Tailoring the growth and proliferation of human dermal fibroblasts by DNA-based polymer films for skin regeneration
Cristiano Ceron Jayme
Department of Chemistry, Center of Nanotechnology and Tissue Engineering Photobiology and Photomedicine Research Group, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto-FFCLRP, University of São Paulo, Ribeirão Preto, São Paulo, Brazil
Search for more papers by this authorCarla Souza
Department of Chemistry, Center of Nanotechnology and Tissue Engineering Photobiology and Photomedicine Research Group, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto-FFCLRP, University of São Paulo, Ribeirão Preto, São Paulo, Brazil
Search for more papers by this authorDaniela Silvestrini Fernandes
Department of Chemistry, Center of Nanotechnology and Tissue Engineering Photobiology and Photomedicine Research Group, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto-FFCLRP, University of São Paulo, Ribeirão Preto, São Paulo, Brazil
Search for more papers by this authorCorresponding Author
Antonio Claudio Tedesco
Department of Chemistry, Center of Nanotechnology and Tissue Engineering Photobiology and Photomedicine Research Group, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto-FFCLRP, University of São Paulo, Ribeirão Preto, São Paulo, Brazil
Correspondence
Antonio Claudio Tedesco, Department of Chemistry, Center of Nanotechnology and Tissue Engineering Photobiology and Photomedicine Research Group, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto-FFCLRP, University of São Paulo, Av. Bandeirantes, 3900, Zip code: 14.040-901, Ribeirão Preto, São Paulo, Brazil.
Email: [email protected]
Search for more papers by this authorCristiano Ceron Jayme
Department of Chemistry, Center of Nanotechnology and Tissue Engineering Photobiology and Photomedicine Research Group, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto-FFCLRP, University of São Paulo, Ribeirão Preto, São Paulo, Brazil
Search for more papers by this authorCarla Souza
Department of Chemistry, Center of Nanotechnology and Tissue Engineering Photobiology and Photomedicine Research Group, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto-FFCLRP, University of São Paulo, Ribeirão Preto, São Paulo, Brazil
Search for more papers by this authorDaniela Silvestrini Fernandes
Department of Chemistry, Center of Nanotechnology and Tissue Engineering Photobiology and Photomedicine Research Group, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto-FFCLRP, University of São Paulo, Ribeirão Preto, São Paulo, Brazil
Search for more papers by this authorCorresponding Author
Antonio Claudio Tedesco
Department of Chemistry, Center of Nanotechnology and Tissue Engineering Photobiology and Photomedicine Research Group, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto-FFCLRP, University of São Paulo, Ribeirão Preto, São Paulo, Brazil
Correspondence
Antonio Claudio Tedesco, Department of Chemistry, Center of Nanotechnology and Tissue Engineering Photobiology and Photomedicine Research Group, Faculty of Philosophy, Sciences and Letters of Ribeirão Preto-FFCLRP, University of São Paulo, Av. Bandeirantes, 3900, Zip code: 14.040-901, Ribeirão Preto, São Paulo, Brazil.
Email: [email protected]
Search for more papers by this authorFunding information: Conselho Nacional de Desenvolvimento Científico e Tecnológico; Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, Grant/Award Number: 88882.317646/2019-01; Financiadora de Estudos e Projetos, Grant/Award Number: 01.10.0758.01; Fundação de Amparo à Pesquisa do Estado de São Paulo, Grant/Award Numbers: #2018/10237-1, #2017/01272-5; Thematic project, Grant/Award Number: 2013/50181-1; National Institute of Science and Technology (INCT) of Nanobiotechnology, Grant/Award Number: 573880/2008-5
Abstract
The use of DNA as a functional biomaterial for therapeutic, diagnostic, and drug delivery applications has been prominent in recent years, but its use as a scaffold for tissue regeneration is still limited. This study aimed to evaluate the biocompatibility and interaction of DNA-based polymeric films (DNA-PFs) with primary human fibroblasts (PHF) for regenerative medicine and wound healing purposes. The morphological characterization of the films was performed by scanning electron microscopy, SEM–energy-dispersive X-ray spectroscopy, and atomic force microscopy analysis. Cell viability, cell cycle kinetics, oxidative stress, and migration studies were carried out at 48 and 72 hr of incubation and compared to control cells. Cell adhesion was impaired in the first 24 hr, DNA-PFs with higher concentrations of DNA (1.0 and 2.0 g/L) this effect was not seen in DNA-PFs (0.5 g/L), explained by the difference in topography and roughness of DNA-PFs, but it was overcome after 48 hr of incubation. PHF seeded on DNA films showed higher proliferation and migration rates than the control after 48 hr of incubation, with the maintenance of cell morphology and lower cytotoxicity and oxidative stress during the evaluation time. Therefore, these results indicate that DNA-PFs are highly biocompatible and provide a suitable microenvironment for dermal fibroblasts to maintain their activity, helping build new and more complex biomaterials suitable for future tissue repair applications.
CONFLICT OF INTEREST
The authors declare no potential conflict of interest.
Open Research
DATA AVAILABILITY STATEMENT
The data that support the findings of this study are available from the corresponding author upon reasonable request.
Supporting Information
| Filename | Description |
|---|---|
| jbma37220-sup-0001-FigureS1.tifTIFF image, 2.7 MB | Figure S1 Schematic diagram of the preparation of DNA-FP |
| jbma37220-sup-0002-FigureS2.tifTIFF image, 3.4 MB | Figure S2 Cell morphology/attachment and cell spreading on DNA-PF. Photomicrographs recorded 24 (A, D, G, J), 48 (B, E, H, K), and 72 hr (C, F, I, L) after seeding PHF on DNA-PFs at different DNA concentrations (0.5, 1.0, and 2.0 g/L) using a Carl Zeiss microscope with ×10 magnification objective coupled to a high resolution Axiocam-40 CFL digital camera. DNA-PFs: DNA polymeric films; PHF: primary human fibroblasts; Neg. Ctrl: cells without DNA-PFs. |
| jbma37220-sup-0003-FigureS3.tifTIFF image, 1 MB | Figure S3 Adhesion cell assay performed by MTT. Human fibroblast (PHF) was seeded on DNA films (at 0.5, 1.0, and 2.0 g/L) and control (without the DNA film) and analyzed after (A) 24 and (B) 48 hr after incubation. The data were subjected to one-way ANOVA with Tukey's post-hoc test. *p < .05. |
| jbma37220-sup-0004-VideoS1.mp4MPEG-4 video, 10.4 MB | Video 1 Migration free healing area from 0 to 72 hr. |
Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.
REFERENCES
- 1Li S, Tian T, Zhang T, Cai X, Lin Y. Advances in biological applications of self-assembled DNA tetrahedral nanostructures. Mater Today. 2019; 24: 57-68. https://doi.org/10.1016/j.mattod.2018.08.002.
- 2Vybornyi M, Vyborna Y, Häner R. DNA-inspired oligomers: from oligophosphates to functional materials. Chem Soc Rev. 2019; 48(16): 4347-4360. https://doi.org/10.1039/C8CS00662H.
- 3Watson JD, Crick FHC. A structure for deoxyribose nucleic acid. Nature. 1953; 171(4356): 737-738. https://doi.org/10.1038/171737a0.
- 4James BD, Guerin P, Iverson Z, Allen JB. Mineralized DNA-collagen complex-based biomaterials for bone tissue engineering. Int J Biol Macromol. 2020; 161: 1127-1139. https://doi.org/10.1016/j.ijbiomac.2020.06.126.
- 5Qi H, Ghodousi M, Du Y, et al. DNA-directed self-assembly of shape-controlled hydrogels. Nat Commun. 2013; 4: 2275. https://doi.org/10.1038/ncomms3275.
- 6Angell C, Xie S, Zhang L, Chen Y. DNA nanotechnology for precise control over drug delivery and gene therapy. Small. 2016; 12(9): 1117-1132. https://doi.org/10.1002/smll.201502167.
- 7Jayme CC, Calori IR, Cunha EMF, Tedesco AC. Evaluation of aluminum phthalocyanine chloride and DNA interactions for the design of an advanced drug delivery system in photodynamic therapy. Spectrochim Acta A Mol Biomol Spectrosc. 2018; 201: 242-248. https://doi.org/10.1016/j.saa.2018.05.009.
- 8Linko V, Ora A, Kostiainen MA. DNA nanostructures as smart drug-delivery vehicles and molecular devices. Trends Biotechnol. 2015; 33(10): 586-594. https://doi.org/10.1016/j.tibtech.2015.08.001.
- 9Jayme CC, Ferreira Pires A, Tedesco AC. Development of DNA polymer films as a drug delivery system for the treatment of oral cancer. Drug Deliv Transl Res. 2020; 10: 1612-1625. https://doi.org/10.1007/s13346-020-00801-9.
- 10Ma W, Shao X, Zhao D, et al. Self-assembled tetrahedral DNA nanostructures promote neural stem cell proliferation and neuronal differentiation. ACS Appl Mater Interfaces. 2018; 10(9): 7892-7900. https://doi.org/10.1021/acsami.8b00833.
- 11Zhou M, Liu N-X, Shi S-R, et al. Effect of tetrahedral DNA nanostructures on proliferation and osteo/odontogenic differentiation of dental pulp stem cells via activation of the notch signaling pathway. Nanomedicine. 2018; 14(4): 1227-1236. https://doi.org/10.1016/j.nano.2018.02.004.
- 12Fukushima T, Kawaguchi M, Hayakawa T, et al. Drug binding and releasing characteristics of DNA/lipid/PLGA film. Dent Mater J. 2007; 26(6): 854-860. https://doi.org/10.4012/dmj.26.854.
- 13Lee Y, Dugansani SR, Jeon SH, et al. Drug-delivery system based on salmon DNA nano-and micro-scale structures. Sci Rep. 2017; 7(1): 1-10. https://doi.org/10.1038/s41598-017-09904-9.
- 14Zhang H, Ma Y, Xie Y, et al. A controllable aptamer-based self-assembled DNA dendrimer for high-affinity targeting, bioimaging, and drug delivery. Sci Rep. 2015; 5:10099. https://doi.org/10.1038/srep10099.
- 15Carrow JK, Gaharwar AK. Bioinspired polymeric nanocomposites for regenerative medicine. Macromol Chem Phys. 2015; 216(3): 248-264. https://doi.org/10.1002/macp.201400427.
- 16Jiang J, Kong X, Xie Y, et al. Potent anti-tumor immunostimulatory biocompatible nano hydrogel made from DNA. Nanoscale Res Lett. 2019; 14(1): 217. https://doi.org/10.1186/s11671-019-3032-9.
- 17DeForest CA, Anseth KS. Cytocompatible click-based hydrogels with dynamically tunable properties through orthogonal photoconjugation and photocleavage reactions. Nature Chem. 2011; 3(12): 925. https://doi.org/10.1038/nchem.1174.
- 18Khademhosseini A, Langer R. A decade of progress in tissue engineering. Nat Protoc. 2016; 11(10): 1775-1781. https://doi.org/10.1038/nprot.2016.123.
- 19Tan H-L, Kai D, Pasbakhsh P, Teow S-Y, Lim Y-Y, Pushpamalar J. Electrospun cellulose acetate butyrate/polyethylene glycol (CAB/PEG) composite nanofibers: a potential scaffold for tissue engineering. Colloids Surf B Biointerfaces. 2020; 188:110713. https://doi.org/10.1016/j.colsurfb.2019.110713.
- 20Guo B, Ma PX. Conducting polymers for tissue engineering. Biomacromolecules. 2018; 19(6): 1764-1782. https://doi.org/10.1021/acs.biomac.8b00276.
- 21D'Souza SM, Ramees M, Sarojini B. Studies on the properties of Nano metal oxides doped DNA-CTMA matrix. Mater Today Proc. 2018; 5(1): 2582-2587. https://doi.org/10.1016/j.matpr.2017.11.042.
10.1016/j.matpr.2017.11.042 Google Scholar
- 22Rhim JW, Mohanty AK, Singh SP, Ng PK. Effect of the processing methods on the performance of polylactide films: Thermocompression versus solvent casting. J Appl Polym Sci. 2006; 101(6): 3736-3742. https://doi.org/10.1002/app.23403.
- 23Grote JG, Diggs DE, Nelson RL, et al. DNA photonics [deoxyribonucleic acid]. Mol Crys Liquid Crys. 2005; 426(1): 3-17. https://doi.org/10.1080/15421400590890615.
- 24Wang L, Yoshida J, Ogata N, Sasaki S, Kajiyama T. Self-assembled supramolecular films derived from marine deoxyribonucleic acid (DNA)− cationic surfactant complexes: large-scale preparation and optical and thermal properties. Chem Mater. 2001; 13(4): 1273-1281. https://doi.org/10.1021/cm000869g.
- 25Jayme CC, Kanicki J, Kajzar F o, Nogueira AF, Pawlicka A. Influence of DNA and DNA-PEDOT: PSS on dye-sensitized solar cell performance. Mol Crys Liquid Crys. 2016; 627(1): 38-48. https://doi.org/10.1080/15421406.2015.1137680.
- 26Varol M. Cell-extracellular matrix adhesion assay. Methods Mol Biol. 2019; 1: 209-217. https://doi.org/10.1007/76512019246.
10.1007/7651_2019_246 Google Scholar
- 27O'Brien J, Wilson I, Orton T, Pognan F. Investigation of the Alamar blue (resazurin) fluorescent dye for the assessment of mammalian cell cytotoxicity. Eur J Biochem. 2000; 267(17): 5421-5426. https://doi.org/10.1046/j.1432-1327.2000.01606.x.
- 28Souza C, Mônico DA, Tedesco AC. Implications of dichlorofluorescein photostability for detection of UVA-induced oxidative stress in fibroblasts and keratinocyte cells. Photochem Photobiol Sci. 2020; 19: 40-48. https://doi.org/10.1039/c9pp00415g.
- 29Liang C-C, Park AY, Guan J-L. In vitro scratch assay: a convenient and inexpensive method for analysis of cell migration in vitro. Nat Protoc. 2007; 2(2): 329-333. https://doi.org/10.1038/nprot.2007.30.
- 30Lemor M, de Bustros S, Glaser BM. Low-dose colchicine inhibits astrocyte, fibroblast, and retinal pigment epithelial cell migration and proliferation. Arch Ophthalmol. 1986; 104(8): 1223-1225.
- 31Okahata Y, Tanaka K. Oriented thin films of a DNA-lipid complex. Thin Solid Films. 1996; 284: 6-8. https://doi.org/10.1016/S0040-6090(96)08830-X.
- 32Andrukhov O, Huber R, Shi B, et al. Proliferation, behavior, and differentiation of osteoblasts on surfaces of different microroughness. Dent Mater. 2016; 32(11): 1374-1384. https://doi.org/10.1016/j.dental.2016.08.217.
- 33Umeda H, Mano T, Harada K, Tarannum F, Ueyama Y. The appearance of a cell-adhesion factor in osteoblast proliferation and differentiation of apatite coating titanium by a blast coating method. J Mater Sci Mater Med. 2017; 28(8): 112. https://doi.org/10.1007/s10856-017-5913-8.
- 34Jayme CC, de Paula LB, Rezende N, Calori IR, Franchi LP, Tedesco AC. DNA polymeric films support cell growth as a new material for regenerative medicine: compatibility and applicability. Exp Cell Res. 2017; 360: 404-412. https://doi.org/10.1016/j.yexcr.2017.09.033.
- 35Anselme K. Osteoblast adhesion on biomaterials. Biomaterials. 2000; 21(7): 667-681. https://doi.org/10.1016/s0142-9612(99)00242-2.
- 36Martin JY, Schwartz Z, Hummert TW, et al. Effect of titanium surface roughness on proliferation, differentiation, and protein synthesis of human osteoblast-like cells (MG63). J Biomed Mater Res A. 1995; 29(3): 389-401. https://doi.org/10.1002/jbm.820290314.
- 37Rampersad SN. Multiple applications of Alamar blue as an indicator of metabolic function and cellular health in cell viability bioassays. Sensors. 2012; 12(9): 12347-12360. https://doi.org/10.3390/s120912347.
- 38Liu Y, Bai H, Wang H, et al. Comparison of hypocrellin B-mediated sonodynamic responsiveness between sensitive and multidrug-resistant human gastric cancer cell lines. J Med Ultrason. 2019; 46(1): 15-26. https://doi.org/10.1007/s10396-018-0899-5.
- 39Geesala R, Bar N, Dhoke NR, Basak P, Das A. Porous polymer scaffold for on-site delivery of stem cells–protects from oxidative stress and potentiates wound tissue repair. Biomaterials. 2016; 77: 1-13. https://doi.org/10.1016/j.biomaterials.2015.11.003.
- 40Belső N, Gubán B, Manczinger M, et al. Differential role of D cyclins in the regulation of cell cycle by influencing Ki67 expression in HaCaT cells. Exp Cell Res. 2019; 374(2): 290-303. https://doi.org/10.1016/j.yexcr.2018.11.030.
- 41Sen S, Basak P, Sinha BP, et al. Anti-inflammatory effect of epidermal growth factor conjugated silk fibroin immobilized polyurethane ameliorates diabetic burn wound healing. Int J Biol Macromol. 2020; 143: 1009-1032. https://doi.org/10.1016/j.ijbiomac.2019.09.219.
- 42Liu J, Wei T, Zhao J, et al. Multifunctional aptamer-based nanoparticles for targeted drug delivery to circumvent cancer resistance. Biomaterials. 2016; 91: 44-56. https://doi.org/10.1016/j.biomaterials.2016.03.013.
Citing Literature
