Calcineurin subunit B is involved in shell regeneration in Haliotis diversicolor

Abalone shells are mainly composed of two major polymorphs of CaCO3 that are distributed in different layers of the shell. The process of shell biomineralization is controlled by genes and proteins expressed within the mantle epithelium. In this present paper, we conducted a shell regeneration experiment to study the role of HcCNA and HcCNB (individual subunits of calcineurin) in shell biomineralization in H. diversicolor. The results of qPCR showed that HcCNB is upregulated to a greater extent than HcCNA in the mantle after shell notching. In vivo study of the effects of rHcCNB injection showed a significantly higher percentage of regenerated shell length, but not area, in the injected group compared to the control group. In addition, SEM observation of the inner surface of the regenerated shells revealed three different zones including prismatic, nacreous, and a distinct transition zone. Changes in the crystal organization and ultrastructure are clearly evident in these three zones, particularly after 3 weeks of rHcCNB administration. We hypothesize that this is due to faster biomineralization rates in the rHcCNB treated group. Taken together, our results demonstrate that HcCNB participates in shell regeneration in H. diversicolor. As calcineurin subunits have also been implicated in shell formation in bivalves, these findings suggest that calcineurin subunits may play important roles in biomineralization in all conchiferans.

In vivo biomimetic calcification of selected organic scaffolds using snail shell regeneration: a new methodological approach

In vivo biomimetic biomineralization using living organisms known as biomineralizers is currently a major research trend. Industrially cultivated terrestrial snails, such as the common garden snail Cornu aspersum, represent a simple model organism that is ideal for use in experiments on the regeneration of the calcified shell after the excavation of a corresponding shell fragment. The mollusk’s artificially damaged shell is regenerated via the formation of an organic regenerative membrane, which serves as a native template for in vivo biocalcification. In this study, for the first time, a special plexiglass device for non-lethal fixation of living snails, enabling real-time monitoring of their ability to regenerate their shells using digital microscopy, has been proposed and tested. As an alternative to natural biomineralization using the mollusk’s own sources, we propose chitin- and collagen-based templates, which have been shown to be effectively calcified by living snails. The results indicate that the type of organic template used for in vivo biomineralization has a substantial effect on the nature of the mineral phases.

Direct control of shell regeneration by the mantle tissue in the pearl oysterPinctada fucatavia accelerating CaCO3 nucleation

Molluscan bivalves rapidly repair the damaged shells to prevent further injury. However, it remains unclear how this process is precisely controlled. In this study, we applied scanning electronic microscopy, transmission electronic microscopy and histochemical analysis to examine the detailed shell regeneration process of the pearl oysterPinctada fucata. It was found that the shell damage caused the mantle tissue to retract, which resulted in dislocation of the mantle zones to their correspondingly secreted shell layers. However, the secretory repertoires of the different mantle zones remained unchanged. As a result, the dislocation of the mantle tissue dramatically affected the shell morphology, and the unusual presence of the submarginal zone on the nacreous layers caused de novo precipitation of prismatic layers on the nacreous layers. Real-time PCR revealed that the expression of the shell matrix proteins (SMPs) were significantly upregulated, which was confirmed by the thermal gravimetric analysis (TGA) of the newly formed shell. The increased matrix secretion accelerated CaCO3nucleation thus promoting shell deposition. Taken together, our study revealed the close relationship between the physiological activities of the mantle tissue and the morphological change of the regenerated shells.

Natural Pearls

A literature less traveled – peaking between 1900-1920 – draws on pre-classical concepts of crystal growth and a trove of field biology, to understand ectopic shell production, the natural source of pearls. By 1907, grafts from the calcifying mantle epithelium on gonads induced nacre mineralization consistently in Pinctada margaritifera, suggesting that anomalously displaced, readily specialized cells are at least a sufficient cause of natural pearl formation. Otherwise, the epithelial sacks wrapping natural nacreous pearls must specialize for nacre production independently from the shell producing mantle – an idea supported by experiments with shell regeneration, but not amenable to a method of inducing pearl formation. At the time, chasing epithelial cell migration was technically unfeasible, signalling was news, stemness was fiction. Boldly, Jameson & Rubbel [1902-1912] marshalled natural pearl nuclei and shell repairs as mineral records of cells specializing de novo into the shell’s secretory regimes. Much of this paper reenacts the historic debate on the origin of pearls: thence bold ideas connect smoothly with new work both on bone or shell. I replicate Jameson’s choice of samples and revisit his proposal to search for an “agency [other than the] shell-secreting mechanism“ acting on ”replacement cells” as the origin of pearls. Much has changed: specialized epithelial cells reportedly migrate; non-differentiated cells remain available throughout and near the calcifying mantle epithelium – both, open possibilities for natural pearl nucleation. Interest in understanding the latter now connects with results sketching the signalling cascade in cell specialization toward bone morphogenesis. Replicating Jameson’s choice of samples, I describe the more spectacular structural changes in the mineralization of pearls associated with two instances of cell specialization: toward producing one shell material – in the event of natural pearl nucleation, or switching between two in later pearl growth. Clusters of cells producing distinctly novel mineralization – nacre over fibrous-prismatic aragonite – could be singled out next to natural pearls by Jameson. The possibility has not been probed in roughly a hundred years. Natural pearl nucleation as a cellular event has never been explored.