Although skeletal muscle has a high potential for self-repair, volumetric muscle loss can result in impairment beyond the endogenous regenerative capacity. There is a clinical need to improve on current clinical treatments that fail to fully restore the structure and function of lost muscle. Decellularized extracellular matrix (dECM) scaffolds have been an attractive platform for regenerating skeletal muscle, as dECM contains many biochemical cues that aid in cell adhesion, proliferation, and differentiation. However, there is limited capacity to tune physicochemical properties in current dECM technologies to improve outcome. In this study, we aim to create a novel, high-throughput technique to fabricate dECM scaffolds with tunable physicochemical properties while retaining proregenerative matrix components. We demonstrate a successful decellularization protocol that effectively removes DNA. We also identified key steps for the successful production of electrospun muscle dECM without the use of a carrier polymer; electrospinning allows for rapid scaffold fabrication with high control over material properties, which can be optimized to mimic native muscle. To this end, fiber orientation and degree of crosslinking of these dECM scaffolds were modulated and the corollary effects on fiber swelling, mechanical properties, and degradation kinetics were investigated. Beyond application in skeletal muscle, the versatility of this technology has the potential to serve as a foundation for dECM scaffold fabrication in a variety of tissue engineering applications.
Impact Statement
This study develops a method to decellularize skeletal muscle and fabricate electrospun scaffolds from the decellularized tissue without the need for a carrier polymer. In addition, the resulting scaffolds have tunable physicochemical properties, including fiber alignment, while retaining important extracellular matrix components for regeneration of skeletal muscle.
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References
1.
TurnerN.J., and BadylakS.F.Regeneration of skeletal muscle. Cell Tissue Res, 347, 759, 2012.
2.
DzikiJ., BadylakS., YabroudiM., et al.An acellular biologic scaffold treatment for volumetric muscle loss: results of a 13-patient cohort study. NPJ Regen Med, 1, 16008, 2016.
3.
GroganB.F., and HsuJ.R.Volumetric muscle loss. J Am Acad Orthop Surg, 19Suppl 1, S35, 2011.
4.
WuX., CoronaB.T., ChenX., and WaltersT.J.A standardized rat model of volumetric muscle loss injury for the development of tissue engineering therapies. Biores Open Access, 1, 280, 2012.
5.
TzuM., LiA., WillettN.J., UhrigB.A., GuldbergR.E., and WarrenG.L.Functional analysis of limb recovery following autograft treatment of volumetric muscle loss in the quadriceps femoris. J Biomech, 47, 2013, 2014.
6.
ShahK., MajeedZ., JonasonJ., and O'KeefeR.J.The role of muscle in bone repair: the cells, signals, and tissue responses to injury. Curr Osteoporos Rep, 11, 130, 2013.
7.
DavisK.M., GriffinK.S., ChuT.G., et al.Muscle-bone interactions during fracture healing. J Musculoskelet Neuronal Interact, 15, 1, 2015.
8.
LiaoJ., GuoX., Grande-AllenK.J., KasperF.K., and MikosA.G.Bioactive polymer/extracellular matrix scaffolds fabricated with a flow perfusion bioreactor for cartilage tissue engineering. Biomaterials, 31, 8911, 2010.
9.
SmoakM.M., PearceH.A., and MikosA.G.Microfluidic devices for disease modeling in muscle tissue. Biomaterials, 198, 250, 2019.
10.
FuY., FanX., TianC., et al.Decellularization of porcine skeletal muscle extracellular matrix for the formulation of a matrix hydrogel: a preliminary study. J Cell Mol Med, 20, 740, 2016.
11.
RaoN., AgmonG., TierneyM.T., et al.Engineering an injectable muscle-specific microenvironment for improved cell delivery using a nanofibrous extracellular matrix hydrogel. ACS Nano, 11, 3851, 2017.
12.
UngerleiderJ.L., JohnsonT.D., RaoN., and ChristmanK.L.Fabrication and characterization of injectable hydrogels derived from decellularized skeletal and cardiac muscle. Methods, 84, 53, 2015.
13.
WassenaarJ.W., BradenR.L., OsbornK.G., and ChristmanK.L.Modulating in vivo degradation rate of injectable extracellular matrix hydrogels. J Mater Chem B, 4, 2794, 2016.
14.
ChoiY.-J., KimT.G., JeongJ., YiH.-G., ParkJ.W., HwangW., and ChoD.-W.3D cell printing of functional skeletal muscle constructs using skeletal muscle-derived bioink. Adv Healthc Mater, 5, 2636, 2016.
15.
HindererS., LaylandS.L., and Schenke-LaylandK.ECM and ECM-like materials - Biomaterials for applications in regenerative medicine and cancer therapy. Adv Drug Deliv Rev, 97, 260, 2016.
16.
WolfM.T., DalyK.A., ReingJ.E., and BadylakS.F.Biologic scaffold composed of skeletal muscle extracellular matrix. Biomaterials, 33, 2916, 2012.
17.
SicariB.M., RubinJ.P., DearthC.L., et al.An acellular biologic scaffold promotes skeletal muscle formation in mice and humans with volumetric muscle loss. Sci Transl Med, 6, 234ra58, 2014.
18.
AgmonG., and ChristmanK.L.Controlling stem cell behavior with decellularized extracellular matrix scaffolds. Curr Opin Solid State Mater Sci, 20, 193, 2016.
19.
RothrauffB.B., YangG., and TuanR.S.Tissue-specific bioactivity of soluble tendon-derived and cartilage-derived extracellular matrices on adult mesenchymal stem cells. Stem Cell Res Ther, 8, 133, 2017.
20.
SarafA., BaggettL.S., RaphaelR.M., KasperF.K., and MikosA.G.Regulated non-viral gene delivery from coaxial electrospun fiber mesh scaffolds. J Control Release, 143, 95, 2010.
21.
PhamQ.P., SharmaU., and MikosA.G.Electrospun poly(ɛ-caprolactone) microfiber and multilayer nanofiber/microfiber scaffolds: characterization of scaffolds and measurement of cellular infiltration. Biomacromolecules, 7, 2796, 2006.
22.
DeitzelJ., KleinmeyerJ., HarrisD., and Beck TanN.The effect of processing variables on the morphology of electrospun nanofibers and textiles. Polymer (Guildf), 42, 261, 2001.
23.
GongW., LeiD., LiS., et al.Hybrid small-diameter vascular grafts: anti-expansion effect of electrospun poly ɛ-caprolactone on heparin-coated decellularized matrices. Biomaterials, 76, 359, 2016.
24.
KishanA.P., NezaratiR.M., RadzickiC.M., et al.In situ crosslinking of electrospun gelatin for improved fiber morphology retention and tunable degradation. J Mater Chem B, 3, 7930, 2015.
25.
LahaA., SharmaC.S., and MajumdarS.Sustained drug release from multi-layered sequentially crosslinked electrospun gelatin nanofiber mesh. Mater Sci Eng C, 76, 782, 2017.
26.
KoJ.H., YinH., AnJ., et al.Characterization of cross-linked gelatin nanofibers through electrospinning. Macromol Res, 18, 137, 2010.
27.
FrancisM.P., SachsP.C., MadurantakamP.A., et al.Electrospinning adipose tissue-derived extracellular matrix for adipose stem cell culture. J Biomed Mater Res A, 100, 1716, 2012.
28.
BaigueraS., Del GaudioC., LucatelliE., et al.Electrospun gelatin scaffolds incorporating rat decellularized brain extracellular matrix for neural tissue engineering. Biomaterials, 35, 1205, 2014.
29.
LevorsonE.J., Raman SreerekhaP., ChennazhiK.P., KasperF.K., NairS.V., and MikosA.G.Fabrication and characterization of multiscale electrospun scaffolds for cartilage regeneration. Biomed Mater, 8, 14103, 2013.
30.
ZhongS., TeoW.E., ZhuX., BeuermanR.W., RamakrishnaS., and YungL.Y.L.An aligned nanofibrous collagen scaffold by electrospinning and its effects on in vitro fibroblast culture. J Biomed Mater Res A, 79, 456, 2006.
SchoenB., AvrahamiR., BaruchL., et al.Electrospun extracellular matrix: paving the way to tailor-made natural scaffolds for cardiac tissue regeneration. Adv Funct Mater, 27, 1700427, 2017.
33.
CoenenA.M.J., BernaertsK.V, HaringsJ.A.W., JockenhoevelS., and GhazanfariS.Elastic materials for tissue engineering applications: natural, synthetic, and hybrid polymers. Acta Biomater, 79, 60, 2018.
34.
DahlinR.L., NiM., MeretojaV.V, KasperF.K., and MikosA.G.TGF-β3-induced chondrogenesis in co-cultures of chondrocytes and mesenchymal stem cells on biodegradable scaffolds. Biomaterials, 35, 123, 2014.
35.
StegemannH., and StalderK.Determination of hydroxyproline. Clin Chim Acta, 18, 267, 1967.
36.
SingelynJ.M., and ChristmanK.L.Modulation of material properties of a decellularized myocardial matrix scaffold. Macromol Biosci, 11, 731–738, 2011.
37.
KammounM., PouletautP., CanonF., et al.Impact of TIEG1 deletion on the passive mechanical properties of fast and slow twitch skeletal muscles in female mice. PLoS One, 11, e0164566, 2016.
38.
ReingJ.E., BrownB.N., DalyK.A., et al.The effects of processing methods upon mechanical and biologic properties of porcine dermal extracellular matrix scaffolds. Biomaterials, 31, 8626, 2010.
39.
KwonI.K., and MatsudaT.Co-Electrospun nanofiber fabrics of Poly(l-lactide-co-ɛ-caprolactone) with type I collagen or heparin. Biomacromolecules, 6, 2096, 2005.
40.
StitzelJ., LiuJ., LeeS.J., et al.Controlled fabrication of a biological vascular substitute. Biomaterials, 27, 1088, 2006.
41.
StankusJ.J., GuanJ., and WagnerW.R.Fabrication of biodegradable elastomeric scaffolds with sub-micron morphologies. J Biomed Mater Res A, 70, 603, 2004.
42.
CrapoP.M., GilbertT.W., and BadylakS.F.An overview of tissue and whole organ decellularization processes. Biomaterials, 32, 3233, 2011.
43.
BadylakS.F., DzikiJ.L., SicariB.M., AmbrosioF., BoningerM.L.Mechanisms by which acellular biologic scaffolds promote functional skeletal muscle restoration. Biomaterials, 103, 128, 2016.
44.
BraunT., and GautelM.Transcriptional mechanisms regulating skeletal muscle differentiation, growth and homeostasis. Nat Rev Mol Cell Biol, 12, 349, 2011.
45.
VlodavskyI., FolkmanJ., SullivanR., FridmanR., Ishai-MichaeliR., SasseJ.and Klagsbrun, M. Endothelial cell-derived basic fibroblast growth factor: synthesis and deposition into subendothelial extracellular matrix. Proc Natl Acad Sci USA, 84, 2292, 1987.
46.
VlodavskyI., MiaoH.-Q., MedalionB., DanagherP., and RonD.Involvement of heparan sulfate and related molecules in sequestration and growth promoting activity of fibroblast growth factor. Cancer Metastasis Rev, 15, 177, 1996.
47.
PhamQ.P., SharmaU., and MikosA.G.Electrospinning of polymeric nanofibers for tissue engineering applications: a review. Tissue Eng, 12, 1197, 2006.
48.
QaziT.H., MooneyD.J., PumbergerM., GeißlerS., and DudaG.N.Biomaterials based strategies for skeletal muscle tissue engineering: existing technologies and future trends. Biomaterials, 53, 502, 2015.
49.
MertensJ.P., SuggK.B., LeeJ.D., and LarkinL.M.Engineering muscle constructs for the creation of functional engineered musculoskeletal tissue. Regen Med, 9, 89, 2014.
50.
PatelZ.S., UedaH., YamamotoM., TabataY., and MikosA.G.In vitro and in vivo release of vascular endothelial growth factor from gelatin microparticles and biodegradable composite scaffolds. Pharm Res, 25, 2370, 2008.
51.
ButtafocoL., KolkmanN.G., Engbers-BuijtenhuijsP., et al.Electrospinning of collagen and elastin for tissue engineering applications. Biomaterials, 27, 724, 2006.
52.
OrrS.B., ChainaniA., HippensteelK.J., et al.Aligned multilayered electrospun scaffolds for rotator cuff tendon tissue engineering. Acta Biomater, 24, 117, 2015.
53.
ChenF., SuY., MoX., HeC., WangH., and IkadaY.Biocompatibility, alignment degree and mechanical properties of an electrospun chitosan-P(LLA-CL) fibrous scaffold. J Biomater Sci Polym Ed, 20, 2117, 2009.
54.
ZhuB., LiW., ChiN., LewisR.V., OsamorJ., and WangR.Optimization of glutaraldehyde vapor treatment for electrospun collagen/silk tissue engineering scaffolds. ACS Omega, 2, 2439, 2017.
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