ALBERT

Description: To understand mineral transport pathways for shell secretion and to assess differences in cellular activity during mineralization, we imaged with TEM and FE-SEM ultrastructural characteristics of outer mantle epithelium (OME) cells. Imaging was carried out on Magellania venosa shells embedded/etched, chemically fixed/decalcified and high-pressure frozen/freeze-substituted samples from the commissure, central shell portions and from puncta. Imaging results are complemented with morphometric evaluations of volume fractions of membrane-bound organelles. At the commissure the OME consists of several layers of cells. These cells form oblique extensions that, incross-section, are round below the primary layer and flat underneath fibres. At the commissure the OME is multi-cell layered, in central shell regions it is single-cell layered. When actively secreting shell carbonate extrapallial space is lacking, because OME cells are in direct contact with the calcite of the forming fibres. Upon termination of secretion, OME cells attach via apical hemidesmosomes to extracellular matrix membranes that line the proximal surface of fibres. At the commissure volume fractions for vesicles, mitochondria and lysosomes are higher relative to single-cell layered regions, whereas for endoplasmic-reticulum and Golgi apparatus there is no difference. FE-SEM, TEM imaging reveals the lack of extrapallial space between OME cells and developing fibres. In addition, there is no indication for an amorphous precursor within fibres when these are in active secretion mode. Accordingly, our results do not support transport of minerals by vesicles from cells to sites of miner-alization, rather by transfer of carbonate ions via transport mechanisms associated with OME cell membranes.

Description: The fibrous calcite layer of modern brachiopod shells is a hybrid composite material and forms a substantial part of the hard tissue. We investigated how cells of the outer mantle epithelium (OME) secrete calcite material and generate the characteristic fibre morphology and composite microstructure of the shell. We employed AFM, FE-SEM, and TEM imaging of embedded/etched, chemically fixed/decalcified and high-pressure frozen/freeze substituted samples. Calcite fibres are secreted by outer mantle epithelium (OME) cells. Biometric analysis of TEM micrographs indicates that about 50% of these cells are attached via hemidesmosomes to an extracellular organic membrane present at the proximal, convex surface of the fibres. At these sites, mineral secretion is not active. Instead, ion transport from OME cells to developing fibres occurs at regions of closest contact between cells and fibres, however only at sites where the extracellular membrane at the proximal fibre surface is not developed yet. Fibre formation requires the cooperation of several adjacent OME cells. It is a spatially and temporally changing process comprising of detachment of OME cells from the extracellular organic membrane, mineral secretion at detachment sites, termination of secretion with formation of the extracellular organic membrane, and attachment of cells via hemidesmosomes to this membrane.