The Spatial and Temporal Expression Patterns of Integrin α9β1 and One of Its Ligands, the EIIIA Segment of Fibronectin, in Cutaneous Wound Healing

The fibronectins (FN) comprise a family of adhesive extracellular matrix proteins thought to mediate important functions in cutaneous wounds. Plasma fibronectin (pFN) extravasates for days from intact hyperpermeable vessels following injury whereas mRNAs encoding the cellular fibronectins (cFN) that include two segments, termed EIIIA (EDA) and EIIIB (EDB), are expressed by wound cells. Wounds in mice null for pFN appear to heal normally whereas those in EIIIA null mice exhibit defects, suggesting that cFN may play a role when pFN is missing. Integrin α9β1, a receptor for several extracellular matrix proteins as well as the EIIIA segment, is expressed normally in the basal layer of squamous epithelia. We report results from immunohistochemistry on healing wounds demonstrating that EIIIA-containing cFN are deposited abundantly but transiently from day 4 to 7 whereas EIIIB-containing cFN persist at least through day 14. Elevated expression of α9β1 is seen in basal and suprabasal epidermal keratinocytes in wounds. The spatial expression patterns of cFN and α9β1 are distinct, but overlap in the dermal–epidermal junction, and both are expressed contemporaneously. These observations suggest a role for α9β1–EIIIA interactions in wound keratinocyte function

The distribution of glycoconjugates in the basal lamina and ECM during esophageal muscle formation in embryos of the starfish Pisaster ochraceus as revealed by lectin hsitochemistry

Morphogenetic events consist of complex interactions of cells and extracellular materials resulting in the movement and rearrangement of groups of cells and their subsequent differentiation to form organs or organ systems. Although we can predict these movements for any given event, we have little understanding of how morphogenesis is controlled. In the starfish Pisaster ochraceus. assembly of mesenchyme cells on one particular region of the larval gut, the oesophagus, and their subsequent differentiation into muscle is an example of a simple morphogenetic event which is readily accessible for study. In their migration to the gut, the mesenchyme cells travel through a rich substrate of ECM. Upon their arrival at the presumptive esophagus, they come to settle on the BL underlying the endodermal epithelium. It is quite possible that interactions between the mesenchymal cells and 'the ECM/basal lamina are important in directing and regulating their differention into muscle. The basal laminae and ECM of vertebrates and invertebrates is rich in glycoconjugates, including glycoproteins, proteoglycans and glycosaminoglycans. Ultrastructural studies of embryos of the asteroid Pisaster ochraceus have demonstrated that at the late gastrula stage, the endodermal basal lamina is thinner and less alcianophilic in the esophageal region. FITC and colloidal gold labelled lectins, which act as specific probes for carbohydrate moities, usually those at the terminal end of oligosaccharide chains, have been used to localize these sugars at the light and electron microscope levels. These studies show that a heterogeneity exists with respect to terminal sugars in the basal lamina, i.e. lectin binding of the basal lamina is not uniform in all regions of the embyro. Specifically, a statistical analysis of lectin binding determined that labelling with the two lectins, Au₂₅-Con A and Au₂₅-LFA was significantly reduced in the esophageal region as compared with the other regions of the embyro, while labelling of the BL with AU₂₅-DBA showed a similar intensity in all areas of the embryo. These results confirmed the alcian blue results described above and suggest that there are some sugar containing molecules, perhaps specific glycoproteins, GAGs and/or proteoglycans, which are present in reduced quantities in this region. In addition, these studies show a distinct labelling pattern of the ECM through which the mesenchymal cells migrate on route to the esophagus. Different lectins label different regions of the ECM, however it can not yet been said whether there is a regionally distinct pattern in the area of the migratory path of mesenchymal cells to the esophagus. Proteoglycans and GAGs are involved in cell movement in vertebrates and sulfated glycoconjugates have been shown to be necessary for mesenchyme cell movement in echinoids. A decrease in proteoglycans and GAGs in the esophageal BL could therefore help to direct movement of the presumptive muscle cells to the esophagus by providing a "stop" signal.Medicine, Faculty ofGraduat

Characterization of starfish yolk and cortical granule proteins, and of a novel extracellular proteoglycan implicated in digestive tract morphogenesis

Extracellular matrix (ECM) is thought to play a major role in morphogenesis by influencing processes such as cell migration and differentiation, although the specific mechanisms involved are poorly understood. This study examined the ECM and egg storage granules of starfish (Pisaster ochraceus) embryos, and attempted to identity components important for digestive tract morphogenesis. Three monoclonal antibodies were developed with specificities for the ECM, yolk and cortical granules in Pisaster eggs and embryos. These antibodies were then used to localize, isolate and characterize the antigens through early development, using immunohistochemistry, immunocytochemistry, immunochemical and biochemical techniques. The first antibody, PM1, binds to a large extracellular proteoglycan, which appears in the blastocoel matrix at mid-gastrulation, and is synthesized only by endodermally-derived tissues. The use of PM1 antibody as a function blocking agent in live embryo cultures suggested that it plays an important role in digestive tract morphogenesis. A second antibody recognizes a protein localized in cortical granules of unfertilized eggs. The majority of these granules are located in the peripheral egg cytoplasm and are released at fertilization. However, a second morphologically identical population of granules remain dispersed throughout the egg cytoplasm, and appear to contribute to ECMs of the developing embryo, including the blastocoel ECM, basement membranes, and the hyaline layer. The function of this protein is currently unknown; however, it has a different storage and secretion profile from the PM1 proteoglycan, suggesting its role in the blastocoel matrix may be different. A third antibody recognizes proteins stored in yolk granules located throughout the egg and cells of the developing embryo, which do not appear to contribute to ECMs during embryogenesis. Partial biochemical characterizations using the anti-yolk antibody revealed that there are several molecular species of yolk proteins present in the oocyte, and that their molecular composition changes during embryogenesis. Depletion of the yolk proteins is not significant until the larval stage, suggesting that they do not play a major role until later in development.Medicine, Faculty ofGraduat

Estimates of anthropogenic carbon uptake from four three-dimensional global ocean models

We have compared simulations of anthropogenic CO$_2$ in the four threedimensional ocean models that participated in the first phase of the Ocean Carbon-Cycle Model Intercomparison Project (OCMIP), as a means to identify their major differences.Simulated global uptake agrees to within $\pm$19%, giving a range of 1.85$\pm$0.35 PgC yr$^{-1}$ for the 1980-1989 average. Regionally, the Southern Ocean dominates the present-day air-sea flux of anthropogenic CO$_2$ in all models, with one third to one half of the global uptake occurring south of 30°S. The highest simulated total uptake in the Southern Ocean was 70% larger than the lowest. Comparison with recent data-based estimates of anthropogenic CO$_2$ suggesthat most of the models substantially overestimate storage in the Southern Ocean; elsewhere they generally underestimate storage by less than 20%. Globally, the OCMIP models appear to bracket the real ocean's present uptake, based on comparison of regional data-basedstimates of anthropogenic CO$_2$ and bomb $^{14}$C. Column inventories of bomb $^{14}$C have become more similar to those for anthropogenic CO$_2$ with the time that has elapsed between the Geochemical Ocean Sections Study (1970s) and Word Ocean Circulation Experiment (1990s) global sampling campaigns. Our ability to evaluate simulated anthropogenic CO$_2$ would improve if systematic errors associated with the data-based estimates could be provided regionally