Comparison of four different transfection reagents for transfection efficiency of H9C2 cells_West China Medical Journal

Authors：

NI Yinyun ¹ , DING Yu ¹ ,  WU Sisi ^1,2

1. Core Facility, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P. R. China;
2. The State Key Laboratory of Biotherapy, Sichuan University, Chengdu, Sichuan 610041, P. R. China;

Corresponding?author：

WU Sisi, Email: wusisi1981@scu.edu.cn

Keywords：

Transfection; Rat heart myoblast cells H9C2; FuGENE HD; DNA-In CRISPR; Lipofectamine 3000; Lipofectamine 2000

DOI：

10.7507/1002-0179.201606200

Video：

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ObjectiveTo compare four different transfection reagents for transfection efficiency of rat heart myoblast cells H9C2, to choose the optimal transfection method.MethodsThe plasmids of enhanced green fluorescent protein (EGFP) gene were transfected as exogenous genes to H9C2 cells from four different transfection regents including FuGENE HD, DNA-In CRISPR, Lipofectamine 3000 and Lipofectamine 2000. Fluorescence intensity was measured by fluorescence microscopy and fluorescence microplate reader to evaluate transfection efficiency. The effects of four transfection reagents on cell viability were measured by Cell Counting Kit-8 (CCK-8) reagents.ResultsTransfection efficiency of Lipofectamine 3000 was the highest (>50%), while that of DNA-In CRISPR was the lowest (<1%). The cytotoxicity of Lipofectamine 3000 was the lowest in the four transfection reagents and the cell viability was 94.55% after 48-hour transfection.ConclusionTransfection regent Lipofectamine 3000 has the relatively high transfection efficiency as well as the lowest cytotoxicity, which is more suitable for use in H9C2 cells by transfection.

Citation： NI Yinyun, DING Yu, WU Sisi. Comparison of four different transfection reagents for transfection efficiency of H9C2 cells. West China Medical Journal, 2017, 32(7): 1024-1028. doi: 10.7507/1002-0179.201606200 Copy

1.	Pari GS, Keown WA. Experimental strategies in efficient transfection of mammalian cells. Calcium phosphate and DEAE-dextran. Methods Mol Biol, 1997, 62(1): 301-306.
2.	Bhattacharya S, Bajaj A. Advances in gene delivery through molecular design of cationic lipids. Chem Commun (Camb), 2009(31): 4632-4656.
3.	Jordan ET, Collins M, Terefe J, et al. Optimizing electroporation conditions in primary and other difficult-to-transfect cells[J]. J Biomol Tech, 2008, 19(5): 328-334.
4.	Yang SH, Agca Y, Cheng PH, et al. Enhanced transgenesis by intracytoplasmic injection of envelope-free lentivirus. Genesis, 2007, 45(4): 177-183.
5.	Kafri T, van Praag H, Gage FH, et al. Lentiviral vectors: regulated gene expression. Mol Ther, 2000, 1(6): 516-521.
6.	Zhang C, Yadava P, Hughes J. Polyethylenimine strategies for plasmid delivery to brain-derived cells. Methods, 2004, 33(2): 144-150.
7.	Karmali PP, Chaudhuri A. Cationic liposomes as non-viral carriers of gene medicines: resolved issues, open questions, and future promises. Med Res Rev, 2007, 27(5): 696-722.
8.	Kimes BW, Brandt BL. Properties of a clonal muscle cell line from rat heart. Exp Cell Res, 1976, 98(2): 367-381.
9.	Zordoky BN, El-Kadi AO. H9c2 cell line is a valuable in vitro model to study the drug metabolizing enzymes in the heart. J Pharmacol Toxicol Methods, 2007, 56(3): 317-322.
10.	Spallarossa P, Altieri P, Garibaldi S, et al. Matrix metalloproteinase-2 and-9 are induced differently by doxorubicin in H9c2 cells: the role of MAP kinases and NAD(P)H oxidase. Cardiovasc Res, 2006, 69(3): 736-745.
11.	Bernuzzi F, Recalcati S, Alberghini A, et al. Reactive oxygen species-independent apoptosis in doxorubicin-treated H9c2 cardiomyocytes: role for heme oxygenase-1 down-modulation. Chem Biol Interact, 2009, 177(1): 12-20.
12.	陳彥祥, 喬衛紅, 劉棟良, 等. 陽離子脂質體的轉染機制及轉染效率影響因素. 生物工程學報, 2007, 23(5): 776-780.
13.	楊云廷, 付崇羅. 陽離子脂質體 Lipofectamine 2000 對 PC12 細胞轉染效率的研究. 生物技術通報, 2011(5): 231-235.
14.	潘寧, 章藹然, 侯穎春. 脂質體介導法轉染腫瘤細胞效率的優化. 生物技術, 2008, 18(6): 47-50.
15.	Liu YC, Lin WY, Jhang YR, et al. Efficiency of DNA transfection of rat heart myoblast cells H9c2(2-1) by either polyethyleneimine or electroporation. Appl Biochem Biotechnol, 2011, 164(7): 1172-1182.
16.	Kiefer K, Clement J, Garidel P, et al. Transfection efficiency and cytotoxicity of nonviral gene transfer reagents in human smooth muscle and endothelial cells. Pharm Res, 2004, 21(6): 1009-1017.
17.	段艷, 王冰, 崔韶輝, 等. 陽離子脂質體轉染 Hela 細胞的實驗. 中國組織工程研究與臨床康復, 2009, 13(11): 2119-2122.
18.	Lappalainen K, J?skel?inen I, Syrj? nen K, et al. Comparison of cell proliferation and toxicity assays using two cationic liposomes. Pharm Res, 1994, 11(8): 1127-1131.
19.	Dalkara D, Chandrashekhar C, Zuber G. Intracellular protein delivery with a dimerizable amphiphile for improved complex stability and prolonged protein release in the cytoplasm of adherent cell lines. J Control Release, 2006, 116(3): 353-359.
20.	Nguyen LT, Atobe K, Barichello JM, et al. Complex formation with plasmid DNA increases the cytotoxicity of cationic liposomes. Biol Pharm Bull, 2007, 30(4): 751-757.
21.	Datiles MJ, Johnson EA, Mccarty RE. Inhibition of the ATPase activity of the catalytic portion of ATP synthases by cationic amphiphiles. Biochim Biophys Acta, 2008, 1777(4): 362-368.

1. Pari GS, Keown WA. Experimental strategies in efficient transfection of mammalian cells. Calcium phosphate and DEAE-dextran. Methods Mol Biol, 1997, 62(1): 301-306.
2. Bhattacharya S, Bajaj A. Advances in gene delivery through molecular design of cationic lipids. Chem Commun (Camb), 2009(31): 4632-4656.
3. Jordan ET, Collins M, Terefe J, et al. Optimizing electroporation conditions in primary and other difficult-to-transfect cells[J]. J Biomol Tech, 2008, 19(5): 328-334.
4. Yang SH, Agca Y, Cheng PH, et al. Enhanced transgenesis by intracytoplasmic injection of envelope-free lentivirus. Genesis, 2007, 45(4): 177-183.
5. Kafri T, van Praag H, Gage FH, et al. Lentiviral vectors: regulated gene expression. Mol Ther, 2000, 1(6): 516-521.
6. Zhang C, Yadava P, Hughes J. Polyethylenimine strategies for plasmid delivery to brain-derived cells. Methods, 2004, 33(2): 144-150.
7. Karmali PP, Chaudhuri A. Cationic liposomes as non-viral carriers of gene medicines: resolved issues, open questions, and future promises. Med Res Rev, 2007, 27(5): 696-722.
8. Kimes BW, Brandt BL. Properties of a clonal muscle cell line from rat heart. Exp Cell Res, 1976, 98(2): 367-381.
9. Zordoky BN, El-Kadi AO. H9c2 cell line is a valuable in vitro model to study the drug metabolizing enzymes in the heart. J Pharmacol Toxicol Methods, 2007, 56(3): 317-322.
10. Spallarossa P, Altieri P, Garibaldi S, et al. Matrix metalloproteinase-2 and-9 are induced differently by doxorubicin in H9c2 cells: the role of MAP kinases and NAD(P)H oxidase. Cardiovasc Res, 2006, 69(3): 736-745.
11. Bernuzzi F, Recalcati S, Alberghini A, et al. Reactive oxygen species-independent apoptosis in doxorubicin-treated H9c2 cardiomyocytes: role for heme oxygenase-1 down-modulation. Chem Biol Interact, 2009, 177(1): 12-20.
12. 陳彥祥, 喬衛紅, 劉棟良, 等. 陽離子脂質體的轉染機制及轉染效率影響因素. 生物工程學報, 2007, 23(5): 776-780.
13. 楊云廷, 付崇羅. 陽離子脂質體 Lipofectamine 2000 對 PC12 細胞轉染效率的研究. 生物技術通報, 2011(5): 231-235.
14. 潘寧, 章藹然, 侯穎春. 脂質體介導法轉染腫瘤細胞效率的優化. 生物技術, 2008, 18(6): 47-50.
15. Liu YC, Lin WY, Jhang YR, et al. Efficiency of DNA transfection of rat heart myoblast cells H9c2(2-1) by either polyethyleneimine or electroporation. Appl Biochem Biotechnol, 2011, 164(7): 1172-1182.
16. Kiefer K, Clement J, Garidel P, et al. Transfection efficiency and cytotoxicity of nonviral gene transfer reagents in human smooth muscle and endothelial cells. Pharm Res, 2004, 21(6): 1009-1017.
17. 段艷, 王冰, 崔韶輝, 等. 陽離子脂質體轉染 Hela 細胞的實驗. 中國組織工程研究與臨床康復, 2009, 13(11): 2119-2122.
18. Lappalainen K, J?skel?inen I, Syrj? nen K, et al. Comparison of cell proliferation and toxicity assays using two cationic liposomes. Pharm Res, 1994, 11(8): 1127-1131.
19. Dalkara D, Chandrashekhar C, Zuber G. Intracellular protein delivery with a dimerizable amphiphile for improved complex stability and prolonged protein release in the cytoplasm of adherent cell lines. J Control Release, 2006, 116(3): 353-359.
20. Nguyen LT, Atobe K, Barichello JM, et al. Complex formation with plasmid DNA increases the cytotoxicity of cationic liposomes. Biol Pharm Bull, 2007, 30(4): 751-757.
21. Datiles MJ, Johnson EA, Mccarty RE. Inhibition of the ATPase activity of the catalytic portion of ATP synthases by cationic amphiphiles. Biochim Biophys Acta, 2008, 1777(4): 362-368.

West China Medical Journal

Comparison of four different transfection reagents for transfection efficiency of H9C2 cells

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