Keywords 

Transcutaneous auricular vagus nerve stimulation · Vagus   nerve stimulation · Inflammatory reflex · Pediatrics

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Dear Editor, 

Idiopathic nephrotic syndrome, which includes minimal change disease, focal segmental glomerulosclerosis,   and membranous nephropathy, is characterized by proteinuria, hypoalbuminemia, edema, and dyslipidemia.  Nephrotic syndrome in childhood has an incidence of  2–7 per 100,000 in the United States [1]. While rare, nephrotic syndrome takes a considerable toll on the healthcare system and the economy, accounting for 12% of incident kidney failure in children [2].

Although the etiology is not clearly understood, most patients with nephrotic syndrome respond to corticosteroid therapy. Unfortunately, relapses are common, with up to 50% of children developing frequently relapsing nephrotic syndrome (FRNS) and 10–20% developing steroid-resistant nephrotic syndrome (SRNS), a designation that carries a poor prognosis (e.g., progression to kidney failure in 35–50% of cases) [3]. Additionally, children with nephrotic syndrome are exposed to large cumulative doses of corticosteroids that place them at risk for developing a multitude of side effects [3]. Prolonged use of steroids leads to growth retardation, osteopenia, mood disorders, cataracts, diabetes mellitus, hypertension, obesity,   and immunosuppression [4]. Furthermore, putative steroid-sparing therapies, such as tacrolimus, mycophenolate mofetil, and rituximab, have their extensive adverse effect profiles [3]. Even with these various therapeutic options, there is still a subset of patients who do not respond. Therefore, there is a pressing need to identify safe and novel therapies that can be used to decrease exposure to corticosteroids and other immunosuppressants.

Stimulation of the vagus nerve is being evaluated as a   novel, effective treatment for many inflammatory and immune-mediated diseases [5–10]. Herein, we discuss what is understood about vagus nerve stimulation (VNS)   and the potential application of this modality to nephrotic syndrome.

Nephrotic Syndrome and the Immune System

The underlying pathogenesis of idiopathic nephrotic syndrome is poorly understood, but it likely involves dysregulation of the immune system. Most nephrotic patients respond to corticosteroid therapy and relapses are often triggered by illness, vaccinations, or allergic episodes, suggesting an immune-mediated etiology [3]. It has been hypothesized that podocyte damage may result from dysfunctional T cells that release circulating factors such as cytokines. This is evidenced by multiple studies which show that pro-inflammatory cytokine levels are elevated during times of relapse (i.e., tumor necrosis factor  [TNF], interleukin [IL]-1, IL-2, IL-4, IL-6, IL-8, IL-12,  IL-13, IL-17, IL-18, interferon-gamma, vascular endothelial growth factor) [11–15]. Recent genomic studies have identified TNF, IL-4, and IL-13 polymorphisms that are associated with susceptibility to nephrotic syndrome, and transcriptomic studies have demonstrated the activation of TNF in children and adults with nephrotic syndrome  [15–17]. Rats infused with TNF and IL-13 developed increases in proteinuria [18]. However, the exact mechanism by which these cytokines are involved is unclear.

Furthermore, the fact that some children with nephrotic syndrome respond to B-cell-depleting therapy with rituximab indicates that T cells may not play a major role in the pathogenesis of this disease [19–21]; B cells may secrete a circulating autoantibody as the inciting permeability factor. Recent developments in animal and human studies have pointed to an antibody to the nephrin podocyte protein as a possible permeability factor [22,  23]. The Weins group discovered circulating anti-nephrin antibodies in children with minimal change disease which correlate with disease activity and IgG in kidney biopsy tissue that colocalizes with nephrin [23].

It is also theoretically possible that the nephrotic syndrome could be a result of some process causing podocyte damage, which has yet to be identified. Regardless of the exact mechanism, dysregulation of the immune system is likely an important feature of idiopathic nephrotic syndrome.

The Inflammatory Reflex

The vagus nerve is the main component of the parasympathetic nervous system, regulating important autonomic functions throughout the body. The left and right vagus nerves are comprised of sensory (afferent) and motor (efferent) fibers that travel from the brainstem to innervate the visceral organs, including the kidneys [24].  The nucleus tractus solitaries (NTS) receive the majority of vagus afferent signals in the brainstem; another brainstem-targeting nucleus is the spinal nucleus. Signals are then transmitted to higher brain structures via the  NTS. Efferent vagus nerve fibers originate in two nuclei,   the dorsal motor nucleus and nucleus ambiguous, in the brainstem medulla oblongata.

The inflammatory reflex, first described 2 decades ago by Tracey et al. [25], is a neural circuit comprised of vagus afferent and efferent signals that modulate the innate immune response. Peripheral inflammatory signals activate afferent action potentials in the vagus nerve that are transmitted to the brain stem nuclei and higher brain structures. Anti-inflammatory signals return via at least two pathways (Fig. 1) [25].

Cholinergic Anti-Inflammatory Pathway

Signals transmitted through efferent vagus nerve fibers inhibit the release of pro-inflammatory cytokines. This neural pathway is termed the cholinergic anti-inflammatory pathway (CAP). The efferent fibers of the vagus nerve relay functional signals to the splenic nerve via the celiac superior mesenteric ganglion complex to release norepinephrine. Norepinephrine then binds to splenic lymphocytes to stimulate the release of acetylcholine (ACh) by a subset of splenic T cells (CD4+ ChAT+ T cells). Pro-inflammatory cells such as macrophages express alpha7 nicotinic  ACh receptors on their surface, which when stimulated by  ACh inhibit the nuclear translocation of the transcription factor NF-κB and the activation of the JAK2-STAT3 pathway [25]. Adrenergic vagus nerve signaling also activates the release of ACh which inhibits the release of pro-inflammatory cytokines including TNF and other mediators [25].  Additionally, the stimulation of the vagus nerve regulates  B-cell trafficking during maturation into antibody-producing cells [25]. In a mouse model of streptococcal infection,  Mina-Osorio et al. showed that VNS resulted in arrested  B-cell migration and decreased antibody secretion, thus demonstrating that CAP regulates antibody production following B-cell exposure to antigens [26].

Anti-Inflammatory Hypothalamic Pituitary Adrenal Axis

In this pathway, inflammatory stimuli cause afferent sensory vagus fibers to stimulate the NTS, which in turn activates neurons in the hypothalamus to release adrenocorticotrophic hormone (ACTH) by the anterior pituitary gland. The adrenal glands then release glucocorticoids to suppress peripheral inflammation [27].

Vagus Nerve Stimulation

Bioelectronic medicine is a growing field that utilizes technology to modulate the nervous system as an approach to the treatment of disease and injury. Implantable  VNS was approved by the Federal Drug Administration in the United States for the treatment of epilepsy in 1997 and treatment-resistant depression in 2005. There is also a growing interest in noninvasive modalities to stimulate the vagus nerve, including transcutaneous auricular VNS  (taVNS) using surface skin electrodes applied to the outer ear to engage the inflammatory reflex by electrically stimulating the auricular branch of the vagus nerve [28].  The cymbal concha is exclusively innervated by the afferent fibers of the vagus nerve, while the vagus nerve branches also innervate the posterior and inferior walls of the ear canal [29]. The auricular branch of the vagus nerve consists of afferent sensory fibers that, when stimulated, send anterograde action potentials to the NTS. Human functional MRI and evoked brainstem potential studies have shown that the electrical stimulation of the cymbal concha activates the NTS in the brain [28]. We initially reported that taVNS in humans decreased cytokine production as measured using endotoxin-stimulated whole-blood organ cultures. We observed taVNS reduced TNF by 80%, IL-6  by 73%, and IL-1β by 50% as compared to pretreatment levels in healthy adults after auricular stimulation of the cymbal concha for 2–5 min [5]. These cytokine changes lasted as long as 24 h after a single session of taVNS, suggesting it may be possible to perform this as a once-daily therapy. However, much remains unknown, including the optimal settings for electrical stimulation: anatomic location, electric current intensity, pulse width, frequency, duty cycle, and session duration. To date, clinical trials using taVNS have employed dozens of variations in these settings and an optimal standard for taVNS is not currently known [28].

In recent years, vagus nerve-mediated inflammatory reflex has become a focus of study for treating chronic immune-mediated illnesses. Multiple animal and human studies have demonstrated that VNS has immunomodulatory effects and leads to improvement in various disease conditions [10]. In a pilot study of Crohn’s disease, seven out of nine participants achieved long-term remission with VNS using a surgically implanted VNS device [7]. In another pilot study, a reduction in disease severity scores was seen in six out of eight participants with Crohn’s disease [8]. In a rheumatoid arthritis trial of 18 patients with an implanted device, VNS led to a decrease in TNF levels and a reduction in symptoms [9].

There are over 200 studies currently registered on clinicaltrials.gov, and many published reports of clinical trials showing the benefit of taVNS for various immune-mediated conditions. One multicenter single-arm study of 30 patients with moderate to severe rheumatoid arthritis indicated that daily taVNS resulted in significant decreases in disease activity scores after 12 weeks [30]. Similarly, a   study of 16 rheumatoid arthritis patients with high disease activity had lower activity scores after 4 days of taVNS; there was no change in the activity score in 20 patients with low disease activity [31]. In 6 patients with rheumatoid arthritis treated with taVNS twice daily for 2 days, there was a reported improvement in disease severity scores and global health assessments with a significant reduction in CRP levels [5]. Additionally, a pilot randomized double-blind study of 18 patients with systemic lupus erythematosus comparing 5 min of taVNS versus sham for 4 days reported improvement in pain, fatigue,   and global health scores in the taVNS treatment group at 12 days [6].

Side Effects of Taverns

Direct surgical stimulation of the efferent vagus nerve has been associated with side effects of bradycardia, gastric motility, vasodilation, and constriction of pupils, but these have not complicated taverns. In a systematic analysis of 1,322 adults treated with taVNS, the most common side effects were skin irritation, nasopharyngitis, headache, and dizziness; there was one report of symptomatic bradycardia [32]. A study examining the effects of taVNS on infants with feeding delay found at taVNS was tolerated well without any adverse effects [33]. Travis has also been used in other pediatric studies without any serious adverse effects [34–36].

VNS and the Kidney

A significant body of preclinical research reveals a link between the immunomodulatory effects of VNS and the kidney [37–44]. Inoue et al. [40] found that VNS ameliorates inflammation and is protective against kidney ischemia-reperfusion injury in mice. Stimulation of the vagus nerve 24 h before insult (but not immediately before insult) resulted in reduced creatinine, improved histology, and reduction in cytokines, the effects of which lasted for 48 h. Experiments showed that the spleen was a necessary component of inflammatory reflex signaling for the immunomodulatory effect [40]. In a similar study of reperfusion injury, Gigliotti et al. [38] used pulsed ultrasound of the spleen in an attempt to trigger the inflammatory reflex. Ultrasound application 24 h before ischemia attenuated kidney injury in mice and reduced circulating cytokines. The effect was not seen in mice that were splenectomized, again demonstrating that inflammatory reflex signaling in the spleen is a critical component of the neuromodulation of the kidney [38].

A seminal paper by Tanaka et al. [45] mapped out two neural pathways involved in the neuro-immune protection of the kidneys with VNS in a mouse model of ischemia-reperfusion injury using optogenetics to selectively stimulate nerve fibers. Anterograde stimulation of both efferent and afferent vagus nerves resulted in kidney protection, and the protection was mediated by the spleen.  The effect was not seen in mice that were splenectomized,   and in another experiment, the adoptive transfer of splenocytes from VNS-treated mice protected the kidneys of naïve mice. The vagus efferent pathway exerted its effect through activating CAP, while the vagus afferent pathway protected the kidneys by activating the vago-sympathetic pathway via the splenic nerve to the spleen, originating from the C1 neurons in the rostral ventrolateral medulla.

taverns in Nephrotic Syndrome

Given that VNS activates the inflammatory reflex,   which affects cytokine release and B and T cell function,   it is plausible that VNS could be a treatment for idiopathic nephrotic syndrome. The successful use of taVNS in immune-mediated conditions in animal and human clinical trials suggests a possible role for taVNS in the treatment of nephrotic syndrome in children.

The work by Tanaka et al. [45] provides a theoretical framework of how taVNS may interact with the kidneys in nephrotic syndrome. We hypothesize that stimulation of the afferent sensory neurons of the auricular branch of the vagus nerve by taVNS leads to (1) stimulation of the  NTS, (2) activation of the C1 neurons in the rostral ventrolateral medulla, (3) activation of CAP via the splenic nerve to the spleen, (4) stimulation of norepinephrine and ACh, (5) inhibition of cytokines, B-cell, and T-cell function, and (6) protection from immune-related injury to the podocytes of the kidney. It is also possible that other mechanisms may be recruited by taVNS (e.g., anti-inflammatory hypothalamic pituitary adrenal axis, renin-angiotensin system, nitric oxide system, or effects on metabolic syndrome and blood pressure mediated by the kidney) and may directly affect podocytes in the kidney,   which requires further investigation. It is also reasonable to consider that taVNS may induce the release from the spleen of T-ChAt, the regulatory T-cell subset which inhibits inflammation by traveling to the kidney to release  ACh. These and other hypothetical mechanisms of VNS   in human nephrotic syndrome warrant further study.

Recently, we reported the results of a small pilot study of taVNS in the treatment of seven children with nephrotic syndrome [46]. In brief, a commercially available transcutaneous electrical nerve stimulation unit delivered electrical stimulation to the auricular branch of the vagus nerve via the left ear cymbal concha for 5 min/day for 26   weeks. In three children with FRNS (not on standing immunosuppression for at least 3 months), all remained relapse-free during the study period. Treatment with taVNS in three children with SRNS (on stable medications for at least 3 months) was accompanied by a 25–76% reduction in proteinuria compared to baseline. For one SRNS participant, the urine protein: creatinine decreased from 2.1   to 0.5 by 26 weeks. (Note: This participant entered complete remission with a urine protein: creatinine of 0.2 at 23   months after publication of the study.) As expected, one participant with a genetic form of congenital nephrotic syndrome did not respond and remained nephrotic. Levels of serum TNF significantly decreased compared to baseline in our study population during taVNS therapy.  There were no adverse events or side effects reported. Results were sustained in three participants who continued with daily taVNS treatment post-study (15–21 months),   whereas relapse/increased proteinuria occurred in those who discontinued therapy.

Future Directions

Travis has several theoretical advantages as an experimental option for nephrotic syndrome therapy in children because it is an at-home, nonpharmacologic modality. Travis is also a steroid-sparing therapy free of the deleterious side effects of immunosuppressant medications in children with nephrotic syndrome. Furthermore, with the poor long-term outcomes associated with SRNS,   taVNS could be utilized as an adjunct proteinuria-lowering therapy, which could slow the progression of chronic kidney disease in these individuals.

In conclusion, the results of our pilot study suggest that taVNS therapy may be useful in the prevention of nephrotic syndrome relapses in patients with FRNS and decreased proteinuria in those with noncongenital SRNS.   Travis is a potentially promising, novel, nonpharmacologic, noninvasive, steroid-sparing approach in the experimental treatment of idiopathic nephrotic syndrome in children. The effectiveness and safety of taVNS for nephrotic syndrome should be further investigated and confirmed with a randomized controlled trial.

Conflict of Interest Statement 

Dr. Sethna was a consultant for Kite Medical.

Funding Sources 

Dr. Sethna received grant funding from the Feinstein Institutes  Advancing Women in Science and Medicine Early Career Development Award. Dr. Chavan received grant funding from the NIH  R01GM132672. Dr. Tracey received grant funding from  R35GM118182-01. Dr. Datta-Chaudhuri received grant funding from General Electric and United Therapeutics. Dr. Zanos received grant funding from General Electric and BARDA. Dr. Seth Na is the site PI for NIH-funded CureGN and NEPTUNE.

Author Contributions

All authors conceptualized and designed the study and critically reviewed the manuscript for important intellectual content.  All authors approved the final manuscript as submitted and agree to be accountable for all aspects of the work. Dr. Christine B. Sethna drafted the initial manuscript.

Data Availability Statement All data generated or analyzed during this study are included in this article. Further inquiries can be directed to the corresponding author.

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46 Merchant K, Zanos S, Datta-Chaudhuri T,  Deutschman CS, Sethna CB. Transcutaneous auricular vagus nerve stimulation (taVNS) for the treatment of pediatric nephrotic syndrome:  a Pilot Study. Bioelectron Med. 2022;8:1.

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