Urothelial Atypia Research Articles

Case Series of Thermal Cautery Artifact in Urine Cytology: An Uncommon Finding but a Potential Pitfall.

Therapeutic modalities used in bladder lesions/neoplasms (electrocautery, fulguration, and laser) could produce morphologic alterations leading to diagnostic challenges. While often seen on histology, they can present in urine cytology as atypia or even raise the concern for malignancy. Herein, we present clinicopathologic findings of four patients with cautery artifact in urine cytology. The authors' lists and a computerized search of urine cytology with features of thermal cautery artifact were collated. Slides and clinicopathologic variables (prior specimens, concurrent and follow-up surgical specimens, age, sex, manner of urine collection, cystoscopy, and treatment procedure during cystoscopy) were reviewed. Four cases were identified. Procedures performed during cystoscopy included laser therapy (n = 2), postbiopsy cauterization (n = 1), and fulguration (n = 1). All cases showed spindled cells with delicate bipolar cytoplasm, "cigar-shaped" nuclei, smooth to slightly irregular nuclear membranes, and arranged singly and loosely cohesive to lamellar stacks. Few atypical well-preserved, undistorted cells were identified in 2/4. The concurrent biopsies showed superficial cellular fragments with thermal artifact (n = 1) and rare atypical cells (n = 2). Follow-up biopsies in two cases had high-grade urothelial carcinoma both of which were diagnosed as atypical urothelial cells on cytology. Thermal cautery artifact in urine cytology is rare and can be seen following laser, electrocautery, and fulguration. The spindling artifact can render some degree of difficulty in assessing for the presence of urothelial atypia both in cytology and biopsy. Familiarity with these cytologic features, knowledge of the treatment procedure, attention to the cytologic findings of well-preserved cells and repeat sampling in cases with atypical urothelial cells in patients with history of malignancy is essential.

Next-generation sequencing has diagnostic utility in challenging small/flat urothelial lesions

Small/flat urothelial lesions are challenging and currently available ancillary immunohistochemistry testing often cannot reliably distinguish between reactive lesions and urothelial carcinoma (UCa). UCa has a characteristic molecular profile, but small/flat urothelial lesions are typically considered too small to perform next generation sequencing (NGS). Herein, we present our institution's experience with utilizing comprehensive DNA-based NGS to evaluate small/flat urothelial lesions (n = 13 cases). NGS was ordered on 7/13 small/flat urothelial lesions initially diagnosed as urothelial atypia, ordered by the pathologist to aid in further diagnosis; the remaining 6/13 cases were diagnosed as urothelial carcinoma in situ (uCIS), ordered by a treating oncologist. The test was considered as adding value if it yielded pathogenic or likely pathogenic alterations previously associated with urothelial carcinoma in the literature. Macroscopic dissection was determined necessary in all cases and obtained either by scraping (7), punch biopsy (5) or scooping (1) of paraffin tissue blocks. In 4/13 cases, tumor content was considered low (<25%); in 2/13 cases, DNA quantity yield was considered below optimal (<250 ng); all cases met required DNA quantity for testing (>50 ng). Mean target coverage ranged: 498 to 985 (optimal >500 reads). NGS testing identified mutations compatible with urothelial carcinoma in all 7 cases initially diagnosed as atypical; and in one case, the tumor recurred as a lung metastasis. All 6 uCIS had NGS testing results concordant with UCa. In conclusion, despite small sample quantity with low tumor content and DNA concentration yield, NGS testing with appropriate methodology can be considered in the setting of small/flat urothelial lesions to aid in diagnosis or per oncologist request and yield interpretable results.

Large-scale validation study of an improved semiautonomous urine cytology assessment tool: AutoParis-X.

Adopting a computational approach for the assessment of urine cytology specimens has the potential to improve the efficiency, accuracy, and reliability of bladder cancer screening, which has heretofore relied on semisubjective manual assessment methods. As rigorous, quantitative criteria and guidelines have been introduced for improving screening practices (e.g., The Paris System for Reporting Urinary Cytology), algorithms to emulate semiautonomous diagnostic decision-making have lagged behind, in part because of the complex and nuanced nature of urine cytology reporting. In this study, the authors report on the development and large-scale validation ofa deep-learning tool, AutoParis-X, whichcan facilitate rapid, semiautonomous examination of urine cytology specimens. The results of this large-scale, retrospective validation study indicate that AutoParis-X can accurately determine urothelial cell atypia and aggregate a wide variety of cell-related and cluster-related information across a slide to yield an atypia burden score, which correlates closely with overall specimen atypia and is predictive of Paris system diagnostic categories. Importantly, this approach accounts for challenges associated with the assessment of overlapping cell cluster borders, which improve the ability to predict specimen atypia and accurately estimate the nuclear-to-cytoplasm ratio for cells in these clusters. The authors developed a publicly available, open-source, interactive web application that features a simple, easy-to-use display for examining urine cytology whole-slide images and determining the level of atypia in specific cells, flagging the most abnormal cells for pathologist review. The accuracy of AutoParis-X (and other semiautomated digital pathology systems) indicates that these technologies are approaching clinical readiness and necessitates full evaluation of these algorithms in head-to-head clinical trials.

Atypical Category of the Johns Hopkins Template Has Higher Risk of Malignancy than the Paris System but the Paris System Is More Applicable for Suspicious Category

Introduction: The aim of this study was to compare performance of individual categories between the Johns Hopkins template and the Paris system for reporting urinary cytology. Methods: Medical records of patients with bladder biopsy and relevant cytology slides were obtained from archived material. Slides were reclassified according to Johns Hopkins template and the Paris system. Results were compared to histological diagnoses. Results: BD SurePath preparations from 205 cases with biopsy follow-up (118 benign, 5 dysplasia, 23 low, and 59 malignant urothelial carcinoma [UC]) were reviewed. There were 2 inadequate specimens in each system. According to the Johns Hopkins template, there were 96 (46.8%) no urothelial atypia or malignancy, 37 (18%) atypical urothelial cells of uncertain significance (AUC-US), 21 (10.2%) atypical urothelial cells, cannot exclude high-grade urothelial carcinoma (HGUC), 38 (18.5%) HGUC, and 11 (5.4%) low-grade urothelial carcinoma (LGUC). The Paris system categorized 111 (54.1%) negative for high-grade urothelial carcinoma, 29 (14.1%) atypical urothelial cells (AUCs), 25 (12.2%) suspicious for HGUC (SHGUC), 36 (17.6%) HGUC, and 2 (1%) LGUC. The Johns Hopkins template had a sensitivity of 95.6%, specificity of 73.6%, positive predictive value of 61.5%, negative predictive value of 96.3, with an overall diagnostic accuracy of 79.8%. The Paris System had a sensitivity of 93.6%, specificity of 77.9%, positive predictive value of 65.6%, negative predictive value of 96.5, with an overall diagnostic accuracy of 82.8%. The risk of malignancy (ROM) for atypical category (AUC-US/AUC) in the Johns Hopkins template was 43.2%, while it has been 24.1% for the Paris System. The ROM for suspicious category was 47.6% and 68.0%, respectively. There were statistically significant differences between negative and atypical, suspicious, and HGUC categories in each system (p < 0.0001). Conclusions: Discrete negative or benign urine cytology had the same sensitivity and specificity between two systems. Although atypical category was associated with a higher ROM with the Hopkins template, the ROM for the suspicious category yielded better result with the Paris system.

Large-scale longitudinal comparison of urine cytological classification systems reveals potential early adoption of The Paris System criteria.

Urine cytology is used to screen for urothelial carcinoma in patients with hematuria or risk factors (eg, smoking, industrial dye exposure) and is an essential clinical triage and longitudinal monitoring tool for patients with known bladder cancer. However, urine cytology is semisubjective and thus susceptible to issues including specimen quality, interobserver variability, and "hedging" towards equivocal ("atypical") diagnoses. These factors limit the predictive value of urine cytology and increase reliance on invasive procedures (cystoscopy). The Paris System for Reporting Urine Cytology (TPS) was formulated to provide more quantitative/reproducible endpoints with well-defined criteria for urothelial atypia. TPS is often compared to other assessment techniques to justify its adoption. TPS results in decreased use of the atypical category and better reproducibility. Previous reports comparing diagnoses pre- and post-TPS have not considered temporal differences between diagnoses made under prior systems and TPS. By aggregating across time, studies may underestimate the magnitude of differences between assessment methods. We conducted a large-scale longitudinal reassessment of urine cytology using TPS criteria from specimens collected from 2008 to 2018, prior to the mid-2018 adoption of TPS at an academic medical center. Findings indicate that differences in atypical assignment were largest at the start of the period and these differences progressively decreased towards insignificance just prior to TPS implementation. This finding suggests that cytopathologists had begun to utilize the quantitative TPS criteria prior to official adoption, which may more broadly inform adoption strategies, communication, and understanding for evolving classification systems in cytology.

Usefulness of the Urine Methylation Test (Bladder EpiCheck®) in Follow-Up Patients with Non-Muscle Invasive Bladder Cancer and Cytological Diagnosis of Atypical Urothelial Cells-An Institutional Study.

Urothelial bladder cancer is a heterogeneous disease and one of the most common cancers worldwide. Bladder cancer ranges from low-grade tumors that recur and require long-term invasive surveillance to high-grade tumors with high mortality. After the initial contemporary treatment in non-muscle invasive bladder cancer, recurrence and progression rates remain high. Follow-up of these patients involves the use of cystoscopies, cytology, and imaging of the upper urinary tract in selected patients. However, in this context, both cystoscopy and cytology have limitations. In the follow-up of bladder cancer, the finding of urothelial cells with abnormal cytological characteristics is common. The main objective of our study was to evaluate the usefulness of a urine DNA methylation test in patients with urothelial bladder cancer under follow-up and a cytological finding of urothelial cell atypia. In addition, we analyzed the relationship between the urine DNA methylation test, urine cytology, and subsequent cystoscopy study. It was a prospective and descriptive cohort study conducted on patients presenting with non-muscle invasive urothelial carcinoma between 1 January 2018 and 31 May 2022. A voided urine sample and a DNA methylation test was extracted from each patient. A total of 70 patients, 58 male and 12 female, with a median age of 70.03 years were studied. High-grade urothelial carcinoma was the main histopathological diagnosis. Of the cytologies, 41.46% were cataloged as atypical urothelial cells. The DNA methylation test was positive in 17 urine samples, 51 were negative and 2 were invalid. We demonstrated the usefulness of a DNA methylation test in the follow-up of patients diagnosed with urothelial carcinoma. The methylation test also helps to diagnose urothelial cell atypia.

Could double stain for p53/CK20 be a useful diagnostic tool for the appropriate classification of flat urothelial lesions?

BackgroundThe differential diagnosis between flat urothelial lesions [reactive urothelial atypia (RUA), atypia of unknown significance (AUS), urothelial dysplasia (UD) and carcinoma in situ (CIS)] has relevant prognostic and therapeutic implications. This crucial distinction could be very challenging but it is currently performed on hematoxylin and eosin (H&E) slides, with a great amount of partially discordant and/or not conclusive findings of the potential adjunctive role of immunohistochemistry. Herein, we tested double staining (DS) for p53/CK20 to verify if p53(+) cells, CK20(+) cells and double-positive cells (DPCs) are differentially expressed among these lesions and if p53/CK20 could be a useful tool in this diagnostic setting. MethodsWe tested 50, 9, 36 and 29 consecutive and retrospectively enrolled cases of RUA, AUS, UD and CIS, respectively. p53(+) cells, CK20(+) cells and DPCs were evaluated and compared by adopting the appropriate statistic tests (Mann-Whitney U and Kruskal-Wallis tests). ResultsWe found that p53(+) cells (p = 0.000), CK20(+) cells (p = 0.000) and DPCs (p = 0.000) showed statistically significant differences among the different flat urothelial lesions. Besides, when dichotomized, both CIS and RUA are easily differentiable from their histological mimickers adopting all these markers; by contrast, AUS and UD did not reach statistically significant differences able to differentiate them from each other [p53(+) cells, p = 0.123; CK20(+) cells, p = 0.567; DPCs, p = 0.409], except if compared to CIS [AUS VS CIS: p53(+) cells, p = 0.013; CK20(+) cells, p = 0.000; DPCs, p = 0.000; UD vs CIS: p53(+) cells, p = 0.000; CK20(+) cells, p = 0.000; DPCs, p = 0.000]. Conclusionsp53(+) cells, CK20(+) cells and DPCs are differently expressed by flat urothelial lesions and p53/CK20 could be a time- and money-saving tool for the appropriate management of these lesions if applied to a routine scenario.

Paris System Has A Higher Cytohistologic Correlation And Reproducibility Than Traditional Urine Cytology Method

Abstract Introduction/Objective The major limitation of urine cytology is the lack of consensus regarding the terminology and diagnostic criteria that should be used for urothelial atypia. The main goal of the newly proposed Paris System for Reporting Urine Cytology is to concentrate primarily on the detection of high grade urothelial carcinoma while minimizing the detection of low grade lesions. This study aimed to apply the criteria and categories of the Paris system in retrospectively collected urine cytology specimens and assess histologic correlation and reproducibility. Methods Two senior pathologists independently reviewed retrospectively collected urine cytology specimens strictly following the Paris system criteria for categorization. Cytologic diagnosis were compared with previous cytology result and histologic diagnosis. Results A total of 67 patients were included in the study. The mean age is 65.8 years (36-89 years), majority were males (73.1%) and African American (89.6%). Urine cytology using traditional method showed mostly atypical results (58.2%), followed by reactive (26.9%), high-grade urothelial carcinoma (11.9%) and suspicious (3.0%). On the other hand, the Paris system had more negative results (62.7%), followed by atypical (19.4%), high-grade urothelial carcinoma (11.9%) and lastly suspicious (6.0%). All of negative cases (18 out of 18) and majority of HGUC cases (7 out of 8) were concordant between the two cytology methods. Traditional urine cytology method only yielded 71.4% histologic concordance, while 100% Paris system results were concordant with bladder histology. Majority of the atypical cases using traditional method were converted to negative, and a few atypical cases were converted to suspicious and high-grade. All results using Paris system were concordant between two general pathologists. Conclusion Using the Paris system in analyzing urine cytology resulted to higher cytohistologic concordance than traditional method. Majority of atypical cases from traditional method were converted to more definitive categories. The cytopathologic analyses from two general pathologists applying the criteria of Paris system had superior reproducibility. Applying the Paris system, therefore, can significantly improve the performance of urine cytopathology.

Vitamin A Rich Diet Diminishes Early Urothelial Carcinogenesis by Altering Retinoic Acid Signaling.

Urinary bladder cancer is one of the leading malignancies worldwide, with the highest recurrence rates. A diet rich in vitamin A has proven to lower the risk of cancer, yet the molecular mechanisms underlying this effect are unknown. We found that vitamin A decreased urothelial atypia and apoptosis during early bladder carcinogenesis induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN). Vitamin A did not alter urothelial cell desquamation, differentiation, or proliferation rate. Genes like Wnt5a, involved in retinoic acid signaling, and transcription factors Pparg, Ppara, Rxra, and Hoxa5 were downregulated, while Sox9 and Stra6 were upregulated in early urothelial carcinogenesis. When a vitamin A rich diet was provided during BBN treatment, none of these genes was up- or downregulated; only Lrat and Neurod1 were upregulated. The lecithin retinol acyltransferase (LRAT) enzyme that produces all-trans retinyl esters was translocated from the cytoplasm to the nuclei in urothelial cells as a consequence of BBN treatment regardless of vitamin A rich diet. A vitamin A-rich diet altered retinoic acid signaling, decreased atypia and apoptosis of urothelial cells, and consequently diminished early urothelial carcinogenesis.

Immunohistochemical staining patterns of Ki-67 and p53 in florid reactive urothelial atypia and urothelial carcinoma in situ demonstrate significant overlap

Flat urothelial lesions with atypia may pose significant diagnostic challenges. Given frequent increased proliferation rates in florid reactive urothelial atypia and limited studies on the interpretation of p53 stains in the urothelium (following current standard guidelines for correlation with P53 mutation status), we sought to further study the discriminatory value of Ki-67 and p53 for florid reactive urothelial atypia versus urothelial carcinoma in situ (CIS). Bladder specimens diagnosed as reactive urothelial atypia (n=40) and CIS (n=40) were assessed by immunohistochemical staining with antibodies for Ki-67, p53, CD44, and CK20. Immunoreactivity was scored based on percent cells positive for Ki-67 and pattern of reactivity with p53 (aberrant: diffuse strong positive or negative; normal: patchy/wild type). CD44 and CK20 reactivity patterns served as adjunctive internal validation controls for reactive urothelial atypia and CIS, as previously described. In reactive urothelial atypia, Ki-67 ranged from 0% to 90% (mean, 34%±26) with 30 cases (75%) having >10%. In CIS, Ki-67 ranged from 5% to 95% (mean, 50%±25) with 17 cases (43%) having >50%. In all 40 cases (100%) of reactive urothelial atypia, p53 expression had a wild-type pattern. In CIS, aberrant p53 expression was identified in 15 cases (37%): 3 cases (7%) were p53 negative (i.e. null phenotype) and 12 cases (30%) showed strong and diffuse nuclear reactivity (in >85% of cells). The remaining 25 cases (63%) of CIS had a p53 wild-type pattern of expression. Cytoplasmic CK20 immunoreactivity in umbrella cells was seen in 34 cases (85%) of reactive urothelial atypia, and 6 cases (15%) were negative. In addition, 35 cases (88%) of reactive urothelial atypia demonstrated full-thickness CD44 expression, while 5 cases (12%) had expression confined to the basal/parabasal layers of the urothelium. Strong and diffuse CK20 positivity was present in 39 cases (98%) of CIS, and patchy positivity was detected in 1 case (2%). None of the CIS cases overexpressed CD44: 16 cases (40%) showed focal expression in the nonneoplastic basal cell layer; 24 cases (60%) demonstrated no staining. In summary, Ki-67 has poor discriminatory value for reactive urothelial atypia versus CIS and adds little to the classic CK20/CD44 immunophenotype. While p53 sensitivity for CIS is relatively low (30%) and interpretation as either wild type or negative may be challenging in a small subset of cases, strong and diffuse nuclear reactivity was 100% specific in the distinction from florid reactive urothelial atypia in this cohort.

Integrated Fourier Transform Infrared Imaging and Proteomics for Identification of a Candidate Histochemical Biomarker in Bladder Cancer

Histopathological differentiation between severe urocystitis with reactive urothelial atypia and carcinoma in situ (CIS) can be difficult, particularly after a treatment that deliberately induces an inflammatory reaction, such as intravesical instillation of Bacillus Calmette-Guèrin. However, precise grading in bladder cancer is critical for therapeutic decision making and thus requires reliable immunohistochemical biomarkers. Herein, an exemplary potential biomarker in bladder cancer was identified by the novel approach of Fourier transform infrared imaging for label-free tissue annotation of tissue thin sections. Identified regions of interest are collected by laser microdissection to provide homogeneous samples for liquid chromatography-tandem mass spectrometry-based proteomic analysis. This approach afforded label-free spatial classification with a high accuracy and without interobserver variability, along with the molecular resolution of the proteomic analysis. Cystitis and invasive high-grade urothelial carcinoma samples were analyzed. Three candidate biomarkers were identified and verified by immunohistochemistry in a small cohort, including low-grade urothelial carcinoma samples. The best-performing candidate AHNAK2 was further evaluated in a much larger independent verification cohort that also included CIS samples. Reactive urothelial atypia and CIS were distinguishable on the basis of the expression of this newly identified and verified immunohistochemical biomarker, with a sensitivity of 97% and a specificity of 69%. AHNAK2 can differentiate between reactive urothelial atypia in the setting of an acute or chronic cystitis and nonmuscle invasive-type CIS.

CK20 and p53 Immunohistochemical Staining Patterns in Urinary Bladder Specimens With Equivocal Atypia.

- Urinary bladder flat carcinoma in situ (CIS) is a worrisome lesion, requiring aggressive surveillance and treatment. Cytokeratin 20 (CK20) and p53 are common immunohistochemical antibodies used to supplement CIS diagnosis in biopsy samples. However, existing data come primarily from unequivocally benign and malignant specimens. - To correlate these markers in specimens with borderline histology with outcomes. - CK20 and p53 immunohistochemistry was analyzed for staining pattern, classified as CIS pattern (both stains yielding strong labeling of the area of concern), discordant (only 1 stain yielding CIS pattern), indeterminate (1 or both stains yielding partial or equivocal labeling), or benign (both stains yielding a benign pattern). - Specimens with equivocal atypia (n = 69) from 65 patients were studied. There were 9 specimens (13%) that had a CIS staining pattern, 18 (26%) were discordant, 31 (45%) were indeterminate, and 11 (16%) were benign. Of the discordant specimens, 13 labeled for CK20 but not p53, whereas 5 showed the opposite. Most specimens (n = 47; 68%) were obtained from patients with a known history of bladder cancer, of which recurrence developed in 27, with an average interval of 37 months (range, 2-216 months). A subset (n = 22; 34%) had no prior history of bladder cancer, from which only 1 patient with CK20-positive/p53-equivocal staining later developed diagnostic carcinoma. - In our cohort of specimens with equivocal urothelial atypia, very few patients without a prior diagnosis of bladder cancer progressed to diagnostic cancer (1 of 22), suggesting that staining results should be interpreted with caution in de novo atypia. Patients with a known history of bladder cancer had a substantial rate of recurrence, independent of staining pattern.

The role of AMARC and cytokeratin 5/6 in differentiation between carcinoma in situ and reactive urothelial atypia in comparison with previously studied markers

The role of AMARC and cytokeratin 5/6 in differentiation between carcinoma in situ and reactive urothelial atypia in comparison with previously studied markers

Prevalence of atypical urine cytology in patients treated with gemcitabine-cisplatin (GC) for urothelial carcinoma (UC).

363 Background: The prevalence of atypical urothelial cells (AUC) in the general population is estimated to range between 2-23% with known variability between individual pathologists’ use of this diagnosis. At our institution, the AUC rate for all urinary tract cytology (UTC) is 6.1%. Increased rates of urothelial atypia have been described in pts receiving certain systemic and intravesical therapies for UC. This may confound the use of UTC for post-therapy surveillance. Our objective is to describe the prevalence of urothelial atypia in pts treated with GC, as it is currently unknown. Methods: Patients who received at least one cycle of GC for UC at a single institution from 1/1/2007-9/30/2014 were identified. Demographics, clinical characteristics, treatment setting, number of cycles, and specimen type were recorded. Urine cytology reports were reviewed and tabulated. Results: Seventy-four pts treated with GC were identified. Median age at treatment was 65 (range 42-80); 58 (78%) were male. Median number of cycles was 4 (range 1-9). Treatment settings were: 15 (20%) neoadjuvant, 38 (51%) adjuvant, and 21 (28%) metastatic. Ten (14%) pts were previously treated with intravesical therapy (9 BCG, 1 mitomycin). Thirty-six (49%) pts had urine cytology available. Twenty-seven (75%) pts had at least 1 urine cytology within 12 months post-chemotherapy, with 44 total specimens. Nine (25%) pts had cytology performed &gt;1-year post treatment. Cytology specimens within 12 months post-chemotherapy were reviewed. Two out of 44 specimens had AUC (5%), with all other cases negative. The pts with AUC were male, received adjuvant therapy, had prior radical cystectomy with orthotopic neobladder reconstruction, and had no intravesical therapy. Conclusions: The prevalence of AUC in pts treated with GC was similar to the general population. Chemotherapy with GC does not appear to morphologically affect urothelial cells. If atypical UTC is seen in this population, the cause should be investigated, as GC does not appear to influence urothelial changes. Additional study is needed to characterize the impact of systemic therapy on urothelial atypia and to guide the use of urine cytology for post-therapy surveillance.

The application of the Johns Hopkins Hospital Template on urine cytology.

To evaluate the utility of the Johns Hopkins Hospital (JHH) template in detection of high-grade urothelial carcinoma (HGUC). A computerized search of our laboratory information system was performed for all urine cytology cases from 2009 to 2011 processed by the SurePath™. We included only cases with correlating surgical pathology within 6 months after the urinary samples were obtained. The original cytologic diagnoses were reclassified according to the JHH template, and these cytolog ic diagnoses were then correlated with the follow-up surgical pathology diagnoses. A total of 273 urine samples with histopathologic follow-up were identified. The reclassified cytologic diagnoses included negative for urothelial atypia or malignancy (NUAM) 110; atypical urothelial cells of undetermined significance (AUC-US) 83; atypical urothelial cells, cannot exclude high-grade urothelial carcinoma (AUC-H) 49; HGUC 29; and low-grade urothelial carcinoma (LGUC) 2. More than one-half of patients (58%) who had biopsy-confirmed high-grade urothelial lesions had a preceding cytologic diagnosis of AUC-H or HGUC. AUC-H and HGUC are associated with high-grade urothelial lesions in 80% and 90% of the cases and show statistical significance when compared with AUC-US or NUAM (P < 0.05). The JHH template is useful and effective in identifying patients with high-grade urothelial lesions who need to undergo cystoscopy.

Histopathological assessment of inflammation and expression of inflammatory markers in patients with ketamine-induced cystitis.

The aim of the current study was to evaluate the histopathological features of inflammation and the expression levels of inflammatory markers in tissue samples from patients with ketamine-induced cystitis. Bladder biopsy samples for histological analysis were obtained from 23 patients (18 men and 5 women) with a self-reported history of ketamine use and who were treated for cystitis at the Tri-Service General Hospital of Taipei, Taiwan. Immunohistochemical staining for cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), matrix metallopeptidase-9 (MMP-9), mammalian target of rapamycin (mTOR), and phosphorylated 40S ribosomal protein S6 (Phos-S6) was performed. The results revealed urothelial atypia in all patients, and intravascular eosinophil accumulation in 22 patients. Histopathological features included denuded urothelial mucosa, ulceration, collagen deposition, smooth muscle degeneration and vessel proliferation. Tissue samples were immunopositive for all of the inflammation markers, including the urothelium, vessel walls, and smooth muscle. COX-2 staining revealed a significant difference between the inflammatory levels in the urothelium and smooth muscle, and iNOS staining differed significantly between inflammatory levels in smooth muscle (p=0.029). A positive correlation was observed between the percentage of Phos-S6-positive cells and the levels of inflammation in the urothelium. These results add to the descriptive literature on the histopathological aspects of ketamine-induced cystitis, emphasizing the inflammatory nature and a possible role for proteins such as COX-2, iNOS and Phos-S6 in the degree of inflammation.

The clinical presentation and outcome of urothelial atypia on biopsy of the bladder

ObjectivesTo examine the presentation and clinical outcome of patients diagnosed with reactive urothelial atypia (RUA) or urothelial atypia of unknown significance (AUS) on tissue biopsy of the bladder. Methods and materialsWe performed a retrospective cohort study of all patients who were diagnosed with either RUA or AUS on biopsy of the bladder at our institution from November 2000 to May 2008. Excluded from the analysis were patients with a history of or a concurrent diagnosis of urothelial carcinoma. A total of 66 patients were available for final analysis. ResultsThe mean patient age at diagnosis was 63 years (range: 18–89 years). There were 46 men (70%) and 20 women (30%). The indication for cystoscopic examination included lower urinary tract symptoms in 29 (44%), gross hematuria in 17 (26%), microscopic hematuria in 12 (18%), urolithiasis in 2 (3%), and hydronephrosis in 6 (9%) patients. On biopsy, 54 (82%) had RUA and 12 (18%) had AUS. The mean follow-up was 36 months (range: 0–271 months). During this period, 37 (56%) patients underwent at least 1 additional cystoscopy with negative result. None of the 66 (0%) patients developed biopsy-proven urothelial carcinoma. ConclusionUrothelial atypia is common in men older than 60 years and often presents with either hematuria or lower urinary tract symptoms. Both RUA and AUS have a similar benign clinical course. Therefore, after diagnosis, further intervention or surveillance is unnecessary.

Utility of a Triple Antibody Cocktail Intraurothelial Neoplasm-3 (IUN-3-CK20/CD44s/p53) and α-Methylacyl-CoA Racemase (AMACR) in the Distinction of Urothelial Carcinoma In Situ (CIS) and Reactive Urothelial Atypia

Urothelial carcinoma in situ (CIS) is a prognostically and therapeutically significant lesion with considerable morphologic overlap with reactive conditions especially in the setting of prior therapy. Various markers including CK20, CD44s, and p53 have been used as an adjunct in making this distinction; however, the utility of these markers in the posttreatment scenario is not fully established. α-Methylacyl-CoA racemase (AMACR) is a tumor-associated marker that is expressed in a subset of high-grade urothelial carcinomas but has not been studied in CIS. This study was undertaken to evaluate the immunoreactivity of CK20, CD44s, and p53 as a triple antibody cocktail intraurothelial neoplasm-3 (IUN-3) in distinguishing CIS from its mimics and to compare its utility with AMACR in the diagnosis of CIS. A total of 135 specimens (7 benign ureters and 128 bladder biopsies-28 reactive, 33 posttherapy reactive, 43 CIS, 24 CIS posttherapy) were included in this study. Immunostaining for p53 (brown, nuclear), CD44s (brown, membranous), and CK20 (red, cytoplasmic and membranous) was performed as a cocktail, and the staining pattern was further classified as: malignant (full-thickness CK20 and/or full-thickness p53 with CD44s negativity), reactive/benign (CK20 limited to the umbrella cell layer, p53 negative, and CD44s positivity ranging from basal to full thickness), and indeterminate (CK20 and p53 positive but not full thickness and/or CD44s positive). AMACR staining was performed in 50 cases. Cytoplasmic staining for AMACR was graded as negative (absent to weak focal staining [<5% cells]) and positive (≥5%). The "IUN-3 malignant" pattern was observed in 84% of cases of CIS without a history of prior therapy and in 71% of the cases of CIS with a history of prior therapy. Cases with posttherapy reactive atypia showed an "IUN-3 reactive" pattern in 84% cases and "IUN-3 indeterminate" pattern in 16% of the cases; the IUN-3 malignant pattern was not identified in any of the cases. Benign and reactive urothelium (with and without a history of therapy) showed an IUN-3 reactive pattern and negative AMACR staining in all the cases (100%). AMACR positivity was observed in 78% of nontreated CIS cases and 50% of CIS posttherapy cases. In these cases, the IUN-3 cocktail showed an IUN-3 malignant pattern in 83% of untreated CIS cases and 88% of CIS posttherapy cases. AMACR positivity is a potentially useful marker of CIS. However, the IUN-3 malignant pattern is a more reliable indicator of CIS compared with AMACR, especially in the posttreatment setting. The simultaneous evaluation of all 3 markers (p53, CD44s, and CK20) in a single slide in the form of a cocktail is advantageous, especially in small biopsy specimens.

The role of immunohistochemistry in the diagnosis of flat urothelial lesions: a study using CK20, CK5/6, P53, Cd138, and Her2/Neu

Although differentiating reactive urothelial atypia from urothelial carcinoma in situ (CIS) relies primarily on histologic evaluation, confirming the morphologic impression using immunohistochemistry (IHC) has been increasingly used in routine clinical practice. The aims of this study are to confirm the utility of commonly used markers (CK20, P53) and to test the performance of CK5/6, CD138, and Her2/Neu in the diagnosis of CIS. Using a tissue microarray comprising 52 cases of normal/reactive urothelium and 45 cases of CIS, the IHC evaluation of 5 markers was undertaken. Although the individual specificity of CK20, P53, and Her2/Neu was high (94%, 90%, and 93%, respectively), their sensitivity for CIS detection was lower, with the most sensitive marker being HER2/Neu (63%). Whereas 78% of CIS shows positivity of at least 2 of those 3 markers, only 1 case of reactive urothelium shows positivity for 2 of those 3 markers. The discriminatory performance of CK5/6 and CD138 was poor. In conclusion, HER2/Neu can be added to a panel of CK20 and P53 to help differentiate reactive atypia from CIS in difficult cases. Positive staining for at least 2 of the 3 antibodies (CK20, P53, and HER2/Neu) is strongly associated with CIS. However, the histologic findings should be a primary determinant in the diagnosis of flat urothelial lesions, with IHC playing a supportive confirmatory role.

SIGNIFICANCE OF FLAT EPITHELIAL LESIONS OF THE URINARY BLADDER IN EARLY DETECTION OF BLADDER CANCER: HISTOPATHOLOGICAL AND IMMUNOHISTOCHEMICAL STUDY

Background: Urothelial carcinoma of the urinary bladder is the second most common malignancy of the genitourinary system, after prostate cancer. The flat lesions are classified into flat lesions without cytological atypia (Flat hyperplasia) and flat lesions with cytological atypia; reactive atypia, atypia of unknown significance, urothelial dysplasia and urothelial carcinoma in situ). The immunohistochemical markers such as cytokeratin 20,CD 44 and Ki 67 may be useful in differentiating CIS from reactive changes in difficult biopsy cases. Aim: This study was conducted to determine the role of cytokeratin 20, CD 44 and Ki 67 in the diagnosis of dysplasia and other flat lesions of the urinary bladder. Methods: Sixty representative cases for flat urothelial lesions of urinary bladder were examined immunohistochemically using antibodies against CD44, cytokeratin 20 and Ki-67. Results: CD 44 were positive in 100%of cases of reactive urothelial atypia, but flat Hyperplasia and CIS cases were negative. However CD 44 were positive in 42.9% of atypia of unknown significance and 14.3% of Dysplasia. CK 20 were positive in (95.5%) of Dysplasia. Non of Flat Hyperplasia were positive for CK 20. However CK 20 was positive in 88.2% of CIS, 57.1 % of atypia of unknown significance and 7.1% of reactive urothelial atypia. ki 67 index were high in 82.4% of cases of CIS, non of flat hyperplasia expressed high ki 67 index. However ki 67 index were high in 57.1% of atypia of unknown significance, 52.4% of dysplasia and 14.3% of reactive urothelial atypia. Conclusion. CD 44 is the most constant marker in the reactive urothelium. CK20 is the most commonly detected marker in the dysplastic urothelium. Nevertheless, the most accurate is application of an immunohistochemical panel composed of the three antibodies (standard CD 44, CK20 and Ki-67) together with correlation with morphology. This is very useful for confirming the presence of dysplastic changes in the urothelium and differentiating reactive urothelium from dysplastic urothelium (dysplasia/CIS). Keywords: Flat epithelial lesions, CD44, cytokeratin ck20, ki67, immunohistochemistry.