Role of neurogenic inflammation in local communication in the visceral mucosa

Abstract

Intense research has focused on the involvement of the nervous system in regard to cellular mechanisms underlying neurogenic inflammation in the pelvic viscera. Evidence supports the neural release of inflammatory factors, trophic factors, and neuropeptides in the initiation of inflammation. However, more recently, non-neuronal cells including epithelia, endothelial, mast cells and paraneurons are likely important participants in nervous system functions. For example, the urinary bladder urothelial cells are emerging as key elements in the detection and transmission of both physiological and nociceptive stimuli in the lower urinary tract. There is mounting evidence that these cells are involved in sensory mechanisms and can release mediators. Further, localization of afferent nerves next to the urothelium suggests these cells may be targets for transmitters released from bladder nerves and that chemicals released by urothelial cells may alter afferent excitability. Modifications of this type of communication in a number of pathological conditions can result in altered release of epithelial-derived mediators, which can activate local sensory nerves. Taken together, these and other findings highlighted in this review suggest that neurogenic inflammation involves complex anatomical and physiological interactions among a number of cell types in the bladder wall. The specific factors and pathways that mediate inflammatory responses in both acute and chronic conditions are not well understood and need to be further examined. Elucidation of mechanisms impacting on these pathways may provide insights into the pathology of various types of disorders involving the pelvic viscera.

1. Introduction

Neurogenic inflammation of the urinary bladder mucosa is a complex process triggered by the release of inflammatory peptides, Substance P (SP), Calcitonin Gene-Related Peptide (CGRP) and, neurokinin A (NKA) from the afferent neurons in response to various stimuli including antigens, bacterial or viral infection, direct stimulation of sensory nerves or other irritants. This process is characterized by an intricate interplay between vascular, immune and nervous systems and results in alterations in all bladder wall components (urothelium, smooth muscle, nerves, blood vessels, and other cells types), leading to dysfunctional voiding and pain.

Clinically, neurogenic inflammation of the urinary bladder mucosa may be present in a percentage of patients with interstitial cystitis/bladder pain syndrome (IC/BPS) [1, 2], prostatitis, urethritis, chronic pelvic pain [3], radiation cystitis [4] and spinal cord injury [5]. Patients experience pain in the pelvic area and voiding dysfunction typically characterized by increased frequency, urgency and nocturia. Although difficult to diagnose, the involvement of neurogenic inflammation in these patients is supported by a) tissue biopsies which present a number of characteristic features of neurogenic inflammation (such as increased mast cells, angiogenesis, which will be discussed in this chapter) and b) functional studies using intravesical instillation of capsaicin/resiniferatoxin (RTX), and/or bladder hydrodistention, that presumably deplete C-fibers from neuropeptides, resulting in improvements in voiding function and pain relief [3, 6, 7].

This chapter will review the structure and function of the bladder mucosa, alterations induced by neurogenic inflammation and potential therapeutic targets.

2. Anatomy of the bladder wall

The bladder wall has three main components: the mucosa, muscularis propria (also termed the detrusor or bladder smooth muscle), and adventitia/serosa (Fig 1) [8]. The mucosa is composed of an urothelium, basement membrane and lamina propria, which will be discussed below in more detail in this chapter. The detrusor is composed of smooth muscle bundles interspaced with collagen and elastin. The muscle bundles run in longitudinal and circumferential directions which help maintain structural integrity during bladder filling and emptying. In humans, there is an inner longitudinal layer that extends into the proximal urethra, a middle circularly arranged layer and an outer longitudinal layer that extends to the bladder neck. The collagen and elastin fibers, which are found also in the lamina propria and adventitia, are the major passive structural components of the bladder that play a significant role in the ability of the bladder to accommodate large urine volumes [9]. The adventitia/serosa is the outmost bladder layer composed of connective tissue with a much less understood role. Bladder innervation consists of efferent and afferent nerves originating in the lumbosacral and thoracic spinal cord. The efferent innervation consists of sympathetic and parasympathetic nerves that perform two different functions of the bladder. The sympathetic nerves are active during bladder filling. They release norepinephrine which relaxes the bladder smooth muscle to allow filling and contracts the urethral smooth muscle to maintain continence. The parasympathetic nerves are activated during voiding and they release acetylcholine (ACh) to contract the bladder and nitric oxide (NO) to relax the urethra to allow flow of urine [10, 11]. The afferent nerves collect sensory information (including distention, pain) and convey this information to the spinal cord.

Figure 1. Anatomy of the bladder wall.

Left: Transverse section through human urinary bladder. Right: cartoon depicting bladder wall components. Adapted with permission from Birder L, Andersson KE. Urothelial signaling. Physiol Rev 2013.

2.1. Urothelium

The uroepithelium, or urothelium, is a highly specialized transitional epithelium that forms the inner lining of the renal pelvis, ureters, bladder, upper urethra, and glandular ducts of the prostate. The urothelium is the interface between the lumen and the underlying vasculature, connective, nervous, and muscular tissues forming a high-resistance barrier to ion, solute and water flux, and pathogens [8]. The urothelium is composed of three cell types arranged in 3–7 layers (depending on the species): a superficial or apical layer composed of large hexagonal cells known as “umbrella cells”, an intermediate layer and a basal cell layer attached to a basement membrane. The umbrella cells are terminally differentiated cells that possess unique properties (e.g. expression of uroplakins and tight junction proteins such as claudins and occludins) which allow them to maintain a tight barrier while undergoing considerable stretch as the bladder fills. The intermediate and basal cells are capable of dividing to regenerate the urothelial structure upon injury [8]. In contrast to the bladder, the type of epithelium in the urethra varies according to the region: from transitional at the bladder neck and proximal urethra, to pseudostratified columnar and stratified columnar in the mid urethra part and to stratified squamous cells at the external urethral orifice [12]. The urothelium expresses a variety of ion channels and receptors, including purinergic (P2X1–7 and P2Y1,2,4), adenosine receptors (A1, A2a, A2b, and A3), α- and β-adrenergic, bradykinin, neurokinin, nicotinic and muscarinic (M1–M5), protease-activated receptors, and transient receptor potential (TRP) channels (TRPV1, TRPV2, TRPV4, and TRPM8). There is evidence that the urothelium can respond to a number of mechanical and chemical stimuli, including bladder distention and irritants present in the urine, resulting in the release of transmitters such as ATP, ACh, adenosine, NO, and prostaglandins [8]. These properties play important roles in neurogenic inflammation.

2.2. Lamina Propria

The lamina propria is the layer between the basement membrane and the detrusor muscle. It is composed of an extracellular matrix consisting of collagen and elastin fibers, rich vascular networks, lymphatic channels, sensory and autonomic nerve endings and several types of cells, including smooth muscle, fibroblasts, adipocytes and interstitial cells (IC) [13]. Different populations of collectively called interstitial cells (IC) (comprising interstitial cells, interstitial cells of Cajal, interstitial Cajal-like cells, myofibroblasts, or telocytes), have been found in the lamina propria close to the urothelium and along the blood vessels. Similar populations have been identified in the urethra. These cells have been described as spindle-shaped cells that form networks interconnected by Cx43 gap junctions and have close contacts with nerves containing small clear vesicles with and without dense cores, implying that they had an efferent and afferent nerve supply. IC cells are excitable, show spontaneous electrical activity, possess receptors for ATP, ACh, prostaglandins and are thought to play a role in cell-cell communication in the bladder wall [13–15].

2.3. Bladder and urethral afferent innervation

The afferent nerves detect sensory information such as bladder fullness and pain from the periphery and convey this information to the spinal cord. They consist of myelinated Aδ-fibers and unmyelinated C-fibers, and terminate near and within the urothelium, lamina propria and the muscle layers of the bladder [10, 11] [16]. In rodents, Aδ-fiber and C-fiber afferents are not distinguishable on the basis of stimulus modality; both types of afferents consist of mechanosensitive and chemosensitive populations which can be differentiated according to the electrophysiological and chemical properties of the dorsal root ganglia (DRG) neurons. The C-fiber population is characterized by sensitivity to capsaicin, associated with the expression of TRPV1 receptors, have high threshold for activation and long duration action potential insensitive to tetrodotoxin (TTX, a Na⁺ channel blocker). Depending on species this population represents ~70% of bladder afferents. The Aδ-fiber population is insensitive to capsaicin, but sensitive to TTX. Under normal conditions in conscious animals micturition is initiated by Aδ-afferents. However, in pathological conditions and in anesthetized animals C-fibers are also involved. Immunostaining studies have shown that bladder afferent neurons contain several neuropeptides including SP and CGRP. Some afferents express more than one peptide, though the extent of overlap between peptides co-expression is not well defined. In the periphery, CGRP and SP expressing fibers are distributed throughout the bladder and urethral wall, inside the urothelium and epithelium of the urethra, in the lamina propria, surrounding blood vessels (both arteries and veins) and along muscle bundles [16–20]. Regional distribution of CGRP positive fibers in the lamina propria and urothelium have indicated the presence of a dense network at the bladder base and proximal urethra compared to the bladder dome [16]. In the spinal cord peptidergic afferents project into Lissauer’s tract at the tip of the dorsal horn and then pass rostrocaudally, giving off collaterals that extend through the superficial layer of the dorsal horn in the lamina I and into the deeper layers (laminae V to VII and X) at the base of the dorsal horn. The lateral pathway, which is the most prominent projection, terminates in the region of the sacral parasympathetic nucleus and also sends projections to the dorsal commissure [10, 11].

Bladder afferents, especially peptidergic C-fibers, express a number of receptors that allow them to detect mechanical (e.g. bladder distention), chemical (e.g. pH, osmolarity) and noxious (e.g. overdistention, irritants) stimuli. Among these, TRP channels (TRPV1, TRPA1, TRPM8 and others), purinergic receptors (P2X2, P2X3, and several P2Y receptors) [10, 11] and receptors for trophic factor (TrkA, TrkB, GRFα1 and GRFα3) are thought to play prominent roles not only in normal bladder function but also in pathological conditions including neurogenic inflammation. Stimuli that activate these receptors have the potential to cause release of peptides from afferent neurons. For example, studies in guinea pig bladder tissue have shown that activation of TRPV1 by capsaicin releases both SP and CGRP [21]. The release of peptides in the bladder wall triggers inflammatory responses, including plasma extravasation or vasodilation (i.e., neurogenic inflammation). The release of peptides in the spinal cord is involved in central sensitization and persistent pain.

3. Physiology of the bladder: the ‘sensory web’

It is well accepted that the afferent limb of micturition, including the urothelium and lamina propria, can modulate the activity of the afferent nerves, influencing afferent sensations, the micturition reflex and perception of pain. For example, during bladder filling, a cascade of urothelial transmitter/mediators (excitatory and inhibitory) are involved in the transduction mechanisms underlying the activation of afferent fibers. Mechanical signals such as pressure, tension in the urothelium or bladder wall, torsion, osmotic swelling and sheer stress can all serve to activate the urothelium. Chemical signals including changes in urine tonicity and osmolarity as well as toxins, bacteria, drugs and irritants present in the urine can also activate the urothelium. In response, these urothelial ‘sensor’ cells release transmitters (such as ATP, ACh, prostaglandins) that can act on afferent nerves and/or other cells in the bladder wall. Interstitial cells in the lamina propria respond to ATP and ACh and this may play a role in the intramural signaling [13]. Thus, the mucosa is likely to play an important role in the complex transfer of information to and from the nervous system. It may act as a modulator to change the gain of the system which is particularly significant in pathology [8].

4. Pathophysiology, animal models and links to human condition

Neurogenic inflammation is triggered by stimuli including antigens, irritants, bacterial or viral infection, or direct stimulation of sensory nerves that cause the release of pro-inflammatory peptides, CGRP, SP and NKA from the sensory nerves. This elicits a host of vascular events, including vasodilation of arterioles, extravasation of plasma from post-capillary venules which causes edema, leukocyte adhesion to endothelial cells in the venules and leukocyte migration in to the affected tissue. These processes result in activation of mast cells, disruption of the tissue architecture and function (i.e. disruption of mucosa and urothelial layers including effects on urothelial mediators and growth factors), increased afferent excitability and smooth muscle contractility, all which underlie dysfunctional voiding and pain.

A number of animal models have been developed and employed for investigating factors that contribute to neurogenic inflammation as well as pathways affected by neurogenic inflammation. These models can be classified into three main groups:

• manipulations that produce direct damage to the bladder, including intravesical instillation of irritants such as mustard oil (2.5%) [22], acetic acid (0.25%) [23], capsaicin [23], Escherichia coli lipopolysaccharide (LPS) [24] in otherwise healthy animals. Intravesical SP, capsaicin or ovalbumin (OVA) have also been used in antigen - sensitized guinea pigs [25, 26] and rats [27].

• manipulations that do not involve direct damage to the bladder, including stimulation of dorsal spinal roots [28], antidromic stimulation of fibers innervating the bladder [22, 23], intravenous injection of SP (0.1 ml; 10⁻⁶ M) [24], virus-induced neurogenic cystitis [29–31], and chronic stress such as immobilization [32], water avoidance stress (WAS) [33] and social stress [34, 35].

• naturally occurring: cats suffering from feline interstitial cystitis (FIC) may exhibit neurogenic inflammation especially during ‘flares’ [36].

These animal models together with findings from human tissue and clinical features revealed many structural and functional changes in the bladder wall, which are discussed below.

4.1. Alterations in neuronal physiology and morphology

Animal models and human studies provided substantial evidence for the involvement of C-fibers, particularly those expressing TRPV1 receptors, in neurogenic inflammation. Initial experiments using capsaicin have established that both SP and CGRP can be released from the bladder afferents [21]. Further, manipulations including stimulation of dorsal spinal roots [28], antidromic stimulation of fibers innervating the bladder [22, 23], intravesical instillation of mustard oil [22], acetic acid or capsaicin [23], E. coli LPS [24], intravenous injection of SP [24] and virus-induced cystitis [31], have been shown to evoke plasma extravasation in rodent bladder. In many of these manipulations, plasma extravasation was diminished and/or prevented by desensitizing C-fibers with systemic capsaicin treatment, performed neonatally or several days before the terminal experiments, or by denervation of the bladder. Mast cell degranulation, which is another prominent feature described in bladder tissue was also reduced by desensitization of C-fibers, as shown in rodent models of immobilization stress [32] or virus induced cystitis [29, 31]. Clinical studies in patients with IC/BPS and neurogenic detrusor overactivity (NDO) due to spinal cord lesions demonstrated that capsaicin/ resiniferatoxin (RTX) instillation into the bladder results in improved bladder function: reduction in the number of micturition episodes per 24 hours, number of episodes of micturition associated with urgency per 24 hours, number of episodes of incontinence and increased bladder capacity. Additionally, in patients with IC/BPS, hydro-distention alone which presumably depletes afferent nerves from neuropeptides, or in combination with intravesical RTX, provides pain relief [3, 6, 7, 37, 38].

Increased afferent nerve hyperexcitability associated with increased nerve density

Neurogenic inflammation is associated with increased afferent nerve excitability and sensitization of primary afferent neurons, as shown in FIC cats [39, 40], WAS rats [41], in rodents following SCI [42] and other inflammatory models (e.g. cyclophosphamide, instillation of bladder with irritants). A number of alterations in the expression and function of K⁺ and Na⁺ ion channels that govern neuronal excitability have been reported, including decrease in Kv1.4, an A-type K⁺ channel and changes in TTX-resistant Na⁺ channels Nav1.8 and Nav1.9 [42–45]. While the precise mechanisms underlying the development of afferent nerve hyperexcitability are complex, excessive release of CGRP, SP and NKA from the afferent nerves can play a role. Electrophysiological studies have shown that CGRP enhances TTX-resistant currents in small and medium diameter DRG neurons [46], and SP increases the number of action potentials in a subset of DRG neurons [47]. SP and NKA activate neurokinin (NK) receptors on afferent nerves. In particular, NK2 receptor activation modulates K⁺ and Ca²⁺ channels that influence the excitability of bladder afferent neurons, primarily C-fibers, by lowering the threshold for triggering an action potential [42, 48]. In addition to increased hyperexcitability, there is evidence for increased nerve density in the bladder wall. In human tissue from IC/BPS patients, there was significantly more PGP9.5 [49] and SP [50] nerve fiber staining especially within the sub-urothelial layers, than in tissue from normal patients. Also, in patients with NDO as a result of spinal cord injury or multiple sclerosis, immunostaining for TRPV1, PGP9.5 and P2X3, which are often co-localized with TRPV1, was increased, especially in the suburothelium/lamina propria area. Interestingly, intravesical RTX instillation in this cohort had positive effects on bladder function in a subset of patients, called ‘the responders’. In biopsies from these patients the density of P2X3, TRPV1 and PGP9.5 nerve fibers decreased after treatment and it was thought that growth factors play a role in these plastic changes [37, 51]. Thus, conditions associated with neurogenic inflammation, can result in long lasting functional and anatomical changes of primary afferent neurons, particularly C-fibers. This includes sensitization, manifested as a decrease in the threshold for activation and an amplification of responses to noxious stimuli, and sprouting. Peripheral sensitization leads to allodynia (when innocuous stimuli, such as bladder filling, can produce pain) and hyperalgesia (when noxious stimuli can produce exaggerated and prolonged response). Together, these peripheral changes can contribute to urgency, increased voiding frequency and pain.

Pain/nociception and central sensitization

Bladder and/or pelvic pain is a prominent feature in patients with IC/BPS and chronic prostatitis [52]. Similarly, animal models of neurogenic inflammation also exhibit pelvic sensitivity/bladder hyperalgesia. For example, mice with virus induced cystitis develop tactile allodynia and pelvic sensitivity [29–31], rats exposed to chronic WAS develop bladder hyperalgesia associated with increased visceromotor reflexes (VMR, an indicator of nociception) in response to cold infusion of the bladder [41, 53, 54], and antigen sensitized guinea pigs demonstrate abdominal licking behavior, suggestive of pain, in response to intravesical OVA instillation [26]. In the virus induced neurogenic cystitis model tactile allodynia and increased pelvic sensitivity are gender dependent, being more pronounced in the females [29–31], similar to the IC/BPS population.

Persistent and/or repetitive peripheral insults resulting in alterations in the periphery (decreased threshold for afferent nerve activation, increased excitability of afferent nerves) can lead to alterations in circuits in the spinal cord and brain, a phenomenon known as central sensitization. This is a state of the central nervous system (CNS) characterized by heightened sensitivity, abnormal responsiveness and increased gain of the nociceptive system. [55] The mechanisms involved in central sensitization are complex, involve neuronal and non-neuronal cells (e.g. microglia, astrocytes) [55] and beyond the scope of this review. However several changes in the CNS that may be connected to bladder neurogenic inflammation are worth mentioning. Cats suffering from FIC show increased expression of the astrocyte marker GFAP and evidence for co-expression with a marker for glia reactive state, nestin, in regions of the spinal cord that receive input from pelvic afferents [56]. Reactive glia cells have been shown to contribute to the maintenance of visceral pain by releasing signals that sensitize spinal cord neurons [57]. Patients with IC/BPS have alterations in the brain white matter that are associated with the severity of the symptom [58]. Also, the activity of brain centers involved in bladder control, bladder sensations and pain such as the motor and sensory cortex, anterior cingulate cortex, amygdala, insula is altered [59]. Thus, neurogenic inflammation of the mucosa can produce long lasting changes not only in the periphery but also in the CNS and these likely contribute to persistent pain states.

4.2. Alterations in the mucosa

4.2.1. Urothelium

Neurogenic inflammation is associated with many changes in the urothelium/mucosa. The events that take place from the release of peptides from the afferent nerves to changes in the urothelial/mucosa structure and function are not well understood. It is unclear whether peptides act directly on urothelial cells or indirectly by altering blood supply and/or other cell types and therefore altering the properties of these cells. However, there is evidence for numerous alterations in the urothelium in animal models of IC/BPS as well as in patients.

ATP is a transmitter with complex actions in the urinary bladder. ATP can be released by urothelial cells, afferent and efferent nerves and can act on multiple subtypes of purinergic receptors P2X and P2Y, located at many sites in the bladder wall. These include the urothelial cells, afferent nerves, myofibroblasts and smooth muscles as well as spinal cord neurons receiving input from the primary afferent neurons. The urothelial cells release ATP in response to various stimuli such as stretch (e.g. bladder distention, hypotonic solution in cultured urothelial cells), activation of muscarinic receptors, TRPs (TRPV4) or P2Y receptors. Once released, ATP can act in a paracrine fashion to modulate the activity of afferent nerves and myofibroblasts. ATP can also act in an autocrine manner to facilitate its own release as well as the release of other mediators from urothelial cells. Experiments mimicking the release of ATP from the urothelium by instilling ATP intravesical have shown stimulation of the micturition reflex in awake rats [60] possibly through activation of receptors on capsaicin-sensitive C-fibers [61]. The use of genetically engineered mice (P2X3 knock-out) as well as the use of antagonists, demonstrated that activation of P2X2 and P2X3 receptors on unmyelinated (capsaicin sensitive) bladder afferent fibers play an important role in mechanosensitivity [62, 63].

There is substantial evidence for alterations in the purinergic system in the urothelium and afferents in pathological conditions involving inflammation (Fig 2). In rodent bladders that are overactive as a result of inflammation [64], bladder tissue from patients suffering from IC/BPS [65] and in urothelial cells from patients or cats suffering from interstitial cystitis [66–68], ATP release is upregulated and/or the expression of urothelial purinergic receptors is altered [69, 70]. The increase in ATP may activate underlying afferent nerves, particularly nociceptive fibers which express P2X2/3 receptors (many of which also co-express TRPV1), and increase the excitability of these nerves (Fig 2). In addition, bladder inflammation (e.g. produced by cyclophosphamide) sensitizes and enhances P2X receptor function on primary afferent neurons [71], potentially contributing to overactivity, urgency and pain. On the other hand, mice lacking the P2X3 receptor show reduced inflammatory pain and marked urinary bladder hyporeflexia with reduced voiding frequency and increased voiding volume [72]. This suggests that purinergic (P2X3 and other subtypes) receptors are involved in mechanosensory transduction in conditions involving inflammatory pain and hyperreflexia [72].

Figure 2. Role of urothelial derived factors in neurogenic inflammation.

Injury to the urothelial barrier results in release of a number of factors including ATP, prostaglandins (PG), nitric oxide (NO), ACh, NGF and others, which excite the afferent nerves (in blue). These nerves then in turn release pro-inflammatory peptides CGRP, SP and NKA, which act back on vascular smooth muscle, cells on the lamina propria and/or urothelial cells, to further augment the release of urothelial-derived factors. This vicious cycle results in hyperexcitability of the afferent nerves and pain.

4.2.2. Lamina propria

As the lamina propria is considered a center for propagation and integration of sensory and mechanical signals, changes induced by pathology in this area are likely to affect voiding and pain. Acute irritation of the bladder (using acetic acid in cats), resulted in edema and infiltration of neutrophils in the lamina propria of the bladder and urethra. [73] Chronic pathological conditions associated with neurogenic inflammation also show edema and infiltration of neutrophils, increased mast cells and changes in IC cells. For example, in tissue from patients with NDO, IC cells were often found in close apposition to lymphocytes, their layer organization was disrupted and their structural features were altered [74]. The lamina propria expresses a number of growth factors, including vascular endothelial growth factor (VEGF), bone morphogenetic protein 4 (BMP4), platelet-derived growth factor receptor (PDGF)-BB, transforming growth factor α (TGFα), β1 (TGFβ1) and epidermal growth factor (EGF), that are thought to play a role in proliferation and response to injury [75]. A major impact of these factors in chronic conditions can be the development of fibrosis. Fibrosis has been reported in patients and animal models of irradiation cystitis [4, 76, 77] and spinal cord injury [78, 79] and has been linked to compromised function of the organ. Recent studies have shown that collagen fibers in the lamina propria play a more prominent role in bladder compliance than fibers in the smooth muscle [9], thus, the development of fibrosis in this area is likely to severely impact bladder function.

4.2.3. Neurotrophins

Urothelial cells, various cells in the mucosa and the smooth muscle produce a number of neurotrophins including nerve growth factor (NGF), brain-derived nerve factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4 (NT-4) [80] (Fig 3). Epithelial cells in inflamed tissue express increased levels of NGF, BDNF, and other neurotrophic factors [81].

Figure 3. Role of mucosa-derived neurotrophins in neurogenic inflammation.

NGF and other neurotrophins (e.g. BDNF) released by urothelial cells, mast cells and other cells in the mucosa. NGF binds to its receptors TrkA and p75 located on afferent neurons. It is thought that the TrkA-NGF complex is internalized and retrogradely transported to cell bodies in lumbosacral DRG, where de novo transcription of TRPV1, voltage gated calcium channels (VGCs), mechanosensitive channels (MSCs) and purinergic P2X3 receptor is initiated. These newly synthesized ion channels are anterogradely transported back to afferent terminals to contribute to peripheral hypersensitivity. Adapted with permission from Ochodnicky P, Cruz CD, Yoshimura N, Cruz F. Nat Rev Urol. 2012 Nov;9(11):628–37. Review. PMID: 23045265

There is ample evidence for a role of NGF in neurogenic inflammation. Intravesical instillation of E. coli LPS increases NGF mRNA and protein in the bladder [82]. In several animal models of chemical, neurogenic and immune-mediated inflammation, as well as in the urine of patients with IC/BPS the levels of NGF in the bladder tissue are elevated, suggesting that an increase in NGF represents a generalized response to bladder inflammation [43, 80, 83–85]. Acute intravesical NGF in rats produces bladder hyperexcitability associated with afferent nerve sensitization [86]. Chronic upregulation of NGF in the bladder wall of rodents (using osmotic mini-pump delivery or intramuscular adenoviral NGF gene transfer) results in bladder hyperexcitability, reduced bladder capacity, increased voiding frequency and increased amplitude of non-voiding contractions [87]. These are associated with increased pain markers (c-fos) and increased CGRP immunoreactivity in the lumbosacral spinal cord areas receiving afferent input from the bladder [87]. The underlying mechanisms may involve uptake of peripheral NGF by TrkA and retrograde transport of the NGF-TrkA complex to soma of the primary afferent neurons in the DRG, where de novo transcription of TRPV1, voltage gated calcium channels, mechanosensitive channels and possibly other sensory ion channels (such as purinergic P2X3 receptors) is initiated (Fig 3). The newly synthesized ion channels are then anterogradely transported to afferent terminals in the bladder where they contribute to peripheral hypersensitivity [80] (Fig 3). Additional studies in transgenic mice overexpression of urothelial NGF show increased nerve density and number of mast cells in the bladder wall, changes in bladder function (reduced urinary bladder capacity, increased number and amplitude of non-voiding contractions) and pelvic hypersensitivity [88]. Not only peripheral elevation of NGF levels results in bladder hyperexcitability, but also central elevation in the spinal cord (achieved by intrathecal NGF delivery in the lumbosacral spinal cord of rats for 2 weeks) results in hyperexcitability of the bladder primary afferent neurons in the DRG. The underlying mechanism involves alterations in K⁺ channels (i.e. decrease in the density of A-type K⁺ current) that govern the excitability of these neurons [89]. Conversely, decreasing NGF levels and/or preventing NGF from binding to its receptors, TrkA and p57, using intravenous injection of an NGF-scavenging recombinant protein, systemic administration of an anti-NGF antibody, intravesical instillation of antisense oligodeoxynucleotides to block NGF synthesis, and intrathecal injection or instillation of k252a, a nonspecific antagonist of Trk receptors, improve bladder function (reducing micturition frequency and non-voiding contractions), and reduce pain in various animal models [80, 83]. Similarly, in patients with IC/BPS, reduction of NGF levels following botulinum toxin A (BoTN/A) injections into the bladder improved the symptoms [80]. Expression of BDNF, particularly in the afferent pathways (DRG neurons), is altered in animals models associated with inflammation triggered by cyclophosphamide or turpentine instillation [83]. This is associated with increase in the proliferative marker KI76 in bladder tissue, suggesting a role in organ hypertrophy following injury. Sequestration of BDNF using TrkB-Ig2 intravenous or intravesical instillation in an inflammatory model (cyclophosphamide), improves bladder function (i.e. reduces micturition frequency) and decreases the expression of nociceptive markers (pERK, c-Fos) in the spinal cord [90]. Thus, the actions of BDNF may be more prominent in neuronal pathways involved in bladder pain.

Artemin is a glial derived growth factor important for the development and repair of a particular class of afferent neurons, best characterized by their expression of TRPA1. In rodent models of inflammation (e.g. cyclophosphamide induced bladder cystitis), artemin expression is increased and is correlated with bladder hyperactivity and visceral hypersensitivity [91]. Reducing artemin levels using an anti-artemin antibody not only blocks but also reverses already established visceral hyperalgesia [91]. Artemin can sensitize primary afferent neurons and contribute to persistent pain by regulating the expression and function of nociceptive receptor channels TRPV1 and TRPA1 [92–95]. Taken together, there is support for artemin as a candidate target for treatment of chronic pain disorders associated with neurogenic inflammation. However little is known about artemin expression or function in either animal models of neurogenic inflammation or human tissue from patients that present with neurogenic inflammation (e.g. IC/BPS or chronic prostatitis).

Much less is known about other neurotrophic factors, such as NT-3 and NT-4 in the bladder. There is evidence for increased expression NT-3 and NT-4 in the bladder wall in models of cystitis or NDO [83] as well as in the urine pf IC/BPS patients [96], however their role is relatively unknown. Together, these studies demonstrate that neurotrophins, and especially NGF, are important contributors to conditions associated with neurogenic inflammation. Furthermore, NGF was proposed as a biomarker for these conditions and it has been suggested that anti-NGF therapies may be beneficial.

Clinical significance of the sensory web

Studies have shown that defects in urothelial sensor molecules and urothelial-cell signaling can contribute to the pathophysiology of bladder diseases. Upon inflammation or in response to irritants, the urothelium can synthetize and release different factors that further amplify the inflammatory response and contribute to symptoms. Conditions involving neurogenic inflammation (IC/BPS, SCI, chemically-induced cystitis) are associated with augmented release of urothelial-derived ATP [64–68], prostaglandins, nitric oxide, ACh, NGF, leukotrienes and others (Fig 2) [97, 98]. ATP excites purinergic receptors on nearby sensory fibers resulting in altered sensations or changes in bladder reflexes [99, 100]. Additionally, a number of cytokines/ chemokines and their receptors are upregulated in inflammatory conditions, including the chemokine/receptor pairs CXCL12/CXCR4 and CCL2/CCR2, the cytokine/receptor pair transforming growth factor (TGF-β)/TGF-β type 1 receptor, IL-6, IFNγ, and others [101, 102]. Thus, the urothelium has the potential to amplify mechanical and chemical stimuli within the urothelium but also involving other cells in the bladder wall. Structural and functional changes induced by neurogenic inflammation can lead to enhanced signaling between the urothelium and underlying cells in the bladder wall, resulting in amplified signals and contributing to a gain of function in sensory processing.

4.2.4. Vasodilation, increased vascular permeability and angiogenesis

Vascular smooth muscle is densely innervated by CGRP and SP expressing nerves [103, 104]. Vasodilation and increased vascular permeability are prominent features of neurogenic inflammation. Vasodilation is primarily mediated by CGRP acting on receptors on the vascular smooth muscle [105, 106]. For example, in the pig bladder, CGRP does not affect the smooth muscle contractility but potently dilates the blood vessels via a direct action on the vascular smooth muscle (i.e. removal of the endothelium or nitric oxide synthase inhibition does not prevent the effects of CGRP)[107]. In addition, SP also causes vasodilation [108] by acting on NK1 receptors on the vascular smooth muscle. Vascular permeability is primarily mediated by SP and NKA acting on NK1 receptors, which are abundant in the endothelial cells and the smooth muscle of the blood vessels. Indeed, NK1R antagonists were able to reduce/block SP-induced plasma extravasation, commonly measured in animal models by the Evans blue method. [108, 109]. Morphological analysis of the tissue from patients and animal models shows areas of leukocyte infiltration, edema, mast cells [25] and bladder wall glomerulations (‘pinpoint bleeding’) suggestive of fragile blood vessels (review [110]). A recent study using intravital microscopy methods imaged in vivo interactions between leukocytes and endothelial cell in the microcirculatory system following capsaicin application to the serosa surface of the bladder [111]. The experiments revealed a rapid (within 15 minutes after application) and long lasting (over 45 min) increase in the dynamic of leukocyte rolling and adhesion to endothelial cells accompanied by increased immunoreactivity for the adhesion proteins E-selectin and ICAM-1. These interactions, which are likely involved in leukocyte infiltration in the tissue, are dependent on the activation of C-fibers (as they were prevented by capsazepine or neonatal systemic capsaicin treatment) and involve CRGP and NK1 receptors. Leukocyte rolling was prevented by a CGRP receptor antagonist (CGRP8-34) but not by an NK1 receptor antagonist (RP67580), while the adhesion was prevented by blocking either CGRP or NK1 receptors. Taken together, these findings suggest that the dynamics and extent of neurogenic inflammation processes depend not only on the density of the TRPV1 positive fibers, but also on other factors such as the density of CGRP receptors and the expression of adhesion molecules in the tissue [111]

Tissue remodeling - role of VEGF

Repetitive injury induces tissue remodeling. This is evidenced by the presence of Von Brunn's nests (e.g. in human bladders with NDO [51] and cats with FIC [112]) and increased angiogenesis [110, 112, 113]. Vascular endothelial growth factor (VEGF) plays a significant role in remodeling of blood vessels as well as exerting protective effects on neurons and other cell types upon injury. Changes in VEGF and/or VEGF receptors/ signaling pathways in the bladder have been reported in patients with IC/BPS [113–115], and rodent models of bladder inflammation [114, 116]. High expression of VEGF in IC/BPS biopsies was associated with immature blood vessels that have insufficient coverage of pericytes, resulting in hemorrhagic vessels [113]. While VEGF is thought to act primarily on endothelial cells on blood vessels, there is evidence that urothelial cells and intramural ganglia cells are also potential targets. VEGF receptors (VEGF-R1, VEGF-R2) and co-receptors, the neuropilins (NRP 1,2, which enhance binding to of VEGF to VEGF-R2) are expressed in these cells and substantial differences were reported in bladders from IC/BPS patients compared to controls. [114] VEGF can alter the expression of tight junctions in epithelial cells [117] and thus increase urothelial permeability. Intravesical instillation of VEGF (for 1 – 4 weeks in mice) has been shown to alter bladder function (reducing the micturition pressure threshold and increasing voiding frequency), and increase nerve density (TRPV1, PGP 9.5) [118].

These findings indicate that IC/BPS (possibly because of neurogenic inflammation) is correlated with upregulation of VEGF system in the bladder which is responsible for alterations in blood vessels, afferent nerves, bladder function and pain. Therefore, blocking the VEGF signaling pathways may represent a potential treatment method for IC/BPS.

4.3. Immune response and involvement of the urothelium

4.3.1. Mast cells

Mast cells are produced by the bone marrow and their accumulation in the tissue is triggered by various factors, including SP. These cells store a number of mediators (e.g. heparin, histamine, 5-HT, proteases, phospholipases, chemotactic substances, and cytokines), and also can synthesize de novo factors (e.g cytokines: IL-6), leukotrienes (LTC4), prostaglandins (PG; e.g., PGD2), platelet-activating factor (PAF), and NO), all of which can impact the urothelium, smooth muscle and nerves. Mast cells also release VIP and TNF-alpha which are vasodilators.

The involvement of mast cells in the genesis and development of neurogenic cystitis is supported by a number of observations. First, there is an increased number of mast cells in bladder tissue from IC/BPS patients [119–123], FIC [36], rats exposed to immobilization stress [32] and rodents with virus induced cystitis [29, 31]. This increase may be related to injury and/or alterations of the urothelial cells that result in production of cytokines and NGF, which can stimulate proliferation and/or activation of mast cells [124]. Second, in mice deficient of mast cells, the severity of neurogenic inflammation induced by intravenous SP or E. Coli LPS is greatly reduced [24]. Third, degranulation resulting in the release of histamine, tryptase and other factors, plays an important role in cystitis. Animal studies have shown that pre-emptive mast cell degranulation prevents virus-induced cystitis in rats [29]. Mast cells are commonly found in the close vicinity of SP positive fibers, suggesting that factors released upon degranulation can easily target afferent nerves [50, 119, 125]. In addition, the urine of rodents with virus-induced cystitis and patients with IC/BPS, contains higher levels of histamine, methylhistamine and tryptase compared to controls [29, 126]. Tryptase can activate protease-activated receptors (PARs) in the urothelium, in particular PAR1, resulting in inflammation and submucosal neuronal hyperexcitability [127, 128]. Taken together, vasoactive and pro-inflammatory factors released by mast cells may act on afferent nerves, urothelial cells and other structures in the bladder, and can greatly contribute to bladder pathology [124].

4.3.2. Bladder microbiota

In recent years, a number of studies suggest that the human urinary tract contains a diverse microbiota that likely plays an important role in bladder health and disease [129]. It is becoming clear that the urine contains different populations of bacteria and the composition of the microbiota is correlated to urinary symptoms, especially urgency and urinary incontinence [130–133], and associated with response to treatment [133, 134]. The urothelial cells are the first line of defense against pathogens. Studies using uropathogenic Escherichia coli (UPEC) infection of the bladder (chronic and acute) have shown apoptosis of the apical urothelial cell layer and inflammatory responses involving release of multiple mediators from the urothelium (such as interleukins and cytokines) [135], which result in structural damage to the urothelial barrier [136–138]. As a result urothelial cells start mounting multiple defense mechanisms to limit inflammatory responses [139]. For example, these cells can secrete pro-inflammatory cytokines including interleukin-1 (IL-1), IL-6 and IL-8 (usually detected in urine following infection), which play important role in the recruitment of phagocytes into the infected bladder or kidney tissue. Additionally, the urothelium may produce antibacterial agents, such as uromodulin (also known as Tamm-Horsfall urinary glycoprotein) which acts to prevent bacteria from interacting with the epithelial cell surface [140], or beta-defensin which restricts bacterial growth [141]. Urinary tract infections (UTIs) are often accompanied by acute and chronic pain. It is believed that the acute pain is related to the release of mediators from the urothelium that produce inflammation and activation of afferent nerves. In addition, there is evidence for both inflammation dependent and independent pain mediated by endotoxin (a glycolipid on the outer membrane of Gram-negative bacteria) interacting with Toll-like receptors 4 [138, 142] and dependent on TRPA1 activation [143]. Chronic pain post UTI develops in some patients and it can also be reproduced in animal models. Experiments in which activation of TRPV1 receptors was prevented using the antagonist capsazepine or in TRPV1 knock-out mice prior to bacterial infection (with the E. Coli stain SΦ874), revealed that allodynia and increased VMR reflexes (an indicator of nociception) triggered by bladder distention, were prevented [142]. Together, these studies demonstrate that the microbiota by engaging urothelial cells and afferent TRPV1 nerves plays an important role in pain after bladder infection.

4.4. Alterations in the smooth muscle: bladder hyperreflexia

SP and NKA can activate NK receptors located on the smooth muscle. In vitro contractility studies in several species (human, rat, hamster, pig) demonstrated NK2 receptor mediated contraction of the bladder smooth muscle [144–149]. In vivo experiments in anesthetized animals (rat, hamster, guinea pig) reported that NKA as well as specific NK2 receptor agonists produce an increase in bladder contractility associated with non-voiding contractions [146, 150–153]. Thus, during neurogenic inflammation, excess release of SP and NKA can lead to increased smooth muscle contractility which can contribute to bladder spasticity, urgency and pain.

4.5. Neurogenic inflammation and the influence of urethral ‘sentinel’ (paraneuron) cells

Neurogenic inflammation can extend beyond the bladder and can affect the urethra, which is densely innervated by CGRP and SP expressing nerve fibers extending into the epithelium [17–20]. Common urethral pathologies such as urethritis, inflammatory polyps, cysts, strictures and in some cases the use of catheters are characterized by pain, inflammation of the mucosa and/or neurogenic inflammation [154–158]. For example, in rats, irritation of the urethra by mechanical stimulation due to catheter insertion resulted in neurogenic inflammation which was dependent on capsaicin sensitive nerves [159–161].

In contrast to the bladder mucosa, much less is known about urethral mucosa and its involvement and response to neurogenic inflammation. The urethral epithelium possesses several classes of specialized cells, termed paraneurons, identified by the neurotransmitter they express (e.g., acetylcholine (ACh), [162] serotonin (5-HT), [162–170] or somatostatin [169]) (Fig 5). These cells reside in the epithelium but can reach the lumen, often expressing microvilli, and may share similarities to GI enterochromaffin cells or chemosensory cells of the trachea or respiratory tract (termed ‘brush cells). ACh expressing paraneurons have been shown to express bitter taste receptors and respond to bitter stimuli (e.g. denatonium) and UPEC by releasing ACh. These cells are located in the proximity of nerve fibers expressing nACh receptors (Fig 5C), presumed afferents and intra-urethral infusion of denatonium reflexively increases bladder activity in anesthetized rats. Based on these experiments, it was hypothesized that ACh expressing paraneurons act as ‘chemosensory sentinels’ to monitor the lumen for potential hazardous content such as those produced by infecting bacteria [162, 171, 172]. The other major group of urethral specialized cells, the 5-HT expressing paraneurons, are anatomically located in close proximity to sensory nerves expressing CGRP (Fig 5B), SP and TRPV1. Experiments mimicking the release of 5-HT from these cells demonstrate that 5-HT activates urethral primary afferent neurons including capsaicin sensitive neurons, presumed C-fibers. Furthermore, intra-urethral 5-HT increases the expression of pain associated markers such as p-ERK in the spinal cord and sensitizes visceromotor reflexes elicited by non-noxious urethral distention [19]. As neurogenic inflammation is associated with increased release of peptides from the afferent nerves, it can be speculated that interaction between nerves and 5-HT expressing paraneurons may increase the release of 5-HT, which can in turn increase neuronal excitability and contribute to pain.

Figure 5. Neurogenic inflammation in the urethra.

A. Cartoon depicting interactions between paraneurons, epithelial cells and nerves in the urethral epithelium. Stimuli are detected by paraneurons and epithelial cells and conveyed to the nerves using transmitters including 5-HT and ACh from paraneurons. Information in then transmitted to the spinal cord and brain. Communication with nerves can be bi-directional. Nerves can release CGRP, SP, NKA which then can act on paraneurons and epithelial cells to modulate their activity. In neurogenic inflammation, increased release of peptides may result in modulation of transmitter levels from paraneurons and epithelial cells and may result in altered lower urinary tract function (LUT) and pain. Adapted from Kullmann et al., Acta Physiol (Oxf). 2017 Jul 18. doi: 10.1111/apha.12919) B. Relationship between a paraneuron expressing 5-HT (red) and nerve fibers expressing CGRP (green). C. Relationship between a paraneuron expressing Villin (yellow) and presumed ACh and nerve fibers expressing nAChRs (green). Adapted with permission from Deckmann et al. Proc Natl Acad Sci U S A. 2014 Jun 3;111(22):8287–92.

While our knowledge of urethral epithelial-afferent signaling is limited, it is clear that ‘sentinel’ cells are able to recognize potentially harmful stimuli in the urethral lumen, urethral epithelium, or released from the nerves and in turn modulate the excitability of afferent neurons, alter lower urinary tract function and pain. (Fig 5)

5. Potential treatments/targets

A variety of treatment modalities have been proposed for reliving the symptoms of conditions associated with neurogenic inflammation, namely bladder overactivity, urgency, frequency and pain.

5.1. TRPV1 agonists

TRPV1 receptors are expressed in the afferent nerves and urothelial cells [8]. While the role of TRPV1 in normal human bladder is not well understood, a role in the pathophysiology and treatment of neurogenic bladder has been well established [173, 174]. It is believed that C-fibers do not participate in the micturition reflex in normal bladder, however after spinal cord injury [42, 43, 48], bladder outlet obstruction [175] or idiopathic detrusor overactivity [176, 177] these fibers play a significant role. Studies in rats with SCI demonstrated reduction of non-voiding contractions after desensitization of TRPV1 fibers using capsaicin. Thus, it has been predicted that detrusor overactivity that develops after SCI and other pathologies, and that contributes to bladder spasticity, urgency and pain, depends on the activation of TRPV1 positive fibers. Consequently, it was proposed that desensitization of these fibers using intravesical capsaicin would be beneficial. Indeed, studies using intravesical capsaicin or RTX in patients with NDO demonstrated positive effect on bladder function, including increased bladder capacity, decrease in voiding frequency and urinary incontinence [176, 177]. Other beneficial effects in reducing nocturia were reported in patients with bladder pain [178]. However, a large meta-analysis suggested that although efficacy has been shown in some studies, the use of vanilloids is not yet recommended for routine use in patients with OAB or BPS, due to lack of strong evidence for efficacy or tolerability, and larger studies are needed to better define their role [179].

6.1. NK1 receptor antagonists

Evidence from preclinical animal models and clinical studies indicate that blocking NK1 receptors, using various antagonists, could be beneficial for reducing the effects of neurogenic inflammation. For example, plasma extravasation elicited by intravesical irritants in OVA-sensitized rats was prevented by SR 140333 (0.1 micromol kg(−1), i.v.) [27] and bladder overactivity induced by intravesical acetic acid was reduced by netupitant (0.1–3 mg/kg, i.v.) and L-733,060 (3–10 mg/kg, i.v.) in guinea pigs [180]. In patients with overactive bladder, serlopitant (0.25 and 4 mg for 8 weeks) [181], aprepitant (160 mg for 8 weeks) [182] and netupitant (50, 100, 200 mg for 8 weeks) [183], demonstrated trends for decrease in the number of urgency episodes, urinary incontinence episodes and micturitions per day. Although not fully explored, the results of these studies hold promise for the development of NK1R antagonists for bladder overactivity.

5.2. Purinergic receptor antagonists

Based on a well-established stimulatory role of ATP on afferent nerves (C-fibers) and findings that reveal ATP signaling is increased in bladders affected by pathological conditions such as IC/BPS, a purinergic P2X3 antagonist, AF-219/MK-7264 was developed and tested in patients with IC/BPS [184, 185]. In a proof-of-concept trial, IC/BPS women treated with AF-219 showed improvement in the key symptoms of IC/BPS: pain scores, urinary urgency and general improvement in patient reported symptoms [185]. These and other studies suggest that P2X3 receptor antagonists may have potential for in the treatment of IC/BPS symptoms.

5.3. Modulation of NGF levels

Evidence from preclinical animal models indicate that decreasing NGF levels and/or blocking NGF binding to its receptors, TrkA and p75, has beneficial effects on bladder function. Preclinical studies using intravenous injection of an NGF-scavenging recombinant protein (REN1820), systemic administration of anti-NGF antibody, intravesical instillation of antisense oligodeoxynucleotides to block NGF synthesis, and intrathecal injection or instillation of k252a, a nonspecific antagonist of Trk receptors, improve bladder function (reducing micturition frequency and non-voiding contractions) and reduce pain [83]. Recent evidence from an irradiation cystitis mouse model suggests that treatment with the p75 modulator LM11A-31 (a drug in clinical trials for the treatment of Alzheimer‘s disease - NCT03069014) ameliorates bladder overactivity and prevents irradiation- induced detrimental changes in the urothelium [186]. Results from clinical trials using biologic antibodies (tanezumab, a monoclonal anti-NGF antibody) to block the NGF effects indicated improvements in urgency episodes and pain in patients with chronic prostatitis/chronic pelvic pain syndrome [187] and patients with IC/BPS [188, 189]. However, adverse effects reported after tanezumab treatment in other clinical studies have led to discontinuation of further testing at this time.

5.4. Modulation of VEGF levels

Preclinical studies have shown that VEGF neutralizing antibodies reduce bladder inflammation and prevent associated increase in nerve density [118]. Clinical studies have shown increased VEGF expression in biopsies of patients with IC/BPS, correlated with the degree of pain [113]. Intravesical OnabotulinumtoxinA injections decrease VEGF expression, reduce bladder inflammation and improve bladder function [190]. Together, there is evidence that modulation of VEGF (perhaps in combination with other therapies) may be beneficial for reducing effects associated with inflammation and tissue remodeling.

5.5. Botulinum toxin A (BoNT/A)

BoNT/A has been shown to inhibit contraction of both smooth and skeletal muscle by blocking neurotransmitter release. The mechanism of action is thought to be due to cleavage of the SNARE protein SNAP-25. As the SNAP-25 proteins are also expressed in the urothelium and possibly afferent nerves, accumulating evidence suggests that BoNT/A exerts effects on afferent pathways. Preclinical models of acute or chronic bladder inflammation have shown that BoNT/A decreases the release of SP and CGRP and release of ATP from the urothelium. It also reduces bladder hyperalgesia and/or pelvic sensitivity [64, 191–195]. Clinical studies in patients with IC/BPS have shown that intravesical BoNT/A injections decreases VEGF expression, reduces bladder inflammation and improves bladder function [190] by decreasing daytime frequency, nocturia, and bladder pain [196, 197]. Repeated injections of BoNT-A reduces chronic inflammation and urothelial apoptotic signaling in patients with IC/BPS. This was associated with clinical symptom improvement [198]. Thus, the mechanism of botulinum toxin A action on IC/BPS includes actions on the afferent system (i.e. decreases CGRP, SP, and ATP release from urothelium, decreases the expression of P2X3 and TRPV1 receptor in the urothelium), an anti-inflammatory effect (i.e. decreases active mast cell number, decreases SP, NGF, and VEGF), and improvement of urothelium dysfunction (decreases apoptosis in the urothelium) [199].

6. Conclusions

Various types of stimuli can result in a neurogenic inflammation, including irritants and bacteria in the urine, peripheral mechanical injury (e.g. catheter insertion) and injury to the CNS (mechanical, vascular, or induced by viruses). Patients suffering from IC/BPS, prostatitis, spinal cord injury, and perhaps other CNS injuries frequently exhibit neurogenic inflammation of the mucosa, manifested as voiding dysfunction and pelvic pain.

The mechanisms underlying neurogenic inflammation are complex and involve a complex level of communication between cell types within the urothelium, lamina propria along with afferent nerves. The initial release of SP and CGRP peptides from TRPV1 positive fibers triggers a multitude of processes including vascular events (vasodilation and increased vascular permeability), the release of mediators from mast cells, urothelial cells, and other cells in the bladder wall, sensitization and resultant hyperexcitability of primary afferent neurons and changes in the CNS. Though their role is not well understood, growth factors (such as NGF, VEGF) and upregulation of purinergic receptors also are understood to play significant roles in the development and maintenance of patient symptoms. New emerging evidence highlights a potential role of ‘sentinel’ cells within the urethral epithelium which are likely to be important in the detection, transmission and amplification of peripheral signals implicated in neurogenic inflammation.

It is conceivable that the effectiveness of some agents currently used in the treatment of various inflammatory disorders may involve urothelial-neural sensory processes. Taken together, pharmacologic interventions aimed at targeting urothelial receptor/ion channels expression of release mechanisms may provide a new avenue of research and a potentially new strategy for the clinical management of such disorders.

Figure 4. Urinary bladder microbiota.

Summary of mechanisms involved in the curtail of inflammatory responses triggered by different pathogens (green). These can include exfoliation of bladder epithelial cells, increased proliferation to regenerate the barrier; mast cell secretion of interleukins (to aid in tissue regeneration); release of chemokines and other factors from macrophages. Adapted with permission from Abraham and Miao, 2015; Nat Rev Immunol. 2015 Oct; 15(10): 655–663.).