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Abstract Purpose of ReviewInterfacial tissue exists throughout the body at cartilage-to-bone (osteochondral interface) and tendon-to-bone (enthesis) interfaces. Healing of interfacial tissues is a current challenge in regenerative approaches because the interface plays a critical role in stabilizing and distributing the mechanical stress between soft tissues (e.g., cartilage and tendon) and bone. The purpose of this review is to identify new directions in the field of interfacial tissue development and physiology that can guide future regenerative strategies for improving post-injury healing. Recent FindingsCues from interfacial tissue development may guide regeneration including biological cues such as cell phenotype and growth factor signaling; structural cues such as extracellular matrix (ECM) deposition, ECM, and cell alignment; and mechanical cues such as compression, tension, shear, and the stiffness of the cellular microenvironment. SummaryIn this review, we explore new discoveries in the field of interfacial biology related to ECM remodeling, cellular metabolism, and fate. Based on emergent findings across multiple disciplines, we lay out a framework for future innovations in the design of engineered strategies for interface regeneration. Many of the key mechanisms essential for interfacial tissue development and adaptation have high potential for improving outcomes in the clinic. 

Abstract Skeletal muscle activation using optogenetics has emerged as a promising technique for inducing noninvasive muscle contraction and assessing muscle function both in vivo and in vitro. Transgenic mice overexpressing the optogenetic fusion protein, Channelrhodopsin 2‐EYFP (ChR2‐EYFP) in skeletal muscle are widely used; however, overexpression of fluorescent proteins can negatively impact the functionality of activable tissues. In this study, we characterized the contractile properties of ChR2‐EYFP skeletal muscle and introduced the ChR2‐only mouse model that expresses light‐responsive ChR2 without the fluorescent EYFP in their skeletal muscles. We found a significant reduction in the contractile ability of ChR2‐EYFP muscles compared with ChR2‐only and WT mice, observed under both electrical and optogenetic stimulation paradigms. Bulk RNAseq identified the downregulation of genes associated with transmembrane transport and metabolism in ChR2‐EYFP muscle, while the ChR2‐only muscle did not demonstrate any notable deviations from WT muscle. The RNAseq results were further corroborated by a reduced protein‐level expression of ion channel‐related HCN2 in ChR2‐EYFP muscles and gluconeogenesis‐modulating FBP2 in both ChR2‐EYFP and ChR2‐only muscles. Overall, this study reveals an intrinsic skeletal dysfunction in the widely used ChR2‐EYFP mice model and underscores the importance of considering alternative optogenetic models, such as the ChR2‐only, for future research in skeletal muscle optogenetics. 

Abstract BackgroundThe growth of most bony tuberosities, like the deltoid tuberosity (DT), rely on the transmission of muscle forces at the tendon‐bone attachment during skeletal growth. Tuberosities distribute muscle forces and provide mechanical leverage at attachment sites for joint stability and mobility. The genetic factors that regulate tuberosity growth remain largely unknown. In mouse embryos with global deletion offibroblast growth factor 9(Fgf9), the DT size is notably enlarged. In this study, we explored the tissue‐specific regulation of DT size using both global and targeted deletion ofFgf9. ResultsWe showed that cell hypertrophy and mineralization dynamics of the DT, as well as transcriptional signatures from skeletal muscle but not bone, were influenced by the global loss ofFgf9. Loss ofFgf9during embryonic growth led to increased chondrocyte hypertrophy and reduced cell proliferation at the DT attachment site. This endured hypertrophy and limited proliferation may explain the abnormal mineralization patterns and locally dysregulated expression of markers of endochondral development inFgf9^nullattachments. We then showed that targeted deletion ofFgf9in skeletal muscle leads to postnatal enlargement of the DT. ConclusionTaken together, we discovered thatFgf9may play an influential role in muscle‐bone cross‐talk during embryonic and postnatal development. 

Skeletal shape depends on the transmission of contractile muscle forces from tendon to bone across the enthesis. Loss of muscle loading impairs enthesis development, yet little is known if and how the postnatal enthesis adapts to increased loading. Here, we studied adaptations in enthesis structure and function in response to increased loading, using optogenetically induced muscle contraction in young (i.e., growth) and adult (i.e., mature) mice. Daily bouts of unilateral optogenetic loading in young mice led to radial calcaneal expansion and warping. This also led to a weaker enthesis with increased collagen damage in young tendon and enthisis, with little change in adult mice. We then used RNA sequencing to identify the pathways associated with increased mechanical loading during growth. In tendon, we found enrichment of glycolysis, focal adhesion, and cell-matrix interactions. In bone, we found enrichment of inflammation and cell cycle. Together, we demonstrate the utility of optogenetic-induced muscle contraction to elicit in vivo adaptation of the enthesis.