H-2RIIBP expressed from a baculovirus vector binds to multiple hormone response elements.

H-2RIIBP is a member of the nuclear hormone receptor superfamily that binds to the region II enhancer of major histocompatibility complex class I genes. Based on its homology with Drosophila XR2C/CF1, H-2RIIBP may play a role in development. By using a baculovirus expression system, a large amount of recombinant H-2RIIBP was produced. The recombinant protein accumulated in the nucleus of insect cells. A series of monoclonal antibodies reacting with the recombinant H-2RIIBP was then generated. A DNA-protein immunoprecipitation assay was developed with these antibodies, enabling the DNA-binding specificity of H-2RIIBP to be distinguished from that of an endogenous region II binding factor expressed in uninfected insect cells. We show that H-2RIIBP binds to estrogen response elements with an affinity comparable to that for the region II enhancer. H-2RIIBP also bound to some, but not all, thyroid hormone response elements and retinoic acid response elements, albeit at a lower affinity. Binding to these elements was demonstrated without exogenous addition of a ligand. The H-2RIIBP binding specificity determined by this assay was in agreement with the specificity assessed by Southwestern and gel mobility shift assays. Furthermore, methylation interference assays indicated that H-2RIIBP recognizes the conserved hormone response motif GG(T/A)CA. Taken together, these data demonstrate that H-2RIIBP is capable of binding to hormone response elements of a variety of genes. They suggest that H-2RIIBP may exert a pleiotropic function.     

Characterization of tissue biomechanics and mechanical signaling in uterine leiomyoma

Leiomyoma are common tumors arising within the uterus that feature excessive deposition of a stiff, disordered extracellular matrix (ECM). Mechanical stress is a critical determinant of excessive ECM deposition and increased mechanical stress has been shown to be involved in tumorigenesis. Here we tested the viscoelastic properties of leiomyoma and characterized dynamic and static mechanical signaling in leiomyoma cells using three approaches, including measurement of active RhoA. We found that the peak strain and pseudo-dynamic modulus of leiomyoma tissue was significantly increased relative to matched myometrium. In addition, leiomyoma cells demonstrated an attenuated response to applied cyclic uniaxial strain and to variation in substrate stiffness, relative to myometrial cells. However, on a flexible pronectin-coated silicone substrate, basal levels and lysophosphatidic acid-stimulated levels of activated RhoA were similar between leiomyoma and myometrial cells. In contrast, leiomyoma cells plated on a rigid polystyrene substrate had elevated levels of active RhoA, compared to myometrial cells. The results indicate that viscoelastic properties of the ECM of leiomyoma contribute significantly to the tumor's inherent stiffness and that leiomyoma cells have an attenuated sensitivity to mechanical cues. The findings suggest there may be a fundamental alteration in the communication between the external mechanical environment (extracellular forces) and reorganization of the actin cytoskeleton mediated by RhoA in leiomyoma cells. Additional research will be needed to elucidate the mechanism(s) responsible for the attenuated mechanical signaling in leiomyoma cells.     

Integrin β1 regulates leiomyoma cytoskeletal integrity and growth

Uterine leiomyomas are characterized by an excessive extracellular matrix, increased mechanical stress, and increased active RhoA. Previously, we observed that mechanical signaling was attenuated in leiomyoma, but the mechanisms responsible remain unclear. Integrins, especially integrin β1, are transmembrane adhesion receptors that couple extracellular matrix stresses to the intracellular cytoskeleton to influence cell proliferation and differentiation. Here we characterized integrin and laminin to signaling in leiomyoma cells. We observed a 2.25 ± 0.32 fold increased expression of integrin β1 in leiomyoma cells, compared to myometrial cells. Antibody-mediated inhibition of integrin β1 led to significant growth inhibition in leiomyoma cells and a loss of cytoskeletal integrity. Specifically, polymerization of actin filaments and formation of focal adhesions were reduced by inhibition of integrin β1. Inhibition of integrin β1 in leiomyoma cells led to 0.81 ± 0.02 fold decrease in active RhoA, and resembled levels found in serum-starved cells. Likewise, inhibition of integrin β1 was accompanied by a decrease in phospho-ERK. Compared to myometrial cells, leiomyoma cells demonstrated increased expression of integrin α6 subunit to laminin receptor (1.91 ± 0.11 fold), and increased expression of laminin 5α (1.52 ± 0.02), laminin 5β (3.06 ± 0.92), and laminin 5γ (1.66 ± 0.06). Of note, leiomyoma cells grown on laminin matrix appear to realign themselves. Taken together, the findings reveal that the attenuated mechanical signaling in leiomyoma cells is accompanied by an increased expression and a dependence on integrin β1 signaling in leiomyoma cells, compared to myometrial cells.     

New potential regulators of uterine leiomyomata from DNA arrays: the ionotropic glutamate receptor GluR2

In the post-Genome era, new concepts emerge about the growth regulation of uterine leiomyomata. Screening of leiomyoma and myometrial tissues with DNA arrays revealed numerous genes up-regulated in leiomyomata that were not known to be expressed in the human uterus. GluR2, a subunit of a ligand-gated cation channel, is up-regulated in leiomyomata relative to myometrium by 15- to 30-fold at the protein and mRNA level and is localized in endothelial cells. GluR2 pre-mRNA in leiomyoma and myometrial tissues is nearly 100% edited at the Q/R site, indicative of low Ca(2+) permeability of the ion channels. In spontaneous leiomyomata in women or leiomyomata induced in the guinea pig model, there is a likely synergism linking increased production of estradiol and all-trans retinoic acid with up-regulation of nuclear receptor PPARgamma and RXRalpha proteins to support tumor growth. GluR2 might be coupled to this synergism directly or via interleukin-17B, kinesin KIF5 or related genes also up-regulated in leiomyomata. GluR antagonists should be tested as inhibitors of leiomyoma growth.     

Precision medicine in human heart modeling

Precision medicine is a new frontier in healthcare that uses scientific methods to customize medical treatment to the individual genes, anatomy, physiology, and lifestyle of each person. In cardiovascular health, precision medicine has emerged as a promising paradigm to enable cost-effective solutions that improve quality of life and reduce mortality rates. However, the exact role in precision medicine for human heart modeling has not yet been fully explored. Here, we discuss the challenges and opportunities for personalized human heart simulations, from diagnosis to device design, treatment planning, and prognosis. With a view toward personalization, we map out the history of anatomic, physical, and constitutive human heart models throughout the past three decades. We illustrate recent human heart modeling in electrophysiology, cardiac mechanics, and fluid dynamics and highlight clinically relevant applications of these models for drug development, pacing lead failure, heart failure, ventricular assist devices, edge-to-edge repair, and annuloplasty. With a view toward translational medicine, we provide a clinical perspective on virtual imaging trials and a regulatory perspective on medical device innovation. We show that precision medicine in human heart modeling does not necessarily require a fully personalized, high-resolution whole heart model with an entire personalized medical history. Instead, we advocate for creating personalized models out of population-based libraries with geometric, biological, physical, and clinical information by morphing between clinical data and medical histories from cohorts of patients using machine learning. We anticipate that this perspective will shape the path toward introducing human heart simulations into precision medicine with the ultimate goals to facilitate clinical decision making, guide treatment planning, and accelerate device design.