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Synthetic Notch (synNotch) receptors are genetically encoded, modular synthetic receptors that enable mammalian cells to detect environmental signals and respond by activating user-prescribed transcriptional programs. Although some materials have been modified to present synNotch ligands with coarse spatial control, applications in tissue engineering generally require extracellular matrix (ECM)-derived scaffolds and/or finer spatial positioning of multiple ligands. Thus, we develop here a suite of materials that activate synNotch receptors for generalizable engineering of material-to-cell signaling. We genetically and chemically fuse functional synNotch ligands to ECM proteins and ECM-derived materials. We also generate tissues with microscale precision over four distinct reporter phenotypes by culturing cells with two orthogonal synNotch programs on surfaces microcontact-printed with two synNotch ligands. Finally, we showcase applications in tissue engineering by co-transdifferentiating fibroblasts into skeletal muscle or endothelial cell precursors in user-defined micropatterns. These technologies provide avenues for spatially controlling cellular phenotypes in mammalian tissues. 

The uterus is susceptible to benign tumors known as fibroids, which have been associated with many pregnancy complications, including preterm labor. However, the impact of fibrotic tissue remodeling on the physiology of the myometrium, the smooth muscle layer of the uterus, is poorly understood, in large part due to a lack of model systems. In this study, we engineered healthy-like and fibrotic-like myometrium by culturing human myometrial smooth muscle cells on polyacrylamide hydrogels micropatterned with fibronectin to independently tune matrix rigidity and tissue alignment, respectively. We then evaluated calcium transients in response to oxytocin stimulation. Isotropic myometrial tissues on stiff substrates (representing fibrotic myometrium) had shorter calcium transients due to shorter decay time compared to aligned myometrial tissues on soft substrates (representing healthy myometrium). Calcium transients in aligned tissues had longer response times and longer decay times than isotropic tissues, irrespective of substrate stiffness. The amplitude of calcium transients was also higher on soft substrates compared to stiff substrates, irrespective of tissue alignment. We also performed RNA sequencing to detect differentially expressed genes between healthy- and fibrotic-like tissues, which revealed that a bitter taste receptor shown to induce smooth muscle relaxation, TAS2R31, was down-regulated in fibrotic-like tissues. Finally, we measured oxytocin-induced calcium transients in response to pre-treatment with progesterone, caffeine, thrombin, and nifedipine to demonstrate applications for our model system in drug screening. Both progesterone and caffeine caused a decrease in calcium transient duration, as expected, while thrombin and nifedipine had less impact. Collectively, our engineered model of the myometrium enables new insights into myometrial mechanobiology and can be extended to identify or screen novel drug targets. 

The myometrium is the smooth muscle layer of the uterus that generates the contractions that drive processes such as menstruation and childbirth. Aberrant contractions of the myometrium can result in preterm birth, insufficient progression of labor, or other difficulties that can lead to maternal or fetal complications or even death. To investigate the underlying mechanisms of these conditions, the most common model systems have conventionally been animal models and human tissue strips, which have limitations mostly related to relevance and scalability, respectively. Myometrial smooth muscle cells have also been isolated from patient biopsies and cultured in vitro as a more controlled experimental system. However, in vitro approaches have focused primarily on measuring the effects of biochemical stimuli and neglected biomechanical stimuli, despite the extensive evidence indicating that remodeling of tissue rigidity or excessive strain is associated with uterine disorders. In this review, we first describe the existing approaches for modeling human myometrium with animal models and human tissue strips and compare their advantages and disadvantages. Next, we introduce existing in vitro techniques and assays for assessing contractility and summarize their applications in elucidating the role of biochemical or biomechanical stimuli on human myometrium. Finally, we conclude by proposing the translation of “organ on chip” approaches to myometrial smooth muscle cells as new paradigms for establishing their fundamental mechanobiology and to serve as next-generation platforms for drug development.