The effects of TNF-alpha blocker and IL-6 blocker therapy on vascular function in a collagen induced arthritis rat model
Date
2020
Authors
Fourie, Serena
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Abstract
Endothelial dysfunction and arterial stiffness are associated with increased cardiovascular
morbidity and mortality in rheumatoid arthritis (RA). Chronic systemic inflammation appears
to play a vital role in the development of endothelial dysfunction and arterial stiffness in
patients with RA. Although biologic disease modifying antirheumatic drugs (DMARDS),
including tumour necrosis factor alpha (TNF-α) blockers and interleukin-6 (IL-6) blockers
have shown reduced inflammatory status and disease progression in RA patients, their effect
on cardiovascular disease risk is inconsistent. The role of biologic DMARDS on endothelial
function and arterial stiffness in the collagen induced arthritis (CIA) rat model, has not been
investigated.
Male (n=47) and female (n=31) Sprague-Dawley rats were randomly divided into the control
(n=22), inflammation (n=26), tumor necrosis factor-alpha blocker (TNF-α blocker; n=15) and
interleukin-6 blocker (IL-6 blocker; n=15) groups. To induce arthritis, rats in the inflammation,
TNF-α blocker and IL-6 blocker groups received a 0.2 ml subcutaneous injection of bovine
type-II collagen emulsified in incomplete Freund’s adjuvant at the base of the tail. Following
the onset of inflammation, the TNF-α blocker group received 10mg/kg intraperitoneal
injections of Etanercept (TNF-α receptor blocker) every three days for six weeks and the IL-6
blocker group received 8mg/kg intraperitoneal injections of Tocilizumab (IL-6 receptor
blocker) once a week for 6 weeks. The control group received a 0.2 ml injection of saline at
the base of the tail. Body weight, food intake, blood pressure, arthritis scores and paw thickness
measurements at the tarsometatarsal and ankle joints were assessed every two weeks. After six
weeks of treatment, arterial stiffness and vascular reactivity in mesenteric and renal arteries
were assessed using the pulse pen and wire-myograph, respectively. Serum concentrations of
TNF-α, IL-6 and C-reactive protein (CRP) were measured by ELISA. Group differences in all
arthritis scores were measured by repeated measures analysis of variance (ANOVA). Group
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differences in inflammatory and vascular function markers were assessed using a two-way
ANOVA with a Tukey post-hoc test. Associations between inflammatory markers and vascular
reactivity changes were determined by Pearson’s correlations.
There were no significant differences in body weight, food intake and blood pressure between
the groups at baseline and at termination (all p>0.05). Arthritis scores and paw thickness
measurements were significantly increased in the inflammation, TNF-α blocker and IL-6
blocker groups compared to the control group at termination (all p<0.01). There were no
differences in arthritis scores or paw thickness measurements between the inflammation, TNF
α blocker and IL-6 blocker groups (p<0.05). Circulating TNF-α concentrations were
significantly higher in the inflammation, TNF-α blocker and IL-6 blocker groups compared to
the control group (p<0.0001, p=0.02, p=0.001 respectively). There were no differences in the
contractile responses in mesenteric or renal arteries between any of the groups (all p>0.05).
Acetylcholine-induced relaxation in mesenteric (p<0.0001) and renal arteries (p<0.0001) was
significantly impaired in the inflammation compared to control group. In the IL-6 blocker
group, ACh-induced relaxation in both mesenteric (p<0.0001) and renal arteries (p<0.0001)
were significantly greater compared to the inflammation group. There were no differences in
ACh-induced relaxation between the inflammation and TNF-α blocker groups in either renal
or mesenteric arteries (all p>0.05). Markers of arterial stiffness were not different between the
groups (all p>0.05). Greater serum concentrations of TNF-α (r=-0.36, p=0.01), IL-6 (r=-0.30,
p=0.02) and CRP (r=-0.34, p=0.01) were associated with impaired KCl-induced contractions
(lower Emax) in mesenteric arteries. Greater serum concentrations of TNF-α (r=-0.68,
p<0.0001), IL-6 (r=-0.54, p=0.001) and CRP (r=-0.68, p<0.0001) were associated with
impaired ACh-induced relaxation (lower Emax) in renal arteries.
In conclusion, chronic high-grade inflammation impairs vascular relaxation, but not
contraction, in resistance and large arteries. Despite unchanged circulating inflammatory
v
marker concentrations, IL-6 blocker treatment reduced the inflammation-induced impairments
in endothelial function in resistance and large arteries. However, the mechanisms underlying
the improved vascular function requires further investigation.
Description
A dissertation submitted in fulfilment of the requirements for the degree of Master of Science in Medicine to the Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, 2020