Biomarkers for lower urinary tract dysfunction

Authors


Correspondence: Joon Chul Kim M.D., Ph.D., Department of Urology, Bucheon St. Mary's Hospital, 2 Sosa-dong, Wonmi-gu, Bucheon City, Kyunggi-do 420-717, Korea. Email: kjc@catholic.ac.kr

Abstract

Lower urinary tract dysfunction is highly prevalent, but has relatively low persistence and compliance with therapy because of poor efficacy. Although urodynamic study is the gold standard for detailed evaluation of lower urinary tract dysfunction, urodynamic study has limitations as a biomarker, such as invasiveness and a lack of reproducibility of symptoms. Thus, many investigations about new biomarkers for lower urinary tract dysfunction have been carried out and reported. For imaging biomarkers, bladder and prostate parameters assessed by ultrasonography have been used to evaluate lower urinary tract dysfunction. For urinary biomarkers, neurotrophins, such as nerve growth factor and brain derived neurotrophics factor, prostaglandins and cytokines, have been analyzed and evaluated. Among these, nerve growth factor is considered one of the key factors in the pathophysiology of lower urinary tract dysfunction, and is researched in various ways. Serum markers have suggested that C-reactive protein and sex hormones have a relationship with lower urinary tract dysfunction. The possibility of genetic biomarkers in lower urinary tract dysfunction has also been raised. Nevertheless, as yet these biomarkers have not shown enough evidence to reflect lower urinary tract dysfunction and require further investigation. This review will discuss promising and potential biomarkers in lower urinary tract dysfunction to date.

Abbreviations & Acronyms
BDNF

brain-derived neurotrophic factor

BOO

bladder outlet obstruction

BPH

benign prostatic hyperplasia

BWT

bladder wall thickness

COX

cyclooxygenase

CRP

C-reactive protein

DO

detrusor overactivity

DWT

detrusor wall thickness

IC/PBS

interstitial cystitis/painful bladder syndrome

ICS

International Continence Society

IL

interleukin

IPP

intravesical prostatic protrusion

LUTD

lower urinary tract dysfunction

LUTS

lower urinary tract symptoms

MCP

monocyte chemotactic protein

NGF

nerve growth factor

OAB

overactive bladder

PG

prostaglandin

Qmax

maximum urinary flow rate

SUI

stress urinary incontinence

TrkA

tyrosine kinase receptor A

TrkB

tyrosine kinase B

TZI

transition zone index

UDS

urodynamic study

US

ultrasonography

Introduction

LUTD is a functional disorder characterized by symptoms of storage, voiding and post-micturition, and may or may not be related to anatomical abnormalities. LUTS are a common problem and increase with advancing age. Population-based surveys in five countries reported that 64.3% of adults experience at least one LUTS.[1] LUTS are associated with significant impairment in health-related quality of life and reportedly have similar effects as a heart attack or stroke.[2] A common LUTD, OAB, had a prevalence of 12.2% in the Korean European Prospective Investigation onto Cancer and Nutrition study.[3] OAB was estimated to affect up to 22.9% of the population aged more than 30 years and also has severe effects on health-related quality of life.[4]

As a result of significant advances in pharmacotherapy and surgery, many patients with LUTD are successfully treated. However, some patients continue to endure OAB because of decreased adherence to medical therapy due to adverse effects and poor efficacy.[5, 6] To determine appropriate therapy in LUTD, a comprehensive assessment of the lower urinary tract, functional impairments and concurrent medical conditions should be completed. However, the clinical diagnosis and definition of LUTD, and in particular OAB, are based on subjective symptoms according to the ICS.[7] UDS are objective tests carried out to evaluate bladder and urethral function during both the filling and voiding phases, and can provide information on the pathophysiology of LUTS. UDS can help identify the cause of urinary dysfunction, and evaluate associated comorbidities and the functional status of the patient. However, UDS might not accurately represent all LUTD-related symptoms, as there is a poor correlation between urinary symptoms and UDS findings.[8] Although UDS findings suggestive of OAB symptoms are attributed to DO, approximately 40% of men and 30% of women with no OAB symptoms were diagnosed with DO on UDS.[9] Assessment of bladder sensations through cystometry is not an objective measurement, because it relies on the compliance, alertness and subjective feelings of the patient. Bladder sensations vary widely at differing volumes, even in young healthy volunteers, during cystometry.[10] In addition, UDS is invasive, time-consuming, expensive and repeated testing is typically not well-tolerated. As a result, an exact, objective assessment of LUTD is limited and there is presently a growing interest in other biomarkers for LUTD.

According to the Biomarkers Definitions Working Group, a biomarker is defined as “a characteristic that is objectively measured and evaluated as an indicator of normal biologic processes, pathogenic processes, or pharmacologic responses to a therapeutic intervention.”[11] Biomarkers for LUTD have greater value in early efficacy and safety evaluations. In addition to UDS, other biomarkers in LUTD patients include imaging findings, such as bladder wall thickness, urinary proteins (neurotrophins, prostaglandins and cytokines), serum biomarkers (C reactive protein and sex hormones) and genetic biomarkers. In the present review, we discuss these identified and potential biomarkers for an improved assessment of LUTD.

Imaging biomarkers

Bladder ultrasonography

US is a simple, rapid and non-invasive examination that can accurately assess parameters of bladder, such as BWT and bladder weight. Sustained isometric contraction of the detrusor muscle against an obstructed bladder outlet might lead to detrusor hypertrophy and wall thickening. Oelke et al. found that the diagnostic accuracy of BOO assessment was greater with ultrasound measurement of DWT than with free uroflowmetry, postvoid residual urine or prostate volume.[12] Another study determined that DWT, evaluated sonographically as 2.9 mm or greater, had a high predictive value for male BOO and could replace pressure flow study in diagnosing BOO.[13] As patients with DO might have frequent detrusor contractions, a relationship between increased BWT and DO can be presumed. Khullar et al. reported that 94% of women with a mean BWT greater than 5 mm with transvaginal US had detrusor instability with UDS.[14] Panayi et al. showed that women with OAB symptoms had a mean BWT value of 5.6 mm through transvaginal US.[15] Another study found that a cut-off BWT value of 6.5 mm had a positive predictive value of 100% for all DO.[16] However, the value of this biomarker seems to be limited. US for BWT is not yet standardized in terms of route, bladder volume at measurement, type and resolution of probe, and number and sites of measurements in the bladder. Bladder volume at measurement might depend on the sonographic approach. In general, transabdominal US measures DWT at a larger bladder volume, whereas transvaginal US requires a nearly empty bladder.[17] Bladder volumes can affect the measurements of BWT or DWT greatly. In addition, Blatt et al. reported no significant difference in BWT among patients with normal UDS, BOO or OAB.[18] They suggested that sonographic findings of BWT were not clinically useful for assessing LUTD, because premature use of ultrasound at this stage could not provide refined data of the gross bladder morphology to aid a clinical diagnosis.

The weight of the bladder, which has hypertrophied detrusor muscle, can be estimated by US. The measurement of bladder weight using US has been studied as a clinical technique that could evaluate LUTD. Bright et al. suggested that US-estimated bladder weight could be a potential non-invasive clinical tool for assessing the lower urinary tract.[19] However, other data showed that US-estimated bladder weight did not present any individual correlation with clinical and urodynamic BOO.[20]

Further studies and technical improvements are required to clarify and standardize US for bladder measurements.

Prostate ultrasonography

IPP by transabdominal or transrectal US could be a useful parameter to predict BOO in male patients. Significant IPP, where the vertical distance from the tip of the protrusion to the bladder circumference at the prostate base was greater than 10 mm, had a significant relationship with higher BOO index, higher incidence of detrusor overactivity and low bladder compliance.[21] The combination of IPP and DWT produced a more accurate assessment of BOO in patients with BPH.[22]

RI of prostate capsular arteries, which was measured using transrectal pulsed wave spectral Doppler US, is a parameter to diagnose BOO in patients with BPH. RI is determined by the formula: (peak systolic velocity – end diastolic velocity) / peak systolic velocity. Zhang et al. reported that RI was significantly higher in patients with BOO and could reflect BOO severity in patients with BPH.[23] Increased vascular resistance would be caused by increased intraprostatic pressure along with BOO.

Measurement of prostate volume by transrectal US could be a parameter to evaluate BOO, especially as the transition zone of the prostate correlated better with clinical BOO than total prostate volume alone. The TZI, which is determined by the formula: transition zone volume / total prostate volume, correlated significantly with BPH and might serve as a useful factor for evaluating worsening BOO.[24] In addition, an increase of the RI of prostate capsular arteries correlated with increases in the TZI.[25]

These parameters would be valued in male patients, especially in patients with BPH, but many other factors besides prostate determined BOO. Thus, they are of limited additional value.

Urinary biomarkers

Neurotrophins

Neurotrophins are a family of proteins that promote the survival and regulate the development and function of neurons.[26] Several neurotrophins are involved in altered lower urinary tract function through reorganization of the micturition reflex. The expression of NGF and BDNF have been determined in the urothelium and detrusor smooth muscle layers of the bladder, and can be quantitatively analyzed in urine.

Nerve growth factor

NGF is a small protein integral to the growth, maintenance and survival of particular target neurons, including those of the sensory pathway. It elicits neurotrophic actions through its high affinity receptor, TrkA.[27] Animal studies have found that the intravesical instillation of NGF acutely induced bladder hyperactivity in rats.[28] Overexpression of NGF in the bladder stimulates neuronal sprouting in the bladder, thereby increasing voiding frequency and somatic hypersensitivity.[29] Lowe et al. reported that women with idiopathic sensory urgency and interstitial cystitis had increased NGF expression in the urothelium.[30] Other studies have shown that urinary NGF levels were significantly increased in patients with OAB compared with normal controls.[31, 32] Furthermore, patients with OAB wet (at least one episode of urge incontinence in the voiding diary) with a higher percentage of DO had significantly higher urinary NGF levels than those with OAB dry.[33] Furthermore, patients with BOO and OAB had significantly greater urinary NGF levels compared with BOO without OAB.[34] In addition, women with mixed urinary incontinence and DO had significantly higher urinary NGF levels than those with pure stress urinary incontinence by urethral incompetence.[35] Taken together, these studies suggest that the expression of NGF in the bladder is closely associated with OAB.

Given the considerable urine NGF levels, it has been suggested that changes in bladder function can be detected by diagnostic urinalysis. Studies have investigated urinary NGF as a biomarker for assessing the therapeutic effects of treatment. Giannantoni et al. showed that intravesical botulinum toxin A reduced NGF bladder tissue levels in patients with neurogenic DO, suggesting that the decreased acetylcholine released at the presynaptic level induces decreased detrusor contractility and NGF production.[36] Elkelini et al. showed that intravesical onabotulinum toxin A significantly lowered NGF concentration in female rats with bladder dysfunction after spinal cord injury.[37] Urine NGF levels were significantly reduced in patients with OAB who had responded to antimuscarinic treatment, but elevated in those who failed to respond to antimuscarinic treatment and had recurrent OAB symptoms after the discontinuation of the treatment.[38] For the assessment of OAB, urinary NGF has been shown to be a better biomarker compared with DWT.[39]

Increased bladder NGF levels were also described in other inflammatory lower urinary tract conditions, such as bacterial cystitis[40] and IC/PBS.[41] However, bacterial cystitis or IC/PBS can be differentiated from OAB through urine cultures or cystoscopy. As mentioned earlier, NGF is not the sole protein responsible for OAB. The clinical studies investigating the role of NGF as a biomarker in LUTD are summarized in Table 1.

Table 1. Clinical studies investigating the role of NGF as a biomarker in LUDT
AuthorsConditionsnResults
Lowe et al.[30]Painful conditions of bladder (idiopathic sensory urgency, IC/PBS)16

The levels of NGF were higher in samples from all three painful bladder conditions than in samples from controls.

Immunostaining showed increased NGF expression in the urothelium, most marked in patients with idiopathic sensory urgency.

Kim et al.[31]Women with OAB65Urinary NGF increased >13-fold vs controls; no correlations with urodynamic parameters
Liu et al.[33]Patients with increased bladder sensation, OAB dry and OAB wet157Lower urinary NGF in controls and patients with increased bladder sensations vs OAB dry; higher urinary NGF in OAB wet vs OAB dry;
Liu et al.[34]Men with BOO with or without OAB153Urinary NGF elevated in BOO with OAB vs BOO without OAB; urinary NGF not different between BOO with OAB vs BOO with DO; urinary NGF reduced in well-treated BOO
Liu et al.[35]Women with mixed urinary incontinence85Lower urinary NGF in controls and women with pure SUI; Higher urinary NGF in women with mixed SUI and DO; Higher urinary NGF in women with de novo DO after antiincontinence surgery
Giannantoni et al.[36]Patients with DO unresponsive to anticholinergic therapy23Urinary NGF decreased after intravesical treatment with botulinum A toxin, persisted at least up to 3 months
Jacobs et al.[41]Patients with OAB and IC/PBS59Urinary NGF elevated in patient with OAB and IC/PBS vs controls

There are several limitations to using urine NGF as a biomarker for LUTD. First, measurement of urinary NGF levels requires specific technical expertise in methods such as enzyme-linked immunosorbent assay. Second, normal values of urinary NGF levels are still not clear and might depend on bladder distention and sex. Third, urinary NGF levels could increase physiologically in healthy people with a strong desire to void or a full, distended bladder.[42] Further studies to standardize urine sample collection and studies with larger control populations are required before urinary NGF can be used as a biomarker in the diagnosis and therapy of LUTD.

Brain derived neurotrophic factor

BDNF regulates the survival and differentiation of neurons by binding to TrkB.[43] TrkB is a neurotrophin receptor expressed in bladder sensory afferents, whereas TrkA has a high affinity to NGF.[44] Pinto et al. reported that intravenous administration of recombinant protein neutralized BDNF activity and effectively reduced the frequency of bladder contractions in rats with chronic cystitis, suggesting that BDNF might play a role in bladder function.[45] BDNF has been postulated to be critical in the development of inflammation-induced sensory hypersensitivity by modulating the sensitivity of primary afferents. Trigonal injection of botulinum toxin A in patients with IC/PBS reduced urinary BDNF significantly, resulting in LUTS improvement.[46] However, BDNF has been determined to be overexpressed in several tumor types including bladder transitional cell carcinoma.[47] Thus, the relationship between BDNF and bladder function remains unclear.

Prostaglandin

PG is a lipid compound that regulates the contraction and relaxation of smooth muscle tissue and mediates inflammatory processes. PG are synthesized locally from the bladder smooth muscle and urothelium by physiological stimulation, such as stretching of the detrusor muscle, urothelium injury, nerve stimulation and reaction to inflammatory mediators.[48] PG have been found to lower the threshold for the micturition reflex by acting through capsaicin-sensitive afferent nerves.[49] Intravesical administration of PGE2, which is produced in significantly larger quantities than other PG subtypes such as PGI2 or PGF in bladder mucosa,[50] induced bladder overactivity accompanied with decreased micturition interval and volume.[51] Urinary PGE2 and PGF were also significantly increased in patients with OAB who did not have any neurological diseases.[31] Moreover, urinary PGE2 was elevated in patients with suprapontine brain pathology and associated with the presence of OAB.[52] Furthermore, there have been several studies investigating the inhibition of PG synthesis and bladder function. COX inhibitors in rats with OAB inhibited isovolumic bladder contractions and increased bladder volume capacity in a dose-dependent manner.[53] In addition, intravesical instillation of COX-2 inhibitors reduced bladder hyperactivity in the rats with DO.[54]

Nevertheless, a role for PG as biomarkers of LUTD is controversial and further research is required. Urinary PGE2 levels have been found to be elevated in patients with urinary tract infections, bladder cancer and inflammatory processes, such as those induced by bacillus Calmette–Guérin. When these diseases are treated, urinary PGE2 levels are subsequently decreased.[55] Liu et al. reported that urinary PGE2 levels in patients with IC/PBS or DO were not significantly elevated, in contrast to urinary NGF levels, suggesting that PGE2 might not be a useful biomarker of LUTD.[56] Of note, clinical evidence indicating the therapeutic benefit of selective COX-2 inhibitors in patients with LUTD or the reduction of PG levels after adequate anticholinergic treatment of OAB have not yet been reported.

Cytokine

Cytokines consist of a group of small proteins secreted by numerous cells and mediate interactions between cells, particularly in inflammatory processes.[57] Cytokines from urothelial cells can be detected in urine, and might reflect bladder inflammation and contribute to LUTD. Elevated cytokine levels were found concurrently with increased urinary frequency in cyclophosphamide-induced cystitis, suggesting that multiplex analysis for urinary cytokines might be beneficial as a non-invasive assessment of inflammatory bladder disease.[58] Although the pathophysiology of OAB is complex and not completely understood, inflammation might have a role in its development, as signs of inflammation in bladder biopsies have been noted in OAB patients.[59] Analysis of urinary cytokines in OAB showed a significant elevation of cytokines, such as MCP-1, soluble fraction of the CD40 ligand, macrophage inflammatory protein, IL-12, IL-5 and IL-10.[60] It was hypothesized that elevated urine cytokines in the bladder might result from an interaction of overactive cholinergics and peptidergic/sensory innervations of the bladder with local immune cells, and contribute to altered sensory processing. Ghoniem et al. also found that some cytokines, such as MCP-1, were exclusively expressed; but others, such as IL-5, IL-6 and IL-7, were decreased in the urine of patients with OAB.[61]

Studies have examined the association between cytokines and IC/PBS. Urine IL-6 was significantly increased in patients with IC/PBS and correlated with symptom severity.[62] Tyagi et al. used urinary cytokines to differentiate ulcerative from non-ulcerative IC/PBS and found a significant increase in CXCL-1, CXCL-10 and IL-6 in ulcerative cases.[63] Corcoran et al. investigated the changes of urine cytokine levels in IC/PBS patients after hydrodistention and found that post-hydrodistention urine levels of MCP-3, IL-3 and tumor necrosis factor-related apoptosis-inducing ligand were significantly decreased, in keeping with clinical improvement.[64]

However, cytokines lack specificity for LUTD, as they might be elevated in other urological diseases, including bladder cancer[65] and bacterial cystitis.[66] Further research is required to carefully delineate the role of cytokines in LUTD.

Serum biomarkers

C-reactive protein

CRP is an acute phase protein, serving as a general, non-specific biomarker of inflammation and tissue damage.[67] The synthesis of CRP is rapidly upregulated by hepatocytes through signaling with cytokines, such as IL-6, originating at the site of pathology.[68] As mentioned earlier, some LUTD subtypes, such as OAB, might be associated with inflammation. The AUA-SI determined that increased CRP levels were associated with increased odds of moderate to severe LUTS (AUA-SI 8).[69] Chung et al. reported that serum CRP levels were significantly higher in patients with OAB and IC/PBS in both sexes.[70] However, those levels were not as high as those in acute systemic inflammation, and only a small portion of OAB and IC/PBS had clinically significant elevation of serum CRP levels (>3 mg/L), suggesting that the inflammation in these chronic bladder disorders was likely local and mild. The Boston Area Community Health Survey showed a consistent association between increasing CRP levels and OAB among both sexes, with a larger magnitude of association in men.[71] Hsiao et al. investigated the role of serum CRP levels in women with LUTD based on voiding diaries, uroflowmetry and selective video UDS.[72] They found that serum CRP levels were significantly higher in OAB wet than those in bladder oversensitivity (patients with storage symptoms, but without urgency, having a normal uroflowmetry results and low bladder capacity) and control groups. In addition, increased serum CRP levels were associated with a low Qmax, but not postvoid residual volumes, suggesting that patients with a low Qmax had bladder dysfunction secondary to inflammation in the detrusor muscles or bladder outlet. Decreased serum CRP in response to therapy for systemic inflammation suggests that serum CRP might be a useful biomarker for monitoring disease status and response to treatment in patients with LUTS.

Chuang et al. evaluated CRP expression in bladder tissue and urine CRP levels in patients with LUTS, and found that urinary CRP was rarely detected, and mRNA expression of CRP was very modest and several fold lower than the expression of housekeeping genes in the detrusor or urothelium.[73] They do not recommend CRP as a biomarker of LUTD. Studies have shown an association between CRP and LUTD, but there is a limit to serum CRP discriminating LUTD subtypes.

Sex hormones

Although the results about the relationship between serum total testosterone and BPH have not been consistent, higher serum dihydrotestosterone level was associated with larger prostate volumes.[74] As sex hormones could be contribute to the development of LUTS secondary to BPH in older men, several studies about the relationship between LUTS and serum sex hormones have been carried out. Favilla et al. showed that the severity of LUTS evaluated using IPSS was associated with serum total testosterone levels.[75] Rohrmann et al. reported that greater circulating concentrations of androstanediol glucuronide and estradiol were associated with an increased risk of LUTS compared with low levels, but no associations with LUTS were noted for circulating testosterone, free testosterone or sex hormone binding globulin concentrations.[76] However normalization of testosterone levels by testosterone administration in men with late onset hypogonadism might improve LUTS/bladder functions by increasing bladder capacity, compliance and decreasing detrusor pressure at maximal flow.[77]

These results mean that abnormally high and low levels of sex hormones might contribute to LUTD. Therefore, better and more studies about sex hormones are required.

Genetic biomarkers

Several studies have provided evidence for a genetic basis in the pathophysiology of LUTD. Family history studies have found a two- to threefold higher prevalence of urinary incontinence among first-degree relatives of incontinent women compared with first-degree relatives of continent women.[78] Furthermore, a population-based survey in a cohort of adult twins showed the strongest genetic effects in urinary incontinence, frequency and nocturia.[79] Blalock et al. analyzed the gene expression in urine in patients with IC/PBS and found that patients with Hunner's lesion IC/PBS had increased pro-inflammatory gene expression in urine sediment.[80] Peripheral blood mononuclear cell studies showed that 16 genes were differentially regulated in all patients with OAB.[81]

Nevertheless, there are currently no definitively identified genetic factors associated with LUTD, but the discovery of genetic biomarkers might help prognosticate LUTD and enable susceptible patients to alter their lifestyles accordingly to prevent or delay disease onset.

Conclusion

Without invasive diagnostic procedures, it is difficult to differentiate LUTD subtypes, such as OAB, BOO and voiding dysfunction. There are no definite, clinically useful biomarkers for the diagnosis of disease. Bacterial cystitis or lower urinary tract neoplasm can be differentiated from other causes of LUTD based on routine diagnostic tools. Further research into new, LUTD-specific biomarkers and easy, rapid methods to detect these biomarkers is required. Future research directions might consider a combination of biomarkers to increase specificity. For instance, serum CRP might be useful in conjunction with urinary inflammatory chemokines as a multivariant biomarker of LUTD.

Conflict of interest

None declared.

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