Handheld instrument for wound-conformal delivery of skin precursor sheets improves healing in full-thickness burns.

The current standard of care for patients with severe large-area burns consists of autologous skin grafting or acellular dermal substitutes. While emerging options to accelerate wound healing involve treatment with allogeneic or autologous cells, delivering cells to clinically relevant wound topologies, orientations, and sizes remains a challenge. Here, we report the one-step in situ formation of cell-containing biomaterial sheets using a handheld instrument that accommodates the topography of the wound. In an approach that maintained cell viability and proliferation, we demonstrated conformal delivery to surfaces that were inclined up to 45° with respect to the horizontal. In porcine pre-clinical models of full-thickness burn, we delivered mesenchymal stem/stromal cell-containing fibrin sheets directly to the wound bed, improving re-epithelialization, dermal cell repopulation, and neovascularization, indicating that this device could be introduced in a clinical setting improving dermal and epidermal regeneration.

Three-dimensional niche stiffness synergizes with Wnt7a to modulate the extent of satellite cell symmetric self-renewal divisions.

Satellite cells (SCs), the resident adult stem cells of skeletal muscle, are required for tissue repair throughout life. While many signaling pathways are known to control SC self-renewal, less is known about the mechanisms underlying the spatiotemporal control of self-renewal during skeletal muscle repair. Here, we measured biomechanical changes that accompany skeletal muscle regeneration and determined the implications on SC fate. Using atomic force microscopy, we quantified a 2.9-fold stiffening of the SC niche at time-points associated with planar-oriented symmetric self-renewal divisions. Immunohistochemical analysis confirms increased extracellular matrix deposition within the basal lamina. To test whether three-dimensional (3D) niche stiffness can alter SC behavior or fate, we embedded isolated SC-associated muscle fibers within biochemically inert agarose gels tuned to mimic native tissue stiffness. Time-lapse microscopy revealed that a stiff 3D niche significantly increased the proportion of planar-oriented divisions, without effecting SC viability, fibronectin deposition, or fate change. We then found that 3D niche stiffness synergizes with WNT7a, a biomolecule shown to control SC symmetric self-renewal divisions via the noncanonical WNT/planar cell polarity pathway, to modify stem cell pool expansion. Our results provide new insights into the role of 3D niche biomechanics in regulating SC fate choice.

Tethered Jagged-1 Synergizes with Culture Substrate Stiffness to Modulate Notch-Induced Myogenic Progenitor Differentiation.

INTRODUCTION: Notch signaling is amongst the key intrinsic mechanisms regulating satellite cell fate, promoting the transition of activated satellite cells to highly proliferative myogenic progenitor cells and preventing their premature differentiation. Although much is known about the biochemical milieu that drives myogenic progression, less is known about the spatial cues providing spatiotemporal control of skeletal muscle repair in the context of Notch signaling. METHODS: Using a murine injury model, we quantified in vivo biophysical changes that occur within the skeletal muscle during regeneration. Employing tunable poly(ethylene glycol)-based hydrogel substrates, we modeled the measured changes in bulk stiffness in the context of Notch ligand signaling, which are present in the regenerative milieu at the time of injury. RESULTS: Following injury, there is a transient increase in the bulk stiffness of the tibialis anterior muscle that may be explained in part by changes in extracellular matrix deposition. When presented to primary myoblasts, Jagged-1, Jagged-2, and Dll1 in a tethered format elicited greater degrees of Notch activity compared to their soluble form. Only tethered Jagged-1 effects were tuned by substrate stiffness, with the greatest Notch activation observed on stiff hydrogels matching the stiffness of regenerating muscle. When exposed to tethered Jagged-1 on stiff hydrogels, fewer primary myoblasts expressed myogenin, and pharmacological inhibitor studies suggest this effect is Notch and RhoA dependent. CONCLUSION: Our study proposes that tethered Jagged-1 presented in the context of transient tissue stiffening serves to tune Notch activity in myogenic progenitors during skeletal muscle repair and delay differentiation.

Biomechanical Origins of Muscle Stem Cell Signal Transduction.

Skeletal muscle, the most abundant and widespread tissue in the human body, contracts upon receiving electrochemical signals from the nervous system to support essential functions such as thermoregulation, limb movement, blinking, swallowing and breathing. Reconstruction of adult muscle tissue relies on a pool of mononucleate, resident muscle stem cells, known as "satellite cells", expressing the paired-box transcription factor Pax7 necessary for their specification during embryonic development and long-term maintenance during adult life. Satellite cells are located around the myofibres in a niche at the interface of the basal lamina and the host fibre plasma membrane (i.e., sarcolemma), at a very low frequency. Upon damage to the myofibres, quiescent satellite cells are activated and give rise to a population of transient amplifying myogenic progenitor cells, which eventually exit the cell cycle permanently and fuse to form new myofibres and regenerate the tissue. A subpopulation of satellite cells self-renew and repopulate the niche, poised to respond to future demands. Harnessing the potential of satellite cells relies on a complete understanding of the molecular mechanisms guiding their regulation in vivo. Over the past several decades, studies revealed many signal transduction pathways responsible for satellite cell fate decisions, but the niche cues driving the activation and silencing of these pathways are less clear. Here we explore the scintillating possibility that considering the dynamic changes in the biophysical properties of the skeletal muscle, namely stiffness, and the stretch and shear forces to which a myofibre can be subjected to may provide missing information necessary to gain a full understanding of satellite cell niche regulation.