Ingrid Ehrén

The antimicrobial peptide cathelicidin protects the urinary tract against invasive bacterial infection

The urinary tract functions in close proximity to the outside environment, yet must remain free of microbial colonization to avoid disease. The mechanisms for establishing an antimicrobial barrier in this area are not completely understood. Here, we describe the production and function of the cathelicidin antimicrobial peptides LL-37, its precursor hCAP-18 and its ortholog CRAMP in epithelial cells of human and mouse urinary tract, respectively. Bacterial contact with epithelial cells resulted in rapid production and secretion of the respective peptides, and in humans LL-37/hCAP-18 was released into urine. Epithelium-derived cathelicidin substantially contributed to the protection of the urinary tract against infection, as shown using CRAMP-deficient and neutrophil-depleted mice. In addition, clinical E. coli strains that were more resistant to LL-37 caused more severe urinary tract infections than did susceptible strains. Thus, cathelicidin seems to be a key factor in mucosal immunity of the urinary tract.

Nitric oxide: a useful gas in the detection of lower urinary tract inflammation.

PURPOSEnLuminal nitric oxide has been shown to be elevated in the bladder of patients with cystitis of various etiologies. We determine whether luminal nitric oxide can be used as a marker to differentiate inflammation, that is interstitial cystitis, from urgency, frequency, nocturia and pain due to noninflammatory disorders, such as outflow obstruction and neurogenic dysfunction.nnnMATERIALS AND METHODSnWe measured luminal nitric oxide in the bladder of patients with urgency due to detrusor instability (6), outflow obstruction (7), sensory urge (19) and interstitial cystitis (8), and controls without urgency symptoms (11). Nitric oxide-free air was incubated in the bladder for 5 minutes and analyzed in a chemiluminescence nitric oxide analyzer.nnnRESULTSnThere was a nearly 20-fold increase in mean bladder nitric oxide concentration in patients with interstitial cystitis (234+/-67 parts per billion) compared to those with detrusor instability (11+/-1), outflow obstruction (9+/-1) and sensory urgency (10+/-1), and controls (13+/-2).nnnCONCLUSIONSnMeasurement of nitric oxide in air from the bladder is a simple, safe and fast method to differentiate urgency due to inflammation from neurogenic disorders or outflow obstruction. The simplicity of this method makes it potentially useful as a screening method for office use.

Effects of l-arginine treatment on symptoms and bladder nitric oxide levels in patients with interstitial cystitis

OBJECTIVESnNitric oxide (NO) is involved in host defense reactions, and NO production is elevated in various inflammatory disorders. We have found very high levels of luminal NO in the urinary bladder of patients with interstitial cystitis. Oral treatment with low doses of L-arginine, the substrate for NO production, has been reported to alleviate symptoms in patients with interstitial cystitis. The aim of our investigation was to evaluate the effect of higher doses of L-arginine in patients with interstitial cystitis and to study the effects of L-arginine on NO production in the bladder.nnnMETHODSnNine women (age 69+/-3 years) with interstitial cystitis were treated daily with 3 or 10 g of L-arginine for 5 weeks. Symptoms were evaluated with an interstitial cystitis symptom score index, and NO production was measured. Patients with stress incontinence (n=18) were used as control subjects for measurement of NO levels.nnnRESULTSnNO concentration in the urinary bladder was markedly elevated in the patients with interstitial cystitis (239+/-60 ppb) compared with the control patients (15+/-2 ppb). NO levels did not change in the patients with interstitial cystitis after oral treatment with L-arginine (189+/-72 ppb). There was no significant change in the symptom scores at either dose after 5 weeks of L-arginine treatment.nnnCONCLUSIONSnL-arginine treatment in the doses used in this study did not change NO production in the urinary bladder in patients with interstitial cystitis. Furthermore, the patients in our study did not notice any relief of their symptoms.

Characteristics of Biofilms from Urinary Tract Catheters and Presence of Biofilm-Related Components in Escherichia coli

Long term catheterization of the urinary tract leads to bacterial colonization of the urine, whereby adherence to the catheter surface is a major determinative factor for colonization. Collection of bacterial isolates from urine and urinary catheters of 45 patients showed multi-species catheter-colonization, while Escherichia coli isolates were frequently found in the urine in high numbers. Biofilm formation of catheter and urine-derived E. coli isolates was associated with the presence of the fluA gene, loss of O-antigen, and expression of type 1 fimbriae. The second messenger cyclic di-GMP (cdiGMP), a major regulator of biofilm formation, regulated adherence to the catheter surface in a selected clinical isolate suggesting that the cdiGMP second messenger pathway may be a target for anti-biofilm therapeutic approaches.

Measurement of luminal nitric oxide in bladder inflammation using a silicon balloon catheter: a novel minimally invasive method.

OBJECTIVESnNitric oxide (NO) measured in the gaseous phase has been shown to be a marker of inflammation in the urinary bladder. The NO content of air incubated in the bladder can be measured in an NO analyzer. The aim of our study was threefold: to evaluate whether NO can be measured in air incubated in a catheter balloon, to determine the optimal time of incubation, and to find the most suitable type of catheter.nnnMETHODSnThe NO concentration in air introduced directly into the bladder and into the catheter balloon was measured in patients with and without bladder infections. The air was incubated for 5 to 60 minutes. NO concentration in the bladder of patients with interstitial cystitis was also analyzed. The diffusion rate of NO through silicon and latex catheters was studied.nnnRESULTSnElevated NO levels were detected in the urinary bladder in patients with bladder inflammation due to infection or interstitial cystitis. A marked increase in NO concentration was found after just 5 minutes of incubation and continued to rise for up to 20 minutes, both in air taken directly from the bladder and from the catheter balloon. The NO diffusion rate into the balloons of silicon catheters was high; the recovery rate in latex catheters was poor.nnnCONCLUSIONSnMeasurement of NO concentration in a silicon balloon catheter inserted into the urinary bladder is a fast, convenient, and reliable method to detect inflammation.

BACILLUS CALMETTE-GUERIN INDUCES LONG-TERM LOCAL FORMATION OF NITRIC OXIDE IN THE BLADDER VIA THE INDUCTION OF NITRIC OXIDE SYNTHASE ACTIVITY IN UROTHELIAL CELLS

PURPOSEnBladder instillation of bacillus Calmette-Guerin (BCG) is effective therapy for recurrent superficial bladder cancer and carcinoma in situ. BCG induces nitric oxide synthase activity in the bladder. Nitric oxide is formed from L-arginine by nitric oxide synthase. We investigated nitric oxide formation and its localization in bladder cancer patients treated with intravesical BCG instillation.nnnMATERIALS AND METHODSnThe L-citrulline conversion assay was done to assess nitric oxide synthase activity in BCG treated T24 human bladder cancer cells and cultured normal human urothelial cells. Nitrite and nitrate in cell culture medium, urine and plasma were measured by capillary electrophoresis. Nitric oxide formation in the bladder was measured by chemiluminescence.nnnRESULTSnA 24-hour treatment with BCG induced calcium independent nitric oxide synthase activity in T24 cells in a dose dependent manner. Nitrite and nitrate production by T24 cells also increased in a dose dependent manner after 24-hour BCG treatment. BCG treatment of cultured normal human urothelial cells resulted in the induction of calcium dependent and independent nitric oxide synthase activity. Nitrite in the urine of patients receiving BCG for the first time was increased 5-fold 24 hours after instillation. Furthermore, BCG increased luminal nitric oxide in the bladder. The increase was noted after a single treatment and sustained for 6 months. No changes in plasma nitrite or nitrate were observed after BCG treatment.nnnCONCLUSIONSnBCG induces the local formation of nitric oxide in the bladder, whereas no evidence for systemic nitric oxide formation was noted. Increased nitric oxide production in the bladder is likely due to the induction of nitric oxide synthase activity in urothelial cells.

Nitric oxide as a marker for evaluation of treatment effect of cyclosporine A in patients with bladder pain syndrome/interstitial cystitis type 3C.

Abstract Objective. Bladder pain syndrome/interstitial cystitis (BPS/IC) is a chronic inflammatory disease and to date few treatments or tools for investigating the activity of the disease are available. This study evaluated whether luminal nitric oxide (NO) could be used as a marker for evaluation of therapeutic outcome in BPS/IC type 3C treated with the immunosuppressive agent cyclosporine A (CsA). Material and methods. Ten patients with BPS/IC type 3C were given CsA for 16 weeks, initially at 3 mg/kg/day, and after 12 weeks the dose was scaled down. Formation of NO was measured in the urinary bladder with a silicone catheter, and symptom and bother score related to the disease were evaluated with the Interstitial Cystitis Symptom and Problem Index, every second week. Results. All patients had elevated NO levels in the bladder initially and NO levels decreased during treatment with CsA. When the dose of CsA was lowered NO formation increased and after 2 weeks without medication, the NO formation was the same as before the study began. Conclusions. The results indicate that measurement of NO is a tool for evaluating the response to anti-inflammatory treatment in patients with BPS/IC type 3C. NO could serve as a marker for assessing the activity of the inflammation.

Measurement of nitric oxide may differentiate between inflammatory and non-inflammatory prostatitis

Objective. The majority of patients with prostatitis have chronic non-bacterial prostatitis/chronic pelvic pain syndrome of inflammatory type (Category IIIA) or non-inflammatory type (Category IIIB), based on the National Institutes of Health classification. The aim of this study was to investigate whether measurement of nitric oxide (NO) formation in the prostatic urethra can be used as a marker for inflammation in the evaluation of patients with chronic prostatitis/pelvic pain syndrome. Material and methods. A total of 25 men with prostatitis were examined. In 8 patients >10 white blood cells/high-power field (WBC/hpf) were found in expressed prostatic secretion (EPS) (Category IIIA), whereas the other 17 had no signs of inflammation (Category IIIB). NO production was measured using a silicon catheter, with the catheter balloon being placed in the prostatic urethra. Room air (5u2009ml) was incubated for 5u2009min and analyzed. NO formation in the urinary bladder was also measured. Results. The NO concentration in the prostatic urethra was significantly higher in the 8 patients with >10u2009WBC/hpf than in those with <10u2009WBC/hpf. The NO concentration in the urinary bladder was low in both groups. Conclusions. We found an elevated NO concentration in the prostatic urethra in patients with >10u2009WBC/hpf in the EPS but not in those with <10u2009WBC/hpf, which supports the theory of different pathogeneses for Categories IIIA and IIIB. Measurement of NO production in the prostatic urethra can be used to discriminate between the two categories and as the method is easy and fast it may represent an attractive alternative to the four-glass test.

Measurement of nitric oxide in the vagina.

Objective. Local measurement of nitric oxide (NO) gas has been used to detect and monitor inflammatory processes in the airways, the colon and in the urinary bladder, but so far NO has not been studied in the lower female genital tract. The objective of this study is to measure NO gas directly in the vaginal lumen of healthy women and in patients with vaginitis. Setting. The outpatient clinic of a university hospital. Methods. Eighteen non‐pregnant women from 19 to 65 years of age with symptoms of vaginitis, eight healthy women in reproductive age with regular menstrual cycles and nine healthy postmenopausal women were enrolled in the study. NO levels were measured in air incubated for five minutes in a catheter balloon in the vagina. Results. In patients with symptoms of vaginitis, NO concentration was almost 100‐fold increased compared to healthy controls (p<0.001) with no individual overlap. Vaginal NO levels were uniformly low among healthy women, both in reproductive age and in menopause. Conclusions. NO gas can be measured directly in the vagina with a fast, simple and safe method. The levels of NO are increased in patients with vaginitis.

Measurement of luminal nitric oxide in the uterine cavity using a silicon balloon catheter

The aim of this pilot case-control study was to measure nitric oxide (NO) gas in air incubated in a catheter balloon in the uterus of healthy women and patients with pelvic inflammatory disease, to determine the optimal time of incubation and to find whether NO level rises after manipulation in the uterine cavity. We measured nitric oxide levels in air incubated for 2-10 min in a catheter balloon in the uterine cavity in 6 non pregnant women from 22 to 50 years of age with lower abdominal pain and 10 healthy women with regular menstrual cycles. After an incubation time of just 2 min, intrauterine nitric oxide levels were significantly increased in patients with diagnosed pelvic inflammatory disease compared to healthy women. Uterine nitric oxide levels did not rise after manipulation in the uterine cavity. In conclusion, NO gas can be measured directly in the uterine cavity with a fast, simple, well-tolerated and safe method. The levels of nitric oxide are increased in women diagnosed with pelvic inflammatory disease already after an incubation time of 2 min.