Cell-based meat (CBM) production is a promising technology that could generate meat without the need of animal agriculture. The generation of tissue requires a three-dimensional (3D) scaffold to provide support to the cells and mimic the extracellular matrix (ECM). For CBM, the scaffold needs to be edible and have suitable nutritional value and texture. Here, we demonstrate the use of textured soy protein—an edible porous protein-based biomaterial—as a novel CBM scaffold that can support cell attachment and proliferation to create a 3D engineered bovine muscle tissue. The media composition was optimized for 3D bovine satellite cell (BSC) proliferation and differentiation by adding myogenic-related growth factors. Myogenesis of several cell combinations was compared, and elevated myogenesis and ECM deposition were shown in co-culture of BSCs with bovine smooth muscle cells and tri-cultures of BSCs, bovine smooth muscle cells and bovine endothelial cells. The expression of proteins associated with ECM gene sets was increased in the co-culture compared with BSC monoculture. Volunteers tasted the product after cooking and noted its meaty flavour and sensorial attributes, achieving the goal of replicating the sensation and texture of a meat bite. This approach represents a step forward for the applied production of CBM as a food product.
This is a preview of subscription content
Subscribe to Nature+
Get immediate online access to the entire Nature family of 50+ journals
Subscribe to Journal
Get full journal access for 1 year
only $9.92 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Tax calculation will be finalised during checkout.
Get time limited or full article access on ReadCube.
All prices are NET prices.
The data that support the findings of this study are available from the corresponding author upon request.
Post, M. & Weele, C. Principles of Tissue Engineering for Food (Elsevier, 2014).
Slade, P. If you build it, will they eat it? Consumer preferences for plant-based and cultured meat burgers. Appetite 125, 428–437 (2018).
Ben-Arye, T. & Levenberg, S. Tissue engineering for clean meat production. Front. Sustain. Food Syst. 3, 46 (2019).
Specht, E. A., Welch, D. R., Rees Clayton, E. M. & Lagally, C. D. Opportunities for applying biomedical production and manufacturing methods to the development of the clean meat industry. Biochem. Eng. J. 132, 161–168 (2018).
Edelman, P. D., McFarland, D. C., Mironov, V. A. & Matheny, J. G. Commentary: in vitro-cultured meat production. Tissue Eng. 11, 659–662 (2005).
Egozi, D. et al. Engineered vascularized muscle flap. J. Vis. Exp. 107, 52984 (2016).
Gholobova, D. et al. Endothelial network formation within human tissue-engineered skeletal muscle. Tissue Eng. Part A 21, 2548–2558 (2015).
Levenberg, S. Engineering blood vessels from stem cells: recent advances and applications. Curr. Opin. Biotechnol. 16, 516–523 (2005).
Shandalov, Y. et al. An engineered muscle flap for reconstruction of large soft tissue defects. Proc. Natl Acad. Sci. USA 111, 6010–6015 (2014).
Listrat, A. et al. How muscle structure and composition influence meat and flesh quality. Sci. World J. 2016, 3182746 (2016).
Vitello, L. et al. Enhancing myoblast proliferation by using myogenic factors: a promising approach for improving fiber regeneration in sport medicine and skeletal muscle diseases. Basic Appl. Myol. 14, 45–51 (2004).
Yin, H., Price, F. & Rudnicki, M. A. Satellite cells and the muscle stem cell niche. Physiol. Rev. 93, 23–67 (2013).
Purslow, P. P. Muscle fascia and force transmission. J. Bodyw. Mov. Ther. 14, 411–417 (2010).
Jockenhoevel, S. et al. Fibrin gel—advantages of a new scaffold in cardiovascular tissue engineering. Eur. J. Cardiothorac. Surg. 19, 424–430 (2001).
Guo, B. et al. Transcriptome analysis of cattle muscle identifies potential markers for skeletal muscle growth rate and major cell types. BMC Genomics 16, 177 (2015).
Jain, R. K., Au, P., Tam, J., Duda, D. G. & Fukumura, D. Engineering vascularized tissue. Nat. Biotechnol. 23, 821–823 (2005).
Christov, C. et al. Muscle satellite cells and endothelial cells: close neighbors and privileged partners. Mol. Biol. Cell 18, 1397–1409 (2007).
Butler, J. M., Kobayashi, H. & Rafii, S. Instructive role of the vascular niche in promoting tumour growth and tissue repair by angiocrine factors. Nat. Rev. Cancer 10, 138–146 (2010).
Rafii, S., Butler, J. M. & Ding, B.-S. Angiocrine functions of organ-specific endothelial cells. Nature 529, 316–325 (2016).
Kyriakopoulou, K., Dekkers, B. & van der Goot, A. J. in Sustainable Meat Production and Processing (ed. Galanakis, C. M.) 103–126 (Academic Press, 2019).
Day, L. Proteins from land plants—potential resources for human nutrition and food security. Trends Food Sci. Technol. 32, 25–42 (2013).
Zeltinger, J., Sherwood, J. K., Graham, D. A., Müeller, R. & Griffith, L. G. Effect of pore size and void fraction on cellular adhesion, proliferation, and matrix deposition. Tissue Eng. 7, 557–572 (2001).
Hayes, J. S., Czekanska, E. M. & Richards, R. G. in Tissue Engineering III: Cell-Surface Interactions for Tissue Culture (eds Kasper, C., Witte, F. & Pörtner, R.) 1–31 (Springer, 2012).
Rodriguez, B. L. & Larkin, L. M. in Functional 3D Tissue Engineering Scaffolds (eds Deng, Y. & Kuiper, J.) 279–304 (Woodhead Publishing, 2018).
Choi, J. S., Lee, S. J., Christ, G. J., Atala, A. & Yoo, J. J. The influence of electrospun aligned poly(epsilon-caprolactone)/collagen nanofiber meshes on the formation of self-aligned skeletal muscle myotubes. Biomaterials 29, 2899–2906 (2008).
Aviss, K. J., Gough, J. E. & Downes, S. Aligned electrospun polymer fibres for skeletal muscle regeneration. Eur. Cells Mater. 19, 193–204 (2010).
Levenberg, S. et al. Engineering vascularized skeletal muscle tissue. Nat. Biotechnol. 23, 879–884 (2005).
Perry, L., Landau, S., Flugelman, M. Y. & Levenberg, S. Genetically engineered human muscle transplant enhances murine host neovascularization and myogenesis. Commun. Biol. 1, 161 (2018).
Specht, L. An Analysis of Culture Medium Costs and Production Volumes for Cell-Based Meat (The Good Food Institute, 2019).
Loh, Q. L. & Choong, C. Three-dimensional scaffolds for tissue engineering applications: role of porosity and pore size. Tissue Eng. Part B Rev. 19, 485–502 (2013).
Du, M., Wang, B., Fu, X., Yang, Q. & Zhu, M.-J. Fetal programming in meat production. Meat Sci. 109, 40–47 (2015).
Ding, S. et al. Maintaining bovine satellite cells stemness through p38 pathway. Sci. Rep. 8, 10808 (2018).
Verbruggen, S., Luining, D., van Essen, A. & Post, M. J. Bovine myoblast cell production in a microcarriers-based system. Cytotechnology 70, 503–512 (2018).
Péault, B. et al. Stem and progenitor cells in skeletal muscle development, maintenance, and therapy. Mol. Ther. 15, 867–877 (2007).
Du, M., Huang, Y., Das, A. K., Yang, Q. & Duarte, M. S. Manipulating mesenchymal progenitor cell differentiation to optimize performance and carcass value of beef cattle. J. Anim. Sci. 91, 1419–1427 (2013).
Chapman, M. A., Meza, R. & Lieber, R. L. Skeletal muscle fibroblasts in health and disease. Differentiation 92, 108–115 (2016).
Krieger, J., Park, B.-W., Lambert, C. R. & Malcuit, C. 3D skeletal muscle fascicle engineering is improved with TGF-β1 treatment of myogenic cells and their co-culture with myofibroblasts. PeerJ. 6, e4939 (2018).
Bauman, T. M. et al. Characterization of fibrillar collagens and extracellular matrix of glandular benign prostatic hyperplasia nodules. PLoS ONE 9, e109102 (2014).
Suvik, A. & Effendy, A. W. M. The use of modified Masson’s trichrome staining in collagen evaluation in wound healing study. Mal. J. Vet. Res. 3, 39–47 (2012).
Mehta, F., Theunissen, R. & Post, M. J. in Myogenesis: Methods and Protocols (ed. Rønning, S. B.) 111–125 (Springer, 2019).
Frey, R. S., Johnson, B. J., Hathaway, M. R., White, M. E. & Dayton, W. R. Growth factor responsiveness of primary satellite cell cultures from steers implanted with trenbolone acetate and estradiol-17β. Basic Appl. Myol. 5, 71–79 (1995).
Lapin, M. R., Gonzalez, J. M. & Johnson, S. E. Substrate elasticity affects bovine satellite cell activation kinetics in vitro. J. Anim. Sci. 91, 2083–2090 (2013).
Lu, R., Chen, Y.-R., Solomon, M. B. & Berry, B. W. Tensile properties and Warner–Bratzler tenderness measurement of raw and cooked beef. Trans. ASAE 41, 1431–1439 (1998).
The authors thank M. Hathaway for providing the BSCs and relevant protocols, Y. Dahan for assistance with bovine cell isolation, I. Redenski for support with micro-CT experimental design, the BCF Bioimaging Center, Faculty of Medicine, Technion, for help with micro-CT imaging and analysis, J. Zavin for assistance with cryosectioning, O. Katovitz for assistance with data quantification, I. Michael for assistance with experimental design and Y. Posen for editorial assistance during preparation of this manuscript. The authors thank Technion’s MIKA Center for support with the SEM measurement and Technion’s Smoler Center for support with proteome measurements and analysis. The research was supported by funding from Aleph Farms.
This research was sponsored by Aleph Farms. S.Levenberg is the chief scientific officer and N.L. is the vice president of research and development of Aleph Farms.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Extended Data Fig. 1 Micro-CT analysis of Porosity, connectivity and surface area per volume of TSP1, TSP2.
Surface area calculation assumes a scaffold density of 0.65 gr/ml. Scaffold density was assessed based on scaffold height and weight measurements, assuming the scaffold diameter is 6 mm.
Extracellular matrix and myogenesis proteins upregulated in co-cultures versus BSC mono-cultures.
Extended Data Fig. 3 Enriched gene ontology (GO) terms of proteins elevated in co-cultures of Bovine satellite cells with bovine smooth muscle cells.
Including all DE proteins.
Extended Data Fig. 4 Number of differentially expressed proteins in each gene ontology (GO) cluster & average fold Change (FC).
In reference to figure 6. Averaging was performed on the log2 scale, to prevent bias towards elevated proteins.
Supplementary Figs. 1–7.
Micro-CT TSP-1 reconstruction.
Fibroblast (red) proliferation on TSP scaffold over 21 days.
A full factorial statistical analysis (2 × 23) of the effect of the proliferation media (LLM1 versus control), IGF-1 in the differentiation medium (+IGF-1 versus −IGF-1) and EGF in the differentiation medium (+EGF versus −EGF) on myotube coverage, average myotube area and myotube complexity.
About this article
Cite this article
Ben-Arye, T., Shandalov, Y., Ben-Shaul, S. et al. Textured soy protein scaffolds enable the generation of three-dimensional bovine skeletal muscle tissue for cell-based meat. Nat Food 1, 210–220 (2020). https://doi.org/10.1038/s43016-020-0046-5
A serum-free media formulation for cultured meat production supports bovine satellite cell differentiation in the absence of serum starvation
Nature Food (2022)
npj Science of Food (2022)
Formation of contractile 3D bovine muscle tissue for construction of millimetre-thick cultured steak
npj Science of Food (2021)
Engineered whole cut meat-like tissue by the assembly of cell fibers using tendon-gel integrated bioprinting
Nature Communications (2021)
Establishment, characterization, and transfection potential of a new continuous fish cell line (CAM) derived from the muscle tissue of grass goldfish (Carassius auratus)
In Vitro Cellular & Developmental Biology - Animal (2021)