Page 73 - Vibrational Spectroscopic Imaging for Biomedical Applications

Algal Cells, Cartilage, and IRENI 49

over the end of the bones, is a matrix-rich tissue composed primarily
of type II collagen and large hydrophilic proteoglycans. Normally
the matrix is unmineralized. During aging and with osteoarthritis,
pathogenic calcium crystals deposit in the pericellular matrix around
chondrocytes.
CPPD crystals cause both an acute inflammatory as well as a
41
chronic noninflammatory polyarticular arthritis. BCP crystals are
associated with severe degenerative arthritis and a variety of nonin-
flammatory articular syndromes such as Milwaukee shoulder syn-
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drome. These crystal-associated syndromes are common, often
underdiagnosed, and produce irreversible joint destruction in elderly
patients. No specific therapies are currently available.

2.4.2 Current Methods of Crystal Identification
Our understanding of how and why calcium crystals form in the nor-
mally unmineralized matrix of articular cartilage has been hampered
by inadequate methods of crystal analysis. This is certainly problem-
atic in the clinic where it results in missed diagnostic opportunities,
but is also a major issue in the research laboratory. Standard methods
of crystal identification in patient samples are tailored to the presence
of relatively large numbers of crystals in synovial fluids. Typically,
CPPD crystals in synovial fluids are identified morphologically under
compensated polarizing light microscopy. CPPD crystals appear as
weakly positively birefringent rhomboid-shaped crystals. In contrast,
BCP crystals have no characteristic features under light microscopy.
Alizarin red staining has been used to identify these crystals in syno-
vial fluid, where they appear as large amorphous reddish-orange
deposits. Unfortunately, alizarin red staining can be difficult to inter-
pret, and is often misread. 43
For research purposes, other more sophisticated and expensive
techniques have been used to validate the presence of calcium-
containing crystals in biologic models of crystal formation. These
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include FTIR spectral analysis and x-ray diffraction. X-ray diffrac-
tion requires relatively pure dry samples. FTIR spectral microanaly-
sis has proven quite useful in biologic samples with abundant
crystals, but is pushed beyond its limits when crystals are very small
and sparse. Using FTIR spectral microanalysis, for example, crystal
identification proved impossible in models using chondrocyte
monolayers, where crystals were rare and mixed with abundant
complex biological material. 45

2.4.3 Biologic Models of Calcium-Containing
Crystal Formation
No animal models of calcium-containing crystal formation currently
exist. Cell and tissue culture models, however, are well described.
Normal articular cartilage can be removed from an adult animal and