Multifactorial bottom‐up bioengineering approaches for the development of living tissue substitutes

Abstract

Bottom‐up bioengineering utilizes the inherent capacity of cells to build highly sophisticated structures with high levels of biomimicry. Despite the significant advancements in the field, monodomain approaches require prolonged culture time to develop an implantable device, usually associated with cell phenotypic drift in culture. Herein, we assessed the simultaneous effect of macromolecular crowding (MMC) and mechanical loading in enhancing extracellular matrix (ECM) deposition while maintaining tenocyte (TC) phenotype and differentiating bone marrow stem cells (BMSCs) or transdifferentiating neonatal and adult dermal fibroblasts toward tenogenic lineage. At d 7, all cell types presented cytoskeleton alignment perpendicular to the applied load independently of the use of MMC. MMC enhanced ECM deposition in all cell types. Gene expression analysis indicated that MMC and mechanical loading maintained TC phenotype, whereas tenogenic differentiation of BMSCs or transdifferentiation of dermal fibroblasts was not achieved. Our data suggest that multifactorial bottom‐up bioengineering approaches significantly accelerate the development of biomimetic tissue equivalents.—Gaspar, D., Ryan, C. N. M., Zeugolis, D. I. Multifactorial bottom‐up bioengineering approaches for the development of living tissue substitutes. FASEB J. 33, 5741–5754 (2019). http://www.fasebj.org

Carrageenan enhances chondrogenesis and osteogenesis in human bone marrow stem cell culture

“…In vivo, the fate of mesenchymal stem cells is controlled by a reciprocal interaction with the surrounding extracellular matrix (ECM). The ECM regulates several cell functions, including attachment, shape, migration and differentiation (Ahmed and Ffrench-Constant, 2016;Gattazzo et al, 2014;Guilak et al, 2009;Kim et al, 2011) and, in response to these signals, cells appropriately remodel the surrounding matrix (Hoshiba et al, 2009;Hoshiba et al, 2010). ECM remodelling that accompanies cell differentiation directly controls stem cell behaviour and fate (Crane and Cao, 2014;Gattazzo et al, 2014;Lu et al, 2011).…”

“…The ECM regulates several cell functions, including attachment, shape, migration and differentiation (Ahmed and Ffrench-Constant, 2016;Gattazzo et al, 2014;Guilak et al, 2009;Kim et al, 2011) and, in response to these signals, cells appropriately remodel the surrounding matrix (Hoshiba et al, 2009;Hoshiba et al, 2010). ECM remodelling that accompanies cell differentiation directly controls stem cell behaviour and fate (Crane and Cao, 2014;Gattazzo et al, 2014;Lu et al, 2011). In vitro, the ECM also controls cell functions and triggers cell differentiation, even in the absence of exogenous inducing factors (Mao et al, 2017;Rao Pattabhi et al, 2014;Tang et al, 2013;Thibault et al, 2010;Young et al, 2013).…”

Engineering the Tenogenic Niche In Vitro with Microenvironmental Tools

“…Low glucose dose has also been shown to promote tenogenic phenotype, as evidenced by upregulated tenogenic marker expression and enhanced collagen expression in tenocyte culture . However, these supplements are not sufficiently tenogenic when used in isolation and are often used in addition to other microenvironmental cues …”

“…For example, carrageenan alone, in tenocyte culture, has been shown to enhance tendon‐specific ECM deposition . Recently, carrageenan was used to crowd dermal fibroblasts, BMSCs, and tenocytes in combination with mechanical loading; it was found that MMC coupled with mechanical stimulation supported the tenogenic phenotype maintenance of tenocytes, but not the differentiation or trans‐differentiation of BMSCs or dermal fibroblasts, respectively, further indicating that multifactorial approaches (see Section 7) should be investigated.…”

“…Ascorbic acid, for example, is a vitamin that prevents free radical formation and protects tenocytes from oxidative stress . It has been used along with various tenogenic inductive treatments, as it facilitates enhanced collagen type I synthesis in vitro . Low glucose dose has also been shown to promote tenogenic phenotype, as evidenced by upregulated tenogenic marker expression and enhanced collagen expression in tenocyte culture .…”