A dynamic cell adhesion surface regulates tissue architecture in growth plate cartilage.

The architecture and morphogenetic properties of tissues are founded in the tissue-specific regulation of cell behaviors. In endochondral bones, the growth plate cartilage promotes bone elongation via regulated chondrocyte maturation within an ordered, three-dimensional cell array. A key event in the process that generates this cell array is the transformation of disordered resting chondrocytes into clonal columns of discoid proliferative cells aligned with the primary growth vector. Previous analysis showed that column-forming chondrocytes display planar cell divisions, and the resulting daughter cells rearrange by ∼90° to align with the lengthening column. However, these previous studies provided limited information about the mechanisms underlying this dynamic process. Here we present new mechanistic insights generated by application of a novel time-lapse confocal microscopy method along with immunofluorescence and electron microscopy. We show that, during cell division, daughter chondrocytes establish a cell-cell adhesion surface enriched in cadherins and ß-catenin. Rearrangement into columns occurs concomitant with expansion of this adhesion surface in a process more similar to cell spreading than to migration. Column formation requires cell-cell adhesion, as reducing cadherin binding via chelation of extracellular calcium inhibits chondrocyte rearrangement. Importantly, physical indicators of cell polarity, such as cell body alignment, are not prerequisites for oriented cell behavior. Our results support a model in which regulation of adhesive surface dynamics and cortical tension by extrinsic signaling modifies the thermodynamic landscape to promote organization of daughter cells in the context of the three-dimensional growth plate tissue.

The anti-estrogenic activity of sediments from agriculturally intense watersheds: assessment using in vivo and in vitro assays.

The goal of the current study was to determine whether sediments from agriculturally intense watersheds can act as a potential source of anti-estrogenic endocrine-disrupting compounds. The specific objectives of the current study were to determine (1) whether female fathead minnows (Pimephales promelas) experience alterations in endocrine function when exposed to sediments collected from agriculturally intense watersheds and (2) if these sediments display anti-estrogenic activity in an in vitro assay. In addition, sediment samples were analyzed for the presence of steroid hormones and pesticides associated with local agricultural practices. To accomplish this, sediments and water were collected from three sites within two agriculturally intense Nebraska watersheds (Bow Creek and the Elkhorn River). In 2009, minnows were exposed to sediment and/or water collected from the two Bow Creek sites (East Bow Creek and the Confluence) in the laboratory, while in 2010, minnows were exposed to sediment and/or water from East Bow Creek, the Confluence and the Elkhorn River. Following the 7-day exposure period, the hepatic mRNA expression of two-estrogen responsive genes, estrogen receptor α (ERα) and vitellogenin (Vtg) was determined. In 2009, females exposed to Confluence sediments, in the presence of laboratory water or Confluence water, experienced significant reductions in ERα expression relative to unexposed and Confluence water-exposed females. The defeminization of these females suggests the presence of a biologically available anti-estrogenic compound in sediments collected from this site. In 2010, sediments were assessed for anti-estrogenic activity on days 0 and 7 of the exposure period using a 4-h yeast estrogen screen. Lipophilic extracts (LEs) of day 0 sediments collected from the Confluence and the Elkhorn River induced significant reductions in the estrogenic reporter activity of treated yeast cultures suggesting the presence of a lipophilic anti-estrogenic compound in these extracts. Chemical analysis revealed the presence of a variety of steroid hormones, including those associated with the production of beef cattle (i.e. ß-trenbolone, α-zearalanol and α-zearalenol), in sediments indicating that compounds utilized by local beef cattle operations are capable of entering nearby watersheds. Overall, the results of this study indicate that an environmentally relevant anti-estrogenic compound is present in sediments from agriculturally intense watersheds and that this compound is bioavailable to fish. Furthermore, the presence of steroid hormones in sediments from these watersheds provides evidence indicating that steroids are capable of sorbing to sediments.