Transposable hypomethylation is associated with metastatic capacity of primary melanomas

Despite the strong progress has been made in the field of melanoma epigenetics, the importance of genome-wide demethylation or hypomethylation remains underestimated. However, this phenomenon might also reflect important epigenetic alterations due to its ability to cause genetic instability. Furthermore, no methylation-based distinction has been drawn among the diverse primary melanoma subtypes. To assess global methylation we measured the methylation level on the 6 CpG sites of LINE1 sequences in 46 primary melanomas in association with patients’ survivals and the clinicopathological characteristics of specimens. We demonstrate that LINE1 hypomethylation is accompanied by the shortened relapse-free survival of melanoma patients; however, Cox regression analysis shows a direct relationship between the overall loss of 5-methylcytosine and metastatic potential of primary melanomas, which is confirmed by Kruskal-Wallis tests with Dunn’s Multiple Comparison Post-test showing that not only the presence but the number of metastases during the 5-year follow-up period is associated with the transposon demethylation. In this study, we demonstrate the strong influence of global DNA demethylation in the metastatic formation of primary melanomas during the follow-up period.     

Micropapillary component of urothelial carcinoma detected in transurethral resection of bladder tumor (TUR-BT) tissues: a case report

A case of urothelial carcinoma (UC) containing a micropapillary carcinoma (MPC) component in the urinary bladder of an 83-year-old man is reported. The MPC component of UC has been reported to be a variant featuring poor prognosis and rapid progression. In the present case, a characteristic MPC component with micropapillary growth, in association with a fine meshwork-like stroma, was observed in less than 10% of fragmented cancer tissues of UC, G3, obtained by transurethral resection of a bladder tumor (TUR-BT). Lymphatic invasion was also detected. UC cancer cells had invaded the prostatic glands and replaced the original epithelial cells. The unique "insideout" feature of the MPC component was immunohistochemically obvious on staining with antibody to epithelial membrane antigen (EMA). On immunohistochemical study, cancer cells of both UC and MPC components were positive for pancytokeratin AE1/AE3 and cytokeratins 7 and 20. Carcinoembryonic antigen (CEA) and CAM5.2 were only focally positive in UC cells. MIB-1(Ki-67) labeling index was high, at 80%-90%, in cancer cells of UC. This was a case of UC, G3 with invasion to the muscularis propria layer of the urinary bladder and also to the prostate. MPC and MPC components in cancers should be recognized as a marker of poor prognosis, even when detected in less than 10% of UC within TUR-BT tissues, as in the present case.     

DNA methylation patterns in hereditary human cancers mimic sporadic tumorigenesis.

Cancer cells have aberrant patterns of DNA methylation including hypermethylation of gene promoter CpG islands and global demethylation of the genome. Genes that cause familial cancer, as well as other genes, can be silenced by promoter hypermethylation in sporadic tumors, but the methylation of these genes in tumors from kindreds with inherited cancer syndromes has not been well characterized. Here, we examine CpG island methylation of 10 genes (hMLH1, BRCA1, APC, LKB1, CDH1, p16(INK4a), p14(ARF), MGMT, GSTP1 and RARbeta2) and 5-methylcytosine DNA content, in inherited (n = 342) and non-inherited (n = 215) breast and colorectal cancers. Our results show that singly retained alleles of germline mutated genes are never hypermethylated in inherited tumors. However, this epigenetic change is a frequent second "hit", associated with the wild-type copy of these genes in inherited tumors where both alleles are retained. Global hypomethylation was similar between sporadic and hereditary cases, but distinct differences existed in patterns of methylation at non-familial genes. This study demonstrates that hereditary cancers "mimic" the DNA methylation patterns present in the sporadic tumors.     

Clinical significance of the reduction of UT-B expression in urothelial carcinoma of the bladder

Urea transporter B (UT-B) is a membrane protein and plays an important role in regulating urea concentration in bladder urothelial cells. It has been reported that UT-B gene mutations were related to bladder carcinogenesis, and UT-B deletion could induce DNA damage and apoptosis in bladder urothelium. However, the functions and clinical significance of UT-B in human bladder cancer remain unknown. The most common type of bladder cancer is urothelial carcinoma (UC). We hypothesized that UT-B expression was related to bladder UC progress. In this study, UT-B was detected using immunohistochemistry in 52 paraffin-embedded specimens of bladder UC and 10 normal urothelium specimens. The results showed that UT-B protein expression in UC tumor cells was significantly lower as compared with normal urothelial cells (P = 0.021). UT-B protein expression was significantly reduced with increasing histological grade (P = 0.010). UT-B protein expression in muscle-invasive stage was significantly lower than in non-muscle-invasive stage (P = 0.014). Taken together, our data suggest that the reduction or loss of UT-B expression may be related to the incidence, progression and invasiveness of bladder UC. UT-B may be a novel diagnostic or prognostic biomarker, as well as a potential therapeutic target in UC of the bladder.     

Reversibility of aberrant global DNA and estrogen receptor-alpha gene methylation distinguishes colorectal precancer from cancer

Alterations in the global methylation of DNA and in specific regulatory genes are two epigenetic alterations found in cancer. However, the significance of epigenetic changes for diagnosis and/or prognosis of colorectal cancer have not been established, although it has been extensively investigated. Recently we have identified a new type of cancer cell called precancerous stem cells (pCSCs) and proposed that cancer may arise from a lengthy development process of tumor initiating cells (TICs) --> pCSCs --> cancer stem cells (CSCs) --> cancer, which is in parallel to histological changes of hyperplasia (TICs) --> precancer (pCSCs) --> carcinoma (CSCs/cancer cells), accompanied by clonal evolutionary epigenetic and genetic alterations. In this study, we investigated whether aberrant DNA methylation can be used as a biomarker for the differentiation between premalignant and malignant lesions in the colorectum. The profile of global DNA and estrogen receptor (ER)-alpha gene methylation during cancer development was determined by analysis of 5-methylcytosine (5-MeC) using immunohistochemical (IHC) staining, dot blot analysis or a quantitative gene methylation assay (QGMA). Herein we show that global DNA hypomethylation and ER-alpha gene hypermethylation are progressively enhanced from hyperplastic polyps (HPs) --> adenomatous polyps (APs) --> adenomatous carcinoma (AdCa). The aberrant methylation can be completely reversed in APs, but not in AdCa by a nonsteroidal anti-inflammatory drug (NSAID) celecoxib, which is a selective inhibitor of cyclooxygenase-2 (Cox-2), suggesting that the epigenetic alterations between colorectal precancer (AP) and cancer (AdCa) are fundamentally different in response to anti-cancer therapy. In normal colorectal mucosa, while global DNA methylation was not affected by aging, ER-alpha gene methylation was significantly increased with aging. However, this increase did not reach the level observed in colorectal APs. Taken together, reversibility of aberrant global DNA and ER-alpha gene methylation distinguishes colorectal precancer from cancer.     

Does p53 immunostaining improve diagnostic accuracy in urine cytology?

The frequent change of the transitional cell carcinoma of the urinary tract accounts for the fact that cytological abnormalities in urinary specimens are often not sufficient to enable a definitive diagnosis of malignancy. The purpose of this work was to evaluate the possible use of p53 protein in increasing the diagnostic accuracy of urinary cytology. The expression of p53 was investigated by immunocytochemistry in two groups of urinary specimens, one cytologically positive and the other cytologically negative for cancer. Immunostaining was carried out using a monoclonal antibody to p53. In the positive group, in which bladder cancer was confirmed by cystoscopy and biopsy (31 cases), positive reaction for p53 was found in 55% of the cases (17 cases). In the negative group (92 cases), presence of cancer was histologically ascertained in 64 cases and in this group 15 cases (23.4%) showed positive p53 staining. In the remaining 28 cases of this group, where TCC was not present, 7 cases showed p53 positivity in non-neoplastic urothelial cells. This result shows that, while immunocytochemical detection of p53 in urinary specimens may be used for prognostic evaluation of patients with bladder cancer, it does not contribute to the diagnostic accuracy in cases with morphologically inconclusive or negative cytology. The sensitivity and specificity of the method in detecting bladder carcinoma were 23.5 and 75%, respectively. Diagn. Cytopathol. 1997;17:436–439. © 1997 Wiley-Liss, Inc.     

Ribonucleotide reductase M2 subunit is a novel diagnostic marker and a potential therapeutic target in bladder cancer

Morikawa T, Maeda D, Kume H, Homma Y & Fukayama M
(2010) Histopathology 57, 885–892 Ribonucleotide reductase M2 subunit is a novel diagnostic marker and a potential therapeutic target in bladder cancer Aims: To examine the immunohistochemical expression and function of ribonucleotide reductase M2 subunit (RRM2), a gemcitabine‐related molecule, in bladder cancer. Methods and results: One hundred and seventeen bladder specimens on a tissue microarray were immunostained for RRM2. Positive RRM2 staining was observed in none of 14 examples of non‐neoplastic urothelium, none of four low‐grade urothelial carcinoma (UC), 69 of 83 (83%) high‐grade UC, three of three (100%) squamous cell carcinoma and 12 of 13 (92%) lymph node metastasis of UC. RRM2 overexpression was associated significantly with muscularis propria invasion in UC patients who had undergone radical cystectomy (P = 0.005). Immunohistochemistry for RRM2 was then applied to small biopsy specimens of 15 cystitis with reactive atypia cases and 25 urothelial carcinoma in‐situ (CIS) cases. Positive RRM2 staining was found in one of 15 (6.7%) cystitis with reactive atypia cases and in 24 of 25 (96%) CIS cases. Finally, UM‐UC‐3 bladder cancer cells were transfected with RRM2 siRNAs and cell growth was evaluated. Knockdown of RRM2 protein markedly inhibited cell growth. Conclusions: We have shown frequent overexpression of RRM2 protein and its possible role in bladder cancer. Our results suggest that RRM2 is a novel diagnostic marker and a potential therapeutic target in bladder cancer.     

Altered global methylation of DNA: an epigenetic difference in susceptibility for lung cancer is associated with its progression

Alterations in global DNA methylation have been observed in many cancers, but whether such alterations represent an epigenetic difference in susceptibility for the disease is unknown. The status of global DNA methylation also has not been reported in intact or specific types of cells involved in the carcinogenic process. To address these issues in lung carcinogenesis, we evaluated the status of global DNA methylation by using a monoclonal antibody specific for 5-methylcytosine (5-mc) in randomly selected lung specimens of 60 cigarette smokers who developed squamous cell carcinoma (SCC) and 30 cigarette smokers who did not. 5-mc immunostaining scores of DNA of SCC (0.61 +/- 0.42) and associated hyperplastic lesions (0.82 +/- 0.27) was significantly lower than those of DNA of histologically normal bronchial epithelial cells (0.99 +/- 0.52) and hyperplastic lesions (1.2 +/- 0.22) of noncancer specimens. The ratio of 5-mc scores between SCC and matched uninvolved bronchial epithelial cells was significantly associated with advanced stage and size of the tumor. The results suggest that alteration in global DNA methylation is an important epigenetic difference in susceptibility for the development of lung cancer. The reduced global DNA methylation in SCC compared with epithelial hyperplasia and its association with tumor size and disease stage is suggestive of its involvement in the progression of SCC. The results also indicate that normal methylation of DNA in epithelial hyperplastic lesions may prevent the transformation of these lesions to invasive cancer. If these results are confirmed, the status of DNA methylation in early lesions such as epithelial hyperplasia could be used to identify smokers who are at risk for the development of SCC.     

The prognostic significance of nuclear CSE1L in urinary bladder urothelial carcinomas

Prognosis of urinary bladder urothelial carcinomas may be challenging; many tumors with similar histopathologic features show significantly different clinical outcomes. CSE1L, the chromosome segregation 1-like protein, is both a cytoplasmic and nuclear protein. We investigated the cytoplasmic/nuclear expression pattern of CSE1L to determine its potential prognostic significance. In immunohistochemical analysis, nonneoplastic urothelium showed faint CSE1L staining, whereas all tumors in the bladder cancer specimens had significant staining for CSE1L (100%, or 38/38). CSE1L cytoplasmic/nuclear staining was defined based on relative staining intensity. A total of 20 (52.6%) of 38 cancer specimens had strong nuclear CSE1L staining, and 44.7.3% (17/38) of the samples had strong cytoplasmic CSE1L staining. Bladder urothelial carcinomas with high CSE1L nuclear staining had a significantly lower overall survival rate (log-rank test, P = .011). CSE1L expression was not correlated with tumor stage, likely reflecting the faultiness of current urothelial carcinoma evaluation methods. Our results suggest that nuclear CSE1L may play an oncogenic role in bladder tumor progression and that immunohistochemical staining of nuclear CSE1L may be useful for the prognosis of bladder urothelial carcinomas.     

Histologic-genetic mapping by allele-specific PCR reveals intraurothelial spread of p53 mutant tumor clones

Common and clinically important features of urothelial carcinomas are multifocality and a high rate of recurrence. Molecular studies demonstrated that multifocal tumors are frequently composed of one tumor clone spreading throughout the urothelial tract. A combination of histologic and genetic mapping of cystectomy specimens from bladder cancer patients is a valuable tool to study bladder carcinogenesis and tumor cell spread by correlating urothelial morphologic features and defined genetic alterations. In the present study, the primary tumors of 14 cystectomy specimens were investigated for p53 protein overexpression by immunohistochemistry and p53 gene mutation by genomic sequencing. Seven tumors showed a strong nuclear staining for the p53 protein. In six of seven tumors, a p53 gene mutation was detected. Allele-specific PCR of defined p53 mutations was established in five of six cases with a p53 mutation. Subsequent screening of the entire urothelial lining of each cystectomy specimen by allele-specific PCR revealed p53-mutant cell clones in urothelial patches with carcinoma in situ and dysplasia, but also frequently in histomorphologically normal urothelium adjacent to the tumor. The pattern of tumor cell spread indicated a continuous intraurothelial growth of the p53-mutant clone. P53 immunohistochemistry visually confirmed the presence of mutant cells in most of these samples. We conclude that allele-specific PCR is a highly sensitive and reliable method for tracking specific p53 mutant clones in the urothelium. Moreover, the detection of p53-mutant cells in histologically normal or preneoplastic urothelial areas in four patients with invasive bladder cancer indicates an extensive intraurothelial tumor cell spread. The excellent correlation of immunohistochemically positive urothelial patches with the presence of a specific mutation highlights the biologic significance of p53-positive cells in the urothelium of tumor patients.     

Identification of genetic alterations in upper urinary tract urothelial carcinoma in end‐stage renal disease patients

Clinical presentations of end‐stage renal disease (ESRD) patients on dialysis with upper urinary tract urothelial carcinoma (UUT‐UC) are different from those with normal renal function. The pathogenesis remains unknown. We investigated the pathogenetic influence of chromosomal aberrations in patient on dialysis with UUT‐UC. The chromosomal aberrations of UUT‐UC specimens from seven dialysis patients were assessed by conventional comparative genomic hybridization (cCGH). Subsequently, we further investigated 20 cases by whole genome and fine‐tiling oligonucleotide array‐based CGH to demonstrate gains and losses, and compared with the clinicopathologic background. The chromosomal aberrations in UUT‐UC specimens from dialysis patients were more complex than in bladder urothelial carcinoma (B‐UC). Our data showed that gains at 5p, 7, 19q, and losses at 4q, 9p, and 15q are common in UUT‐UC of ESRD patients. Gains in regions associated with DNA repair genes were noted in this study. High‐stage and high‐grade tumors displayed more copy number variants. In addition, female ESRD patients with UUT‐UC had more frequent chromosomal aberrations than their male counterparts. In conclusion, unique chromosomal aberrations were indentified in UUT‐UC in ESRD patients. © 2010 Wiley‐Liss, Inc.     

膀胱尿路上皮癌中候选生物标志物转录因子21的甲基化机制及意义

目的 探讨膀胱尿路上皮癌中候选生物标志物转录因子21（TCF21）甲基化的意义。 方法 选择2016年10月~2017年10月间疑似膀胱癌患者142例,其中经病理学检测确诊为膀胱癌80例为研究组,经病理学检测确诊为非膀胱癌共62例为对照组。另选择同期进行体检的健康者尿标本40例作为健康组。检测研究组膀胱癌组织、癌旁组织、对照组病灶组织中TCF21甲基化水平,并检测研究组、对照组和健康组尿液中TCF21甲基化水平,分别比较膀胱癌组织、尿液中TCF21甲基化水平与临床病理特点间的关系,并分析两者对于膀胱癌的诊断效能。 结果 膀胱癌组织TCF21甲基化水平显著高于癌旁组织和对照组（P<0.05）,研究组尿液TCF21甲基化水平显著高于对照组和健康组（P<0.05）。男性、年龄>60岁、高的TNM分期和高分级的膀胱癌患者膀胱癌组织和尿液中TCF21甲基化水平更高（P<0.05）。膀胱癌组织与尿液中TCF21甲基化水平对膀胱癌均有较高的诊断意义,且两者的诊断意义差异无显著性（P>0.05）。 结论 TCF21基因在膀胱癌组织和膀胱癌患者尿液中均具有较高的甲基化水平,并且与病理特点相关,膀胱癌组织和膀胱癌患者尿液对于膀胱癌均具有较高的诊断意义。     

Global hypomethylation is common in prostate cancer cells: a quantitative predictor for clinical outcome?

This study was designed to determine if cytological detection of 5-methylcytosine (5MC) was feasible on prostate tumor sections and to determine if levels of 5MC differed in malignant compared to normal prostate tissue. We further sought to see if 5MC levels correlated with any clinical outcome data. Thirty prostate tumor sections were obtained from patients who underwent radical prostatectomies from 1988 to 1995; these represented a mix of low to high grade tumors. Clinical data were maintained for each of these patients with a minimum of 7 years of follow up. Sections were stained with a commercially available antibody to 5MC and immunocytochemistry levels were subsequently quantified using a computer-assisted true-color imaging system. Tumor and benign regions of the same archived sections were compared, in addition to a series of 12 normal prostate samples. Prostate cancer cells exhibited a pronounced global decrease in methylation compared with benign and normal tissue. This was observed in 29 of 30 patients (96.7%) studied and densitometric scanning of methylation staining indicated that this value was quantifiable. Overall, higher methylation values were detected in men who had positive surgical margins and recurrent disease. These data suggest that loss of methylation is a feature of prostate cancer, and partial gain of methylation (presumably at promoters of specific genes) is associated with clinical outcome and is measurable using whole-cell assays.     

Immunohistochemical expression of pi class glutathione S-transferase in the basal cell layer of benign prostate tissue following chronic treatment with finasteride

BACKGROUND: Glutathione S-transferases (GST) may prevent carcinogenesis through inactivation of reactive electrophiles by conjugation to reduced glutathione. Treatment directed at the induction or preservation of GST-pi expression in normal epithelium could have a profound impact on the prevention of prostate neoplasia. Finasteride, a 5-alpha-reductase inhibitor, is used as a chemopreventive agent because it blocks the conversion of testosterone to its byproduct which promotes prostate tumour growth. OBJECTIVE: To investigate GST-pi expression immunohistochemically in benign prostate tissue from untreated patients and from patients chronically treated with finasteride. MATERIALS: Immunostaining with anti-GST-pi antibody was performed on 10 (cysto-) prostatectomy, eight simple prostatectomy, and three transurethral prostatectomy specimens. The first set of 10 prostates was from untreated patients operated on for bladder cancer. The other cases were from patients with benign prostatic hyperplasia and chronically treated with finasteride. None of the specimens in either group showed prostatic cancer, prostatic intraepithelial neoplasia, urothelial carcinoma, or chronic prostatitis. Specimens were evaluated for the presence, intensity, and distribution of immunostaining. RESULTS: Diffuse cytoplasmic immunostaining was observed in the basal cell layer of the untreated specimens. Some variability in the expression of GST-pi was seen within each zone and also between the prostate zones. Only a minority of the secretory cells was stained weakly, mainly in the subnuclear region of the cells facing an uninterrupted basal cell layer. Staining was more homogeneously diffuse in the cytoplasm of the luminal cells facing the basement membrane directly. In the benign epithelium of the finasteride treated specimens the circumferential staining of the basal cells appeared to be more continuous than in the untreated cases, the gaps in the stained basal cell layer being fewer, shorter, or even absent in some ducts and acini. There was no variability in the intensity of staining of the basal cell layer, all the cells being intensely stained in a uniform way. The intensity of staining of the secretory cells was not influenced by finasteride treatment. CONCLUSIONS: Following chronic treatment with finasteride the immunohistochemical expression of pi class glutathione S-transferase in the benign prostate ducts and acini is upregulated in relation to an expanded basal cell layer. This could indicate that finasteride acts as a GST-pi inducer.     

Undifferentiated small-cell carcinoma of the urinary bladder: Report of two cases with a primary urinary cytodiagnosis

Two undifferentiated small-cell carcinomas of the urinary bladder are reported. The patients, 68- and 55-yr-old men, respectively, presented with painless hematuria. In the first case, numerous small, lymphocyte-like cells with coarse chromatin, sometimes with small nucleoli, and high nuclear/cytoplasmatic ratios were found in cytologic urine specimens. A cytodiagnosis of undifferentiated small-cell cancer was made. In the second case, urine samples showed rare aggregates of small, undifferentiated cells in association with malignant urothelial cells. The cytodiagnosis of mixed tumor composed of undifferentiated small cell and transitional carcinoma was confirmed by histologic examination. The presence of focal reactivity with anti-chromogranin antibody and neurosecretory granules via electron microscopy supports a neuroendocrine differentiation for the small neoplastic cells. The patients died 13 and 8 mo after diagnosis, respectively. © 1995 Wiley-Liss, Inc.     

Genome-wide methylation patterns in normal and uniparental early mouse embryos.

In the normal diploid mouse embryo, active demethylation of the paternal genome but not of the maternal genome occurs within only a few hours and in a highly coordinated fashion as the zygote proceeds through the first G1 phase. This zygotic demethylation may be necessary to reprogram the sperm genome for somatic development. Immunofluorescence staining with an antibody against 5-methylcytosine shows that the cellular machinery of the fertilized egg cannot demethylate the second maternal genome in parthenogenetic, gynogenetic and triploid digynic embryos or remethylate the additional (already demethylated) paternal genome in androgenetic and triploid diandric embryos. This suggests that differential zygotic demethylation results from differences in the remodeling of paternal and maternal chromatin structures after fertilization, i.e. sperm nuclear decondensation and protamine-histone exchange. A proportion of embryos derived from normal matings display abnormal methylation patterns some of which are indistinguishable from those in androgenetic or gynogenetic embryos. We conclude that methylation reprogramming defects in mammalian zygotes contribute to the high incidence of early pregnancy failure.     

Atypical squamous cells in the urine revealing endometrioid adenocarcinoma of the endometrium with squamous cell differentiation: A case report

Urine cytology is mainly used to detect urothelial carcinoma (UC), especially for high‐grade lesions including urothelial carcinoma in situ. Benign squamous cells are often seen in the urine specimens of women, they are either exfoliated from the trigone area of the bladder, the urethra, or the cervicovaginal region. However, abnormal squamous cells in the urine raise concerns of abnormalities of the urinary tract and cervicovaginal area which range from squamous metaplasia of the urothelium, a cervicovaginal squamous intraepithelial lesion, condyloma acuminatum of the bladder, UC with squamous differentiation, and squamous cell carcinoma. We present here a unique case of atypical squamous cells (ASCs) in the urine subsequently leading to the diagnosis of endometrioid adenocarcinoma of the endometrium with squamous differentiation. The presence of ASCs in voided urine is a rare finding that may indicate an underlying malignancy. Careful evaluation of squamous cells in the urine is an important part of our daily cytopathology practice. Diagn. Cytopathol. 2015;43:49–52. © 2014 Wiley Periodicals, Inc.     

Hepatocyte nuclear factor-1β expression in clear cell adenocarcinomas of the bladder and urethra: diagnostic utility and implications for histogenesis

The histogenesis of clear cell adenocarcinoma of the bladder/urethra is uncertain. Hepatocyte nuclear factor-1β is a homeodomain protein that has been reported to be frequently overexpressed in ovarian clear cell adenocarcinoma in comparison with rare or no expression in other types of epithelial ovarian tumors. We assessed the expression of hepatocyte nuclear factor-1β in a series of 18 clear cell adenocarcinomas of the bladder and urethra and compared it with that of invasive high-grade transitional/urothelial carcinoma (n = 35); adenocarcinomas of the bladder, urethra, and paraurethral glands (n = 21); as well as nephrogenic adenomas of the bladder (n = 8). Staining intensity and extent were evaluated using a 4-tiered grading system (0-3). A case was considered positive for hepatocyte nuclear factor-1β if 10% or more of tumor cells showed at least weak nuclear staining or if any moderate or strong nuclear staining was observed. All 18 clear cell adenocarcinomas exhibited nuclear staining in at least 50% of tumor cells (16 strong, 1 moderate, and 1 weak with focal strong nuclear staining) in comparison with positive nuclear staining (moderate) in 1 of 21 bladder adenocarcinoma, 1 of 35 invasive high-grade transitional/urothelial carcinoma (weak to moderate staining), and 2 of 8 nephrogenic adenomas (1 weak and 1 moderate to strong staining). We concluded that hepatocyte nuclear factor-1β is a useful marker in differentiating clear cell adenocarcinomas of the bladder/urethra from invasive high-grade transitional/urothelial carcinoma and other types of bladder adenocarcinomas and to a lesser extent from nephrogenic adenomas. Hepatocyte nuclear factor-1β is of no diagnostic utility in discriminating primary bladder/urethral clear cell adenocarcinomas from metastatic clear cell adenocarcinomas of the female genital tract to the bladder/urethra. From a histogenesis standpoint, although the expression of hepatocyte nuclear factor-1β in both gynecologic and urologic tract clear cell adenocarcinomas may point to a Müllerian derivation/differentiation, this immunohistochemical evidence is insufficient to completely exclude an urothelial association.     

MTHFR C677T polymorphisms are associated with aberrant methylation of the IGF-2 gene in transitional cell carcinoma of the bladder

The purpose of this study was to determine the relationship between methylation status of the insulin-like growth factor 2 (IGF-2) gene and methylenetetrahydrofolate reductase (MTHFR) C677T gene polymorphisms in bladder transitional cell carcinoma tissues in a Chinese population.The polymorphisms of the folate metabolism enzyme gene MTHFR were studied by restrictive fragment length polymorphism (RFLP).PCR-based methods of DNA methylation analysis were used to detect the CpG island methylation status of the IGF-2 gene.The association between the methylation status of the IGF-2 gene and clinical characteristics,as well as MTHFR C677T polymorphisms,was analyzed.Aberrant hypomethylation of the IGF-2 gene was found in 68.3% bladder cancer tissues and 12.4% normal bladder tissues,respectively,while hypomethylation was not detected in almost all normal bladder tissues.The hypomethylation rate of the IGF-2 gene in cancer tissues was significantly higher in patients with lymph node metastasis than in those without lymph node metastasis (46.3% vs 17.2%,P=0.018).No association was found between aberrant DNA methylation and selected factors including sex,age,tobacco smoking,alcohol consumption and green tea consumption.After adjusting for potential confounding variables the variant allele of MTHFR C677T was found to be associated with hypomethylation of the IGF-2 gene.Compared with wildtype CC,the odds ratio was 4.33 (95% CI=1.06-10.59) for CT and 4.95 (95% CI=1.18-12.74) for TT.MTHFR 677 CC and CT genotypes might be one of the reasons that cause abnormal hypomethylation of the IGF-2 gene,and the aberrant CpG island hypomethylation of the IGF-2 gene may contribute to the genesis and progression of bladder transitional cell carcinoma.