Volumetric muscle loss (VML) is characterized as the loss of muscle tissue that exceeds the muscle’s self-repair mechanism, resulting in incomplete restoration of muscle mass and function. Existing treatment modalities, including muscle grafts or autologous muscle transfers, are limited by constraints such as tissue availability and donor site morbidity. Moreover, the inadequate recovery of muscle may lead to fibrosis within the VML site, impeding the process of muscle regeneration and resulting in permanent deficits. Emerging therapeutics, such as hydrogels, show promise in addressing the limitations of current therapeutics and have the potential to significantly reduce fibrosis and facilitate the restoration of muscle form and function following VML injury and repair. This study evaluated the therapeutic potential of repairing a 30% VML injury in the rat tibialis anterior muscle with engineered skeletal muscle units (SMUs), alone, and in combination with a hyaluronic acid-based hydrogel (HyA-HG). Following 1- or 3-months post-implantation, muscle structure and function were assessed. The results indicated that the incorporation of HyA-HG in combination with our SMUs resulted in improvements in force production for VML injuries repaired for 1 month. However, over extended recovery periods (3 months), sustained superior improvements in muscle function with the combination therapy were not observed compared with the repair with just an SMU. Moreover, histological analyses revealed that muscle treated with SMUs and HyA-HG exhibited a greater cross-sectional area and force production in the early stages of recovery (1-month post-surgery) compared with untreated VML sites or those treated with HyA-HG only. However, after 3 months, muscle mass and force production in all experimental groups reached comparable levels, suggesting a transient benefit of the combination therapy. Our findings highlight the potential of HyA-HG and SMU combination therapy to enhance early functional recovery following VML.
Impact Statement
Volumetric muscle loss (VML) is a clinically relevant problem that faces significant limitations in effective treatment methods. Tissue-engineered skeletal muscle and hydrogels are promising therapeutic options. This study aimed to assess the therapeutic potential of a hyaluronic acid-based hydrogel in combination with our tissue-engineered skeletal muscle units for treating VML in a rat model following 1-month and 3-month recovery periods.
CoronaBT, RiveraJC, OwensJG, et al.Volumetric muscle loss leads to permanent disability following extremity trauma. J Rehabil Res Dev, 2015; 52(7):785–792; doi: 10.1682/JRRD.2014.07.0165
2.
GargK, WardCL, HurtgenBJ, et al.Volumetric muscle loss: Persistent functional deficits beyond frank loss of tissue. J Orthop Res, 2015; 33(1):40–46; doi: 10.1002/jor.22730
3.
TestaS, FornettiE, FuocoC, et al.The War after War: Volumetric muscle loss incidence, implication, current therapies and emerging reconstructive strategies, a comprehensive review. Biomedicines, 2021; 9(5):564; doi: 10.3390/biomedicines9050564
4.
CoronaBT, WenkeJC, WardCL. Pathophysiology of volumetric muscle loss injury. Cells Tissues Organs, 2016; 202(3–4):180–188; doi: 10.1159/000443925
5.
GroganBF, HsuJR, Skeletal Trauma Research C. Volumetric muscle loss. J Am Acad Orthop Surg, 2011; 19(Suppl 1):S35–S37; doi: 10.5435/00124635-201102001-00007
6.
GreisingSM, RiveraJC, GoldmanSM, et al.Unwavering pathobiology of volumetric muscle loss injury. Sci Rep, 2017; 7(1):13179; doi: 10.1038/s41598-017-13306-2
7.
LiuJ, SaulD, BokerKO, et al.Current methods for skeletal muscle tissue repair and regeneration. Biomed Res Int, 2018; 2018:1984879; doi: 10.1155/2018/1984879
8.
LakhianiC, DeFazioMV, HanK, et al.Donor-site morbidity following free tissue harvest from the thigh: A systematic review and pooled analysis of complications. J Reconstr Microsurg, 2016; 32(5):342–357; doi: 10.1055/s-0036-1583301
9.
BasurtoIM, PassipieriJA, GardnerGM, et al.Photoreactive hydrogel stiffness influences volumetric muscle loss repair. Tissue Eng Part A, 2022; 28(7–8):312–329; doi: 10.1089/ten.TEA.2021.0137
10.
DienesJ, BrowneS, FarjunB, et al.Semisynthetic hyaluronic acid-based hydrogel promotes recovery of the injured tibialis anterior skeletal muscle form and function. ACS Biomater Sci Eng, 2021; 7(4):1587–1599; doi: 10.1021/acsbiomaterials.0c01751
11.
Silva GarciaJM, PanitchA, CalveS. Functionalization of hyaluronic acid hydrogels with ECM-derived peptides to control myoblast behavior. Acta Biomater, 2019; 84:169–179; doi: 10.1016/j.actbio.2018.11.030
12.
PassipieriJA, BakerHB, SiriwardaneM, et al.Keratin hydrogel enhances in vivo skeletal muscle function in a rat model of volumetric muscle loss. Tissue Eng Part A, 2017; 23(11–12):556–571; doi: 10.1089/ten.TEA.2016.0458
13.
BakerHB, PassipieriJA, SiriwardaneM, et al.Cell and growth factor-loaded keratin hydrogels for treatment of volumetric muscle loss in a mouse model. Tissue Eng Part A, 2017; 23(11–12):572–584; doi: 10.1089/ten.TEA.2016.0457
14.
DickerKT, GurskiLA, Pradhan-BhattS, et al.Hyaluronan: A simple polysaccharide with diverse biological functions. Acta Biomater, 2014; 10(4):1558–1570; doi: 10.1016/j.actbio.2013.12.019
15.
XuX, JhaAK, HarringtonDA, et al.Hyaluronic acid-based hydrogels: From a natural polysaccharide to complex networks. Soft Matter, 2012; 8(12):3280–3294; doi: 10.1039/C2SM06463D
16.
GriecoM, UrsiniO, PalamaIE, et al.HYDRHA: Hydrogels of hyaluronic acid. New biomedical approaches in cancer, neurodegenerative diseases, and tissue engineering. Mater Today Bio, 2022; 17:100453; doi: 10.1016/j.mtbio.2022.100453
17.
VanDusenKW, SyverudBC, WilliamsML, et al.Engineered skeletal muscle units for repair of volumetric muscle loss in the tibialis anterior muscle of a rat. Tissue Eng Part A, 2014; 20(21–22):2920–2930; doi: 10.1089/ten.TEA.2014.0060
18.
NutterGP, VanDusenKW, FloridaSE, et al.The effects of engineered skeletal muscle on volumetric muscle loss in the tibialis anterior of rat after three months in vivo. Regen Eng Transl Med, 2020; 6(4):365–372; doi: 10.1007/s40883-020-00175-x
19.
RodriguezBL, FloridaSE, VanDusenKW, et al.The maturation of tissue-engineered skeletal muscle units following 28-day ectopic implantation in a rat. Regen Eng Transl Med, 2019; 5(1):86–94; doi: 10.1007/s40883-018-0078-7
20.
NovakovaSS, RodriguezBL, Vega-SotoEE, et al.Repairing volumetric muscle loss in the ovine peroneus tertius following a 3-month recovery. Tissue Eng Part A, 2020; 26(15–16):837–851; doi: 10.1089/ten.TEA.2019.0288
21.
RodriguezBL, NovakovaSS, Vega-SotoEE, et al.Repairing volumetric muscle loss in the ovine peroneus tertius following a 6-month recovery. Tissue Eng Part A, 2022; 28(13–14):606–620; doi: 10.1089/ten.TEA.2021.0187
22.
SuEY, KennedyCS, Vega-SotoEE, et al.Repairing volumetric muscle loss with commercially available hydrogels in an ovine model. Tissue Eng Part A, 2024; 30(9–10):440–453; doi: 10.1089/ten.TEA.2023.0240
23.
MisraS, HascallVC, MarkwaldRR, et al.Interactions between hyaluronan and its receptors (CD44, RHAMM) regulate the activities of inflammation and cancer. Front Immunol, 2015; 6:201; doi: 10.3389/fimmu.2015.00201
24.
LengY, AbdullahA, WendtMK, et al.Hyaluronic acid, CD44 and RHAMM regulate myoblast behavior during embryogenesis. Matrix Biol, 2019; 78–79:236–254; doi: 10.1016/j.matbio.2018.08.008
25.
MarinhoA, NunesC, ReisS. Hyaluronic acid: A key ingredient in the therapy of inflammation. Biomolecules, 2021; 11(10):1518; doi: 10.3390/biom11101518
26.
Vega-SotoEE, RodriguezBL, ArmstrongRE, et al.A 30% volumetric muscle loss does not result in sustained functional deficits after a 90-day recovery in rats. Regen Eng Transl Med, 2020; 6(1):62–68; doi: 10.1007/s40883-019-00117-2
27.
Institute of Laboratory Animal Resources (U.S.). Committee on Care and Use of Laboratory Animals. Guide for the care and use of laboratory animals. U.S. Dept. of Health and Human Services, Public Health Service: Bethesda, MD; 2011.
28.
WilliamsML, KostrominovaTY, ArrudaEM, et al.Effect of implantation on engineered skeletal muscle constructs. J Tissue Eng Regen Med, 2013; 7(6):434–442; doi: 10.1002/term.537
29.
WeistMR, WellingtonMS, BermudezJE, et al.TGF-beta1 enhances contractility in engineered skeletal muscle. J Tissue Eng Regen Med, 2013; 7(7):562–571; doi: 10.1002/term.551
30.
HarbersGM, GambleLJ, IrwinEF, et al.Development and characterization of a high-throughput system for assessing cell-surface receptor-ligand engagement. Langmuir, 2005; 21(18):8374–8384; doi: 10.1021/la050396y
31.
HarbersGM, HealyKE. The effect of ligand type and density on osteoblast adhesion, proliferation, and matrix mineralization. J Biomed Mater Res A, 2005; 75(4):855–869; doi: 10.1002/jbm.a.30482
32.
RezaniaA, HealyKE. Biomimetic peptide surfaces that regulate adhesion, spreading, cytoskeletal organization, and mineralization of the matrix deposited by osteoblast-like cells. Biotechnol Prog, 1999; 15(1):19–32; doi: 10.1021/bp980083b
33.
JhaAK, MathurA, SvedlundFL, et al.Molecular weight concentration of heparin in hyaluronic acid-based matrices modulates growth factor retention kinetics and stem cell fate. J Control Release, 2015; 209:308–316; doi: 10.1016/j.jconrel.2015.04.034
34.
JhaAK, TharpKM, YeJ, et al.Enhanced survival and engraftment of transplanted stem cells using growth factor sequestering hydrogels. Biomaterials, 2015; 47:1–12; doi: 10.1016/j.biomaterials.2014.12.043
35.
JhaAK, TharpKM, BrowneS, et al.Matrix metalloproteinase-13 mediated degradation of hyaluronic acid-based matrices orchestrates stem cell engraftment through vascular integration. Biomaterials, 2016; 89:136–147; doi: 10.1016/j.biomaterials.2016.02.023
36.
BrowneS, HossainyS, HealyK. Hyaluronic acid macromer molecular weight dictates the biophysical properties and in vitro cellular response to semisynthetic hydrogels. ACS Biomater Sci Eng, 2020; 6(2):1135–1143; doi: 10.1021/acsbiomaterials.9b01419
37.
LarkinLM, DavisCS, Sims-RobinsonC, et al.Skeletal muscle weakness due to deficiency of CuZn-superoxide dismutase s associated with loss of functional innervation. Am J Physiol Regul Integr Comp Physiol, 2011; 301(5):R1400–R1407; doi: 10.1152/ajpregu.00093.2011
38.
BurkholderTJ, FingadoB, BaronS, et al.Relationship between muscle-fiber types and sizes and muscle architectural properties in the mouse hindlimb. J Morphol, 1994; 221(2):177–190; doi: 10.1002/jmor.1052210207
39.
LieberRL, WardSR. Cellular mechanisms of tissue fibrosis. 4. Structural and functional consequences of skeletal muscle fibrosis. Am J Physiol Cell Physiol, 2013; 305(3):C241–C252; doi: 10.1152/ajpcell.00173.2013
40.
JarvinenTA, JarvinenTL, KaariainenM, et al.Muscle injuries: Optimising recovery. Best Pract Res Clin Rheumatol, 2007; 21(2):317–331; doi: 10.1016/j.berh.2006.12.004
41.
HuardJ, LiY, FuFH. Muscle injuries and repair: Current trends in research. J Bone Joint Surg Am, 2002; 84(5):822–832.
42.
BasurtoIM, MuhammadSA, GardnerGM, et al.Controlling scaffold conductivity and pore size to direct myogenic cell alignment and differentiation. J Biomed Mater Res A, 2022; 110(10):1681–1694; doi: 10.1002/jbm.a.37418
43.
PfaffMR, WagueA, DaviesM, et al.Viscoelastic HyA hydrogel promotes recovery of muscle quality and vascularization in a murine model of delayed rotator cuff repair. Adv Healthc Mater, 2025; 14(10):e2403962; doi: 10.1002/adhm.202403962
44.
ReveteA, AparicioA, CisternaBA, et al.Advancements in the use of hydrogels for regenerative medicine: Properties and biomedical applications. Int J Biomater, 2022; 2022:3606765; doi: 10.1155/2022/3606765
45.
HighleyCB, PrestwichGD, BurdickJA. Recent advances in hyaluronic acid hydrogels for biomedical applications. Curr Opin Biotechnol, 2016; 40:35–40; doi: 10.1016/j.copbio.2016.02.008
46.
CoronaBT, RiveraJC, DalskeKA, et al.Pharmacological mitigation of fibrosis in a porcine model of volumetric muscle loss injury. Tissue Eng Part A, 2020; 26(11–12):636–646; doi: 10.1089/ten.TEA.2019.0272
47.
De PaolisF, TestaS, GuarnacciaG, et al.Long-term longitudinal study on swine VML model. Biol Direct, 2023; 18(1):42; doi: 10.1186/s13062-023-00399-1
48.
GreisingSM, CoronaBT, McGannC, et al.Therapeutic approaches for volumetric muscle loss injury: A systematic review and meta-analysis. Tissue Eng Part B Rev, 2019; 25(6):510–525; doi: 10.1089/ten.TEB.2019.0207
49.
DolanCP, DearthCL, CoronaBT, et al.Retrospective characterization of a rat model of volumetric muscle loss. BMC Musculoskelet Disord, 2022; 23(1):814; doi: 10.1186/s12891-022-05760-5
50.
LangridgeB, GriffinM, ButlerPE. Regenerative medicine for skeletal muscle loss: A review of current tissue engineering approaches. J Mater Sci Mater Med, 2021; 32(1):15; doi: 10.1007/s10856-020-06476-5
