Stemolecule™ Y27632

Y27632 is a cell-permeable small molecule Rho-associated kinase (ROCK) inhibitor¹. Y27632 has been found to prevent apoptosis as well as enhance the survival and cloning efficiency of dissociated human embryonic stem (ES) cells without affecting their self-renewal properties or pluripotency². This molecule has also been shown to enhance survival during the transplantation of ES cell-derived neural precursors³. Y27632 in combination with Pifithrin-u significantly improves cell recovery after cryopreservation⁴.

Stemolecule Y27632 is also available as a 10 mM DMSO Stock Solution (Cat. No. 04-0012-02).

Y27632 is widely used in naive stem cell protocols⁵, such as the 5i/L/A/ protocol.

Stemgent conveniently provides all the components comprising the 5i/L/A media supplement necessary to support naive pluripotent stem cell applications.

5i (inhibitors):

• Stemolecule PD0325901 (04-0006),
• Stemolecule Y27632 (04-0012),
• Stemolecule WH-4-023 (04-0079),
• Stemolecule SB590885 (04-0080),
• Stemolecule IM-12 (04-0081) or alternatively, Stemolecule CHIR99021 (04-0004)

L: Stemfactor LIF, Human Recombinant (03-0016), and

A: Stemfactor Activin A, Human Recombinant (03-0001)

Specifications

Product Name: Stemolecule Y27632

Catalog Number: 04-0012

Size:

• 2 mg (Cat. No. 04-0012)
• 10 mg (Cat. No. 04-0012-10)
• 2 mg in 10 mM DMSO (Cat. No. 04-0012-02)

Alternate Name(s):

• Y-27632
• (1R,4R)-4-((R)-1-aminoethyl)-N-(pyridine-4-yl)cyclohexanecarboxamide dihydrochloride

Chemical Formula: C₁₄H₂₁N₃O·2HCl

Molecular Weight: 320.3

CAS Number: 146986-50-7

Purity: Greater than 97 % by HPLC analysis

Formulation: White solid

Solubility: Reconstitute in DMSO to the desired concentration. For reconstitution instructions please reference product specifications sheet.

Storage and Stability: Store powder at 4 °C protected from light. Following reconstitution, store aliquots at −20 °C. Stock solutions are stable for 6 months when stored as directed.

Quality Control: The purity of Y27632 was determined by HPLC analysis. The accurate mass was determined by mass spectrometry. Cellular toxicity of Y27632 was tested on mouse embryonic stem cells.

References

1. Ishizaki, T., Uehata, M., Tamechika, I., Keel, J., Nonomura, K., Maekawa, M., and Narumiya, S. Pharmacological properties of Y-27632, a specific inhibitor of rho-associated kinases. Mol Pharmacol 57: 976-983 (2000).
2. Watanabe, K., Ueno, M., Kamiya, D., Nishiyama, A., Matsumura, M., Wataya, T., Takahashi, J.B., Nishikawa, S., Nishikawa, S., Muguruma, K., and Sasai, Y. A ROCK inhibitor permits survival of dissociated human embryonic stem cells. Nat Biotechnol 25: 681-686 (2007).
3. Koyanagi, M., Takahashi, J., Arakawa, Y., Doi, D., Fukuda, H., Hayashi, H., Narumiya, S., and Hashimoto, N. Inhibition of the Rho/ROCK pathway reduces apoptosis during transplantation of embryonic stem cell-derived neural precursors. J Neurosci Res 86: 270-280 (2008).
4. Xu, X., Cowley, S., Flaim, C.J., James, W., Seymour, L.W., Cui, Z. Enhancement of cell recovery for dissociated human embryonic stem cells after cryopreservation. Biotechnol Prog 26: 781-788 (2010).
5. Fonseca, SAS; Costas, RM; Pereira, LV. Searching for naive human pluripotent stem cells. World Journal of Stem Cells, 7(3), 649 (2015).

Additional Publications

• Michielin F; Giobbe GG; Luni C; Hu Q; Maroni I; Ofrord MR; Manfredi A; Di Filipp9; David AL; Cacchiarelli D; De Coppi P Eaton S; Elvassore N. The Microfluidic Environment Reveals a Hidden Role of Self-Organizing Extracellular Matrix in Hepatic Commitment and Organoid Formation of hiPSCs. Cell Rep 33:108453 (2020).
• Ryan SK; Jordan-Sciutto KL; Anderson SA. Protocol for Tri-culture of hiPSC-Derived Neurons, Astrocytes, and Microglia. STAR Protocols 1:100190 (2020).
• Akiyama H; Jalloh S; Park S; Lei M; Mostoslavski G; Gummuluru S. Expression of HIV-1 intron-containing RNA in Microglia Induces Inflammatory Responses. J Virol in press:doi:10.1128/JVI.01386-20 (2020).
• Barbar L; Rusielewica T; Zimmer M; Kalpana K; Fossati V. Isolation of Human CD49f+ Astrocytes and In Vitro iPSC-Based Neurotoxicity Assays. STAR Protocols :doi.org/10.1016/j.xpro.2020.100172 (2020).
• Scoger E; Argyiou L; Cyganke L; Zelarayán. Generation of homozygous CRISPRa human induced pluripotent stem cell (hiPSC) lines for sustained endogenous gene activation. Stem Cell Res :101944 (2020).
• Wong C-K; Luk HK-H; Lai W-H; Lau Y-M; Zhang RR; Wong AC-P; Lo GC-S; Chan K-H; Hung IF-N; Tse H-F; Woo PC-Y; Lau SK-P; Siu C-W. Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes Platform to Study SARS-CoV-2 Related Myocardial Injury. Circulation J :doi:10.1253/circj.CJ-20-0881 (2020).
• Ambrosini YM; Park Y; Jergens AE; Shin W; Min S; Atherly T; Borcherding DC; Jang J; Allenspach K; Mochel JP; Kim HJ. Recapitulation of the accessible interface of biopsy-derived canine intestinal organoids to study epithelial-luminal interactions. PLoS ONE 15:e231423 (2020).
• Chu J; Fitzgerald ML; Sehgal M; Manley W; Fitzgerald S; Naung H; Goldberg EM; Anderson SA. Enhanced maturation of human stem cell derived interneurons by mTOR activation . bioRxiv :dx.doi.org/10.1101/777714 (2019).
• Chen D; Sun N; Hou L; Kim R; Faith J; Aslanyan M; Tao Y; Zheng Y; Fu J; Liu W; Kellis M; Clark A. Human Primordial Germ Cells Are Specified from Lineage-Primed Progenitors. Cell Rep 29:4568-4582.e5 (2019).
• Koutmani Y; Gampierakis IA; Polissidis A; Ximerakis M: Koutsoudaki PN; Polyzos A; Agrogiannis G; Karaliota S; Thomaidou D; Rubin LL; Politis PK; Karakis KP. CRH Promotes the Neurogenic Activity of Neural Stem Cells in the Adult Hippocampus. Stem Cell Rep 28:932-945.e7 (2019).
• Nagaraja S; Quezada MA; Gillespie SM; Arzt M; Lennon JJ; Woo PJ; Hovestadt V; Kambhampati M; Filbin MG; Suva ML; Nazarian J; Monje M. Histone Variant and Cell Context Determine H3K27M Reprogramming of the Enhancer Landscape and Oncogenic State. Molecular Cell 76:dev.138982 (2019).
• Trautsch I; Heta E; Soong PL; Levent E; Nikolaev VO; Bogeski I ; Katschinski DM; Mayr M; Zimmermann W-H. Optogenetic Monitoring of the Glutathione Redox State in Engineered Human Myocardium. Front Physiol 10:272 (2019).
• LI S; Zhang H; Wang A; Liu Y; Yue F; Abulati X; Zhang C; Li L. Differentiation of adult human retinal pigment epithelial cells into dopaminergic-like cells in vitro and in the recipient monkey brain. Molecular Med 25:9 (2019).
• Talbot NC; Shannon AE; Garrett WM. Pancreatic duct-like cell line derived from pig embryonic stem cells: Expression of uroplakin genes in pig pancreatic tissue. In vitro Cell Develop Biol - Animal 2019:1 (2019).
• Wadhwani AR; Affaneh A; Van Gulden S; Kessler JA. Neuronal Apolipoprotein E4 increases cell death and p‐tau release in Alzheimer's disease. Ann Neurol doi: 10.1002/ana.25455.: (2019).
• Kim JJ; Savas JN; Miller MT; Hu X; Carromeu C; Lavallée-Adam M; Freitas BCG; Muotri AR; Yates III JR; Ghosh A. Proteomic analyses reveal misregulation of LIN28 expression and delayed timing of glial differentiation in human iPS cells with MECP2 loss-of-function. Plos One 14:e0212553 (2019).
• Dai S-B; Shen T; Zheng T-T; Pu J-L; Chen X-Z. Plasmid-based generation of neural cells from human fibroblasts using non-integrating episomal vectors. Neural Regen Res 14:510-505 (2019).
• Scavuzzo MA; Chmielowiec J; Yang D; Wamble K; Chaboub LS; Duraine L; Tepe B; Glasgow SM; Arenkiel BR; Brou C; Deneen B; Borowiak M; . Pancreatic Cell Fate Determination Relies on Notch Ligand Trafficking by NFIA. Cell Reports 25:3811-3827.e7 (2018).
• Chen D; Liu W: Zimmerman J; Pastor WA; Kim R;  Hosohama L; Ho J; Aslanyan M; Gell JJ; Jacobsen SE; Clark AT. The TFAP2C-Regulated OCT4 Naive Enhancer Is Involved in Human Germline Formation. Cell Reports 25:3591-3602.e5 (2018).
• Worley KE; Chin AS; Wan LQ. Lineage-Specific Chiral Biases of Human Embryonic Stem Cells during Differentiation. Stem Cells Intl 2018:1848605 (2018).
• Kilander MB; Wang C-H; Chang C-H; Nestor JE; Herold K; Tsai J-W; Nestor MW; Lin Y-C. A rare human CEP290 variant disrupts the molecular integrity of the primary cilium and impairs Sonic Hedgehog machinery. Sci Rep 8:17335 (2018).
• Javed AO; Li Y; Muffat J; Su K-C; Cohen MA; Lungjangwa T; Auborg P; Cheeseman IM; Jaenisch R. Microcephaly Modeling of Kinetochore Mutation Reveals a Brain-Specific Phenotype. Cell Rep 25:368-382.e5 (2018).
• Memon B; Karam M; Al-Khawaga S; Abdelalim EM. Enhanced differentiation of human pluripotent stem cells into pancreatic progenitors co-expressing PDX1 and NKX6.1. Stem Cell Res Therap 9:15 (2018).
• Yucer N; Holzapfel M; Jenkins Vogel T; Lenaeus L; Ornelas L; Laury A; Sareen D; Barrett R; Karlan BY; Svendsen CN. Directed Differentiation of Human Induced Pluripotent Stem Cells into Fallopian Tube Epithelium. Sci Rep 7:10741 (2017).
• Lin VJT; Zolekar A; Shi Y; Koneru B; Dimitrijevich S; Di Pasqua AJ; Wang C-Y. Potassium as a pluripotency-associated element identified through inorganic element profiling in human pluripotent stem cells. Sci Reports 7:5005 (2017).
• Donegan JJ; Tyson JA; Branch SY; Beckstead MJ; Anderson SA; Lodge DJ. Stem cell-derived interneuron transplants as a treatment for schizophrenia: preclinical validation in a rodent model. Mol Psychiatry 22:1942-1501 (2017).
• McCracken KW; Aihara E; Martin B; Crawford CM; Broda T; Treguier J; Zhang X; Shannon JM; Montrose MH; Wells JM. Wnt / β-catenin promotes gastric fundus specification in mice and humans. Nature 541:182-187 (2017).
• Noack C; Haupt LP; Zimmerman W-H; Streckfuss-Bomeke; Zelarayan LC. Generation of a KLF15 homozygous knockout human embryonic stem cell line using paired CRISPR/Cas9n, and human cardiomyocytes derivation. Stem Cell Research 23:127-131 (2017).
• Badenes SM; Fernandes TG; Cordeiro CSM; Boucher S; Kuninger D; Vemuri MC; Diogo MM; Cabral JMS. Defined Essential 8 medium and vitronectin efficiently support xeno-free expansion of human induced pluripotent stem cells in stirred microcarrier culture systems. PLoS ONE 11:e0155296 (2016).
• Koehler KR; Ine J; Longworth-Mills E; Liu X-P; Lee J; Holt JR; Hashino E. Generation of inner ear organoids with functional hair cells from pluripotent stem cells. Nat Biotechnol 35:583-589 (2017).
• Du F; Chen C-H; Li Y; Hu Y; An L-Y; Yang L; Zhang J; Chen EY; Xu Jie. Derivation of rabbit embryonic stem cells from vitrified-thawed embryos. Cellular Reprog 17:453 (2015).
• DeRosa BA; Belle KC; Thomas BJ; Cukier HN; Pericak-Vance MA; Vance JM; Dykxhoom DM. hVGAT-mCherry: A novel molecular tool for analysis of GABAergic neurons derived from human pluripotent stem cells. Mol Cell Neurosci 68:244 (2015).
• Zimmerlan L; Park TS; Huo JS; Verma K; Pather SR; Talbot CC; Agarwal J; Steppan D; Zhang YW; Considine M; Guo H; Zhong X; Gutierrez C; Cope L; Canto-Soler MV; Friedman AD; Baylin SB; Zambdis ET. Tankyrase inhibition promotes a stable human naive pluripotent state with improved functionality. Development 2016:dev.138982 (2016).
• Tan H-K; Toh C-XDl Ma D; Yang B; Liu TM; Lu J; Wong C-W; Tan T-K; Li H; SYn C; Tan E-L; Lim B; Lim Y-P; Cook SA; Loh Y-H. Human finger-prick induced pluripotent stem cells facilitate the development of stem cell banking. Stem Cells Trans Medicine 3:586 (2014).
• Prè D; Nestor MW; Sproul AA; Jacob S; Koppensteiner P; Chinchalongpom V; Zimmer M; Yamamoto A; Noggle SA; Arancio O. A time course analysis of the electrophysiological properties of neurons differentiated from human induced pluripotent stem cells (iPSCs). PLoS ONE 9(7): e103418 (2014).
• Borenstrom C; Simonsson S; Enochson L; BIgdeli N; Branstsing C; Ellerstrom C; Hyllner J; Lindahl A. Footprint-Free Human Induced Pluripotent Stem Cells From Articular Cartilage With Redifferentiation Capacity: A First Step Toward a Clinical-Grade Cell Source. Stem Cells Translat Med 3:433-447 (2014).
• Youn H; Kim S-H; Choi KA; Kim S. Characterization of Oct4-GFP spermatogonial stem cell line and its application in the reprogramming studies. J Cell Biochem 114:920 (2013).
• Lian, X., Hsiao, C., Wilson, G, Zhu, K., Hazeltine, L.B., Azarin, S.M., Raval, K.K., Zhang, J., Kamp, T.J., and Palecek, S.P. (2012) Robust cardiomyocyte differentiation from human pluripotent stem cells via temporal modulation of canonical Wnt signaling. Proc Natl Acad Sci USA 109: E1848 (2012).