No description available.

<p> Uterine fibroids (UFs) are benign myometrial neoplasms that produce clinically-relevant symptoms in > 25% of all women and represent the leading cause of hysterectomy. Despite their importance, the mechanisms that promote UF growth are still not well-understood. Accumulating evidence indicates that the mechanical environment is altered and this may be a driver of fibroid pathobiology. There has been a proven regulatory mechanism for cellular proliferation with the Hippo pathway and the Yes-associated protein (YAP)/transcriptional coactivator with PDZ-binding domain (TAZ) in lung fibrosis which shares a response to gonadal steroids and serves as a model for this process. Here we assess the differences in YAP/TAZ nuclear localization and responsiveness to mechanical and hormonal cues in primary cells from UF compared to myometrium (Myo). Matched samples of UF and Myo were obtained from premenopausal women with IRB approval. Atomic force microscopy (AFM) was used to determine in-situ stiffness (N = 7). Immunohistochemistry (N = 5) was performed on OCT embedded tissue with YAP/TAZ antibody. For cell culture, UF and Myo were minced and digested with collagenase type 4. Cells (passages 1&ndash;4) were plated sparsely on hydrogels of five defined stiffness or at confluence on tissue culture plastic (N = 4 for both). Immunostaining for YAP/TAZ, Fibronectin (FN), Collagen I (COLI), and Collagen III (COLIII) were performed (N = 4&ndash;5). To perform siRNA, cells were incubated with control or YAP1 (YAP)/WWTR1 (TAZ) siRNA for 72 hours prior to analysis. RNA expression for Endothelin 1 (ET-1) and Connective Tissue Growth Factor (CTGF) was determined. Paired t-test and Wilcoxon sign-rank test were used as appropriate. AFM-measured stiffness was significantly higher in UF compared to Myo (p = 0.03). In freshly isolated cells cultured at confluence on tissue culture plastic, baseline YAP/TAZ nuclear localization was increased in UF when compared to Myo (p = 0.03). When seeded sparsely, decreasing substrate stiffness significantly (p = 0.05) reduced YAP/TAZ nuclear localization for both Myo and UF. Stimulation of confluent cells with 10nM estradiol and 100nM progesterone increased YAP/TAZ nuclear localization, but only in Myo (p = 0.01) as UF cells were constitutively active under these conditions. UFs also exhibited a trend toward increased FN, COLI, and COLIII deposition when compared to Myo. Following siRNA using YAP1 and WWTR1, ET-1 and CTGF were found to be statistically decreased which are known regulators of COLI and FN. These results demonstrate a possible feedback mechanism by which UFs progress by upregulation of matrix deposition, increase in tissue stiffness, and an increase in YAP/TAZ nuclear localization. </p><p>

In squamate reptiles, the evolutionary transition from oviparity to viviparity is accompanied by loss of the calcareous outer eggshell, which suggests significant implications for the role of calcium during embryonic development (Packard et al., 1977). An experiment was designed to evaluate the impact of reduced calcium availability during development in the oviparous corn snake, Pantherophis guttatus (Stewart and Ecay, 2013). Results from that study showed significant decreases in the mass and length of hatchling corn snakes when the outer calcareous eggshell layer was removed during development. In vertebrates, variation in total body length reflects skeletal differences---primarily differences in the number or sizes of vertebrae and/or differences in length of the skull. Skeletal components obviously are affected by the availability of calcium during development. My study was designed to determine the anatomical and developmental bases for the smaller size observed in hatchling snakes subjected to reduction of eggshell calcium during embryonic development. My hypotheses were, as follows: 1) Differences in mass reflected differences in overall length of hatchlings; 2) Differences in length resulted from decreased size of vertebrae---rather than decreased number---and/or decreased length of skulls in hatchlings with reduced calcium. Hatchlings were prepared for skeletal analysis via clearing-and-staining. Vertebrae of each hatchling were counted and size measurements obtained for separate spinal regions (cervical, thoracic, and caudal). Results demonstrate a significant treatment effect on size of vertebrae in the thoracic and anterior caudal regions, as well as length of the skull. These findings suggest that reduced developmental calcium, comparable to the condition in viviparous species, impacts ossification and growth of skeletal elements in late development.

No description available.

No description available.

No description available.

No description available.

No description available.

No description available.

No description available.