Biomaterials Research (생체재료학회지)

Korean Society for Biomaterials (한국생체재료학회)
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Multiphoton imaging of myogenic differentiation in gelatin-based hydrogels as tissue engineering scaffolds

  • Kim, Min Jeong (Department of Cogno-Mechatronics Engineering, College of Nanoscience & Nanotechnology, Pusan National University) ;
  • Shin, Yong Cheol (Department of Cogno-Mechatronics Engineering, College of Nanoscience & Nanotechnology, Pusan National University) ;
  • Lee, Jong Ho (Department of Cogno-Mechatronics Engineering, College of Nanoscience & Nanotechnology, Pusan National University) ;
  • Jun, Seung Won (Department of Cogno-Mechatronics Engineering, College of Nanoscience & Nanotechnology, Pusan National University) ;
  • Kim, Chang-Seok (Department of Cogno-Mechatronics Engineering, College of Nanoscience & Nanotechnology, Pusan National University) ;
  • Lee, Yunki (Department of Molecular Science and Technology, Ajou University) ;
  • Park, Jong-Chul (Department of Medical Engineering, Cellbiocontrol Laboratory, Yonsei University College of Medicine) ;
  • Lee, Soo-Hong (Department of Biomedical Science, CHA University) ;
  • Park, Ki Dong (Department of Molecular Science and Technology, Ajou University) ;
  • Han, Dong-Wook (Department of Cogno-Mechatronics Engineering, College of Nanoscience & Nanotechnology, Pusan National University)
  • Received : 2015.11.04
  • Accepted : 2016.01.04
  • Published : 2016.03.01
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Abstract

Background: Hydrogels can serve as three-dimensional (3D) scaffolds for cell culture and be readily injected into the body. Recent advances in the image technology for 3D scaffolds like hydrogels have attracted considerable attention to overcome the drawbacks of ordinary imaging technologies such as optical and fluorescence microscopy. Multiphoton microscopy (MPM) is an effective method based on the excitation of two-photons. In the present study, C2C12 myoblasts differentiated in 3D gelatin hydroxyphenylpropionic acid (GHPA) hydrogels were imaged by using a custom-built multiphoton excitation fluorescence microscopy to compare the difference in the imaging capacity between conventional microscopy and MPM. Results: The physicochemical properties of GHPA hydrogels were characterized by using scanning electron microscopy and Fourier-transform infrared spectroscopy. In addition, the cell viability and proliferation of C2C12 myoblasts cultured in the GHPA hydrogels were analyzed by using Live/Dead Cell and CCK-8 assays, respectively. It was found that C2C12 cells were well grown and normally proliferated in the hydrogels. Furthermore, the hydrogels were shown to be suitable to facilitate the myogenic differentiation of C2C12 cells incubated in differentiation media, which had been corroborated by MPM. It was very hard to get clear images from a fluorescence microscope. Conclusions: Our findings suggest that the gelatin-based hydrogels can be beneficially utilized as 3D scaffolds for skeletal muscle engineering and that MPM can be effectively applied to imaging technology for tissue regeneration.

Keywords

Acknowledgement

Supported by : National Research Foundation of Korea (NRF)

References

  1. Jeon BH, Chae YG, Hwang SS, Kim DK, Oak C, Park EK, et al. Multimodal imaging of sarcopeni using optical coherence tomography and ultrasound in rat model. J Opt Soc Korea. 2014;18:55-9. https://doi.org/10.3807/JOSK.2014.18.1.055
  2. Conchello JA, Lichtman JW. Optical sectioning microscopy. Nat Methods. 2005;2:920-31. https://doi.org/10.1038/nmeth815
  3. Weber M, Huisken J. Light sheet microscopy for real-time developmental biology. Curr Opin Genet Dev. 2011;21:566-72. https://doi.org/10.1016/j.gde.2011.09.009
  4. Gao L, Shao L, Higgins CD, Poulton JS, Peifer M, Davidson MW, et al. Noninvasive imaging beyond the diffraction limit of 3D dynamics in thickly fluorescent specimens. Cell. 2012;151:1370-85. https://doi.org/10.1016/j.cell.2012.10.008
  5. Wicker K, Heintzmann R. Interferometric resolution improvement for confocal microscopes. Opt Express. 2007;15:12206-16. https://doi.org/10.1364/OE.15.012206
  6. Kobat D, Horton NG, Xu C. In vivo two-photon microscopy to 1.6-mm depth in mouse cortex. J Biomed Opt. 2011;16:106014. https://doi.org/10.1117/1.3646209
  7. Denk W, Strickler JH, Webb WW. Two-photon laser scanning fluorescence microscopy. Science. 1990;248:73-6. https://doi.org/10.1126/science.2321027
  8. Richard KPB, David WP. Two-photon excitation microscopy for the study of living cells and tissues. Cell Biol. 2014;4:1124-59.
  9. Ammasi P, Paul S, Colten N, Raymond K. An evaluation of two-photon excitation versus confocal and digital deconvolution fluorescence microscopy imaging in xenopus morphogenesis. Microsc Res Techniq. 1999;47:172-81. https://doi.org/10.1002/(SICI)1097-0029(19991101)47:3<172::AID-JEMT3>3.0.CO;2-A
  10. Karel S, Ryohei Y. Principle of two-photon excitation microscopy and its applications to neuroscience. Neuron. 2006;50:823-39. https://doi.org/10.1016/j.neuron.2006.05.019
  11. Majewska A, Yiu G, Yuste R. A custom-made two-photon microscope and deconvolution system. Pflugers Arch. 2000;441:398-408. https://doi.org/10.1007/s004240000435
  12. Thorling CA, Crawford D, Burczynski FJ, Liu X, Liau I, Roberts MS. Multiphoton microscopy in defining liver function. J Biomed Opt. 2014;19:90901. https://doi.org/10.1117/1.JBO.19.9.090901
  13. Wang JW, Wong AM, Flores J, Vosshall LB, Axel R. Two-photon calcium imaging reveals an odore-voked map of activity in the fly brain. Cell. 2003;112:271-82. https://doi.org/10.1016/S0092-8674(03)00004-7
  14. Oertner TG. Functional imaging of single synapses in brain slices. Exp Physiol. 2002;87:733-6. https://doi.org/10.1113/eph8702482
  15. Lee HS, Lee HD, Jeong MY, Kim CS. Wavelength-swept cascaded Raman fiber laser around 1300 nm for OCT imaging. J Opt Soc Korea. 2015;19:154-8. https://doi.org/10.3807/JOSK.2015.19.2.154
  16. Rose CR, Kovalchuk Y, Eilers J, Konnerth A. Two-photon $Na^+$ imaging in spines and fine dendrites of central neurons. Pflugers Arch. 1999;439:201-7.
  17. Chatterjee K, Lin-Gibson S, Wallace WE, Parekh SH, Lee YJ, Cicerone MT, et al. The effect of 3D hydrogel scaffold modulus on osteoblast differentiation and mineralization revealed by combinatorial screening. Biomaterials. 2010;31:5051-62. https://doi.org/10.1016/j.biomaterials.2010.03.024
  18. Nagai Y, Yokoi H, Kaihara K, Naruse K. The mechanical stimulation of cells in 3D culture within a self-assembling peptide hydrogel. Biomaterials. 2012;33:1044-51. https://doi.org/10.1016/j.biomaterials.2011.10.049
  19. Wang LS, Du C, Chung JE, Kurikawa M. Enzymatically cross-linked gelatin-phenol hydrogels with a broader stiffness range for osteogenic differentiation of human mesenchymal stem cells. Acta Biomater. 2012;8:1826-37. https://doi.org/10.1016/j.actbio.2012.02.002
  20. Sahu A, Choi WI, Tae G. A stimuli-sensitive injectable graphene oxide composite hydrogel. Chem Commun. 2012;48:5801-940. https://doi.org/10.1039/c2cc90181a
  21. Lee Y, Bae JW, Oh DH, Park KM, Chun YW, Sung HJ, et al. In situ forming gelatin-based tissue adhesives and their phenolic content-driven properties. J Mater Chem B. 2013;1:2407-14. https://doi.org/10.1039/c3tb00578j
  22. Park KM, Jun I, Joung YK, Shin H, Park KD. In situ hydrogelation and RGD conjugation of tyramine-conjugated 4-arm PPO-PEO block copolymer for injectable bio-mimetic scaffolds. Soft Matter. 2011;7:986-92. https://doi.org/10.1039/C0SM00612B
  23. Lee Y, Bae JW, Lee JW, Suh W, Park KD. Enzyme-catalyzed in situ forming gelatin hydrogels as bioactive wound dressings: effects of fibroblast delivery on wound healing efficacy. J Mater Chem B. 2014;2:7712-8. https://doi.org/10.1039/C4TB01111B
  24. Wang LS, Boulaire J, Chan PPY, Chung JE, Kurikawa M. The role of stiffness of gelatin-hydroxyphenylpropionic acid hydrogels formed by enzyme-mediated crosslinking on the differentiation of human mesenchymal stem cell. Biomateirals. 2010;31:8608-16. https://doi.org/10.1016/j.biomaterials.2010.07.075
  25. Liu XH, Ma PX. Phase separation, pore structure, and properties of nanofibrous gelatin scaffolds. Biomaterials. 2009;30:4094-103. https://doi.org/10.1016/j.biomaterials.2009.04.024
  26. Silva SS, Mano JF, Reis RL. Potential applications of natural origin polymerbased systems in soft tissue regeneration. Crit Rev Biotechnol. 2010;30:200-21. https://doi.org/10.3109/07388551.2010.505561
  27. Huang S, Fu XB. Naturally derived materials-based cell and drug delivery systems in skin regeneration. J Control Release. 2010;142:149-59. https://doi.org/10.1016/j.jconrel.2009.10.018
  28. Neffe AT, Loebus A, Zaupa A, Stoetzel C, Muller FA, Lendlein A. Gelatin functionalization with tyrosine derived moieties to increase the interaction with hydroxyapatite fillers. Acta Biomater. 2011;7:1693-701. https://doi.org/10.1016/j.actbio.2010.11.025
  29. Yuan SJ, Xiong G, Roguin A, Choong C. Immobilization of gelatin onto poly(glycidyl methacrylate)-grafted polycaprolactone substrates for improved cell-material interactions. Biointerphases. 2012;7:30. https://doi.org/10.1007/s13758-012-0030-1
  30. Ratner BD, Hoffman AS, Schoen FJ, Lemons JE. Biomaterials science: an introduction to materials in medicine. 3rd ed. San Diego: Academic Press; 2004.
  31. Hoffman AS. Hydrogels for biomedical applications. Adv Drug Delivery Rev. 2002;54:3-12. https://doi.org/10.1016/S0169-409X(01)00239-3
  32. Kytai TN, Jennifer LW. Photopolymerizable hydrogels for tissue engineering applications. Biomaterials. 2002;23:4307-14. https://doi.org/10.1016/S0142-9612(02)00175-8
  33. Serafim A, Dragusin DM, Zecheru T, Dubruel P, Petre D, Ciocan LT, et al. Gelatin hydrogels: effect of ethylene oxide based synthetic crosslinking agents on the physico-chemical properties. Dig J Nanomater Bios. 2013;8:101-10.
  34. Park S, Lee KS, Bozoklu G, Cai W, Nguyen ST, Ruoff RS. Graphene oxide papers modified by divalent ions-enhancing mechanical properties via chemical cross-linking. ACS Nano. 2008;2:572-8. https://doi.org/10.1021/nn700349a
  35. Shin YC, Lee JH, Jin L, Kim MJ, Kim YJ, Hyun JK, et al. Stimulated myoblast differentiation on graphene oxide-impregnated PLGA-collagen hybrid fibre matrices. J Nanobiotechnol. 2015;13:21. https://doi.org/10.1186/s12951-015-0081-9
  36. Yang T, Liu L, Liu J, Chen M, Wang J. Cyanobacterium metallothionein decorated graphene oxide nanosheets for highly selective adsorption of ultra-trace cadmium. J Mater Chem. 2012;22:21909-16. https://doi.org/10.1039/c2jm34712a
  37. Liu Z, Jiang L, Galli F, Nederlof I, Olsthoorn RCL, Lamers GEM, et al. A graphene oxide streptavidin complex for biorecognition - towards affinity purification. Adv Funct Mater. 2010;20:2857-65. https://doi.org/10.1002/adfm.201000761
  38. Shin YC, Lee JH, Jin L, Kim MJ, Kim C, Hong SW, et al. Cell-adhesive matrices composed of RGD peptide-displaying M13 bacteriophage/poly(lactic-co-glycolic acid) nanofibers beneficial to myoblast differentiation. J Nanosci Nanotechnol. 2015;15:7907-12. https://doi.org/10.1166/jnn.2015.11214
  39. Shin YC, Lee JH, Kim MJ, Hong SW, Oh JW, Kim CS, et al. Stimulating effect of graphene oxide on myogenesis of C2C12 myoblasts on PLGA/RGD nanofiber matrices. J Biol Eng. 2015;9:22. https://doi.org/10.1186/s13036-015-0020-1
  40. Shin YC, Lee JH, Jin OS, Lee EJ, Jin L, Kim C, et al. RGD peptide-displaying M13 bacteriophage/PLGA nanofibers as cell-adhesive matrices for smooth muscle cells. J Korean Phys Soc. 2015;66:12-6. https://doi.org/10.3938/jkps.66.12
  41. Shin YC, Jin L, Lee JH, Jun S, Hong SW, Kim CS, et al. Graphene oxideincorporated PLGA-collagen fibrous matrices as biomimetic scaffolds for vascular smooth muscle cells. Sci Adv Mater. 2015;7:1-6. https://doi.org/10.1166/sam.2015.2002

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