POSTER ABSTRACTS - ISAKOS

cultures of ATDC5 cells (cultured for 28 days in the
differentiation medium).
(3) Analysis of mRNA expression by cDNA
micraorray and real-time PCR procedures:
Microarray analysis was performed as described
previously [4]. To control for labeling differences,
experiment was carried out in duplication in
which the fluorescent dyes were switched during
cDNA synthesis for each experiment. Real-time
PCR analysis was carried out to verify the
microarray data using a Lightcycler fluorescence
temperature rapid-air cycler (Roch Molecular
Biochemicals). PCR was performed using SYBR
Green PCR system according to the manufacture’s
instructions (Roch Molecular Biochemicals).
(4) Statistical analysis: To examine statistical
significance for frequencies of functionally
categorized genes in each cluster, values of the
other groups in the relevant cluster, and values of
other clusters in the relevant group were analyzed
with Fisher’s exact test by computing with the
statistical program StatView (SAS, Cary, NC)
RESULTS:
516 genes were up- or down-regulated 1.5-fold or
more at least at one time point after chondrogenic
induction. The 516 genes were subjected to
clustering analysis based on expression patterns,
and were classifierd into 5 clusters. Each cluster
was characterized as follows: cluster A, gradual or
lagging decrease; cluster B, transient decrease;
cluster C, lagging increase; cluster D, gradual
increase; and cluster E, transient increase. In
cluster A, genes for the category RNA turnover and
protein turnover were significantly frequent
gradual or lagged decreases in expression of
genes for RNA turnover and protein turnover may
reflect decreasing proliferation rates of ATDC5
cells during differentiation. In cluster C, genes for
the category RNA turnover and cytokines and
growth factors were significantly frequent. Lagged
increases in expression of genes for cytokines and
growth factors in the later chondrogenic
differentiation stages appear to be related to
complexity of the differentiation processes. 9
genes for cytokines and growth factors were upregulated
at least at one time point after
chondrogenic induction. 8 of these genes were
categorized in cluster C, 2 of these genes were
categorized in cluster D. In cluster C included
these for osteoglycin (Ogn, accession No.
AI596220), transforming growth facter beta 2
(Tgfb2, No X57413), interferon activated gene 203
(Ifi203, No BC008167), lymphocyte antigen 86
(Ly86, No AB007599), rcd1 (required for cell
differentiation) homolog 1 (S. pombe) (Rqcd1, No
D87957), pre-B-cell colony-enhancing factor (Pbefpending,
No U02020), chemokine (C-C motif)
ligand 9 (Ccl9, No U15209) and tumor necrosis
factor receptor superfamily, member 12a
(Tnfrsf12a, No AK013438).
DISCUSSION:
In the postgenome sequencing era, the
development of functional genome resources is
essential to understand and use information
generated from the genome sequencing projects.
Full-length cDNA cloning and sequencing most
significantly contributes to the development of
this technology because of clarification of coding
and noncoding mRNA and because of suitability
for functional analysis. Therefore, it is important
to construct full-length enriched cDNA library and
to obtain full-length cDNA clones. Our library is
expected to be a useful tool to analyze gene
functions and interactions among the genes in
chondrogenesis. In this study, many candidate
genes associated with chondrogenesis were
screened rapidly. In these genes, expression
patterns of some important genes were confirmed
with the real-time PCR method. Thus, our inhouse
cDNA microarray system is evaluated as an
excellent tool to analyze the expression status of
many genes in this field. The large number of
differentially expressed genes identified from this
analysis and the characterization of these genes
will provide valuable information to precisely
understand the biology of chondrogenesis.
E-poster #731
A New Evaluation Method for Measuring the
Mechanical Properties of Meniscus Using
Ultrasound
Yasuyuki Mizuno, Kyoto, JAPAN, Presenter
Takashi Nakamura, Kyoto, JAPAN
Yasuaki Nakagawa, Kyoto, JAPAN
Makoto Takenaka, Kyoto, JAPAN
Keiji Ando, Kyoto, JAPAN
Hiroshi Kuroki, Kyoto, JAPAN
Koji Mori, Kyoto, JAPAN
Takashi Suzuki, Kyoto, JAPAN
Ken Ikeuchi, Kyoto, JAPAN
Sadami Tsutsumi, Kyoto, JAPAN
Kyoto University, Kyoto, JAPAN
Purpose: The object of this study was to examine
the usefulness of ultrasound for assessing the
mechanical properties and quality of the
meniscus (degree of degeneration).