Effect of fleroxacin on mouse bladder carcinogenesis with N-butyl-N- (4-hydroxybutyl) nitrosamine.

We recently reported fleroxacin significantly affected cell proliferation in a dose-dependent manner in transitional cell carcinoma cell lines. In this study, we investigated the effect of fleroxacin on mouse bladder carcinogenesis with N-butyl-N-(4-hydroxybutyl) nitrosamine (BHBN). Five-week-old C57BL/6 female mice were divided into three groups. The forced oral administrations of fleroxacin (10 or 50 mg/kg/day) were done for 1 week, then the 0.05% BHBN was given for 8 or 12 weeks. Fleroxacin treatments were continued until sacrifice. The mice were sacrificed at 4 weeks or 8 weeks latent period after BHBN administration, and the histological changes in the bladder were examined. Our study suggested that fleroxacin tended to suppress the development of bladder carcinoma or the malignant changes induced by a shorter period of BHBN administration (8 weeks) in mice. However, in the group of the BHBN administration for 12 weeks, there were no significant effects on the prevention of bladder carcinoma in both of the treatment groups with fleroxacin (10 or 50 mg/kg/day). This study did not make it clear that oral administration of fleroxacin suppressed the development of bladder carcinoma induced by BHBN in mice. However, it is very preliminary data and further experiments need to be done with a much higher dose of the fleroxacin or with other kinds of fluoroquinolone antibiotics.     

Stimulation of urinary bladder tumorigenesis by carcinogen-exposed stroma

There is evidence to suggest that control mechanisms, either growth-stimulatory, inhibitory or inductive, may play a role in carcinogenesis. To test the hypothesis that treatment of rat urinary bladder with carcinogen induces alterations in the stroma which result in modified epithelial-stromal interactions, experiments were conducted using a rat model specifically designed for the study. Following exposure of Fischer F344 rats in drinking water to the urinary bladder carcinogen, N-butyl-N-(4-hydroxybutyl)nitrosamine (BHBN) for four weeks, bladders were removed and subjected to a brief detergent treatment to completely remove epithelium. The bladders without epithelium ("stroma" bladder) were heterotopically transplanted to syngeneic recipients. Four days later, the denuded mucosa surface was resurfaced with intraluminal instillation of urothelial cells, either untreated or treated with BHBN for six weeks (6w-BHBN) or 10 weeks (10w-BHBN). Examination at 12 weeks posttransplant of the "stroma" bladders that had received 6w-BHBN urothelial cells showed a higher tumor incidence of carcinoma in the BHBN-exposed "stroma" bladders as compared with the incidence in the carcinogen-unexposed "stroma" bladders (p less than 0.05). Examination at 18 weeks posttransplant showed 100% incidence of tumors in all "stroma" bladders irrespective of the lengths of BHBN exposure of urothelial cells. However, among the bladders that had received 6w-BHBN urothelial cells, carcinogen-exposed "stroma" bladders proved to be better "soil" for neoplastic cells to proliferate; the mean tumor volume as well as the mean total tumor volume per bladder were significantly higher than in the control "stroma" bladders (p less than 0.01 for each comparison). Similarly, among the bladders that had been resurfaced with 10w-BHBN urothelial cells, the mean total tumor volume per bladder was greater in the carcinogen-treated "stroma" bladders than in the controls (p less than 0.05). No proliferative or neoplastic changes were observed in the BHBN exposed "stroma" bladders which had been resurfaced with normal urothelial cells. Our data indicate that neoplastic growth of carcinogen treated urothelium is enhanced when such cells interact with the stroma which has also been exposed to carcinogen.     

Suppression of rat urinary bladder carcinogenesis by alpha-difluoromethylornithine

The inhibitory effect of oral administration of alpha-difluoromethylornithine (DFMO) on urinary bladder carcinogenesis induced by N-butyl-N-(4-hydroxybutyl)-nitrosamine (BHBN) was explored. Since DFMO in the drinking water at 0.5% and 0.2% had been demonstrated to inhibit tumorigenesis, lower doses (0.2%, 0.1%, 0.03% and 0.01%) of DFMO were examined in the present study. After six-week treatment with the drinking water containing 0.05% BHBN, water containing DFMO at 0.2%, 0.1%, 0.03%, 0.01% or 0% was administered during the subsequent 34 weeks. Incidence of bladder carcinoma was 15/35 (43%), 14/35 (40%), 21/35 (60%), 20/35 (57%) and 27/35 (77%) in the 0.2%, 0.1%, 0.03%, 0.01% and 0% DFMO groups, respectively. Stastistical analysis indicated significant tumor suppression in the 0.2% and 0.1% DFMO groups. Tumor multiplicity and size were not affected by DFMO treatment. No untoward effects were demonstrated in body weight gain or examination of pertinent organs. Our data indicate that bladder carcinogenesis induced by six-week exposure to 0.05% BHBN is significantly inhibited by daily administration of DFMO at the level of 0.1% or higher in drinking water.     

BASEMENT MEMBRANE AND CARCINOGENESIS: ULTRASTRUCTURAL OBSERVATIONS IN THE BASEMENT MEMBRANE OF THE BLADDER EPITHELIUM IN RATS TREATED WITH N-BUTYL-N-(4-HYDROXYBUTYL)NITROSAMINE (BBN)

This study investigated the structural alterations in the basement membrane (BM) of the bladder epithelium in rats treated with N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) using transmission electron microscopy. Following administration of BBN, thickening of the BM of the bladder epithelium was observed and remained almost constant from 4 to 20 weeks, although the pathological changes in the rat bladder epithelium induced by BBN altered over the same period of 20 weeks. The reason for this phenomenon can be explained by the increased interfacial area between the basal epithelial cells and the BM of the rat bladder epithelium due to an increase in the number and size of the microvilli on the basal cell surfaces adjacent to the BM. Our results also showed that the frequency of hemidesmosomes increased progressively during the period of carcinogenesis, especially in the lesions of noninvasive transitional cell carcinoma (TCC) in the rat bladder. It is suggested that the neosynthesis of BM components can be carried out both by benign hyperplastic cells and by noninvasive TCC cells of rat bladder. The alterations in the BM thickness may be affected by the changes in the number and size of the microvilli occurring on the basal cell surfaces adjacent to the BM. Both an increased frequency of hemidesmosomes and the neosynthesis of BM are closely related to cell proliferation during carcinogenesis.     

β-catenin的表达在乳腺癌发生发展中的作用

目的 观察β-catenin的表达在乳腺癌发生发展中的作用.方法 采用免疫组织化学Envision+法检测β-catenin在不同病理阶段的乳腺组织中的表达状况.乳腺囊性增生病27例,导管内乳头状瘤25例,囊性增生病伴异型26例,导管内癌(或伴早期浸润)29例,浸润性导管癌58例.结果 β-catenin在乳腺导管内乳头状瘤、囊性增生病、囊性增生病伴异型、导管内癌(或伴早期浸润)和浸润性导管癌中的阳性表达率分别为4.0%、3.7%、23.1%、72.4%和79.3%.各组间表达的差异有统计学意义(P<0.01).β-catenin的单纯膜表达与肿瘤直径的大小、TNM分期和淋巴结转移状况显著相关,直径小的表达率更高(P<0.01).膜表达与肿瘤的组织学分级(P>0.05)、患者年龄均无明显相关(P>0.05).β-catenin的膜浆共表达与淋巴结是否转移显著相关(P<0.05),与临床分期、组织学分级等无明显相关(P>0.05).β-catenin膜浆表达的淋巴结转移率高于膜表达组(P<0.05).结论 β-catenin在囊性增生病伴异型、导管内癌(或伴早期浸润)和浸润性导管癌中的阳性表达率明显高于囊性增生病和导管内乳头状瘤,它的作用可能始于癌变的早期并持续存在.β-catenin的膜浆表达与淋巴结转移呈正相关,提示β-catenin膜浆表达的乳腺癌易发生转移,两者可能是预后不良的因素.β-catenin的膜表达与肿瘤直径的大小、TNM分期和淋巴结转移状况呈负相关,可能提示预后较好.     

Anti-glomerular basement membrane disease and dual positivity for antineutrophil cytoplasmic antibody in a patient with membranous nephropathy

We present the case of a 50-year-old man who underwent kidney biopsy for nephrotic syndrome. In addition to a membranous pattern, anti-glomerular basement membrane (anti-GBM) staining was noted before manifestations of anti-GBM disease. Hematuria and renal failure ensued 2 weeks later. In addition, he had simultaneous circulating levels of anti-GBM antibody and both perinuclear (P-) and cytoplasmic (C-) antineutrophil cytoplasmic antibody (ANCA).     

Immunohistochemical localization of epithelial membrane antigen, carcinoembryonic antigen and secretory component in urinary bladder cancer

To clarify the relationship between the immunohistochemical distribution pattern of epithelial antigens in transitional cell carcinomas and their histopathological grading and staging, epithelial membrane antigen (EMA), carcinoembryonic antigen (CEA) and secretory component (SC) were localized. Formalin-fixed paraffin-embedded sections from 55 patients with bladder carcinoma were stained by the indirect immunoperoxidase method. In normal transitional epithelium, EMA was found on the luminal side of the plasma membrane of a few surface layers, and in the cytoplasm of the superficial cells. In the lower grade and stage of transitional cell carcinoma, only the luminal surface of superficial cells was positively stained. Membrane and cytoplasmic staining of EMA was frequently found in the intermediate and basal layers of the carcinoma, and the incidence of cytoplasmic staining increased with the higher grade and stage. CEA was not detected in normal epithelium. Cytoplasmic staining of CEA was progressively more frequent in the higher grade and stage of transitional cell carcinoma. In normal epithelium SC was observed on the apical surface plasma membrane and in the cytoplasm of the superficial cells, as shown in the immunohistochemical staining for EMA. The correlation of immunohistochemical detection of SC with the grade or stage was not as good as the correlations for EMA or CEA. These findings suggest that immunohistochemical examination for EMA, CEA and SC in bladder carcinoma could provide valuable information for grading or staging in pathological diagnosis.     

Lysozyme Expression Can be Useful to Distinguish Mammary Analog Secretory Carcinoma from Acinic Cell Carcinoma of Salivary Glands

Lysozyme is an enzymatic marker of acinar and intercalated duct cells of normal salivary glands. The aim of this study was to verify whether lysozyme expression could be useful to distinguish acinic cell carcinoma (ACC) from its main mimic, mammary analog secretory carcinoma (MASC). For comparison, DOG1 expression was analyzed as well. Seventeen cases of ACC, 15 MASC, and 125 other salivary tumors were studied. Lysozyme expression was found in tumor cells as well as in secreted material of MASC (86.6 % of cases) and in ductal cells of epithelial–myoepithelial carcinoma (EMC-53.8 %), pleomorphic adenoma (PA-29.1 %) and polymorphous low-grade adenocarcinoma (PLGA-23.8 %). However, in ACC, lysozyme was not expressed. Three patterns of DOG1 staining were seen: apical–luminal, cytoplasmic, and mixed cytoplasmic/membranous. The apical–luminal pattern was detected in ductal cells of ACC (58.8 % of cases), EMC (38.4 %), adenoid-cystic carcinoma (AdCC-35.3 %), PA (8.3 %), and PLGA (4.8 %). These tumors also showed mixed membranous/cytoplasmic staining for DOG1. MASC, mucoepidermoid, and salivary duct carcinomas exhibited only DOG1 cytoplasmic staining. In conclusion, lysozyme cannot be used as a marker of acinar differentiation in salivary tumors. However, lysozyme expression can be helpful to distinguish MASC from ACC due to its high frequency in the former and absence in ACC. It is likely that in MASC, lysozyme expression may reflect a lactational-like secretory differentiation since lysozyme belongs to breast milk proteins. Regarding DOG1 expression, the apical–luminal pattern is related to acinar and intercalated duct differentiation whereas the cytoplasmic staining does not seem to be associated with a specific cellular phenotype.     

膀胱癌组织中蛋白激酶C及其抑制物活性的研究

目的　探讨蛋白激酶C(PKC)及其抑制物(PKCI)在膀胱移行细胞癌发生过程中的分子生物学机制。方法　采用Takai法检测20例膀胱移行细胞癌及癌周正常膀胱粘膜中PKC及其内源性抑制物(PKCI)的活性。结果　膀胱癌与癌周正常膀胱粘膜相比,胞膜（8.67±2.07,10.66±1.24）、胞浆（7.68±3.27,20.63±12.77）及总体（7.71±4.52,14.34±7.47）PKC活性(pmol     

Effects of α-difluromethylornithine on the development of deeply invasive urinary bladder carcinomas in mice

Summary -Difluoromethylornithine (DFMO), was examined for its ability to suppress the development of invasive urinary bladder carcinoma in C3H/He male mice. Continuous administration of 0.2% DFMO in water following carcinogen treatment (0.05% N-bytyl-N-(4-hydroxybutyl) nitrosamine, BHBN, in drinking water for 8 weeks) was effective in suppressing urinary bladder carcinomas (P<0.05) as compared with the control group. However, when comparison was made based on tumors involving the entire urinary tract, protective effects could not be demonstrated. Coadministration of DFMO (0.2%) and BHBN (0.01%) did not alter tumor induction by the latter. These results were in sharp contrast to the protective effects in rats. Since bladder tumors in rats are of low grade and superficial whereas those in mice are of high grade and deeply invasive, our data indicate that DFMO has little to no effects against the development of aggressive forms of bladder carcinoma.Supported by NIH Grant CA 33511     

Apoptosis and Carcinogenesis: Morphologic Observations in the Rat Bladder Treated with N-Butyl-N-(4-hydroxybutyl)nitrosamine

Background: We investigated the role of apoptosis in rat bladder carcinogenesis induced by N-butyl-N- (4-hydroxybutyl)nitrosamine (BBN).
Methods: Apoptosis was detected in BBN-induced rat bladder lesions by routine hematoxylin and eosin (H&E) staining, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL), and transmission electron microscopic examinations.
Results: After administration of BBN, various pathologic changes were observed in the rat bladder after intervals of between 4 and 28 weeks. Through a comparative study in serial sections, the incidence of apoptosis, detected by either routine H&E staining or TUNEL staining, displayed marked elevation in BBN-induced rat bladder lesions, as compared to that in the normal epithelium of the rat bladder, particularly in invasive transitional cell carcinoma. Apoptotic cells with different morphologic features were ultrastructurally demonstrated in a series of rat bladder lesions. Significant group differences in the incidence of apoptosis were seen among the BBN-induced lesions and the normal epithelium of the rat bladder, and among the invasive transitional cell carcinomas and other lesions, before the tumor cells were seen to invade the rat bladder.
Conclusion: Our data suggestthat apoptosis may be closely related to carcinogenesis and tumor invasion of the rat bladder induced by BBN, and that the comparative study of serial sections, each stained by H&E and TUNEL, combined with electron microscopic observation, may provide the most reliable informations in semiquantitative assessment of apoptosis.     

Focal positive prostate-specific membrane antigen (PSMA) expression in ganglionic tissues associated with prostate neurovascular bundle: Implications for novel intraoperative PSMA-based fluorescent imaging techniques

ObjectiveProstate specific membrane antigen (PSMA) is primarily expressed in glandular prostatic tissue and is frequently utilized to detect primary or metastatic prostatic adenocarcinoma (CaP). A purported novel application of PSMA detection is the intraoperative real-time identification of CaP using radioimmunoscintigraphy to define the extension of the surgical resection. Considering that PSMA expression has been reported in other tissues, we evaluated its immunoexpression in prostatic neurovascular bundle elements to assess the convenience and safety of the aforementioned procedure.Materials and methodsTwenty consecutive specimens of radical prostatectomy (RP) were retrieved from our surgical pathology archives. PSMA immunoexpression (Clone 3E6, DAKO) was assessed in a representative section from each specimen containing neurovascular bundle elements.ResultsPSMA expression was documented in 20/20 of examined CaP slides. Most cases exhibited an apical/cytoplasmic or cytoplasmic with membranous accentuation pattern of staining. Focal weak to moderate cytoplasmic staining was detected in associated ganglionic tissue in 3/15 of the examined RP. In all cases, staining was cytoplasmic, less extensive, and weaker than the pattern observed in CaP. None of the peripheral nerve sheath cells or lymphovascular components of the examined neurovascular bundles were positive for PSMA.ConclusionsWe found focal positive PSMA expression in the ganglionic cells of the prostatic neurovascular bundle. Our results suggest that the radioimmunoscintigraphic detection of radiolabeled PSMA antibodies might not be entirely specific for prostatic cells; this observation must be taken into account should an intraoperative PSMA-based fluorescent imaging technique be used to define the extension of the surgical procedure.     

Increased phospho-AKT is associated with loss of the androgen receptor during the progression of N-methyl-N-nitrosourea-induced prostate carcinogenesis in rats

BACKGROUND: Characterization of molecular events during N-methyl-N-nitrosourea (MNU)-induced rat prostate carcinogenesis enhances the utility of this model for the preclinical assessment of preventive strategies. Androgen independence is typical of advanced human prostate cancer and may occur through multiple mechanisms including the loss of androgen receptor (AR) expression and the activation of alternative signaling pathways. METHODS: We examined the interrelationships between AR and p-AKT expression by immunohistochemical staining during MNU-androgen-induced prostate carcinogenesis in male Wistar-Unilever rats. Histone nuclear staining and image analysis was employed to assess parallel changes in chromatin and nuclear structure. RESULTS: The percentage of AR positive nuclei decreased (P < 0.01) as carcinogenesis progressed: hyperplasia (92%), atypical hyperplasia (92%), well-differentiated adenocarcinoma (57%), moderately-differentiated adenocarcinoma (19%), and poorly-differentiated adenocarcinoma (10%). Conversely, p-AKT staining increased significantly during carcinogenesis. Sparse staining was observed in normal tissues (0.2% of epithelial area) and hyperplastic lesions (0.1%), while expression increased significantly (P < 0.001) in atypical hyperplasia (7.6%), well-differentiated adenocarcinoma (16.7%), moderately-differentiated adenocarcinoma (19.6%), and poorly-differentiated adenocarcinoma (17.4%). In parallel, nuclear morphometry revealed increased nuclear size, greater irregularity, and lower DNA compactness as cancers became more poorly differentiated. CONCLUSIONS: In the MNU model, the progressive evolution of dominant tumor cell populations showing an increase in p-AKT in parallel with a decline in AR staining suggests that activation of AKT signaling may be one of several mechanisms contributing to androgen insensitivity during prostate cancer progression. Our observations mimic findings suggested by human studies and support the relevance of the MNU model in preclinical studies of preventive strategies.     

Failure to activate Epstein-Barr virus genome in a latently-infected human lymphoblastoid cell line with urine containing a bladder carcinogenesis promoter

We assessed the possibility to detect promoters of bladder carcinogenesis by a short-term in vitro assay utilizing the activation of Epstein-Barr virus (EBV) expression in EBV genome-carrying human lymphoblastoid cells. This system is composed of EBV-nonproducer Raji cells as the indicator, n-butyrate as the EBV-inducer and the test substance. None of eight known bladder carcinogens (2-naphthylamine, 2-acetylaminofluorene, N-butyl-N-(4-hydroxybutyl)nitrosamine, N-butyl-N-(3-carboxypropyl) nitrosamine, N-methyl-N-nitrosourea, N-[4-(5-nitro-2-furyl)-2-thiazolyl] formamide, benzidine, cyclophosphamide) or five promoters (sodium saccharin, sodium cyclamate, urea, allopurinol, DL-tryptophan) were detected by this system. Normal urine from 20 rats did not activate the EBV genome. Neither the 0.45 micron membrane filtrates nor ether extracts of urine obtained from rats given one of 4 bladder carcinogens, 4 promoters or phorbol 12-myristate 13-acetate reacted with this system, whereas those from rats given either vincristine or vinblastine did, although their activity for promoting bladder carcinogenesis is not known. Urine samples from 42 patients with bladder cancers, when tested as filtrates, ether extracts or XAD-2 resin adsorbates, did not activate EBV expression. These results suggest that EBV-activating substances contribute little to promotion of bladder carcinogenesis and that this test might not be applicable for screening of tumor promoters in urine.     

The effect of pregnancy and delivery on the function and ultrastructure of the rat bladder and urethra

OBJECTIVE: To examine the effect of pregnancy and delivery on the function and ultrastructure of the bladder and urethra in rats. Material and methods The study comprised six virgin and 18 pregnant rats; both groups underwent cystometry (at the 19th day of gestation, and 2 days and 6 weeks after parturition). Tissues from the bladder and urethra were collected for electron microscopy, western blotting and immunostaining for caveolin-1 and caveolin-3. RESULTS: The bladder capacity was greater and the modified leak-point pressures lower in pregnant and 2-day postpartum rats than in virgin and 6-week postpartum rats. The residual volume was significantly higher in the pregnant group. Electron microscopy showed more sarcolemmal caveolae in the smooth muscle cells of both the bladder and urethra of virgin rats than in the other groups. Lipid droplets and subsarcolemmal mitochondria accumulated in pregnant and 2-day postpartum rats. Caveolin-1 protein was detected in the cytoplasmic membrane of urethra and bladder smooth muscle cells. Caveolin-3 was detected in the membrane of striated muscle in the intrinsic sphincter. Western blotting showed increased caveolin-1 protein expression in the bladder and urethra of 2-day postpartum rats; in contrast, levels of caveolin-1 were lower in pregnant rats than in virgin and 6-week postpartum rats. CONCLUSION: s During pregnancy there was a significant decrease in sarcolemmal caveolae and caveolin-1 in the smooth muscle cells of the rat bladder and urethra. The changes in caveolae and the membrane protein caveolin may play a role in the functional changes associated with pregnancy and after delivery.     

Blood group isoantigen and lectin binding studies in the human bladder cancer

The purpose of this study is to investigate changes of glycoconjugates in the cancer cells of the urinary bladder by means of immunohistochemical methods. The normal bladder epithelium, cancerous lesions, and non-malignant epithelia of the tumor bearing bladder were examined by staining by avidin-biotin-peroxidase complex methods using blood group isoantigens (BGA) and lectins. The materials were obtained from 48 cystectomy specimens in our hospital in these seven years. Anti-A, B and H monoclonal antibodies were used for detecting BGA. GSI-A4, UEA-1, LTA-M, BPA, DBA and PNA were used as probes for lectins. The changes of glycoconjugates in the cancer cells of the bladder and in non-malignant epithelia of the tumor bearing bladder were studied by using six kinds of lectins. Each lectin binding rates of the primary lesions and the metastasized lesions was comparatively investigated. Positive rates for BGA of the high grade tumor was lower than that of the low grade. In the high grade tumor, GSI-A4, PNA and BPA showed high binding rates, while the DBA binding rate was low. The correlation between the histopathological grading of bladder tumor and the changes of glycoconjugates in the cells was suggestive of cancerous process.     

Sperm membrane dynamics assessed by changes in lectin fluorescence before and after capacitation

Sperm capacitation is correlated with acquisition of fertilizing ability, and the molecular events underlying this process are only beginning to be understood. A number of membrane changes associated with capacitation have been documented. In this study we used lectin probes to identify changes in glycoprotein localization as a result of capacitation of mouse sperm. Eight lectins (LEA, PSA, PNA, AAA, UEA-1, WGA, STA, and TPA) stained regions of the mouse sperm head, tail, or both. No changes in tail staining patterns were detected when sperm were incubated under capacitating conditions. In contrast, 7 of 8 lectins tested showed clear shifts in staining patterns in the sperm head as a result of incubation under capacitating conditions. When staining patterns were quantified, a distinct heterogeneity within the sperm population was observed. Each lectin displayed 3 distinct staining patterns in both uncapacitated and capacitated sperm samples. The least common pattern represented the acrosome-reacted (AR) pattern, as independently assessed by lectin staining of ionophoretreated sperm that were >95% AR as judged by Coomassie staining. However, a reciprocal shift in the two predominant staining patterns was correlated with capacitation and suggests that changes in distribution of cell surface proteins during capacitation constitute part of the molecular changes which result in changes in sperm function acquired during this process.     

Epithelial membrane antigen as an immunohistochemical marker for transitional cell carcinoma of the urinary bladder

Epithelial membrane antigen (EMA) was immunohistochemically localized in transitional cell carcinomas of the bladder to clarify EMA staining pattern's relationship to the histological grading and staging of tumors and patient prognosis. Formalin-fixed, paraffin-embedded sections from 101 patients with bladder carcinoma were stained by the indirect immunoperoxidase method. In the lower histological grade and stage of transitional cell carcinoma, the localization of EMA was restricted to the luminal surface of the superficial cells. Membrane and cytoplasmic staining of EMA was frequently found in the intermediate and basal layers of the tumor cells, and the incidence of cytoplasmic staining increased with advancing grade and stage. Stromal staining was frequently observed in cases of higher grade and stage. In addition, these distribution patterns of EMA were well correlated with patient survival. Thus, we differentiated three types of EMA distribution: a luminal type, with very good prognosis; a cytoplasmic type, with fair prognosis; and a stromal type, with relatively poor prognosis. These findings suggest that EMA distribution in bladder cancer could be a valuable indicator for histological grading or staging in pathological diagnosis and for predicting the survival of bladder cancer patients.