Visible and UVA light as a potential means of preventing Escherichia coli biofilm formation in urine and on materials used in urethral catheters.

Catheter-associated urinary tract infections are the most common hospital-acquired infection, for which Escherichia coli is the leading cause. This study investigated the efficacy of 385nm and 420nm light for inactivation of E. coli attached to the silicone matrix of a urinary catheter. Using urine mucin media, inactivation of planktonic bacteria and biofilm formation was monitored using silicone coupons. Continuous irradiance with both 385nm and 420nm wavelengths with starting cell density population 103CFU ml-1 reduced planktonic suspensions of E. coli to below the detection level after 2h and 6h, respectively. Bacterial attachment to silicone was successfully prevented during the same treatment. Inactivation by 385nm and 420nm was found to be dependent on media, cell density and oxygen, with less inhibition on planktonic suspensions when higher starting cell densities were used. In contrast to planktonic suspensions in PBS, continuous irradiance of pre-established biofilms showed a greater reduction in survival compared to urine mucin media after 24h. Enhanced inhibition for 385nm and 420nm light in urine mucin media was associated with increased production of reactive oxygen species. These findings suggest 385nm and 420nm light as a promising antimicrobial technology for the prevention of biofilm formation on urethral catheters.

[Pseudomyxoma peritonei arising from urachal carcinoma].

A 77-year-old man was diagnosed with urachal cyst, but had not been followed-up because of his severe dementia. He visited our department complaining of mucinuria. Ultrasonography showed a tumor with heterogeneous contents above the bladder. Mucinousfluid was found by cystoscopic examination. After the fluid was removed, a non-papillary tumor with mucin secretion was identified at the dome of the bladder. Computed tomography revealed that the peritoneal cavity was filled with a mucinous tumor. Peritonitis was suspected because of the existence of free air and abdominal rebound tenderness, and emergent surgery was performed. Bowel perforation was not found. Tumors could not be removed and the tissue biopsy was performed. It was pathologically diagnosed as pseudomyxoma peritonei arising from urachal carcinoma.

Renal pelvic villous adenoma presented with mucusuria: report of a case and literature review.

Villous adenomas of the urinary tract are an uncommon condition, and appear mostly in patients where the disease occurred in the lower urinary tract. In contrast, upper urinary tract villous adenomas are a rare condition. Currently, just three cases of villous adenoma in the renal pelvis have been published. Herein, we present the fourth case of a renal pelvic villous adenoma, along with muconephrosis and mucusuria. A 73-year-old man presented with abdominal discomfort and a palpable abdominal mass. He had a history of bilateral anatrophic nephrolithotomy, 8 years and 6 years earlier. The preoperative radiographic investigation showed severe right hydronephrosis. A right nephrectomy was carried out and the intraoperative finding showed severe perinephric adhesion and a great deal of mucus in the renal pelvis. The pathological examination showed a villous adenoma and urothelial metaplasia in the kidney.

Reflex ImmunoCyt testing for the diagnosis of bladder cancer in patients with atypical urine cytology.

BACKGROUND: ImmunoCyt/uCyt (Scimedx, Denville, NJ, USA) is a well-established urinary marker assay with high sensitivity for the diagnosis of urothelial carcinoma (UC) and can function as a second-level test to arbitrate atypical reads of urine cytology. OBJECTIVE: To determine the utility of uCyt as a reflex test for atypical cytology in patients undergoing a hematuria evaluation or surveillance with a history of UC. DESIGN, SETTING, AND PARTICIPANTS: The uCyt assay was performed as a second-level reflex test on all voided urine cytology tests read as atypical between January 2007 and June 2010 in an academic medical center. Records were retrospectively reviewed. Three hundred twenty-four patients underwent a total of 506 uCyt assays. INTERVENTION: Reflex uCyt assay on atypical urine cytology. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: The uCyt test characteristics include sensitivity, specificity, negative predictive value (NPV), and positive predictive value (PPV). RESULTS AND LIMITATIONS: Reflex uCyt was performed on 506 atypical voided urine samples that were followed by cystoscopy within 90 d. Reflex uCyt with a history of UC showed a sensitivity of 73%, a specificity of 49%, and an NPV of 80%. In those with a history of low-grade UC, reflex uCyt had a sensitivity of 75%, a specificity of 50%, and an NPV of 82%, while in those with a history of high-grade UC, it had a sensitivity of 74%, a specificity of 44%, and an NPV of 79%. Without prior history of UC, reflex uCyt had a sensitivity of 85%, a specificity of 59%, and an NPV of 94%. This study's limitations include its retrospective design and interobserver variability inherent to cystoscopy, which was used as the reference test. CONCLUSIONS: When used as a reflex test on atypical urine cytology, negative uCyt may predict a negative cystoscopy in select patients and modulate the urgency and further work-up in those with no prior history or low-grade disease.

Analysis of the effect of surgical lung biopsy on serum KL-6 Levels in patients with interstitial pneumonia: surgical lung biopsy does not elevate serum KL-6 levels.

OBJECTIVE: It is well known that the serum level of KL-6 can be an indicator of disease activity in patients with interstitial pneumonia (IP). However, surgical lung biopsy is often required for the diagnosis of IP, although this can result in IP exacerbation. METHODS: The effect of surgical lung biopsy on the serum level of KL-6 in patients with IP was analyzed. Thirty-two cases of IP were examined in this study. There were no cases showing exacerbation of IP. RESULTS: The serum level of KL-6 demonstrated 1067+/-550 U/ml (mean+/-SD) before lung biopsy, 991+/-471 U/ml a day, 824+/-377 U/ml 4 days and 826+/-384 U/ml 7 days after lung biopsy. The serum KL-6 levels on the 1st, 4th, 7th day after the lung biopsy were significantly lower than that before the lung biopsy (P<0.05, P<0.01 and P<0.01, respectively). The percent decrease of the serum KL-6 levels on the 4th day (the lowest level) was dependent on the urine volume, and the analysis of the urinary levels of KL-6 showed a transient increase in urinary KL-6 excretion, suggesting that the decrease in serum KL-6 levels associated with surgical lung biopsy may be caused by this increase in urinary KL-6 excretion. CONCLUSION: Surgical lung biopsy of patients with IP has little effect on the increase in serum KL-6 levels. An elevation of serum KL-6 after surgical lung biopsy may indicate exacerbation of IP.

Performance of urine test in patients monitored for recurrence of bladder cancer: a multicenter study in the United States.

PURPOSE: We assessed the performance of the ImmunoCyt immunocytochemical test for detecting bladder cancer recurrence in patients with prior superficial bladder cancers compared with cystoscopic and histological findings. MATERIALS AND METHODS: A total of 341 patients with a history of bladder cancer undergoing monitoring were evaluated at 4 sites. The results of cytology and/or ImmunoCyt were analyzed for sensitivity and specificity compared with biopsy confirmed cancer. RESULTS: The overall sensitivity of cytology alone, ImmunoCyt alone and the 2 methods combined was 23%, 81% and 81%, respectively. The specificity of cytology alone, ImmunoCyt alone and of the 2 methods combined was 93%, 75% and 73%, respectively. The immunocytochemical test was more sensitive than cytology for detecting grades 1 and 2, and stages Ta, T1, and T2 urothelial carcinoma, and it was equally sensitive for detecting grade 3 cancers and carcinoma in situ (CIS). The sensitivity of the combined tests for grades 1 to 3/CIS was 79%, 90% and 82%, while for stages Ta, T1, T2+ and CIS it was 83%, 75%, 100% and 100%, respectively. The overall positive and negative predictive values of the combined tests were 37% and 95%, respectively. Importantly the immunocytochemical test could detect 71% of small (less than 1 cm) tumors. CONCLUSIONS: ImmunoCyt is a sensitive test for detecting bladder cancer. Because of its high sensitivity for detecting small tumors, even those of low histological grade, and its high negative predictive value, this test may have a role in decreasing the frequency of cystoscopic examinations for monitoring patients with low risk bladder cancer.

The bladder does not appear to have a dynamic secreted continuous mucous gel layer.

PURPOSE: We determined whether the nature of any protective barrier in the bladder is composed of a secreted mucous gel layer. MATERIALS AND METHODS: We collected 24-hour urine samples for analysis from 8 healthy 22 to 49-year-old volunteers and 5, 19 to 59-year-old patients treated with bladder reconstruction, in addition to scrapings from 100 freshly slaughtered pig bladders. Samples were subjected to homogenization, dialysis, freeze-drying, papain digestion, gel chromatography, equilibrium density gradient centrifugation, periodic acid-Schiff assay and amino acid analysis. Normal human bladder, pig bladder, normal ileum and transposed intestinal segments were studied for the presence of a mucous layer using a new method of histological analysis. RESULTS: Mucin content in normal urine is 2.7 mg/24 hours, meaning that less than 0.6% of nondialyzable material in normal urine is mucin. The mucin content of urine from reconstructed bladders amounted to 86 mg/24 hours (5.2% of nondialyzable material). We observed that glycosaminoglycans accounted for 41% of the peak total elution volume of PAS positive material in normal urine. Mucin estimation in urine can be grossly overestimated if contaminating glycoconjugates are not removed. Biochemical analysis of material scraped off the pig bladder surface demonstrated that the maximum thickness of a continuous layer that could be achieved was 13.6 mum. While we could visualize an obvious mucous layer on control ileal samples and biopsies of transposed ileal segments from patients with bladder reconstruction, we were unable to note a distinct, measurable mucous layer lining the bladder surface in humans or pigs. CONCLUSIONS: Mucin levels in normal human and pig urine would be enough for slow turnover of a thin barrier but the large increase in mucin in the urine of patients with transposed intestinal segments demonstrates that any layer in normal bladder is much different than that lining the transposed intestinal segment. The most likely constituents of this barrier are membrane bound rather than secreted mucins along with the proteoglycan components of the glycocalix.

ImmunoCyt/uCyt+ improves the sensitivity of urine cytology in patients followed for urothelial carcinoma.

ImmunoCyt/uCyt is a fluorescent test combining three monoclonal antibodies. In this study, it has been tested as a complement to cytology in the detection of urothelial carcinoma in urine. It has been performed simultaneously with standard cytology and cystoscopy on 870 urine analyses from one hospital. In 136 cases, one or more bladder tumors were found. Overall sensitivity of cytology, ImmunoCyt/uCyt and combined analyses reached 29, 74 and 84%, respectively, and overall specificity was 98, 62 and 61%. The negative predictive value of cytology, ImmunoCyt/uCyt and both analyses was 88, 93 and 95%, respectively, and the positive predictive value was 70, 26 and 29%. The sensitivity of cytology for low malignant potential neoplasms, low- and high-grade papillary carcinomas was 6, 18 and 53%, while it reached 71, 79 and 93% when combined with ImmunoCyt/uCyt. The sensitivity of cytology for stages Ta, T1, T2 and over and Tis tumors was 12, 67, 47 and 50%, while it reached 78, 83, 79 and 100% when combined with ImmunoCyt/uCyt. In the absence of tumor on cystoscopy but with positive ImmunoCyt/uCyt, 18% of patients developed a tumor, 2-6 months later. Of the 109 cases diagnosed as suspicious for malignancy by cytology, a tumor was present in 30 cases and ImmunoCyt/uCyt was positive in 22 (73%) of them. In conclusion, ImmunoCyt/uCyt may be used to postpone cystoscopies in patients followed for bladder cancer and may help to save cytologist and pathologist screening time.

[ImmunoCyt--a new urine test in diagnosis of bladder cancer]. / Urinzytologie beim Harnblasenkarzinom. Diagnostische Wertigkeit eines neuen immunologischen Fluoreszenztestes.

ImmunoCyt is a new immunocytologic fluorescence test promising a higher diagnostic sensitivity, esp. for TaG1 carcinomas. The aim of the study was to evaluate the sensitivity of the test in diagnosis of bladder cancer as compared to both urinary cytology and histopathology. A total of 121 spontaneous urine samples of 92 patients (age range 28 to 86, mean 62.5 years) was examined. 41 of the samples were of patients suspicious of transitional cell carcinoma, 46 of patients in whom symptoms were suggestive of tumor recurrence, and 34 of patients who were part of a follow-up protocol. Cystoscopy was performed in all patients. The ImmunoCyt-test was carried out according to the manufacturers protocol using 3 fluorescent monoclonal antibodies. A total of 113 specimens could be evaluated. In 87 cystoscopy and/or histology was negative (control group). There was histologic evidence of 7 pTaG1, 4 pTaG2, 8 pT1G2/G3, and 7 pT2G2/G3 bladder cancers. As for ImmunoCyt and cytology specificity was 83.9% and 91.9%, resp. The combined specificity was 81.6%. Sensitivity amounted to 38.5% and 34.6%, resp., the combined sensitivity to 53.8%. Sensitivity for TaG1 carcinomas was 14.3% each, for TaG2 carcinomas 25% and 50%, for T1G2/G3 carcinomas 37.5% each, and for T2G2/G3 carcinomas 71.4% and 42.9%, resp. In our study the ImmunoCyt test did not show the expected increase in the detection of TaG1 bladder cancers. Because of false-positive results the test should only be used in combination with urinary cytology which reveals a higher specificity. In conclusion the ImmunoCyt test can not replace cystoscopy (with biopsy) in diagnosis and monitoring of bladder cancer.

Immunocyt test improves the diagnostic accuracy of urinary cytology: results of a French multicenter study.

PURPOSE: The limitations of urinary cytology and the invasiveness of cystoscopy generate an increasing interest in noninvasive diagnostic tools for the management of transitional cell carcinoma. We assess the clinical performance of ImmunoCyt (DiagnoCure, Inc., Saint-Foy, Canada) in the detection of bladder cancer in a 10-center French trial. MATERIALS AND METHODS: From October 2000 to April 2001, 694 patients undergoing cystoscopy were prospectively included in the study. Of the patients 458 (66%) had been previously treated for superficial transitional cell carcinoma and 236 (34%) were referred for symptoms suggestive of bladder cancer. All patients underwent ImmunoCyt test and standard urinary cytology from voided urine as well as a complete evaluation including cystoscopy and transurethral resection or biopsy of suspicious lesions. Sensitivity and specificity values of urinary cytology and ImmunoCyt whether or not combined were calculated using cystoscopy as the gold standard and histopathology when available. RESULTS: A total of 85 recurrent and 58 newly diagnosed bladder tumors were diagnosed by cystoscopy and histologicaly confirmed. Overall sensitivity of urinary cytology was 17.9%, 46.3% and 63.8% respectively, for G1, G2 and G3 transitional cell carcinoma, whereas that of ImmunoCyt was 60.7%, 75.6% and 76.8%. Sensitivity of the combined tests was 66.7%, 78% and 87%, respectively. Moreover, 10 of 55 (18.2%) new pT1 and pT2 or greater tumors were diagnosed by ImmunoCyt alone. Specificity of urinary cytology was 94.5%, whereas that of ImmunoCyt was 84.2%. Specificity of the combined tests was 80.7%. Marked variations in urinary cytology sensitivity were observed among the different centers (27.3% to 66.7%), whereas combined assays (urinary cytology and ImmunoCyt) enhanced the overall sensitivity in the 80% range at most centers. CONCLUSIONS: This prospective multicenter series confirmed a marked increase in sensitivity without significant loss in specificity when including ImmunoCyt in standard urinary cytology protocol. This increased sensitivity was observed in high grade lesions (with 100% sensitivity for carcinoma in situ) as well in low grade, low stage tumors.

Mucin 7 and cytokeratin 20 as new diagnostic urinary markers for bladder tumor.

PURPOSE: We determine the sensitivity and specificity of cytokeratin 20 (CK-20) and mucin 7 (MUC7) gene expression in voided urine samples taken from patients with bladder tumor and from control groups to investigate putative, noninvasive urinary markers for bladder tumor detection and monitoring. MATERIALS AND METHODS: Voided urine samples were collected from 50 patients with histologically proven bladder neoplasms (pTaN0M0G1-3 in 19 and pTisN0M0G3-pT4pN1M1G3 in 31), 20 patients with urolithiasis, 20 patients with urinary tract infection, 20 patients with other urological neoplasms and 20 healthy volunteers. Total RNA was extracted from exfoliated cells collected from 200 ml. voided urine. All RNA samples were investigated by a specific CK-20 and MUC7 nested reverse transcriptase polymerase chain reaction. RESULTS: The overall sensitivity of CK-20 gene expression in voided urine samples for the detection of bladder neoplasms was 78%. In contrast, voided urine samples from control patients and healthy volunteers showed a high rate of false-positive CK-20 detection resulting in a low specificity of 36%. The overall sensitivity of the MUC7 test for all bladder tumor cases was 66%. The sensitivity for papillary urothelial neoplasms (pTaN0M0G1-3) was 42% whereas analysis of the carcinoma in situ and invasive bladder cancer group (pTisN0M0G3-pT4pN1M1G3) yielded a sensitivity of 81%. The overall specificity of the MUC7 nested reverse transcriptase polymerase chain reaction method in the control groups was 80%. CONCLUSIONS: A high positive CK-20 detection rate was found not only in voided urine samples from patients with bladder tumor, but also in urine specimens from control groups. Therefore, CK-20 is not a reliable urinary tumor marker for bladder neoplasms. In contrast to CK-20, analysis of MUC7 demonstrated a high sensitivity and high specificity for carcinoma in situ and invasive bladder cancer, thus fulfilling the criteria of a urinary tumor marker.

Reducing mucus production after urinary reconstruction: a prospective randomized trial.

PURPOSE: After transposition into the urinary tract, intestinal segments continue to produce mucus. We determine the effectiveness of muco-regulatory drugs, including N-acetylcysteine, aspirin and ranitidine, in reducing mucus secretion and urine viscosity in patients with transposed segments. MATERIALS AND METHODS: Our trial was a prospective randomized, double-blind placebo controlled crossover study involving 12 patients who underwent ileal conduit and 31 who underwent bladder reconstruction. Each treatment lasted 3 weeks with a 2-week washout. Pretreatment and posttreatment 24-hour urine samples were analyzed for mucin and viscosity after papain digestion, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and periodic acid-Schiff assay. A disease specific questionnaire and SF-36 quality of life survey were completed. RESULTS: According to the questionnaire, mucus production did not decrease with time in 67% of patients. Mucin comprised 3% of the total nondialyzable material in urine (65 mg./24-hour for ileal conduit and 60 mg./24-hour for bladder reconstruction). Analysis of questionnaires and laboratory results failed to demonstrate any benefit of taking muco-regulatory agents compared with placebo. CONCLUSIONS: The use of N-acetylcysteine, aspirin and ranitidine did not result in a reduction in mucin production, urine viscosity or improvement in quality of life.

Aspirin changes the secretion rate and amino acid composition of human small intestinal mucin in subjects with ileal conduits.

The effect of aspirin on the rate of secretion and amino acid composition of human ileal mucin was studied, using subjects with ileal conduits as a model system in which mucin secreted from the ileal conduit tissue is flushed out in the urine and can be measured and analysed. Aspirin (600 mg per day, administered orally) increased the daily mucin output by 37-104% in subjects by days 3 or 4, but thereafter the mucin output declined to below the baseline level by day 10. Mucin samples, purified from the ileal conduit urine during the control period and during aspirin administration, were compared. There were no discernible changes in the degree of polymerisation or the density, but during aspirin administration the amino acid composition was significantly changed, and in particular threonine and proline were enriched. One possible explanation, consistent with the compositional analyses, is that the N- and C-terminal regions of the mucin subunits have been cleaved off and lost during aspirin administration. The observed changes in mucin secretion may have implications for the mechanism of the toxic effects of aspirin on the small intestine by altering the barrier properties of the mucus layer.

Structures of mucin-type sugar chains on human erythropoietins purified from urine and the culture medium of recombinant Chinese hamster ovary cells.

Less is known about the mucin-type sugar chains attached to human erythropoietin as compared with N-linked sugar chains which structures and function have been well studied. In this study, we purified urinary human erythropoietin from three independent groups of aplastic anemia patients, and analyzed the structures of mucin-type sugar chains as well as that obtained from recombinant human erythropoietin produced by Chinese hamster ovary cells. Unlike the N-linked sugar chains, the mucin-type sugar chains are totally different between the urinary and the recombinant erythropoietins. All of the three independent samples of urinary erythropoietin contained oligosaccharides with the structures of +/- Neu5Ac alpha 2-->6GalNAc, while recombinant human erythropoietin contained those with the structures of Neu5Ac alpha 2-->3Gal beta 1-->3(Neu5Ac alpha 2-->6)GalNAc, Neu5Ac alpha 2-->3Gal beta 1-->3GalNAc, and Gal beta 1-->3(Neu5Ac alpha 2-->6)GalNAc.

Topographical investigations of human ovarian-carcinoma polymorphic epithelial mucin by scanning tunnelling microscopy.

Human polymorphic epithelial mucin is a high-molecular-mass glycoprotein that associates to provide protection to the epithelial-cell surface and may afford the malignant cell a selective advantage for growth. The scanning-tunnelling-microscopy micrographs obtained in the present study identify the purified human ovarian-carcinoma polymorphic epithelial mucin glycoproteins as rod-shaped molecules of mixed length. The dimensions of the individual molecules range from 25 to 45 nm in length and are 3-4 nm in width. The images further suggest that lateral association of the rods occurs.

Immunological and structural features of the protein core of human polymorphic epithelial mucin.

The protein core of high mol. wt polymorphic epithelial mucin (PEM--approximately 400 kDa glycoprotein) which is associated with breast carcinomas, consists of a repeating 20 amino acid peptide motif [Gendler et al. (1988) J. biol. Chem. 263, 12,820-12,823]. Monoclonal antibodies C595 (anti-urinary mucin) and NCRC-11 (anti-breast carcinoma cells), and other antibodies against human milk fat globule membranes, were found to recognize determinants present within this 20 amino acid peptide. A model of the peptide was developed based on hydropathicity and structure prediction calculations and these indicated that the repeated structure is dominated by a hydrophilic domain of seven amino acids, extending into two flanking beta turns. NMR analysis of the 20 amino acid peptide was undertaken to probe the secondary structure. Epitope mapping experiments involving solid phase synthesis of overlapping heptapeptides in the repeat unit identified the minimum structures for antibody binding as Arg-Pro-Ala-Pro and Arg-Pro-Ala for the C595 and NCRC-11 antibodies, respectively. These determinants were found within the predicted hydrophilic turn region domain of the peptide. The epitopes for six other PEM-reactive monoclonal antibodies were also determined to reside within the predicted hydrophilic turn domain. This evidence is in accord with the disposition of this region of the PEM peptide core being at the exterior of the glycoprotein where it would be accessible to antibody recognition and binding events.

Mucins obtained from patients with enterocutaneous urinary diversions.

The ureteroenterocutaneostomy is a surgical procedure for urinary diversion by which an intestinal segment is used as a conduit or a reservoir for urine. Histological studies on colonic segments have shown a well preserved mucosal surface with numerous goblet cells and urine from these patients contains a mucus-like sediment. The present study demonstrates that such gels are composed of mucins occurring predominantly as a glycoprotein complex which is 'insoluble' in 6 M guanidinium chloride. The complex was brought into solution by reduction of disulphide bonds and the fragments (subunits) obtained behaved as typical mucins when subjected to density-gradient centrifugation in CsCl/4 M guanidinium chloride. High-Mr glycopeptides obtained after trypsin digestion of subunits expressed the ABH and Lewis antigens, in accordance with the patients' blood-group phenotype and with the regional distribution known from previous immunohistochemical studies on normal colonic mucosa. A heterogeneous population of mucin glycopeptides was revealed by using high-performance ion-exchange chromatography and the major carbohydrate-containing peak did not coincide with that expressing the blood-group activity.