Abstract
There has been a marked improvement in the treatment of rheumatoid arthritis (RA), but most patients do not achieve disease remission. Therefore, there is still a need for new treatments. By screening an adenoviral short hairpin RNA library, we discovered that knockdown of the nicotinic acetylcholine receptor type 7 (α7nAChR) in RA fibroblast-like synoviocytes results in an increased production of mediators of inflammation and degradation. The α7nAChR is intimately involved in the cholinergic anti-inflammatory pathway (CAP). This led us to study the effects of α7nAChR activation in an animal model of RA, and we could show that this resulted in reduced arthritis activity. Accordingly, stimulation of the CAP by vagus nerve stimulation improved experimental arthritis. Conversely, we found aggravation of arthritis activity after unilateral cervical vagotomy as well as in α7nAChR-knockout mice. Together, these data provided the basis for exploration of vagus nerve stimulation in RA patients as a novel anti-inflammatory approach.
Introduction
Rheumatoid arthritis (RA) is a chronic autoimmune disease, which is characterized by pain, swelling, and stiffness of joints, due to synovial inflammation. During active disease, the joints are limited in motion and function, and persistence of synovial inflammation leads to the development of bone erosions and, finally, joint deformities . The signs and symptoms of this condition can be reduced by treatment with synthetic and biological disease-modifying antirheumatic drugs (sDMARDs and bDMARDs, respectively). Treatment of RA is usually initiated with methotrexate, a sDMARD, and can be combined with corticosteroids and other sDMARDs. Biological DMARDs are indicated if there is insufficient response to the initial sDMARD treatment or if there are unfavorable prognostic factors present, such as very active disease, early joint damage, or presence of (high levels) of autoantibodies, immunoglobulin M (IgM) rheumatoid factor (RF), and/or anti-citrullinated protein antibodies (ACPAs) . Despite the fact that there are many types of DMARDs available, there are still many RA patients who do not improve sufficiently. Besides the lack of response to therapy as a reason for the discontinuation of treatment, there are also patients who discontinue medication because of side effects, or because they do not want to take chronic medication. As a result, the need for the development of new therapeutic strategies remains.
Cholinergic anti-inflammatory pathway
Inflammation in peripheral tissues, as observed in RA, can be detected by the afferent vagus nerve and this information is signaled towards the brain. Peripheral administration of the pro-inflammatory mediators lipopolysaccharide (LPS) or interleukin-1-beta (IL1-beta) in rats normally elicits fever, but after bilateral subdiaphragmatic vagotomy the fever response is abated . Later, there was the surprising finding that the vagus nerve could not only sense inflammation but also influence it. Activation of the vagus nerve, which is a part of the parasympathetic nervous system, was found to dampen inflammatory processes. Rats with carrageenan-induced hind paw edema (acute inflammation model) were injected intracerebroventricularly (i.c.v) with a very low dose (noneffective if given systemically ) of the anti-inflammatory drug CNI-1493, and there was a significant decrease in paw edema. The anti-inflammatory mechanism of action of i.c.v. CNI-1493 was, at the time, unknown, but after bilateral cervical vagotomy the drug was no longer able to reduce paw edema. In combination with the finding that an i.c.v. injection of CNI-1493, but not saline, could increase efferent vagus nerve activity, this led to the conclusion that activation of the vagus nerve by i.c.v. CNI-1493 treatment had an anti-inflammatory effect . These were the first indications that efferent vagus nerve activation could inhibit inflammation in an animal model. The combination of sensing peripheral inflammation by the afferent vagus nerve and the subsequent anti-inflammatory response of the efferent vagus nerve is currently known as the cholinergic anti-inflammatory pathway (CAP) .
The CAP can also be activated by electrical vagus nerve stimulation (VNS) or stimulation of the nicotinic acetylcholine receptor type 7 (α7nAChR) . The parasympathetic neurotransmitter acetylcholine is the anti-inflammatory mediator of the CAP, which activates the α7nAChR. Acetylcholine can be produced by the vagus nerve, but it can also be produced by nonneuronal cells, for instance, in the spleen. Several studies have shown that the spleen is essential for the anti-inflammatory effect of the vagus nerve, because after splenectomy VNS is no longer capable of reducing inflammation . After VNS, the anti-inflammatory reflex appears to travel through the sympathetic splenic nerve towards the spleen. The splenic nerve produces norepinephrine, which triggers choline acetyltransferase-positive (CHAT+) T cells in the spleen to produce acetylcholine ( Fig. 1 ) . CHAT-positive cells are also found in the synovium of the RA joint , which suggests that acetylcholine can also be produced locally in the joint. Joints are not known to be innervated by the vagus nerve, but there appear to be sympathetic fibers in the RA synovium . It is, therefore, conceivable that norepinephrine-producing sympathetic fibers in the joint activate CHAT+ T cells to produce acetylcholine, but this has not been studied yet.
Cholinergic anti-inflammatory pathway
Inflammation in peripheral tissues, as observed in RA, can be detected by the afferent vagus nerve and this information is signaled towards the brain. Peripheral administration of the pro-inflammatory mediators lipopolysaccharide (LPS) or interleukin-1-beta (IL1-beta) in rats normally elicits fever, but after bilateral subdiaphragmatic vagotomy the fever response is abated . Later, there was the surprising finding that the vagus nerve could not only sense inflammation but also influence it. Activation of the vagus nerve, which is a part of the parasympathetic nervous system, was found to dampen inflammatory processes. Rats with carrageenan-induced hind paw edema (acute inflammation model) were injected intracerebroventricularly (i.c.v) with a very low dose (noneffective if given systemically ) of the anti-inflammatory drug CNI-1493, and there was a significant decrease in paw edema. The anti-inflammatory mechanism of action of i.c.v. CNI-1493 was, at the time, unknown, but after bilateral cervical vagotomy the drug was no longer able to reduce paw edema. In combination with the finding that an i.c.v. injection of CNI-1493, but not saline, could increase efferent vagus nerve activity, this led to the conclusion that activation of the vagus nerve by i.c.v. CNI-1493 treatment had an anti-inflammatory effect . These were the first indications that efferent vagus nerve activation could inhibit inflammation in an animal model. The combination of sensing peripheral inflammation by the afferent vagus nerve and the subsequent anti-inflammatory response of the efferent vagus nerve is currently known as the cholinergic anti-inflammatory pathway (CAP) .
The CAP can also be activated by electrical vagus nerve stimulation (VNS) or stimulation of the nicotinic acetylcholine receptor type 7 (α7nAChR) . The parasympathetic neurotransmitter acetylcholine is the anti-inflammatory mediator of the CAP, which activates the α7nAChR. Acetylcholine can be produced by the vagus nerve, but it can also be produced by nonneuronal cells, for instance, in the spleen. Several studies have shown that the spleen is essential for the anti-inflammatory effect of the vagus nerve, because after splenectomy VNS is no longer capable of reducing inflammation . After VNS, the anti-inflammatory reflex appears to travel through the sympathetic splenic nerve towards the spleen. The splenic nerve produces norepinephrine, which triggers choline acetyltransferase-positive (CHAT+) T cells in the spleen to produce acetylcholine ( Fig. 1 ) . CHAT-positive cells are also found in the synovium of the RA joint , which suggests that acetylcholine can also be produced locally in the joint. Joints are not known to be innervated by the vagus nerve, but there appear to be sympathetic fibers in the RA synovium . It is, therefore, conceivable that norepinephrine-producing sympathetic fibers in the joint activate CHAT+ T cells to produce acetylcholine, but this has not been studied yet.
CAP in experimental arthritis
In the past decade, the anti-inflammatory effect of the vagus nerve has been shown in animal models for sepsis, acute pancreatitis, colitis, postoperative ileus, and acute respiratory distress syndrome . We were the first to show the effect of activation of the α7nAChR using the specific agonist AR-R17779 and the non-specific agonist nicotine in an animal model of RA, and demonstrated that this approach resulted in reduced clinical signs of arthritis, synovial inflammation, serum cytokine levels, and bone erosions. These results have been confirmed by others using the partially specific agonist GTS-21 and nicotine . Conversely, we found a higher incidence of arthritis and more severe arthritis in mice lacking the α7nAChR . The α7nAChR is present not only on many immune cell types, such as monocytes, macrophages, T lymphocytes, B lymphocytes, and dendritic cells but also on fibroblast-like synoviocytes (FLSs) in the RA synovium . FLSs of RA patients pretreated with acetylcholine, nicotine, or AR-R17779 (α7nAChR activation) reduced the production of IL-6 and IL-8 significantly , indicating that acetylcholine could also have an anti-inflammatory effect in the RA synovium. Besides FLSs, macrophages and monocytes also produce less pro-inflammatory cytokines after α7nAChR activation . Thus, human biology studies were in line with the results obtained in animal models of RA, supporting the notion that stimulation of the CAP could have a beneficial effect in RA.
Stimulation of the CAP by electrical VNS is challenging in the experimental arthritis model, because daily electrical stimulation is needed for a prolonged period rather than single electrical stimulation in the acute mouse models. As an alternative approach, vagus suspension to the sternocleidomastoid muscle was performed, leading to activation of the vagus nerve as a result of head movement, after arthritis induction. This led to reduced arthritis activity, decreased serum levels of tumor necrosis factor (TNF), and protection against bone destruction associated with a decrease in the number of osteoclasts after 3 months of follow-up . When we applied electrical stimulation of the vagus nerve for 60 s a day using an implanted vagus nerve stimulator in the collagen-induced arthritis model of RA in rats, we observed amelioration of arthritis and decreased histological signs of synovial inflammation as well as cartilage damage after 7 days of stimulation . Taken together, activation of the CAP has a consistent anti-inflammatory effect in experimental models of RA.
Effect of nicotine-containing substances in RA patients
As discussed above, nicotine can stimulate the CAP through activation of the α7nAChR. However, despite cigarette smoking being a well-defined risk factor for the development of RA , a beneficial effect of nicotine on experimental arthritis and an anti-inflammatory effect on RA FLS are observed . How can we reconcile these data? First, it is important to note that, obviously, smoking of cigarettes results in exposure to many other compounds in addition to nicotine. In particular, in RA patients who have a specific genetic background, being positive for the shared epitope, smoking of cigarettes is associated with the development of ACPAs . Indeed, several studies have shown that RA patients who smoke cigarettes have higher disease activity compared to nonsmokers , although not all data are conclusive . The response to therapy in RA patients who are current smokers is lower in most of the longitudinal cohort studies . If the negative effect of smoking of cigarettes is related to inhaling compounds other than nicotine, then lessons may be learned from the use of nicotine-containing “snuff” (smokeless tobacco) in RA patients. Interestingly, there was a lower disease activity in RA patients who used snuff compared to those who never smoke and previous smokers . Consistent with our hypothesis that substances in smoked tobacco other than nicotine are associated with an increased risk of active RA in shared epitope-positive subjects, the use of snuff is not associated with the development of RA . Obviously, RA patients who are current smokers should be advised to stop, also because of the increased risk of cardiovascular disease and cancer associated with smoking . The use of snuff is also not harmless, because it contains carcinogenic substances and users have a higher risk of developing oral cancer and cardiovascular disease . However, the observations described above do support the notion that stimulation of the CAP may result in a beneficial effect on RA.
Activity of the parasympathetic nervous system in RA
The vagus nerve, the main component of the parasympathetic nervous system, is most active when the body is in a resting state, and at that time it has a dominant influence on breathing, heart rate, and digestion. In a more active “flight” state, the sympathetic nervous system becomes dominant in controlling these basic body functions. The balance in the autonomic nervous system can be measured by measuring heart rate variability (HRV) . Individuals with a normal HRV are able to respond adequately to changes in blood pressure and breathing, and have therefore a well-functioning autonomic nervous system. Various observational studies have demonstrated that RA patients have lower vagus nerve tone shown by reduced HRV compared to age-matched controls . This phenomenon has also been observed in other autoimmune diseases, such as systemic lupus erythematosus , ankylosing spondylitis , and chronic inflammatory bowel diseases . Parasympathetic activity can be increased by different types of exercise, like cardiac training , yoga , meditation , daily 5-min diaphragmatic breathing , and possibly acupuncture . One study showed that low parasympathetic and high sympathetic activity in RA patients predicts a poor therapeutic response to anti-TNF therapy compared to RA patients with a more balanced autonomic nervous system . Taken together, these data show that chronic inflammatory diseases are associated with reduced parasympathetic and increased sympathetic activity. The autonomic balance could potentially be restored by electrical stimulation of the vagus nerve.
VNS in patients with depression or epilepsy
VNS therapy has been approved as a treatment for epilepsy in Europe in 1994 and in the US in 1997 ; in the US, VNS has also been approved for the treatment of depression . VNS can be performed after neurosurgical implantation of a vagus nerve stimulator; in 2012, 100,000 vagus nerve stimulators had been implanted . The device consists of two parts: a pulse generator and a lead with electrodes. The pulse generator contains the battery and the stimulation system, and is positioned subcutaneously below the left clavicle on the pectoral muscle. It is connected to the left vagus nerve in the neck via the lead, with three helices at the end: one positive electrode, one negative electrode, and an anchor tether. The three helices are placed around the vagus nerve to deliver the electrical pulse of the pulse generator ( Fig. 2 A and B). During surgery, the vagus nerve is electrically stimulated to test the impedance and functionality of the device, which can be accompanied by bradycardia and short-lasting asystole, which is a rare adverse event occurring in about 0.1% of patients; it has only been reported as a consequence of the first stimulation during surgery and is resolved when stimulation is stopped . Adverse events after implantation include infections of the operated area (3–6% of patients) and vocal cord paresis (<1% of patients) . After implantation, VNS therapy can be initiated starting at a low dosage of stimulation with an output current of 0.25 mA. The dosage is increased slowly with steps of 0.25 mA to a maximum output current of 3.5 mA, because toleration to the stimulation is built up with use of the VNS device. However, the final maximum tolerated dosage of the patient can be lower than 3.5 mA. Other stimulation conditions that can be selected are pulse width (130–1000 μs), frequency (10–30 Hz), and duration of stimulation (7–60 s). The VNS settings can be changed with an external programmer and information about performed stimulations (compliance) can be retrieved ( Fig. 2 C and D). During electrical stimulation, >10% of patients report hoarseness, sore throat, shortness of breath, and coughing, but in general the stimulation is well tolerated and these symptoms do not require adjustments of VNS device settings . Patients normally only experience side effects if the VNS device is activated. Epilepsy patients receive stimulation several times per hour (the standard setting is 30 s on/5 min off) and have the option to give an additional stimulation with a magnet activation to prevent an upcoming seizure .
Related posts:
Concepts of pathogenesis and emerging treatments for inflammatory arthritis
Molecular and cellular heterogeneity in the Rheumatoid Arthritis synovium: Clinical correlates of synovitis
How undifferentiated arthritis evolves into chronic arthritis
Small-molecule therapeutics in rheumatoid arthritis: Scientific rationale, efficacy and safety
Inhibitors of angiogenesis: Ready for prime time?
Seronegative rheumatoid arthritis: Pathogenetic and therapeutic aspects
Stay updated, free articles. Join our Telegram channel
Full access? Get Clinical Tree
