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Updated: Jun 25, 2025
A Semi-Automated and Reproducible Biological-Based Method to Quantify Calcium Deposition In Vitro
Published on: June 2, 2022
Dystrophic calcinosis: structural and morphological composition, and evaluation of ethylenediaminetetraacetic acid
Phillip Lee1, Lorraine Green2,3,4, Bartosz Marzec1
1School of Chemistry, University of Leeds, Woodhouse Lane, Leeds, LS2 9JT, UK.
Background:
To perform a detailed morphological analysis of the inorganic portion of two different clinical presentations of calcium-based deposits retrieved from subjects with SSc and identify a chemical dissolution of these deposits suitable for clinical use.
Methods:
Chemical analysis using Fourier Transform IR spectroscopy ('FTIR'), Raman microscopy, Powder X-Ray Diffraction ('PXRD'), and Transmission Electron Microscopy ('TEM') was undertaken of two distinct types of calcinosis deposits: paste and stone. Calcinosis sample titration with ethylenediaminetetraacetic acid ('EDTA') assessed the concentration at which the EDTA dissolved the calcinosis deposits in vitro.
Results:
FTIR spectra of the samples displayed peaks characteristic of hydroxyapatite, where signals attributable to the phosphate and carbonate ions were all identified. Polymorph characterization using Raman spectra were identical to a hydroxyapatite reference while the PXRD and electron diffraction patterns conclusively identified the mineral present as hydroxyapatite. TEM analysis showed differences of morphology between the samples. Rounded particles from stone samples were up to a few micron in size, while needle-like crystals from paste samples reached up to 0.5 µm in length. Calcium phosphate deposits were effectively dissolved with 3% aqueous solutions of EDTA, in vitro. Complete dissolution of both types of deposit was achieved in approximately 30 min using a molar ratio of EDTA/HAp of ≈ 300.
Conclusions:
Stone and paste calcium-based deposits both comprise hydroxyapatite, but the constituent crystals vary in size and morphology. Hydroxyapatite is the only crystalline polymorph present in the SSc-related calcinosis deposits. Hydroxyapatite can be dissolved in vitro using a dosage of EDTA considered safe for clinical application. Further research is required to establish the optimal medium to develop the medical product, determine the protocol for clinical application, and to assess the effectiveness of EDTA for local treatment of dystrophic calcinosis.
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