Osteoarthritis (OA), a chronic degenerative condition of the diarthrodial joints is
most predominant amongst the middle aged and elderly population, where it is
associated with progressive disability and deformity of the joints. Whilst the exact
mechanism of OA pathogenesis is currently unknown, chondrocyte death by
apoptosis is seen as one of the main contributors to the disease process. It is
increasingly recognised that the overproduction of mediators such as Nitric Oxide
(NO) in Osteoarthritic cartilage contributes to the disease pathology partly by
inducing chondrocyte apoptosis. Therefore, agents, which protect against NOinduced
mitochondrial injury, may have therapeutic potential in Osteoarthritis.
The CRF related peptide, Urocortin I (UCNI), has been shown to protect the
human chondrocyte cell line, C-20/A4, from SNAP/NO induced apoptotic death,
but little is yet known about its mechanism of action. Therefore the aim of this
project is to investigate the potential mechanism through which UCNI exerts its
observed protective effects. The studies documented here, have demonstrated
the endogenous expression of this cytoprotective peptide, UCNI, by C-20/A4
chondrocytes and have shown that treatment of chondrocytes with SNAP (a NO
donor) resulted in a significant increase in apoptosis which was abrogated by the
addition of exogenous UCNI. Therefore, an investigation into the potential
mechanism for UCNI mediated protection against apoptosis in chondrocytes (i.e.
chondroprotection) detected the mRNA expression of both CRFR1 and CRFR2
receptors specifically the CRFR1? and CRFR2? splice variants by the C-20/A4
chondrocytes. However the CRFR antagonist, ??helical CRH failed to induce
apoptosis or abrogate the protective effects of UCNI against SNAP/NO induced
apoptosis. RT-PCR studies have also demonstrated C-20/A4 mRNA expression
of the Kir6.1, Kir6.2, SUR1 and SUR2B subunits of the KATP channels suggesting
that functional KATP channels are also present in these cells, which represent a
further putative mechanism of action for UCNI.
Quantitative PCR studies were performed to determine regulation of CRFR and
Kir6.1 expression by UCNI. These studies showed that treatment of chondrocytes
with UCNI and SNAP alone and concurrently resulted in no significant change in
mRNA expression of either CRF receptors or kir6.1 (the pore forming subunit of
the mitochondrial KATP channel). Treatment of chondrocytes with Diazoxide, a
mitochondrial KATP channel opener, resulted in a reduction in SNAP induced
apoptosis, mimicking that seen for UCNI, suggesting a similar protective role for
these channels in chondrocytes, to that proposed in other cell types and
supporting these channels as potential targets for UCNI in chondroprotection.

Tagliamonte A. Tryptophan

Papers

Urocortin: A Novel Player in Cardiac Control