Identification of inflammatory mediators in spinal liquor in relation to synovial nerve sprouting and pain in osteoarthritis

Osteoarthritis (OA) is a progressing degenerative joint disorder, characterized
by joint pain and functional impairment, leading to disability. Pain is the
most important defining clinical presentation of OA. This persistent pain has a
major impact of the quality of life, but is difficult to treat. Current
treatment provides modest relief at best and inadequately controlled pain is a
major reason for total joint replacement. Moreover, in approximately 20% of
patient that have had total joint replacement pain remains. The efficacy of
'first-line' agents such as paracetamol is hard to distinguish from placebo,
and our most common effective therapies, NSAIDs and opioids have very small
effect sizes on OA pain. Therefore, better understanding of the mechanisms
driving persistent OA pain is required to develop new therapeutic approaches
that specifically target deteriorating pain conditions in OA.
Local joint inflammation,altered cartilage and bone turnover in OA have been
implicated to lead to a range of molecular mediators that mediate OA pain.
However, clinically there is disparity between the degree of pain perception
and the extent of joint damage in subjects with OA. It remains largely unknown
why a relatively poor correlation exists in OA between the radiologic signs of
OA (eg, joint-space narrowing, erosive changes) and the severity of pain as
well as the specific mechanisms that drive spontaneous (at rest) versus
movement (at activity)-evoked joint pain in OA. Research with animals has
expanded our understanding of these potential mechanisms that might drive OA
pain independently of joint damage. However, translation of these findings to
humans with OA is very limited and void the development of novel analgesics to
treat pain in OA.

Pain can arise as a result from changes in the sensory nervous system at
several levels: 1) central mechanism including activation of spinal cord glial
cells to produce pro-inflammatory cytokines and neuronal plasticity in spinal
cord and brain circuits processing sensory signals; 2) ongoing generation of
peripheral stimuli due to damage or inflammatory reactions in the joint leading
to long-lasting changes in primary sensory neurons that detect sensory stimuli.
A growing body of research involving pain mechanisms in OA indicate involvement
of central pain mechanisms, including neuroinflammation6. Microglia, the
resident macrophages (macrophages of the central nervous system) and astrocytes
modulate chronic pain in different rodent models for chronic pain induced by
inflammation or damage. In these chronic pain models, including models for OA
pain, resident microglia and astrocytes in the spinal cord switch from a
quiescent inactive state to an activated phenotype associated with inflammatory
mediator production and hypersensitivity of the pain system. In a rodent model
of OA, induced by intra-articular injection of mono-iodoacetate (MIA), spinal
cord microglia and astrocyte are activated. Moreover, in this rodent model of
OA multiple pro-inflammatory cytokines and chemokines such as IL-1, CCL5, CINC
2*/*, IL-13, IL-17, TNF*, and VEGF are increased in the spinal cord, whilst the
anti-inflammatory cytokines IL4 and IL10, the chemokine CX3CL1 (fractalkine)
and growth factor GM-CSF are reduced11. The fact that neuro-inflammation plays
an important role in rodent models for chronic pain and is a potential target
for treating OA pain is highlighted by the fact that inhibition of
anti-inflammatory IL-10 signalling in the spinal cord using a neutralising
IL-10 antibody strongly prolongs inflammatory pain. In addition,
anti-inflammatory cytokines can dampen chronic pain. In the Laboratory for
Translational Immunology (LTI) and department of rheumatology & clinical
immunology we have developed a fusion protein of the anti-inflammatory cytokine
IL-4 and IL-10, called IL4-10 synerkine. Intriguingly, intrathecal injection of
IL4-10 completely blocks hyperalgesia in rodent models of chronic pain induced
by peripheral paw inflammation or nerve damage whilst combined IL-4 and IL-10
treatment is only partially effective. Similarly, local administration of
IL4-10 synerkine in the knee joint attenuates pain in a dog model of OA.
Whether spinal neuro-inflammation plays a role in human subjects with OA pain
is unknown.
One other potential mechanism that may explain the dissociation between disease
progression and pain in OA is active and ectopic sprouting of sensory and
sympathetic neurons in synovial tissue drivingchronic pain. Inappropriate
remodelling of sensory and sympathetic nerve fibers in peripheral tissues is
associated with chronic pain such as in patients suffering from endometriosis
and irritable bowel syndrome and in rodent models for rheumatoid arthritis and
bone cancer pain. Most importantly, in a mouse model of OA, pain is associated
with increased sensory and sympathetic sprouting in the affected joint. Until
now it remains to be determined if similar changes in sympathetic and sensory
neuron innervation of the knee synovium occur in patients with OA pain.

There is clear evidence of an important role of spinal cord neuro-inflammation
and peripheral neuron sprouting in the development of chronic pain in rodent
models for chronic pain and those specifically for OA. However, until now it is
not known whether such mechanisms truly translate to the human situation. This
knowledge is crucial to consolidate the next step to identify therapeutic
possibilities to specifically treat OA pain.

Our key objective of this proposal is to elucidate potential peripheral and
central mechanisms of chronic pain in humans with OA.

Primary Objective:
Identify if specific inflammatory mediators are different in the spinal liquor
of knee OA patients with severe pain compared to those with relatively little
pain.

Secondary Objective(s):
Asses different aspects of pain (intermittent/constant aspects of pain,
neuropathic component, severity) in knee OA patients and elucidate whether
these aspects are associated with ectopic sprouting of sensory or sympathetic
nerve fibers in the knee and or specific profiles of inflammatory mediators in
spinal liquor of OA knee patients.


Determine if a relation exists between these inflammatory mediators in the
spinal cord and extent of peripheral nerve innervation in the knee: Do they
have a seperate or combined effect on pain perception?

Design: This is a multicenter observational study, with invasive (spinal
cerebral fluid sampling) measurements and collection of knee tissues that have
been removed during total knee replacement in regular practice.

Setting: Collection of spinal cerebral fluid and knee synovial, bone, and
cartilage tissues will be performed at the St Antonius Ziekenhuis. Fluid and
tissue analyses will be performed at the University Medical Center Utrecht.

Duration: Inclusion is estimated to take 12 months.

After informed consent (see 9.1 for exact detail on recruitment and consent),
patients are asked to fill out the ICOAP (intermittent and Constant
Osteoarthritis Pain ), KOOS (pain stiffness and physical function) and
PAINDETECT (neuropathic pain components) questionnaires to Asses the extent of
different elements of pain before the day of surgery.

On the day of surgery, spinal cerebral fluid(300 uL) will be collected prior to
the spinal anesthesia that is part of the standard operation procedures for
total knee replacement. Knee tissue consisting of bone, cartilage and synovial
tissue, that has been removed during total knee replacement will be collected,
fixed and stored for further analysis.
The spinal cerebral fluid will be used to measure several factors (see 8.1.1)
associated with spinal cord neuroinflammation. Knee synovial tissue will be
used to assess the extent of infiltration of inflammatory cells and
innervation. Cartilage tissue will be used to assess the actual cartilage
degeneration in the knee joint.

Benefit: Patients have no direct benefit of participating in this study.
Results will elucidate the underlying mechanisms of osteoarthritis pain and may
provide insights for novel pain treatment strategies.
Burden: Prior to spinal anaesthesia, 300 ul cerebral spinal fluid will be
obtained in a syringe. Since the aspiration can be performed with the same
needle as injection of the epidural anaesthesia requires no additional burden
is introduced. The aspiration will take approximately maximum 5 minutes.
Risks: Postdural headache or infection is negligible