Control of maleic acid-propylene diepoxide hydrogel for 3D printing application for flexible tissue engineering scaffold with high resolution by end capping and graft polymerization

다국어 초록 (Multilingual Abstract)

Background: Control of 3D printing of highly tough hydrogel inks with adequate printability, scaffold fidelity and mechanical properties are highly desirable for biomedical and tissue engineering applications. However, developing a biocompatible tough ink with high-resolution printability, biodegradability, self-healing, adhesion, and integration with surrounding tissues is a big challenge in 3D printing. The aim of this study was to develop extrusion-based 3D printing of viscous hydrogel composing of maleic acid and propylene diepoxide by controlling continuous mechanisms of condensation and radical polymerization.
Methods: The molecular weight of highly adhesive propagating poly(malate-co-propylene oxide) copolymer was controlled by capping its growing chain with mono-functional lipoic acid with different compositions during condensation reaction to form lipoic acid capped gel (LP-capped gel). Poly(ethylene oxide)-diacrylate, PEGDA, is graftpolymerized to the LP-capped backbone polymer (MPLE gel) by UV irradiation during 3D printing process to control the properties of gel printability, mechanical properties, and cell adhesiveness and post-printing fidelity of the printed scaffolds with high resolution and mechanical properties (MPLE scaffold). The scaffolds in complex geometries have been printed out in diverse forms with addition of model drugs with different molecular weights and chemical structures.
Both the highly adhesive LP-capped gel and printing-controlled MPLE gel/scaffolds are diversely characterized and compared with for their applications to the extrusion-based printability, including biocompatibility, self-healing, drug releasing, adhesiveness, multi-layered high-resolution printing. Further in vitro/in vivo tests were done to observe cytotoxicity, immune response and tissue formation by using different cells in mice model.
Results: LP-capped hydrogel from maleic acid and propylene diepoxide gel showed control of gel properties with lipoic acid with one function group of thiol during condensation reaction, and the ratio at 1:0.3 (w/v) between LPcapped gel and PEGDA was chosen for the optimal results during radical polymerization process for 3D printing at high resolution (90-140 μm in strut thickness) with various complex geometries (lattice, rhombus, and honeycomb). The hydrogel showed excellent properties of self-healing, mechanical strength, biocompatibility, etc. In addition, the long-term release profiles of bioactive molecules were well-controlled by incorporating drugs of high molecular bovine serum albumin (BSA, 21 days, 98.4 ± 0.69%), or small molecule ornidazole (ORN, 14 days, 97.1 ± 1.98%) into the MPLE gel scaffolds for the tests of potential therapeutic applications. More importantly, the MPLE gels represents excellent in vitro cyto-compatibility against osteoblast-like cells (MC3T3) with viability value at 96.43% ± 7.48% over 7 culturing days. For in-vivo studies, the flexible MPLE scaffolds showed significant improvement on angiogenesis with minor inflammatory response after 4-week implantation in mice.
Conclusion: The MPLE gel inks was well-controlled for the fabrication of flexible complex tissue engineering scaffold with high resolutions, shear-thinning, 3D printability and post-printing fidelity, by modulating the composition of the highly adhesive LP-capped gel and inert PEGDA as well as end capping of lipoic acid to the propagating poly(malate-co-propylene oxide) copolymer. The gel ink demonstrated its excellent printability, in vitro/in vivo biocompatibility and mechanical properties as well as sustained drug release from the gel.

참고문헌 (Reference)

• 1 da Silva PA, "Vapor pressure curve determination of α-lipoic acid raw material and capsules by dynamic thermogravimetric method" 544 : 95-98, 2012
• 2 Zheng Z, "Triblock copolymers containing UCST polypeptide and poly(propylene glycol) : synthesis, thermoresponsive properties, and modification of PVA hydrogel" 115 : 244-250, 2019
• 3 LeTuyen Nguyen ; Sumi Bang ; Insup Noh, "Tissue Regeneration of Human Mesenchymal Stem Cells on Porous Gelatin Micro-Carriers by Long-Term Dynamic In Vitro Culture" 한국조직공학과 재생의학회 16 (16): 19-28, 2019
• 4 Kim SD, "Three-dimensional printed design of antibiotic-releasing esophageal patches for antimicrobial activity prevention" 27 (27): 1490-1502, 2021
• 5 Hospodiuk M, "The bioink : a comprehensive review on bioprintable materials" 35 (35): 217-239, 2017
• 6 Das D, "Synthesis and characterizations of alginate-α-tricalcium phosphate microparticle hybrid film with flexibility and high mechanical property as a biomaterial" 13 (13): 025008-, 2018
• 7 Pillai MM, "Symbiotic culture of nanocellulose pellicle : A potential matrix for 3D bioprinting" 119 : 111552-, 2021
• 8 Malekmohammadi S, "Smart and biomimetic 3D and 4D printed composite hydrogels : opportunities for different biomedical applications" 9 (9): 1537-, 2021
• 9 Janarthanan G, "Self-crosslinking hyaluronic acid–carboxymethylcellulose hydrogel enhances multilayered 3D-printed construct shape integrity and mechanical stability for soft tissue engineering" 12 (12): 045026-, 2020
• 10 Roy P, "Removal of toxic metals using superabsorbent polyelectrolytic hydrogels" 122 (122): 2415-2423, 2011
• 11 자나르다난 고피나단 ; 노인섭, "Recent trends in bioinks for 3D printing" 한국생체재료학회 22 (22): 105-119, 2018
• 12 Askari M, "Recent progress in extrusion 3D bioprinting of hydrogel biomaterials for tissue regeneration : a comprehensive review with focus on advanced fabrication techniques" 9 (9): 535-573, 2021
• 13 Zheng SY, "Programmed deformations of 3D-printed tough physical hydrogels with high response speed and large output force" 28 (28): 1803366-, 2018
• 14 Soltan N, "Printability and cell viability in bioprinting alginate dialdehyde-gelatin scaffolds" 5 (5): 2976-2987, 2019
• 15 Wu H-L, "Preparations and properties of maleic acid and maleic anhydride functionalized multiwall carbon nanotube/poly(urea urethane)nanocomposites" 67 (67): 1854-1860, 2007
• 16 Saimani S, "Polyethylene glycol diacrylate and thermoplastic polymers based semi-IPNs for asymmetric membranes" 15 (15): 781-797, 2008
• 17 Munim SA, "Poly(lactic acid)based hydrogels : formation, characteristics and biomedical applications" 26 (26): 881-901, 2019
• 18 van Bochove B, "Photo-crosslinked synthetic biodegradable polymer networks for biomedical applications" 30 (30): 77-106, 2019
• 19 Wei H, "Orthogonal photochemistryassisted printing of 3D tough and stretchable conductive hydrogels" 12 (12): 1-10, 2021
• 20 Gao T, "Optimization of gelatin–alginate composite bioink printability using rheological parameters : a systematic approach" 10 (10): 034106-, 2018
• 21 Bhattacharyya A, "Nano-biomaterials for designing functional bioinks towards complex tissue and organ regeneration in 3D bioprinting" 37 : 101639-, 2021
• 22 Guo Z, "Mussel-inspired naturally derived double-network hydrogels and their application in 3D printing : from soft, injectable bioadhesives to mechanically strong hydrogels" 6 (6): 1798-1808, 2020
• 23 아미타바 바타차리아 ; Gopinathan Janarthanan ; Taeyang Kim ; Shiva Taheri ; Jisun Shin ; Jihyeon Kim ; Hyun Cheol Bae ; 한혁수 ; 노인섭, "Modulation of bioactive calcium phosphate micro/nanoparticle size and shape during in situ synthesis of photo-crosslinkable gelatin methacryloyl based nanocomposite hydrogels for 3D bioprinting and tissue engineering" 한국생체재료학회 26 (26): 983-999, 2022
• 24 Janarthanan G, "Manufacturing of self-standing multi-layered 3D-bioprinted alginate-hyaluronate constructs by controlling the cross-linking mechanisms for tissue engineering applications" 14 : 035013-, 2022
• 25 Zhang J, "Hydrogel-based preparation of cell aggregates for biomedical applications" 20 : 100747-, 2020
• 26 Unagolla JM, "Hydrogel-based 3D bioprinting : a comprehensive review on cell-laden hydrogels, bioink formulations, and future perspectives" 18 : 100479-, 2020
• 27 Lee JY, "Fundamentals and applications of 3D printing for novel materials" 7 : 120-133, 2017
• 28 Ullah S, "Fabrication, applications and challenges of natural biomaterials in tissue engineering" 20 : 100656-, 2020
• 29 Das D, "Fabrication of alginate-based stimuliresponsive, non-cytotoxic, terpolymric semi-IPN hydrogel as a carrier for controlled release of bovine albumin serum and 5-amino salicylic acid" 98 : 42-53, 2019
• 30 Bang S, "Evaluation of MC3T3 cells proliferation and drug release study from sodium hyaluronate-1, 4-butanediol diglycidyl ether patterned gel" 7 (7): 328-, 2017
• 31 Celebioglu A, "Encapsulation and stabilization of α-lipoic acid in cyclodextrin inclusion complex electrospun nanofibers : antioxidant and fast-dissolving α-lipoic acid/cyclodextrin nanofibrous webs" 67 (67): 13093-13107, 2019
• 32 Yang X, "Electrospun and photocrosslinked gelatin/dextran–maleic anhydride composite fibers for tissue engineering" 113 : 142-147, 2019
• 33 Massoumi B, "Electrically conductive nanofibrous scaffold composed of poly(ethylene glycol)-modified polypyrrole and poly(ε-caprolactone)for tissue engineering applications" 98 : 300-310, 2019
• 34 Zolfagharian A, "Effects of topology optimization in multimaterial 3D bioprinting of soft actuators" 6 (6): 260-, 2020
• 35 Das D, "Effects of molar ratios of two immiscible monomers toward development of an amphiphilic, highly stretchable, bioadhesive, self-healing Copolymeric hydrogel and its mineralactive cellular behavior" 21 (21): 892-902, 2020
• 36 Rajam K, "Effect of caffeine-Zn2+ system in preventing corrosion of carbon steel in well water" 2013 : 1-11, 2013
• 37 Baghayeri M, "Direct electrochemistry and electrocatalysis of hemoglobin immobilized on biocompatible poly(styrenealternative-maleic acid)/functionalized multi-wall carbon nanotubes blends" 188 : 227-234, 2013
• 38 Fellows MD, "Cytotoxicity in cultured mammalian cells is a function of the method used to estimate it" 22 (22): 275-280, 2007
• 39 Xu L, "Conjoined-network rendered stiff and tough hydrogels from biogenic molecules" 5 (5): eaau3442-, 2019
• 40 Das D, "Characterizations of hyaluronate-based terpolymeric hydrogel synthesized via free radical polymerization mechanism for biomedical applications" 170 : 64-75, 2018
• 41 Harikrishna R, "Cationic photopolymerization kinetics of neat coating formulations involving poly(propylene glycol)diglycidyl ether and glycerol diglycidyl ether" 122 (122): 1445-1454, 2015
• 42 AM DC, "Caffeine-catalyzed gels" 170 : 127-135, 2018
• 43 Murphy CA, "Biopolymers and polymers in the search of alternative treatments for meniscal regeneration : state of the art and future trends" 12 : 51-71, 2018
• 44 Bhattacharyya A, "Bioink homogeneity control during 3D bioprinting of multicomponent micro/nanocomposite hydrogel for even tissue regeneration using novel twin screw extrusion system" 415 : 128971-, 2021
• 45 Puppi D, "Biodegradable polymers for biomedical additive manufacturing" 20 : 100700-, 2020
• 46 Moore E, "Bioactive poly(ethylene glycol)acrylate hydrogels for regenerative engineering" 5 (5): 167-179, 2019
• 47 Liu P, "A novel poly(vinyl alcohol)/poly(ethylene glycol)scaffold for tissue engineering with a unique bimodal open-celled structure fabricated using supercritical fluid foaming" 9 (9): 1-12, 2019
• 48 이재후 ; 김규언 ; 방수미 ; 노인섭 ; 이치범, "A Desktop Multi-Material 3D Bio-Printing System with Open-Source Hardware and Software" 한국정밀공학회 18 (18): 605-612, 2017
• 49 Bakarich SE, "3D printing of tough hydrogel composites with spatially varying materials properties" 14 : 24-30, 2017
• 50 Chen Z, "3D printing of multifunctional hydrogels" 29 (29): 1900971-, 2019
• 51 Seymour AJ, "3D printing of microgel scaffolds with tunable void fraction to promote cell infiltration" 10 (10): 2100644-, 2021
• 52 Hong S, "3D printing of highly stretchable and tough hydrogels into complex, cellularized structures" 27 (27): 4035-4040, 2015
• 53 Janarthanan G, "3D printing of bioinspired alginate-albumin based instant gel ink with Electroconductivity and its expansion to direct four-Axis printing of hollow porous tubular constructs without supporting materials" 31 (31): 2104441-, 2021
• 54 Sultan S, "3D printed scaffolds with gradient porosity based on a cellulose nanocrystal hydrogel" 10 (10): 4421-4431, 2018
• 55 노인섭 ; 김나혜 ; Hao Nguyen Tran ; 이재후 ; Chibum Lee, "3D printable hyaluronic acid-based hydrogel for its potential application as a bioink in tissue engineering" 한국생체재료학회 23 (23): 75-83, 2019
• 56 Janarthanan G, "3D printable and injectable lactoferrin-loaded carboxymethyl cellulose-glycol chitosan hydrogels for tissue engineering applications" 113 : 111008-, 2020
• 57 Kim W, "3D bioprinting of functional cell-laden bioinks and its application for cell-alignment and maturation" 19 : 100588-, 2020