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IntroductionVolumetric muscle loss (VML) is characterized by permanent tissue impairment resulting from critically-sized muscle loss. We performed time-series transcriptomic and proteomic analyses to reveal key mediators of irreversible pathological remodeling after induction of VML in mice. MethodsThe dynamics of gene and protein expression patterns were analyzed for up to 3 weeks after muscle injury. ResultsRNA Sequencing revealed transcriptional patterns that show rapid upregulation or downregulation shortly after injury, among which a subset of genes failed to return to pre-injury levels within 3 weeks after VML. Time-series analysis revealed gene clusters with sustained upregulation after 3 weeks, including those associated with extracellular matrix remodeling and inflammation, whereas the gene clusters having sustained downregulation were associated with mitochondrial function and metabolism. We further identifiedSPI1andSP1as novel molecular mediators of the pathological remodeling process. DiscussionThis work demonstrates the utility of time-series analysis to reveal dysregulated pathways in the setting of VML. 

Abstract Despite the therapeutic potential of mesenchymal stromal cells (MSC), there is limited understanding of optimal extracellular matrix (ECM) environments to manufacture these cells. We developed tissue chips to study the effects of multi-factorial ECM environments under manufacturable stiffness ranges and multi-component ECM compositions. Manufacturing qualities of cell expansion potential, immunomodulation, and differentiation capacity were examined. The results show stiffness effects, with 900 kPa substrates supporting higher proliferation and osteogenic differentiation, along with anti-inflammatory IL-10 expression, whereas 150 kPa substrates promoted adipogenic differentiation at 150 kPa, suggesting that optimal ECM environments may differ based on manufacturing goals. ECM biochemistries containing fibronectin and laminin further modulated MSC manufacturing qualities across various stiffnesses. Proteomic and transcriptomic analyses revealed unique ECM combinations that induced higher levels of angiogenic and immunomodulatory cytokines, compared to single factor ECMs. These findings demonstrate that optimized ECM environments enhance MSC manufacturing quality. 

Peripheral artery disease (PAD) affects 10 million people in the United States and >230 million worldwide.1,2 It is associated with impaired vascular perfusion to the lower extremities, leading to limbthreatening amputation in severe cases of critical limb ischemia. Patients with PAD have an inferior prognosis and reduced limb function, when compared with patients without PAD.3 Very few effective therapies have been identified, in part, because the key biologic pathways associated with functional impairment remain unclear. Current treatments include supervised and home-based walking exercise to improve the mobility in patients with PAD.4,5 A better understanding of the underlying pathological mechanisms of skeletal muscle damage underlying PAD may help identify new therapeutic opportunities 

Murine myoblasts cultured on combinatorial extracellular matrix (ECM) proteins are exposed to uniaxial strain. The combined effects of ECMs and strain on myogenesis are investigated by transcriptomic and protein analyses. 

Abstract Traumatic muscle injuries associated with volumetric muscle loss (VML) are characterized by muscle loss beyond intrinsic regeneration capacity, leading to permanent functional impairment. Experimental therapies to augment muscle regeneration, such as cell injection, are limited by low cell transplantation capacity, whereas conventional engineered muscle tissue transplants lack geometric customization to conform to the shape of the muscle defect. Here, a facile approach to engineer scaffold‐free high‐density muscle tissues in customizable geometric shapes and sizes with high cell viability and integration potential is developed. Using a facile mold‐based approach to engineer scaffold‐free modular units, transcriptional profiling is performed to uncover the role of pre‐formed cell–cell interactions within scaffold‐free muscle bioconstructs on myogenesis, an the efficacy of muscle bioconstructs in a mouse model of VML is then evaluated. RNA sequencing revealed that pre‐formed cell–cell interactions supported myogenic pathways related to muscle contraction and myofibril assembly, unlike dissociated monodisperse cells. This work further demonstrates the therapeutic efficacy of 3D rectangular solid‐shaped scaffold‐free transplants in improving muscle function and vascular regeneration. Finally, toward clinical translation, the feasibility of this technology to integrate with medical imaging and artificial intelligence‐driven customized bioconstruct design and assembly for intraoperative use is illustrated. 

Children are said to be a product of both nature and nurture – of their genes and the environment in which they are raised. The cells of the growing liver are not so different in this sense. As the liver of a fetus develops, immature cells called liver progenitors mature to become one of two types of adult cells: the hepatocytes that form the bulk of the liver, or the biliary cells that make up the bile duct. The traditional view is that genetic factors mainly control which cell type the progenitor cells become. However, recent research suggests that the environment around the cells matters more in this process than once thought. Cells can respond to the physical properties of their environment, such as the structure and stiffness of the surrounding tissue. These properties change as the liver develops, and can also be altered by disease. For example, damaged liver cells can spit out proteins that harden and form stiff scars. This raises a question: do changes in stiffness affect how progenitor cells behave? To answer this question, Kaylan et al. printed collagen in circular patterns and grew liver progenitor cells on them. The cells at the edges of the circular patterns matured into bile duct cells, while those in the center became hepatocytes. The stiffness felt by the cells was then determined by measuring the level of mechanical stress that they experienced. This revealed that the cells at the edge of the collagen pattern – the cells that became bile duct cells – were under most stress. In addition, more bile duct cells formed when progenitor cells were grown on a stiffer collagen pattern. Overall, the results reported by Kaylan et al. suggest that the stiffness of the environment, and the resulting stresses on a progenitor cell, can influence how it matures. As well as helping us to understand how the liver develops, this knowledge could also help us to treat a group of diseases called cholangiopathies, in which the bile ducts become inflamed. These diseases are thought to be caused by certain cells (which are similar to liver progenitor cells) maturing to become incorrect cell types. Future studies could determine if preventing changes in stiffness in the environment of these cells, or slowing their response to such changes, would help patients.