Translating musculoskeletal bioengineering into tissue regeneration therapies

Abstract

Musculoskeletal injuries and disorders are the leading cause of physical disability worldwide and a considerable socioeconomic burden. The lack of effective therapies has driven the development of novel bioengineering approaches that have recently started to gain clinical approvals. In this review, we first discuss the self-repair capacity of the musculoskeletal tissues and describe causes of musculoskeletal dysfunction. We then review the development of novel biomaterial, immunomodulatory, cellular, and gene therapies to treat musculoskeletal disorders. Last, we consider the recent regulatory changes and future areas of technological progress that can accelerate translation of these therapies to clinical practice.

Fig. 1. Musculoskeletal injury response.

Immune and tissue-specific progenitor cell regulation of musculoskeletal injury response in vivo. (A) Following injury, neutrophils (NP) and monocytes (M0) infiltrate the injury site to phagocytose damaged tissue and secrete factors that control fate of infiltrating immune cells. Initially, proliferation of immune and resident progenitor cells is stimulated by a pro-inflammatory microenvironment created by cytokine secretion from macrophages (M1) and T cells (T_h1 and T_h17). Subsequent tissue regeneration and remodeling are orchestrated by a switch to an anti-inflammatory microenvironment created by cytokine secretion from macrophages (M2) and T cells (T_regs and T_h2). (B) Skeletal muscle regeneration is orchestrated by muscle resident satellite cells (SCs) that in uninjured tissue are quiescent and express the transcriptional factor PAX7. Upon injury, mechanical disruption and the pro-inflammatory microenvironment stimulate SC activation, proliferation, and MYOD expression. Activated SCs then fuse together to form de novo myofibers, fuse into regenerating myofibers, or return to quiescence by loss of MYOD. (C) Bone remodeling is characterized by an initial hematoma formation and pro-inflammatory microenvironment that recruits circulating MSCs. These MSCs initially differentiate into chondroblasts and fibroblasts to generate a fibrocartilaginous callous, which is further remodeled into bone tissue by MSC-derived and resident osteoblasts. Successful remodeling relies on the balanced synthetic and resorption activities of osteoblasts and osteoclasts, respectively. (D) In response to injury, cartilage undergoes a weak pro-inflammatory response that results in no-to-limited recruitment and proliferation of cartilage-derived progenitor cells (CPCs). Consequently, cartilage does not regenerate and instead undergoes progressive degeneration and degradation.

Fig. 2. Immunomodulatory biomaterials for musculoskeletal regeneration.

(A) Multiple biomaterial modifications including changes to surface topography, surface charge, wettability, and incorporation of bioactive molecules and immunomodulatory drugs can be used to regulate immune-mediated regenerative responses to tissue damage. (B) Decellularized extracellular matrices (dECMs) retain multiple biophysical cues which upon implantation stimulate immune cell infiltration and a pro-inflammatory response. Subsequent degradation of the implanted dECM induces release of growth factors and matrix-bound nanovesicles (MBVs) that promote immune cell conversion to an M2 phenotype and stimulate neighboring stem cell recruitment and, ultimately, regeneration via de novo tissue formation.

Fig. 3. Bioengineering approaches for cell-based musculoskeletal therapies.

(A) Traditional cell culture platforms using tissue culture plastic (TCPS) poorly retain stem cell characteristics. Next generation culture platforms retain stem cell characteristics and facilitate cell expansion by better replicating the stem cell niche microenvironment. (B) Next generation single-cell sequencing and CRISPR-edited reporter lines allow development of more efficient differentiation protocols for derivation of biomimetic musculoskeletal progenitor cells from hiPSCs. (C) Tissue-engineering methods allow in vitro fabrication of functional three-dimensional tissues using: porous scaffolds that initially provide structural and mechanical support to seeded cells and are subsequently remodeled in vitro and in vivo; Cell sheets that are detached from extracellular matrix (ECM)- or thermoresponsive polymer-coated dishes and subsequently stacked; Self-assembly of highly dense cell condensates that initially secrete an immature ECM, followed by cell and matrix maturation and acquisition of native-like mechanical properties; and 3D bioprinting of cells and bioinks to recreate complex tissue architecture and cell composition, which, however, does not lead to native tissue functionality.

Fig. 4. Bioengineering approaches for gene-based musculoskeletal therapies.

(A) Antisense oligonucleotides mask exons from splicing machinery and restore functional gene expression. (B) Gene replacement via use of ubiquitous, tissue-specific, or inflammatory-responsive promoters controls the expression of full-length or modified versions of the gene of interest. (C) Growth factor secretion by ex vivo or in vivo transduced cells creates a pro-regenerative microenvironment at the injury site. (D) CRISPR-Cas9 editing induces double-stranded breaks (DSBs) and gene knock-out by nonhomologous end joining (NHEJ). Alternatively, homology-directed repair (HDR) with inclusion of a DNA template allows for gene knock-in. (E) Systemic gene delivery is accomplished by AAV vector or non-viral polymeric or lipid nanoparticle (NP) systems. Alternatively, ex vivo gene modifications are performed by transduction or transfection of autologous or allogeneic cells prior to transplantation.