubmed: 32053639google scholar: lookup
  54. Choudhary S, Alander C, Zhan P, Gao Q, Pilbeam C, Raisz L. Effect of deletion of the prostaglandin EP2 receptor on the anabolic response to prostaglandin E2 and a selective EP2 receptor agonist.. Prostaglandins Other Lipid Mediat 2008 Jun;86(1-4):35-40.
    doi: 10.1016/j.prostaglandins.2008.02.001pmc: PMC2755544pubmed: 18406186google scholar: lookup
  55. Ball BA, Scoggin KE, Troedsson MH, Squires EL. Characterization of prostaglandin E2 receptors (EP2, EP4) in the horse oviduct.. Anim Reprod Sci 2013 Nov 1;142(1-2):35-41.
    doi: 10.1016/j.anireprosci.2013.07.009pubmed: 24035156google scholar: lookup
  56. Delville M, Soheili T, Bellier F, Durand A, Denis A, Lagresle-Peyrou C, Cavazzana M, Andre-Schmutz I, Six E. A Nontoxic Transduction Enhancer Enables Highly Efficient Lentiviral Transduction of Primary Murine T Cells and Hematopoietic Stem Cells.. Mol Ther Methods Clin Dev 2018 Sep 21;10:341-347.
    doi: 10.1016/j.omtm.2018.08.002pmc: PMC6125771pubmed: 30191160google scholar: lookup
  57. Schomann T, Mezzanotte L, Lourens IM, de Groot JC, Frijns JH, Huisman MA. Lentiviral transduction and subsequent loading with nanoparticles do not affect cell viability and proliferation in hair-follicle-bulge-derived stem cells in vitro.. Contrast Media Mol Imaging 2016 Nov;11(6):550-560.
    doi: 10.1002/cmmi.1717pubmed: 27976505google scholar: lookup
  58. Hizaki H, Segi E, Sugimoto Y, Hirose M, Saji T, Ushikubi F, Matsuoka T, Noda Y, Tanaka T, Yoshida N, Narumiya S, Ichikawa A. Abortive expansion of the cumulus and impaired fertility in mice lacking the prostaglandin E receptor subtype EP(2).. Proc Natl Acad Sci U S A 1999 Aug 31;96(18):10501-6.
    doi: 10.1073/pnas.96.18.10501pmc: PMC17918pubmed: 10468638google scholar: lookup
  59. Tilley SL, Audoly LP, Hicks EH, Kim HS, Flannery PJ, Coffman TM, Koller BH. Reproductive failure and reduced blood pressure in mice lacking the EP2 prostaglandin E2 receptor.. J Clin Invest 1999 Jun;103(11):1539-45.
    doi: 10.1172/JCI6579pmc: PMC408376pubmed: 10359563google scholar: lookup
  60. Biswas S, Bhattacherjee P, Paterson CA, Tilley SL, Koller BH. Ocular inflammatory responses in the EP2 and EP4 receptor knockout mice.. Ocul Immunol Inflamm 2006 Jun;14(3):157-63.
    doi: 10.1080/09273940600665879pubmed: 16766399google scholar: lookup
  61. Yun SP, Ryu JM, Jang MW, Han HJ. Interaction of profilin-1 and F-actin via a β-arrestin-1/JNK signaling pathway involved in prostaglandin E(2)-induced human mesenchymal stem cells migration and proliferation.. J Cell Physiol 2011 Feb;226(2):559-71.
    doi: 10.1002/jcp.22366pubmed: 20717968google scholar: lookup
  62. Khan GA, Bhagat S, Alam MI. PGE(2) -induced migration of human brain endothelial cell is mediated though protein kinase A in cooperation of EP receptors.. J Leukoc Biol 2019 Apr;105(4):705-717.
    doi: 10.1002/JLB.2A0918-361Rpubmed: 30835912google scholar: lookup
  63. De Schauwer C, Meyer E, Van de Walle GR, Van Soom A. Markers of stemness in equine mesenchymal stem cells: a plea for uniformity.. Theriogenology 2011 May;75(8):1431-43.
    doi: 10.1016/j.theriogenology.2010.11.008pubmed: 21196039google scholar: lookup
  64. Barberini DJ, Freitas NP, Magnoni MS, Maia L, Listoni AJ, Heckler MC, Sudano MJ, Golim MA, da Cruz Landim-Alvarenga F, Amorim RM. Equine mesenchymal stem cells from bone marrow, adipose tissue and umbilical cord: immunophenotypic characterization and differentiation potential.. Stem Cell Res Ther 2014 Feb 21;5(1):25.
    doi: 10.1186/scrt414pmc: PMC4055040pubmed: 24559797google scholar: lookup
  65. De Schauwer C, Piepers S, Van de Walle GR, Demeyere K, Hoogewijs MK, Govaere JL, Braeckmans K, Van Soom A, Meyer E. In search for cross-reactivity to immunophenotype equine mesenchymal stromal cells by multicolor flow cytometry.. Cytometry A 2012 Apr;81(4):312-23.
    doi: 10.1002/cyto.a.22026pubmed: 22411893google scholar: lookup
  66. Radcliffe CH, Flaminio MJ, Fortier LA. Temporal analysis of equine bone marrow aspirate during establishment of putative mesenchymal progenitor cell populations.. Stem Cells Dev 2010 Feb;19(2):269-82.
    doi: 10.1089/scd.2009.0091pmc: PMC3138180pubmed: 19604071google scholar: lookup

Citations

This article has been cited 5 times.
  1. Hisey EA, Ross PJ, Meyers S. Genetic Manipulation of the Equine Oocyte and Embryo. J Equine Vet Sci 2021 Apr;99:103394.
    doi: 10.1016/j.jevs.2021.103394pubmed: 33781418google scholar: lookup
  2. Xiang Y, Dong J, Shao L, Chen S. Chimeric antigen receptor natural killer cell therapy for solid tumors: mechanisms, clinical progress, and strategies to overcome the tumor microenvironment. Exp Biol Med (Maywood) 2025;250:10841.
    doi: 10.3389/ebm.2025.10841pubmed: 41488873google scholar: lookup
  3. Han J, Ganguly R, Yi JY, Yun H, Jung SY, Sung C, Lee CS. Osmotically Tunable Microdroplets Enable Amplification-Free CRISPR Detection of Gene Doping. Adv Sci (Weinh) 2025 Dec;12(48):e15861.
    doi: 10.1002/advs.202515861pubmed: 41084992google scholar: lookup
  4. Han AR, Shin HR, Kwon J, Lee SB, Lee SE, Kim EY, Kweon J, Chang EJ, Kim Y, Kim SW. Highly efficient genome editing via CRISPR-Cas9 ribonucleoprotein (RNP) delivery in mesenchymal stem cells. BMB Rep 2024 Jan;57(1):60-65.
    doi: 10.5483/BMBRep.2023-0113pubmed: 38053293google scholar: lookup
  5. Jiang P, Iqbal A, Wang M, Li X, Fang X, Yu H, Zhao Z. Transcriptomic Analysis of Short/Branched-Chain Acyl-Coenzyme a Dehydrogenase Knocked Out bMECs Revealed Its Regulatory Effect on Lipid Metabolism. Front Vet Sci 2021;8:744287.
    doi: 10.3389/fvets.2021.744287pubmed: 34557544google scholar: lookup
Subscribe to our newsletter
Join 99,344 horse owners like you who receive our email updates on horse health and nutrition.