This study aims to investigate whether adipose tissue inflammation plays a role in PAR2-mediated joint damage in osteoarthritis. The animal model used is destabilization of the medial meniscus in mice. Preliminary results show that PAR2 may be expressed in the infrapatellar fat pad in mice and is upregulated in human chondrocytes stimulated by inflammatory cytokines. Conditioned media from human fat tissue explants upregulated PAR2 expression and MMP1 levels in human chondrocyte cell lines, suggesting inflammatory factors from adipose tissue may contribute to joint inflammation and osteoarthritis progression. The study will further examine the role of PAR2 in osteoarthritis pathology using cell culture, tissue explants, and a mouse

1. • Osteoarthritis (OA) is major joint disease (12%
prevalence) and healthcare burden
• OA traditionally thought of as passive ‘wear and tear’
disease
• No disease-modifying treatments available
• Inflammation now recognised to play role in OA
• Obesity is major risk factor for OA (not only due to
weight bearing)
• Chronic, low level inflammation associated with
obesity
• Molecular links between OA, obesity and inflammation
not fully understood
Research which leads to potentially novel
approaches to treatment are of fundamental
importance as the costs of treating obesity and
associated diseases continue to rise.
Preliminary Results – In Vitro
Figure 2: a) the tissue and components of a normal joint including synovium, meniscus of the knee and
smooth cartilage on the surface of the long bones, (femur and tibia in knee) in comparison to b) a joint
showing mild arthritic changes such as an inflamed and thickened synovium, the appearance of
osteophytes and cartilage which has become rough and thin.
http://www.arthritisresearchuk.org/arthritis-information/conditions/arthritis/what-is-arthritis.aspx
Figure 1: Diagram showing activation of PAR2 (McIntosh et al, 2003)
Protease Activated Receptor 2 (PAR2)
a b
Preliminary Results – In Vivo
PAR2 +ve staining
suggestive of mast cells
or macrophages
Immunohistochemistry
Wild type DMM 14 days
Infrapatellar Fat Pad
SAM-11 – PAR2
Figure 3: Knee joint showing
normal structures of a hinge joint
and indicating the proximity of the
synovial membrane, meniscus,
articular cartilage and
Infrapatellar Fat Pat IPFP within
the join structure
www.interactive-biology.com
Osteoarthritis, Obesity & Inflammation
To investigate if:
• PAR2 is present in IPFP (and thus
may contribute to joint inflammation)
• PAR2 activating proteases are
present in IPFP
• PAR2 is more abundant in obese
than lean animals
• Experimental OA progression is
accelerated during obesity
• Blocking PAR2 ameliorates OA
progression in obesity
Aim of ProjectJoint Tissue
IPFP
Adipocytes
Animal Model
Figure 6 a & b: Murine
Infrapatellar Fat Pad stained
with SAM11 antibody and
exhibiting first evidence that
PAR2 may be expressed in
IPFP.
a
b
Does adipose tissue inflammation play a role in PAR2 mediated joint damage in osteoarthritis?
Margaret Fegen, UWS, School of Science & Sport, Paisley Campus.
Director of Studies : Litherland, G., Co-supervisors: Crilly, A., Lockhart, J., Ferrell, W.
IPFP: The IPFP is the pad of fat which is situated below and to
the back of the patella in the knee joint. In such close proximity
to the other key tissues in the joint such as bone, cartilage and
synovium, there may be some inflammatory cytokines arising
from the adipose tissue which may have an impact on disease
progression.
Figure 5: Structures in the knee involved in DMM
(Glasson, Blanchet and Morris, 2007)
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Methods
n.s. n.s.
The animal model used for this study is Destabilisation
of the Medial Meniscus (DMM). The stability of the
knee is dependent upon the tendons and ligaments in
and around the joint to support movement and body
weight. OA can be induced as a result of surgical
incision into the Medial Meniscus (MM) which results in
destabilisation of the joint to the medial component of
the joint.
Cell Culture:
Human Articular
Chondrocytes
(OA patients),
SW1353 cell
line
Explants
Human
Cartilage
Human
Adipose
tissue
Figure4: Resources including the use of
Primary human cells (a), fat (c) and
cartilage tissue (b) from OA patients,
chondrocyte cell line and an established
animal model of DMM (d).Experimental
techniques will include cell and tissue
culture, Immunohistochemistry, qPCR,
RNA isolation.
Results from human studies will be used to
validate and complement the information
gained from in vivo model.
DMM
Figure 7: Figure 7: Detection of PAR2 by qPCR on human OA primary chondrocytes shows that
PAR2 expression is undetectable in basal conditions in vitro, except when cells are stimulated by
the cytokine oncostatin M (OSM) or the phorbol ester PMA (a). MMP1 (a protease involved in
extracellular matrix breakdown) was upregulated in the presence of inflammatory cytokines but
not PAR2 activatory peptide (SLIG) (b). Statistical analysis by student T-Test, n=1 with 6
replicates of each condition.
a
b
c
d
Figure 8: Using cell line SW1353 which were treated with media that has been conditioned
by human fat tissue explants from OA patients showing that there are factors which show
expression of PAR2 (a) and also significant levels of MMP1 being expressed. Statistical
analysis by student T-Test with n=10 with 6 replicates of each condition.
a
a b
b
PAR2
• Part of 4 member family of G-protein coupled
receptors
• Unique protease-dependent activation involving
unmasking of ‘tethered ligand’
• Multiple pathological roles described, especially
in inflammatory pathologies
• CMS researchers have uncovered important
roles for PAR2 in pathogenesis of rheumatoid
arthritis and OA
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n.s.
n.s.
n.s.
n.s.
n.s.
n.s. n.s.