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Urinary Tract Cytology

INTRODUCTION

The urinary tract is composed of the kidneys, the ureters, urinary bladder and the urethra. The basal portion of the urinary bladder contains the trigone, a triangular area with the apex directed forward. The urethra leaves the bladder at the apex of the trigone. The female urethra has a short-course opening into the vestibule behind the clitoris. The male urethra runs across the prostate and the penis.

The renal pelvis, the ureters, bladder and the urethra are lined by a highly specialized and unique epithelium "the urothelium" also known as transitional epithelium. The urothelium is composed of a basal layer of cuboidal cells resting on the basement membrane, intermediate layers, and a superficial layer of cells referred to as umbrella cells that are fairly large and can have multiple nuclei. The urothelium has the unique characteristic of forming a barrier between the toxic urine, preventing leakage, and has the capacity to contract and expand relative to normal physiologic function of the bladder.

The trigone in the bladder in 50% of adult women and in some proportion of the male bladder, can be lined by squamous epithelium. Areas of mucus-producing epithelium can also be noted.

Specimen Collection
Voided urine: is the most readily obtained urinary specimen. Voided urine may be:
  • Randomly voided
  • Voided after hydration
  • Collected over 24 hours
  • Collected from intestinal conduits
The most useful urine specimen for the detection of neoplastic cells is random voided urine, at least 3 specimens collected over 2 weeks. Intestinal conduit specimens must be as fresh as possible.

Washings
Washings can be obtained from the bladder, ureters, renal pelvis, blind-ended urethras, neo-bladders and intestinal conduits. The procedure involves lavage with 50ml of saline before bladder manipulation or biopsy.

Brushings
Brushings can be obtained from the bladder or from urothelial lined structures. After sampling, the brush can be directly applied to the slide (direct smears) or rinsed in Cytolyt® for ThinPrep®preparation.

Washings and brushings tend to be more cellular than voided urine samples, although diagnostic elements may only appear in voided urine.

Normal Elements in Urinary Cytology
Urinary cytologyis usually requested in patients with unexplained hematuria, irritation voiding symptom, patients suspected of bladder cancer, and in patients monitored for recurrent bladder cancer.

Normal Urothelial Cells
Normal urothelial cells are the predominant cellular element varying greatly in numbers, sizes, and shapes. These cells may appear as mononuclear cells that may be columnar, cuboidal, and parabasal-like or polyhydral. The umbrella cells are large, may be multinucleated and may show one flat or convex surface corresponding to the lumen of the bladder. The cytoplasm may take on variable staining characteristics ranging from blue to grey and occasionally shows fine vacuolization. The nuclei are centrally located with finely granular chromatin and small nucleoli. The nuclei may vary in shape from round to ovoid. Bi-nucleation is common. Giant umbrella cells are usually seen in specimens from the ureter or renal pelvis. Columnar shaped cells are usually encountered in bladder washings and are believed to be urothelial in origin. These cells show thin cytoplasmic tails and may occur singly or in clusters.

Urothelial clusters and papillary fragments.
Approximately 20% of normal voided urine samples may contain urothelial clusters. This finding is considerably enhanced in bladder washings, catheterized urines, and brushings due to the propensity of urothelium to exfoliate. The interpretation of papillary urothelial neoplasm should be made with caution and needs to be correlated with other findings. It needs to be kept in mind that urothelial clusters are frequent findings in instrumented catheterized urines, bladder washings, and ureteral and renal pelvis washings and brushings. The cells in the clusters show nuclei that may appear hyperchromatic or pale and may contain one or more nucleoli. Occasionally one may see a cap of umbrella cells on one side of these clusters. The presence of these clusters may be a diagnostic pitfall, and a diagnosis of cancer should not be made in the absence of altered nuclear/cytoplasmic ratio and chromatin texture.

Squamous and Glandular Cells
Squamous cells may be present as contaminants from external genitalia or may appear as a component of normal bladder being shed from the trigone. The numbers may vary. Secretory columnar cells and cells from intestinal metaplasia may be seen as part of normal metaplastic change or as a component of cystitis glandularis.

Miscellaneous Findings
Urinary samples are normally free of blood and inflammation. Their presence may indicate trauma, infection or tumor. The inflammation is reduced on the ThinPrep and the blood cells may appear lysed in the background. The presence of eosinophils may be associated with drug induced interstitial cystitis.

Renal Tubular Cells and Renal Casts
Hyaline and granular casts may be seen even in patients without overt evidence of renal pathology. Hyaline casts are composed of amorphous, eosinophilic proteinaceous material while the granular casts are composed of degenerated renal tubular cells. Renal tubular cells are small columnar cells occurring in sheets or as single cells.

Prostatic and Seminal Vesicle Cells
Prostatic cells occur in small sheets of cuboidal cells with rounded nuclei and occasional small nucleoli. The seminal vesicle cells may appear markedly atypical and contain golden brown pigment in the cytoplasm. One may also see sperm and corpora amylacea in this setting. Corpora amylacea are laminated non-calcified bodies.

Globular or Hyaline Inclusion Bodies
Red or blue globular inclusions are seen frequently in urothelial cells. These represent degenerative changes and are not viral in origin.

Crystals may be present in urine samples as distractions. Specific identification of these crystals is not required for urine cytology, and is a component of routine urinalysis.

Ileal Conduit Urine
Ileal conduit urine is usually obtained for surveillance. Since the colonic mucosa is exposed to a hostile and toxic environment, degenerative changes predominate. The cells resemble macrophages, there is karyorrhexis, pyknosis, and abundant red inclusions in the cytoplasm. Cytoplasmic debris and bacteria are seen in the background. Detection of malignancy may be challenging in this setting. Diagnosis should be based on cells with characteristic features of malignancy.

Urinary Cytology in Benign Conditions
Infection
Inflammatory conditions such as bacterial infection may be associated with increased numbers of neutrophils, reactive changes in urothelial cells such as nuclear enlargement, prominent nucleoli, and even background bacteria. Necrosis and degeneration may occur.

Granulomatous Inflammation
It is extremely rare to see well defined granulomas. Granulomas may be seen in the setting of tuberculosis, BCG therapy and after surgery. Granulomas may show palisaded, elongated histiocytes with or without giant cells and necrosis.

Fungal Infections
Candida is the most common fungal infection and is seen as pseudo-hyphae or spores. It may occur as a contaminant from the vagina in female patients. However, the presence of fungal organisms in cases of renal transplant or immunosuppression denotes true infection, and requires appropriate management.

Viral Infection
Common viral infections seen in urinary tract cytology are Cytomegalovirus, herpes virus and polyoma virus occurring in a setting of immunosuppression due to AIDS, renal transplant or cancer therapy.
  • CMV: is characterized by large basophilic intra-nuclear inclusions surrounded by a halo and margination of chromatin. This is accompanied by cytomegaly and cytoplasmic inclusions.
  • Herpes virus: results in multinucleated cells, with molded ground glass nuclei and occasional eosinophilic intranuclear inclusion.
  • Human Polyoma Virus: Infection with the polyoma virus is acquired early in life. Activation occurs in the setting of immunosupression due to transplantion, chemotherapy, AIDS, diabetes, etc. The infected cells vary in size. The inclusions of polyoma virus are largely basophilic opaque intranuclear inclusions that fill the nucleus. Other nuclei may show a reticular pattern of chromatin due to leaching of viral particles. Polyoma virus changes can be mistaken for urothelial cancer and therefore these cells are called "decoy cells." Polyoma virus may coexist with cancer.
  • Parasitic Infections: Schistosome eggs may be seen, especially in endemic areas or in the immigrant population of the United States, and is associated with squamous metaplasia of the urothelium (leukoplakia) and squamous cell carcinoma.


  • Reactive/Reparative Changes
  • Urinary Lithiasis: The presence of stones can result in large urothelial clusters and papillary fragments with marked variation in the shape and size of the urothelial cells and hyperchromasia. Sometimes the atypia associated with lithiasis is so severe that further work-up to exclude malignancy is necessary. Cytologically, there is nuclear enlargement, pleomorphism, increased N/C ratio, coarse dense chromatin, prominent nucleoli, occasional mitoses, degeneration and necrosis.
  • Chemotherapy Effect: Cyclophosphamide produces marked cytologic atypia due to the concentration and excretion of the drug in the urine. This causes marked cellular enlargement with enlarged eccentric nuclei, irregular nuclear membranes, hyperchromasia and coarse chromatin. Large and irregular nucleoli may be seen. Multinucleation is common. The cytoplasm is vacuolated. There is nuclear pyknosis and karyorrhexis. Busulfan may cause some cytologic atypia and can mimic high grade urothelial cancer. Intravesical agents such as thiotepa, mitomycin, and BCG have relatively mild effects on urothelial cells and can result in multinucleation, smooth nuclear contours, and multiple small nucleoli. The cytoplasm shows vacuoles.
  • Radiation Changes: Radiation changes are associated with cytomegaly with a low N/C ratio. Multinucleation and macronucleoli can be seen. There is nuclear and cytoplasmic vacuolization. Cytoplasmic eosinophilia and polychromasia are characteristic. This may be accompanied by inflammatory cells and debris in the background.
  • Laser Therapy or Cautery Effect: These treatments can make the cells appear spindled and elongated.

Urothelial Tumors
Urothelial tumors may be papillary or non-papillary and can arise from the urothelium in the bladder, ureters or the renal pelvices. Regardless of the mechanisms of origin, all have common characteristics. It is not uncommon to have patients in whom all areas of the urinary tract are involved by synchronous or metachronous tumors. The current WHO/ISUP classification classifies urothelial neoplasms into:
  • Papilloma
  • Papillary urothelial neoplasm of low malignant potential (PUNLMP)
  • Low grade urothelial carcinoma
  • High grade urothelial carcinoma

High Grade Urothelial Carcinomas
Cytologically, high grade urothelial carcinomas are relatively easy to diagnose with a high degree of sensitivity and specificity due to the presence of anaplastic cells. One may see cellular preparations with abundant atypical urothelial clusters and single malignant cells in the background. Occasionally the samples may be sparse. The cells have high N/C ratios, with marked pleomorphism. The nuclei are often eccentric with hyperchromatic coarse chromatin and large irregular nucleoli. The nuclear membrane is thickened and occasional mitoses may be seen. The cytoplasm is poorly demarcated and cyanophilic.

Low Grade Urothelial Carcinoma
Overall, low grade urothelial carcinoma has a low diagnostic sensitivity and specificity. These tumors are cytologically characterized by increased cellularity, and the presence of an increased number of urothelial clusters which may or may not be papillary. The cells in these clusters have high N/C ratios with nuclei bulging out of the cytoplasm. Nuclei are irregular and may appear to have notches or grooves. The chromatin is granular and evenly distributed. Nucleoli are indistinct or absent.

Papillary urothelial neoplasm of low malignant potential (PUNLMP)
There are no reliable cytologic criteria for diagnosing these tumors. There may be abundant urothelial clusters and papillary fronds with cytologic features indistinguishable from low grade urothelial cancer, instrumentation or stone induced artifact.

Carcinoma in Situ(CIS)
CIS is a flat lesion and is the precursor to most invasive urothelial cancers. Cytology plays an important role in the detection of urothelial carcinoma in situ, since these lesions may be multifocal and not visualized on cystoscopy. Essentially the cytologic features are those of a high grade TCC where the cells appear to be dispersed or form small clusters. The cells are large with high N/C ratios and eccentric hyperchromatic nuclei with prominent nucleoli. Invasion can be suspected when tumor cells are associated with diathesis, but it is to be emphasized that an unequivocal diagnosis of CIS versus invasion cannot be rendered by cytology alone.

Other Types Of Malignancies
Squamous Carcinoma Squamous carcinoma is relatively rare in the United States. It has strong association with Schistosomiasis. In this setting, the tumors are usually well-differentiated. Oftentimes, urothelial carcinoma can have focal squamous differentiation with one observing only malignant squamous cells in urine cytology.

One may see anucleated squamous cells associated with nucleated and markedly atypical squamous cells. The cells may show marked nuclear pleomorphism with dense hyperchromatic nuclei and macronucleoli. The cytoplasm may be abundant forming spindle and tadpole shapes. Intercellular bridges and keratohyaline granules may be appreciated along with keratin pearls. The non-keratinized squamous carcinomas can be more difficult to separate from urotheolial carcinoma.

Adenocarcinoma
Primary adenocarcinomas of the bladder are rare, constituting less than 2% of all bladder cancers. The cells have typical features of malignancy with large eccentrically placed nuclei with open chromatin and prominent nucleoli. The cytoplasm may be abundant and may show mucin vacuoles. Adenocarcinoma cells may be a component of a high grade urothelial carcinoma.

Other Tumors
On occasion one may see small cell carcinomas or lymphomas. The diagnosis is based on traditional morphologic criteria.

Secondary Tumors
On rare occasions, cells of prostatic adenocarcinoma may be seen in bladder washings and urine cytology. The cells are usually arranged in clusters and show prominent nucleoli. Similarly, renal cell carcinomas on rare occasions can shed cells that may appear in the urine or washings. The cells are degenerated, have vacuolated cytoplasm, and large nuclei with prominent nucleoli.

Tumors of the female genital tract such as squamous carcinoma and adenocarcinoma of the cervix, and high grade epithelial tumors of the ovary and colon may also be occasionally seen due to direct extension of the tumors through the bladder wall.

References
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  16. Wojcik EM, Bassler TJ, Orozco R: DNA polidy of seminal vesicle cells. A potential diagnostic pitfall in urine cytology. Analyt Quant Cytol Histol 1999; 21: 29-34.
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