Objective—To establish an in vitro assay and determine
the differential suppressive activity of non
steroidal anti-inflammatory drugs (NSAID) on cyclooxygenase
(COX)-1 and COX-2 isoenzymes in dogs.
Procedure—COX activity was evaluated in the presence
and absence of 4 NSAID (meloxicam, tolfenamic
acid, carprofen, and ketoprofen), using a canine
monocyte/macrophage cell line that constitutively
expresses COX-1, but can be induced to express
COX-2 when incubated with lipopolysaccharide.
Inhibition of prostaglandin E2 (PGE2) synthesis by
each NSAID was measured by enzyme immunoassay
and attributed to specific COX-1 or COX-2 activity
through assessment of COX messenger RNA expression
by use of northern blot analysis and reverse transcription-
polymerase chain reaction (RT-PCR). The
COX selectivity of each drug was evaluated from
dose-response curves by calculating a ratio (COX-
1:COX-2) of inhibitory concentration values on the
basis of concentrations that reduced PGE2 by 50% in
each COX model.
Results—Meloxicam and tolfenamic acid preferentially
inhibited COX-2, with meloxicam inhibiting COX-2
activity 12 times more effectively than COX-1 activity.
Carprofen was only 1.75 times more selective for
COX-2 than for COX-1, and ketoprofen was slightly
more selective for COX-1.
Conclusions—COX-1 and COX-2 were differentially
sensitive to inhibition in vitro by NSAID.
Meloxicam and tolfenamic acid were selective for
COX-2. Effects of carprofen and ketoprofen
approached equipotency against both isoenzymes.
Selective COX-2 inhibitors are a new class of drugs
with anti-inflammatory effects similar to conventional
NSAID but with fewer adverse effects.
Development of these agents for veterinary use
would be facilitated by the convenience of using a
canine cell line as a model system to screen COX-
1 and COX-2 inhibitor activities in vitro. (Am J Vet
Objective—To examine cyclooxygenase (COX)
expression in canine platelets and Madin-Darby
canine kidney (MDCK) cells in culture.
Sample Population—Canine platelets and MDCK cells.
Procedure—Total RNA was recovered from isolated
canine platelets and MDCK cells. Northern blot analysis
and reverse transcription-polymerase chain reaction (RTPCR),
using complementary DNA probes and primers
designed from the human COX sequences, were used
to determine COX-1 and -2 (cyclooxygenase isoforms 1
and 2) messenger RNA (mRNA) expression.
Results—Following northern blot analysis, canine
platelets were found to express only the 2.8-kb COX-
1 transcript; COX-2 was not detected. Canine MDCK
cells expressed the 4.5-kb COX-2 transcript, in addition
to the 2.8-kb COX-1 transcript. A single DNA
band of 270 base pairs was identified following gel
electrophoresis of the product obtained from RT-PCR
of mRNA from canine platelets. Sequencing revealed
that this PCR product was 90% homologous to a portion
of the human COX-1 gene (Genbank M59979).
Conclusions and Clinical Relevance—Detection of
COX-1 by RT-PCR of RNA obtained from canine
platelets is a novel finding. The 90% homology of the
PCR product with the human sequence suggests
strong conservation between the canine and human
COX-1 gene. Cloning and sequencing of the canine
gene will be required to fully characterize homologous
regions. Because of the importance of COX in the
inflammatory process and as a potential target of currently
available nonsteroidal anti-inflammatory drugs
(NSAID), a better understanding of canine COX may
improve our ability to use NSAID appropriately,
achieve efficacy, and avoid potential adverse drug
effects in dogs. (Am J Vet Res 2000;61:1512–1516)