A tri-layer tissue engineering heart valve scaffold based on atelocollagen, hyaluronic acid, and elastin

Abstract

Aim: This study aims to fabricate and characterise a novel tri-layer scaffold based on type I atelocollagen, hyaluronic acid (HA), and a novel fibrillar elastin gel, mimicking the native heart valve leaflets in structure, composition, and mechanical properties, among which, the bending anisotropic behaviour in both the with curvature (WC) and the against curvature (AC) directions, is the most desired. The use of atelocollagen is of significant importance in highlighting the non-antigenic potential of the design. Methods: Porous scaffolds were freeze-dried, then crosslinked using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS). The fibrillogenesis occurrence and the scaffold microstructure were imaged using scanning electron microscopy (SEM). Fourier transform infrared spectroscopy (FITR) investigated the effect of fabrication and crosslinking on the backbone structure. Dynamic mechanical analysis (DMA) characterised the compressive and bending properties of the scaffolds in hydrated and non-hydrated states. Three-point bending and a “self-deflection” test were performed on tri-layer scaffolds in both WC and AC directions. Results: Atelocollagen-based scaffolds were successfully produced, rendering this study the first to report a tri-layer structure using atelocollagen, HA, and elastin fibrillar gel. The scaffolds’ porosity was tailored to accommodate potential future biological studies and the transition between layers appeared seamless. FITR unveiled effective crosslinking and the backbone structure preservation. The scaffolds exhibited lightly crosslinked polymer resembling mechanical responses when non-hydrated, and the desired J-curve stress-strain response was observed when hydrated. The tri-layer scaffolds showed anisotropic bending behaviour with a bending modulus of 5.41 ± 1.14 kPa (WC) and 7.98 ± 2.22 kPa (AC). Conclusions: The tri-layer scaffolds fabricated resemble the native aortic valve leaflets in structure and composition, and successfully introduced bending anisotropy in physiological conditions. Together with the suitable microstructure and promising mechanical properties, the design is reckoned to be a potential tissue engineering heart valve candidate.

Keywords

Type I atelocollagen / tissue engineering heart valve / tri-layer / bending anisotropy

References

[1] Nkomo VT, Gardin JM, Skelton TN, Gottdiener JS, Scott CG, Enriquez-Sarano M. Burden of valvular heart diseases: a population-based study. Lancet. 2006; 368: 1005-11.

[2] Saurav A, Alla VM, Kaushik M, Hunter CC, Mooss AV. Outcomes of mitral valve repair compared with replacement in patients undergoing concomitant aortic valve surgery: a meta-analysis of observational studies. Eur J Cardiothorac Surg. 2015; 48: 347-53.

[3] Kheradvar A, Groves EM, Dasi LP, Alavi SH, Tranquillo R, Grande-Allen KJ, et al. Emerging trends in heart valve engineering: Part I. Solutions for future. Ann Biomed Eng. 2015; 43: 833-43.

[4] Fioretta ES, Dijkman PE, Emmert MY, Hoerstrup SP. The future of heart valve replacement: recent developments and translational challenges for heart valve tissue engineering. J Tissue Eng Regen Med. 2018; 12: e323-35.

[5] Oveissi F, Naficy S, Lee A, Winlaw DS, Dehghani F. Materials and manufacturing perspectives in engineering heart valves: a review. Mater Today Bio. 2019; 5: 100038.

[6] Alsoufi B. Aortic valve replacement in children: Options and outcomes. J Saudi Heart Assoc. 2014; 26: 33-41.

[7] Nishimura RA, Warnes CA. Anticoagulation during pregnancy in women with prosthetic valves: evidence, guidelines and unanswered questions. Heart. 2015; 101: 430-5.

[8] Langer R, Vacanti JP. Tissue engineering. Science. 1993; 260: 920-6.

[9] Hinderer S, Seifert J, Votteler M, Shen N, Rheinlaender J, Schäffer TE, et al. Engineering of a bio-functionalized hybrid off-the-shelf heart valve. Biomaterials. 2014; 35: 2130-9.

[10] Vesely I, Noseworthy R. Micromechanics of the fibrosa and the ventricularis in aortic valve leaflets. J Biomech. 1992; 25: 101-13.

[11] Lee JM, Boughner DR, Courtman DW. The glutaraldehyde-stabilized porcine aortic valve xenograft. II. Effect of fixation with or without pressure on the tensile viscoelastic properties of the leaflet material. J Biomed Mater Res. 1984; 18: 79-98.

[12] Vyavahare N, Ogle M, Schoen FJ, Zand R, Gloeckner DC, Sacks M, et al. Mechanisms of bioprosthetic heart valve failure: fatigue causes collagen denaturation and glycosaminoglycan loss. J Biomed Mater Res. 1999; 46: 44-50.

[13] Robi K, Jakob N, Matevz K, Matjaz V. The Physiology of Sports Injuries and Repair Processes. In: Hamlin M, Draper N, Kathiravel Y, editors. Current Issues in Sports and Exercise Medicine. IntechOpen; 2013.

[14] Sachlos E, Reis N, Ainsley C, Derby B, Czernuszka JT. Novel collagen scaffolds with predefined internal morphology made by solid freeform fabrication. Biomaterials. 2003; 24: 1487-97.

[15] Salvatore L, Calò E, Bonfrate V, Pedone D, Gallo N, Natali ML, et al. Exploring the effects of the crosslink density on the physicochemical properties of collagen-based scaffolds. Polym Test. 2021; 93: 106966.

[16] Chen Q, Bruyneel A, Carr C, Czernuszka J. Bio-mechanical properties of novel bi-layer collagen-elastin scaffolds for heart valve tissue engineering. Procedia Eng. 2013; 59: 247-54.

[17] Chen Q, Bruyneel A, Carr C, Czernuszka J. Trilayer scaffold with cardiosphere-derived cells for heart valve tissue engineering. J Biomed Mater Res B Appl Biomater. 2020; 108: 729-37.

[18] Nazir R, Bruyneel A, Carr C, Czernuszka J. Collagen type I and hyaluronic acid based hybrid scaffolds for heart valve tissue engineering. Biopolymers. 2019; 110: e23278.

[19] Nazir R, Bruyneel A, Carr C, Czernuszka J. Mechanical and Degradation Properties of Hybrid Scaffolds for Tissue Engineered Heart Valve (TEHV). J Funct Biomater. 2021; 12: 20.

[20] Merryman WD, Huang HS, Schoen FJ, Sacks MS. The effects of cellular contraction on aortic valve leaflet flexural stiffness. J Biomech. 2006; 39: 88-96.

[21] Lynn AK, Yannas IV, Bonfield W. Antigenicity and immunogenicity of collagen. J Biomed Mater Res B Appl Biomater. 2004; 71: 343-54.

[22] Miyata T, Taira T, Noishiki Y. Collagen engineering for biomaterial use. Clin Mater. 1992; 9: 139-48.

[23] Abbasi N, Hamlet S, Love RM, Nguyen NT. Porous scaffolds for bone regeneration. J Sci Adv Mater Devices. 2020; 5: 1-9.

[24] Lien S, Ko L, Huang T. Effect of pore size on ECM secretion and cell growth in gelatin scaffold for articular cartilage tissue engineering. Acta Biomater. 2009; 5: 670-9.

[25] Rashidi N. Fabrication of a Novel Bilayered Osteochondral Scaffold Using a Bottom up Approach [dissertation]. Oxford: Oxford University; 2018.

[26] Iftekhar S. Isolation of Fibrillar Elastin Gel (FEG) and its application in heart valve tissue engineering [dissertation]. Oxford: Oxford University; 2020.

[27] Baer E, Cassidy JJ, Hiltner A. Hierarchical Structure of Collagen Composite Systems. In: Glasser WG, Hatakeyama H, editors. Viscoelasticity of Biomaterials. American Chemical Society; 1992. pp. 2-23.

[28] Bozec L, van der Heijden G, Horton M. Collagen fibrils: nanoscale ropes. Biophys J. 2007; 92: 70-5.

[29] Borrego-González S, Rico-Llanos G, Becerra J, Díaz-Cuenca A, Visser R. Sponge-like processed D-periodic self-assembled atelocollagen supports bone formation in vivo. Mater Sci Eng C Mater Biol Appl. 2021; 120: 111679.

[30] Wu Y, Lee T, Chiu K, Shaw S, Yang C. A comparative study of the physical and mechanical properties of three natural corals based on the criteria for bone-tissue engineering scaffolds. J Mater Sci Mater Med. 2009; 20: 1273-80.

[31] Stani C, Vaccari L, Mitri E, Birarda G. FTIR investigation of the secondary structure of type I collagen: New insight into the amide III band. Spectrochim Acta A Mol Biomol Spectrosc. 2020; 229: 118006.

[32] Ji Y, Yang X, Ji Z, Zhu L, Ma N, Chen D, et al. DFT-Calculated IR Spectrum Amide I, II, and III Band Contributions of N-Methylacetamide Fine Components . ACS Omega. 2020; 5: 8572-8.

[33] Machado AAS, Martins VCA, Plepis AMG. Thermal and rheological behavior of collagen: Chitosan blends. J Therm Anal Calorim. 2002; 67: 491-8.

[34] Lee Y, Chiang C, Huang P, Chung C, Huang TD, Wang C, et al. Evidence of preserved collagen in an Early Jurassic sauropodomorph dinosaur revealed by synchrotron FTIR microspectroscopy. Nat Commun. 2017; 8: 14220.

[35] Wang XH, Li DP, Wang WJ, Feng QL, Cui FZ, Xu YX, et al. Crosslinked collagen/chitosan matrix for artificial livers. Biomaterials. 2003; 24: 3213-20.

[36] Schmidt FN, Zimmermann EA, Campbell GM, Sroga GE, Püschel K, Amling M, et al. Assessment of collagen quality associated with non-enzymatic cross-links in human bone using Fourier-transform infrared imaging. Bone. 2017; 97: 243-51.

[37] Peppas NA, Merrill EW. Crosslinked poly(vinyl alcohol) hydrogels as swollen elastic networks. J Appl Polym Sci. 1977; 21: 1763-70.

[38] Takallu S, Mirzaei E, Azadi A, Karimizade A, Tavakol S. Plate-shape carbonated hydroxyapatite/collagen nanocomposite hydrogel via in situ mineralization of hydroxyapatite concurrent with gelation of collagen at pH = 7.4 and 37°C. J Biomed Mater Res B Appl Biomater. 2019; 107: 1920-9.

[39] Lai VK, Nedrelow DS, Lake SP, Kim B, Weiss EM, Tranquillo RT, et al. Swelling of Collagen-Hyaluronic Acid Co-Gels: An In Vitro Residual Stress Model. Ann Biomed Eng. 2016; 44: 2984-93.

[40] Tonndorf R, Gossla E, Aibibu D, Lindner M, Gelinsky M, Cherif C. Wet spinning and riboflavin crosslinking of collagen type I/III filaments. Biomed Mater. 2018; 14: 015007.

[41] Monfregola L, Bugatti V, Amodeo P, De Luca S, Vittoria V. Physical and water sorption properties of chemically modified pectin with an environmentally friendly process. Biomacromolecules. 2011; 12: 2311-8.

[42] Grover CN, Gwynne JH, Pugh N, Hamaia S, Farndale RW, Best SM, et al. Crosslinking and composition influence the surface properties, mechanical stiffness and cell reactivity of collagen-based films. Acta Biomater. 2012; 8: 3080-90.

[43] Madaghiele M, Calò E, Salvatore L, Bonfrate V, Pedone D, Frigione M, et al. Assessment of collagen crosslinking and denaturation for the design of regenerative scaffolds. J Biomed Mater Res A. 2016; 104: 186-94.

[44] Harley BA, Leung JH, Silva ECCM, Gibson LJ. Mechanical characterization of collagen-glycosaminoglycan scaffolds. Acta Biomater. 2007; 3: 463-74.

[45] Xu Y, Šavija B. 3D auxetic cementitious-polymeric composite structure with compressive strain-hardening behavior. Eng Struct. 2023; 294: 116734.

[46] Xu D, Craig SL. Strain Hardening and Strain Softening of Reversibly Cross-linked Supramolecular Polymer Networks. Macromolecules. 2011; 44: 7478-88.

[47] Liao T, Yang X, Zhao X, Tang Y, Jiang Z, Men Y. Gaussian and Non-Gaussian Distributions of Fracture Properties in Tensile Stretching of High-Density Polyethylene. Macromolecules. 2021; 54: 8860-74.

[48] Zhao JH, Xie ZL, Zhong T, Sun T, Fezzaa K, Cai Y, et al. Strain rate effects on the mechanical behavior of porous titanium with different pore sizes. Mater Sci Eng A. 2021; 821: 141593.

[49] Pissis P, Kyritsis A. Hydration studies in polymer hydrogels. J Polym Sci Part B Polym Phys. 2013; 51: 159-75.

[50] Huang Y, Li X, Lu Z, Zhang H, Huang J, Yan K, et al. Nanofiber-reinforced bulk hydrogel: preparation and structural, mechanical, and biological properties. J Mater Chem B. 2020; 8: 9794-803.

[51] Suchý T, Šupová M, Bartoš M, Sedláček R, Piola M, Soncini M, et al. Dry versus hydrated collagen scaffolds: are dry states representative of hydrated states? J Mater Sci Mater Med. 2018; 29: 20.

[52] Varley MC, Neelakantan S, Clyne TW, Dean J, Brooks RA, Markaki AE. Cell structure, stiffness and permeability of freeze-dried collagen scaffolds in dry and hydrated states. Acta Biomater. 2016; 33: 166-75.

[53] Davidenko N, Schuster CF, Bax DV, Raynal N, Farndale RW, Best SM, et al. Control of crosslinking for tailoring collagen-based scaffolds stability and mechanics. Acta Biomater. 2015; 25: 131-42.

[54] Wang X, Li Q, Hu X, Ma L, You C, Zheng Y, et al. Fabrication and characterization of poly(L-lactide-co-glycolide) knitted mesh-reinforced collagen-chitosan hybrid scaffolds for dermal tissue engineering. J Mech Behav Biomed Mater. 2012; 8: 204-15.

[55] Xu S, Zhou Z, Liu Z, Sharma P. Concurrent stiffening and softening in hydrogels under dehydration. Sci Adv. 2023; 9: eade3240.

[56] Li Z, Liu Z, Ng TY, Sharma P. The effect of water content on the elastic modulus and fracture energy of hydrogel. Extrem Mech Lett. 2020; 35: 100617.

[57] Pelech AN. Murmurs. In: Kliegman RM, Lye PS, Bordini BJ, Toth H, Basel D, editors. Nelson Pediatric Symptom-Based Diagnosis. Elsevier; 2018. pp. 116-43.e2.

[58] Calvert JW, Lefer DJ. Chapter 6 - Overview of Cardiac Muscle Physiology. In: Hill JA, Olson EN, editors. Academic Press; 2012. pp. 57-66.

[59] Butcher JT, Nerem RM. Valvular endothelial cells and the mechanoregulation of valvular pathology. Philos Trans R Soc Lond B Biol Sci. 2007; 362: 1445-57.

[60] Salem AK, Stevens R, Pearson RG, Davies MC, Tendler SJB, Roberts CJ, et al. Interactions of 3T3 fibroblasts and endothelial cells with defined pore features. J Biomed Mater Res. 2002; 61: 212-7.

[61] Yannas IV, Lee E, Orgill DP, Skrabut EM, Murphy GF. Synthesis and characterization of a model extracellular matrix that induces partial regeneration of adult mammalian skin. Proc Natl Acad Sci U S A. 1989; 86: 933-7.

[62] Gezmis H. The influence of collagen-based scaffolds on cardiac differentiation and endothelial growth [dissertation]. Oxford: Oxford University; 2023.

[63] Cao R, Xu Y, Xu Y, Brand DD, Zhou G, Xiao K, et al. Development of Tri-Layered Biomimetic Atelocollagen Scaffolds with Interfaces for Osteochondral Tissue Engineering. Adv Healthc Mater. 2022; 11: e2101643.

[64] Brazile B, Wang B, Wang G, Bertucci R, Prabhu R, Patnaik SS, et al. On the bending properties of porcine mitral, tricuspid, aortic, and pulmonary valve leaflets. J Long Term Eff Med Implants. 2015; 25: 41-53.

[65] Lam TV. The Mechanical Properties of Native Porcine Aortic and Pulmonary Heart Valve Leaflets [dissertation]. Pittsburgh: University of Pittsburgh; 2005.