PEG-Anthracene Hydrogels as an On-Demand Stiffening Matrix To Study Mechanobiology.

There is a growing interest in materials that can dynamically change their properties in the presence of cells to study mechanobiology. Herein, we exploit the 365 nm light mediated [4+4] photodimerization of anthracene groups to develop cytocompatible PEG-based hydrogels with tailorable initial moduli that can be further stiffened. A hydrogel formulation that can stiffen from 10 to 50 kPa, corresponding to the stiffness of a healthy and fibrotic heart, respectively, was prepared. This system was used to monitor the stiffness-dependent localization of NFAT, a downstream target of intracellular calcium signaling using a reporter in live cardiac fibroblasts (CFbs). NFAT translocates to the nucleus of CFbs on stiffening hydrogels within 6 h, whereas it remains cytoplasmic when the CFbs are cultured on either 10 or 50 kPa static hydrogels. This finding demonstrates how dynamic changes in the mechanical properties of a material can reveal the kinetics of mechanoresponsive cell signaling pathways that may otherwise be missed in cells cultured on static substrates.

Creation of a non-Western humanized gnotobiotic mouse model through the transplantation of rural African fecal microbiota.

IMPORTANCE: There is increasing evidence that microbes residing within the intestines (gut microbiota) play important roles in the well-being of humans. Yet, there are considerable challenges in determining the specific role of gut microbiota in human diseases owing to the complexity of diverse internal and environmental factors that can contribute to diseases. Mice devoid of all microorganisms (germ-free mice) can be colonized with human stool samples to examine the specific contribution of the gut microbiota to a disease. These approaches have been primarily focused on stool samples obtained from individuals in Western countries. Thus, there is limited understanding as to whether the same methods used to colonize germ-free mice with stool from Western individuals would apply to the colonization of germ-free mice with stool from non-Western individuals. Here, we report the results from colonizing germ-free mice with stool samples of Malian children.

Myoblast mechanotransduction and myotube morphology is dependent on BAG3 regulation of YAP and TAZ.

Skeletal muscle tissue is mechanically dynamic with changes in stiffness influencing function, maintenance, and regeneration. We modeled skeletal muscle mechanical changes in culture with dynamically stiffening hydrogels demonstrating that the chaperone protein BAG3 transduces matrix stiffness by redistributing YAP and TAZ subcellular localization in muscle progenitor cells. BAG3 depletion increases cytoplasmic retention of YAP and TAZ, desensitizing myoblasts to changes in hydrogel elastic moduli. Upon differentiation, muscle progenitors depleted of BAG3 formed enlarged, round myotubes lacking the typical cylindrical morphology. The aberrant morphology is dependent on YAP/TAZ signaling, which was sequestered in the cytoplasm in BAG3-depleted myotubes but predominately nuclear in cylindrical myotubes of control cells. Control progenitor cells induced to differentiate on soft (E' = 4 and 12 kPa) hydrogels formed circular myotubes similar to those observed in BAG3-depleted cells. Inhibition of the Hippo pathway partially restored myotube morphologies, permitting nuclear translocation of YAP and TAZ in BAG3-depleted myogenic progenitors. Thus, BAG3 is a critical mediator of dynamic stiffness changes in muscle tissue, coupling mechanical alterations to intracellular signals and inducing changes in gene expression that influence muscle progenitor cell morphology and differentiation.

Injury-mediated stiffening persistently activates muscle stem cells through YAP and TAZ mechanotransduction.

The skeletal muscle microenvironment transiently remodels and stiffens after exercise and injury, as muscle ages, and in myopathic muscle; however, how these changes in stiffness affect resident muscle stem cells (MuSCs) remains understudied. Following muscle injury, muscle stiffness remained elevated after morphological regeneration was complete, accompanied by activated and proliferative MuSCs. To isolate the role of stiffness on MuSC behavior and determine the underlying mechanotransduction pathways, we cultured MuSCs on strain-promoted azide-alkyne cycloaddition hydrogels capable of in situ stiffening by secondary photocrosslinking of excess cyclooctynes. Using pre- to post-injury stiffness hydrogels, we found that elevated stiffness enhances migration and MuSC proliferation by localizing yes-associated protein 1 (YAP) and WW domain-containing transcription regulator 1 (WWTR1; TAZ) to the nucleus. Ablating YAP and TAZ in vivo promotes MuSC quiescence in postinjury muscle and prevents myofiber hypertrophy, demonstrating that persistent exposure to elevated stiffness activates mechanotransduction signaling maintaining activated and proliferating MuSCs.