Publication: Malachite green and phthalocyanine-silver reactions
reveal acidic phospholipid involvement in calcification
of porcine aortic valves in rat subdermal model
Authors
Ortolani, F. ; Petrelli, L. ; Nori, S.L. ; Gerosa, G. ; Spina, M. ; Marchini, M.
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Publisher
Murcia : F. Hernández
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DOI
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info:eu-repo/semantics/article
Description
Abstract
Subdermal implant models are helpful in the
study of calcification "in vivo” and for testing
anticalcific treatments. After implantation of porcine
aortic valve leaflets in rat subcutis, we previously found
that glutaraldehyde-Cuprolinic blue reactions (GA-CB)
at low pH induce favourable tissue unmasking from
mineral deposits, and visualize peculiar, electrondense
layers that outline the calcifying cells and matrix vesiclelike
structures. The layer-forming material seemed to
consist of acidic phospholipids because of its anionic
nature and differential susceptibility to
chemical/enzymatic extractivity. In the present
investigation, pre-embedding glutaraldehyde-Malachite
green (GA-MG) reactions and subsequent osmium postfixation
were compared with pre-embedding GA-CB
reactions, combined with post-embedding von Kossa
silver staining (GA-CB-S), to assess whether the layerforming
material is actually composed of acidic
phospholipids and exhibits calcium-binding properties.
After lowering standard pH, GA-MG reactions also
caused sample demineralization and the appearance of
pericellular osmium-MG-reactive layers comparable to
CB-reactive ones. Moreover, GA-CB-S reactions
showed that major silver precipitation was superimposed
to the CB-reactive layers, whereas minor metal extraprecipitation
occurred at three distinct, additional sites.
These results demonstrate that a unique process of cell
degeneration occurs in this calcification model, in which
acidic phospholipids accumulate at cell surface,
replacing cell membrane and acting as major apatite
nucleator. However, the overall observations are
consistent with the hypothesis that certain phases are
common to the various types of normal and/or abnormal
calcification.
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