DNA hypermethylation in papillary renal cell carcinoma

What’s known on the subject? and What does the study add? Epigenetic alterations play an essential role during carcinogenesis, and earlier studies suggested that global histone modification levels are predictive for patients’ outcome in various tumour entities (e.g. prostate, lung, breast and kidney cancer). So far, only smaller studies on histone H3K9 acetylation in patients with papillary urothelial neoplasm of low malignant potential (PUNLMP) and histone H2AX phosphorylation in non‐muscle invasive bladder cancer exist; both studies indicate prognostic relevance for bladder cancer. We demonstrate that histone methylation levels (H3K4, H4K20) decrease from normal tissue over non‐muscle invasive and muscle invasive bladder cancer to bladder cancer metastases. Histone modifications are correlated to advanced pathological stage in non‐muscle and muscle invasive bladder cancer. Furthermore, H4K20me3 appeared to be predictive for bladder cancer specific survival in patients with muscle‐invasive bladder cancer undergoing radical cystectomy.

OBJECTIVE

To investigate the pattern of DNA CpG island hypermethylation in papillary renal cell carcinoma (pRCC).

MATERIAL AND METHODS

DNA from pRCC (n= 32) and adjacent normal tissue (n= 15) was isolated. A quantitative methylation‐specific PCR was performed to analyse the methylation pattern at APC (actin beta), CDH1 (E‐cadherin), GSTP1 (glutathione S‐transferase pi 1), RASSF1A (Ras association domain family member 1A) and TIMP3 (TIMP metallopeptidase inhibitor 3); a sequence of ACTB without CpG was used to normalize for DNA input and to calculate the relative amount of methylated DNA (normalized index of methylation, NIM).

RESULTS

RASSF1A hypermethylation was observed in most pRCC and normal samples (100 vs 94.4%), but the median NIM was significantly higher in pRCC samples (2.11 vs 0.61; P < 0.001). RASSF1A hypermethylation allowed discrimination of pRCC and normal tissue with a sensitivity of 87.5% and a specificity of 73.3% as determined via receiver operator characteristic analysis (area under curve = 0.814). Hypermethylation at APC (3.0 vs 6.7%), CDH1 (15.6 vs 0%), GSTP1 (21.9 vs 6.7%) and TIMP3 (6.3 vs 0%) was infrequent in pRCC and normal tissue. CDH1 was significantly correlated with pathological stage (P= 0.015), and patients with methylated CDH1 methylation showed a trend towards shorter recurrence‐free survival (log‐rank P= 0.057). The number of methylated gene sites was correlated with pathological stage (P= 0.007) and lymph node metastasis (P= 0.008).

CONCLUSIONS

DNA hypermethylation at RASSF1A is common in pRCC tissue irrespective of the histological subtype, but also frequently seen at lower levels in normal adjacent tissue. Aberrant hypermethylation could be a prognostic marker for pRCC.     

High Histone Deacetylase 7 (HDAC7) Expression Is Significantly Associated with Adenocarcinomas of the Pancreas

Background  Alterations in HDACs gene expression have been reported in a number of human cancers. No information is available concerning the status of HDACs in pancreatic cancer tumors. The aim of the present study was to evaluate the expression levels of members of class I (HDAC1, 2,, 3), class II (HDAC4, 5, 6, and 7), and class III (SIRT1, 2, 3, 4, 5, and 6) in a set of surgically resected pancreatic tissues. Methods  Total RNA was isolated from 11 pancreatic adenocarcinomas (PA): stage 0 (n = 1), IB (n = 1), IIB (n = 6), III (n = 1), IV (n = 2), one serous cystadenoma (SC), one intraductal papillary mucinous tumor of the pancreas (IMPN), one complicating chronic pancreatitis (CP), and normal pancreas (NP) obtained during donor liver transplantation. Moreover, six other control pancreatic were included. HDACs gene expression was conducted using quantitative real-time polymerase chain reaction (qPCR). Protein expression levels were analyzed by Western blot and their localization by immunohistochemistry analyses of cancer tissues sections. Results  Remarkably, 9 of the 11 PA (approximately 81%) showed significant increase of HDAC7 mRNA levels. In contrast to PA samples, message for HDAC7 was reduced in CP, SC, and IMPN specimens. The Western blot analysis showed increased expression of HDAC7 protein in 9 out of 11 PA samples, in agreement with the qPCR data. Most of the PA tissue sections examined showed intense labeling in the cytoplasm when reacted against antibodies to HDAC7. Conclusion  The data showed alteration of HDACs gene expression in pancreatic cancer. Increased expression of HDAC7 discriminates PA from other pancreatic tumors.     

Chorionic Gonadotropin β-Subunit and Core Fragment in Bladder Cancer: mRNA and Protein Expression in Urine, Serum and Tissue

Objectives: Many transitional cell carcinomas (TCC) of the bladder express the β-subunit (CGβ) of chorionic gonadotropin (CG), and elevated serum levels occur especially in advanced disease. We have compared the diagnostic utility of various methods for detecting CG and CGβ expression at the protein and mRNA level.Methods: We used RT-PCR to detect CGβ mRNA in urinary cells and highly sensitive immunoassays to determine CG and CGβ in serum and the core fragment of CGβ (CGβcf) in urine from patients under follow-up for bladder cancer. Tissue expression was studied by immunohistochemistry.Results: CGβ mRNA was detected in urinary cells in 50% (n=84) of the cancer cases and in none of the healthy controls (n=15). Positive staining for CGβ in tissue samples was observed not only in 30% (n=96) of the TCC cases, but also in 5 of 20 histologically benign samples from TCC patients, and in 10 of 21 samples from benign bladder diseases. Serum and urinary concentrations of CGβ were elevated in 29% (n=66) and 8% (n=72), respectively, while serum CG was elevated in 18% of the TCC patients. Urinary CGβcf concentrations were higher in invasive (T1–T4) than superficial (T in situ and Ta) tumors (p=0.037), in cases positive for CGβ mRNA (p=0.03) and cases with suspicious or malignant urinary cytology (p=0.002). The ratio of urinary to serum concentration of CGβ showed the strongest correlation with tumor stage (p<0.00001), grade (p<0.00001), and staining for CGβ (p=0.019).Conclusions: Although CGβ expression may occur in benign bladder epithelium, CGβ mRNA in urinary cells is a potential marker of bladder cancer. Urinary and serum CGβ have low sensitivity in early disease, but the urine/serum ratio appears to indicate local release of CGβ into urine. Further studies are needed to evaluate the clinical usefulness of different forms of CGβ expression.     

Cyclooxygenase-2 Expression Increases with the Stage and Grade in Transitional Cell Carcinoma of the Urinary Bladder

Objectives: Experimental models of carcinogenesis show that non-steroidal anti-inflammatory drugs (NSAIDs) increase apoptosis, inhibit angiogenesis and reduce metastases. A linkage between the activity of prostaglandin synthase enzyme cyclooxygenase-2 (COX-2), a known mediator of inflammation, and cancer angiogenesis is implicated. We investigated the expression of COX-2 in bladder cancer tissue specimens using immunohistochemistry. Methods: The immunohistochemical expression of COX-2 in bladder cancer was evaluated by scoring the intensity of immunoreactivity from 0 to 3. Further, the degree of COX-2 expression was correlated with the tumor grade and depth of invasion (T stage). Result: Fifty eight percent patients (n=22) had superficial bladder tumors, while 42% (n=16) were invasive bladder cancers. Overall, COX-2 immuno-positivity was seen in 84.2% (32/38) patients. COX-2 expression was positive in 76.4% (13/17) cases with pTa tumors, 100% (5/5) of pT1 tumors, 86.6% (13/15) of pT2 tumors and in 100% (1/1) of pT3 tumor. The higher stage tumors stained more intensely; this correlation wassignificant(p=0.01987; χ2=19.6977). With reference to the grade of tumors, a positive expression was seen in 81.25% (13/16) of the low-grade tumors and 89% (17/19) of the high-grade tumors. The differential COX-2 expression relative to the grade of tumor was found to be statistically significant (p=0.05; χ2=15.8612). Conclusion: The degree of COX-2 expression is significantly increased with advancing grade and T stage of disease (p < 0.05).     

Quantification of the survival benefit of early versus deferred cystectomy in high-risk non-muscle invasive bladder cancer (T1 G3)

Objectives  To review understaging and survival of patients who underwent early versus deferred radical cystectomy (RCX) for high-risk non-muscle invasive bladder cancer (NMIBC; T1 G3). Methods  The results of 1,521 RCXs including 1,420 for bladder cancer were reviewed: (1) A total of 114 patients with high-risk NMIBC underwent a single TUR-BT followed by immediate RCX to estimate the understaging rate. (2) As much as 260 patients with NMIBC had long-term follow-up before RCX to determine the upgrading and upstaging over time. (3) We compared survival in patients with initial T1 G3 bladder cancer (BC) treated with early RCX (n = 175) versus deferred RCX (n = 99) for recurrent T1 G3. Results  (1) Our understaging rate was 20.2%. (2) Allowing NMIBC to upgrade portents a 19% survival disadvantage. (3) The 10 years cancer-specific survival rate was 78.7% in early and 64.5% in deferred RCX. Conclusions  Early, as compared to deferred RCX, has a distinct survival advantage for high-risk NMIBC. Patients should be counselled accordingly.     

Prognostic value of immunocytochemistry of the primary tumour in patients with metastatic breast cancer

The immunoreactivity of monoclonal antibodies to six tumour-associated antigens (CEA, HMFG1, HMFG2,115D8, DF3, NCRC-11) has been assessed on the primary breast tumours of 85 consecutive patients with metastatic breast cancer. The immunoreactivity of each antigen was correlated with response to endocrine therapy, probability of disease progression and post-metastases survival. No significant correlations were observed between tumour antigen immunoreactivity and any of the clinical outcomes measured. Oestrogen receptor (ER) status correlated with probability of disease progression (P=0.03) and post-metastases survival (P=0.007). This is in keeping with previous reports on ER. However, from our results immunoreactivity of tumour associated antigens in primary breast tumours is of no prognostic value after diagnosis of metastases.     

A methylation assay for the detection of non-muscle-invasive bladder cancer (NMIBC) recurrences in voided urine

What's known on the subject? and What does the study add? Multiple studies report on the detection of methylation in voided urine samples as a possible approach for the follow‐up of non‐muscle invasive bladder cancer patients. Previous studies analyze methylation gene panels in a mixture of primary and recurrent tumours. As primary tumours are larger than recurrent tumours and thus easier to detect in urine, validation of methylation markers in urine samples from patients with primary tumours will result in a test sensitivity that does not reflect the true sensitivity of the assay. This study is the first to select a subset of genes specifically methylated in non‐muscle invasive bladder cancer recurrences and validates the gene panel in two independent sets of urine samples from recurrent patients, thus simulating the disease course according to the clinical presentation.

OBJECTIVE

• ? To develop a methylation‐specific multiplex ligation‐dependent probe amplification (MS‐MLPA) assay for the detection of non‐muscle invasive bladder cancer (NMIBC) recurrences in voided urine.

PATIENTS AND METHODS

• ? Genes frequently methylated in NMIBC tumours (n= 37) were selected to develop a BC‐specific MS‐MLPA assay.
• ? Genes methylated in blood from patientswith BC (n= 29) and genes methylated in urine from patients with no history of BC (n= 46) were excluded.
• ? A four‐gene panel with the highest predictive value was selected from the initial assay. This four‐gene panel was tested and validated on urine from patients with a histologically confirmed recurrence (n= 68 test set; n= 49 validation set) and urine samples from patients without BC (n= 91, test set) and urine from recurrence‐free BC (rec‐free BC) patients (n= 60, validation set).
• ? A model was developed to predict the probability of having a recurrence based on methylation of the four‐gene panel and a threshold probability with the highest sensitivity and specificity was determined.
• ? The outcome of the model was validated on BC urine samples (n= 65) and on urine samples from rec‐free BC patients (n= 29).

RESULTS

• ? The BC MS‐MLPA assay consisted of 23 methylation probes. The selected four‐gene panel included: APC_a, TERT_a, TERT_b, and EDNRB. This panel reached an area under the receiver operating characteristic curve (AUC) of 0.82 (test set) and AUC 0.69 (validation set). Sensitivity and specificity for the detection of a concomitant tumour were 63.3% and 58.3% respectively (test set) and 72.3% and 55.2%, respectively (validation set).

CONCLUSIONS

• ? We have developed a methylation detection assay specifically for the detection of recurrences in patients with NMIBC in voided urine.
• ? The findings are promising and improvement of this test could eventually contribute to a more individualized patient friendly surveillance.

     

Association of steroid and xenobiotic receptor (SXR) and multidrug resistance 1 (MDR1) gene expression with survival among patients with invasive bladder carcinoma

What’s known on the subject? and What does the study add? SXR and MDR1 are known as responsible for chemo and radiotherapy resistance in some cancers, like kidney cancer (MDR1). Invasive bladder cancer is an aggressive disease, with different behaviour upon its tumoral stage, and also within the same tumoral stage, therefore molecular markers are sought. This study shows a new molecular marker, which has shown as a predictor for bad prognosis cancers, therefore, allowing us for a better patient selection for aggressive therapies.

OBJECTIVE

? To investigate the prognostic value of steroid and xenobiotic receptor (SXR) and multidrug resistance 1 (MDR1) gene expression in relation to survival among patients with invasive bladder cancer.

PATIENTS AND METHODS

? The prospective study included 67 patients diagnosed with invasive bladder cancer and treated with radical cystectomy at one of two institutions. ? SXR and MDR1 gene expression was assessed by real‐time quantitative polymerase chain reaction (RT‐PCR) in tumoral and normal tissue from frozen surgical specimens.

RESULTS

? Patients were followed for a mean of 29 months; 31 patients (46%) had progression. ? In univariate analysis, significant predictors of overall survival (OS) were pathological stage, lymph node (LN) status, histological grade, vascular‐lymphatic invasion, and SXR expression. ? In multivariate analysis, independent predictors of OS were LN status (odds ratio [OR], 2.96; P= 0.034), vascular‐lymphatic invasion (OR, 2.50; P= 0.029), and SXR expression (OR, 1.05, P= 0.03). ? Among the 51 patients with negative LNs (pN0), univariate predictors of OS were SXR expression, MDR1 expression, and pathological stage. In multivariate analysis, SXR expression (OR, 1.06; P= 0.01) and MDR1 expression (OR, 3.27; P= 0.03) were independently associated with survival. ? Within the pN0 group, patients with SXR expression had shorter progression‐free survival than did those without expression (P= 0.004). This association persisted in the N0 subgroup with stage pT3–pT4 disease (P= 0.028). However, in the pN1 group SXR expression did not have any influence.

CONCLUSIONS

? For patients with invasive bladder cancer, SXR expression has value as a predictor of survival independent of the standard pathological predictors. ? Its maximum importance appears to be in patients with stage pT3–pT4 pN0 disease.     

Development and evaluation of an enzyme-linked immunoassay for the prostate: specific antigen utilizing two monoclonal antibodies

Summary PSA is an important tumor-marker for prostatic cancer disease. We developed a sensitive, simple and inexpensive Sandwich ELISA for PSA with two monoclonal antibodies. The precision and reliability of the assay are reflected in the low inter-and intraassay coefficient of variation. PSA was not detectable in sera from normal females (n=50). Sera from males with different serum levels of PSA (normal males, patients with prostate hypertrophy, prostate cancer patients, n=79) and 15 prostate cancer patients treated with Zoladex were measured by our ELISA and by a commercially available RIA. The correlation coefficient between these both testsystems was close to 1 (r=0.97).     

Long-term follow-up of T1 high-grade bladder cancer after intravesical bacille Calmette-Guérin treatment

Study Type – Therapy (cohort) Level of Evidence 2b What’s known on the subject? and What does the study add? High‐grade non muscle invasive bladder cancer is a very aggressive disease, potentially lethal if not managed adequately, because of the ability of these tumours to invade surrounding tissues and become metastatic. Treatment with intravesical BCG has been shown to delay progression to muscle invasive or/and metastatic disease, preserve the bladder, and decrease the risk of death from bladder cancer. However, most studies have analyzed patients with short follow‐up, and long‐term data about the real efficacy of BCG to prevent tumour recurrence, progression and impact mortality are lacking. This study has analyzed a large series of patients with high‐grade non muscle invasive bladder cancer treated with intravesical BCG in two University Institutions (Toronto and Rotterdam), with a central pathology review by a very experienced uro‐pathologist. It provides further insight into the long‐term risks of progression of patients harbouring high‐grade T1 bladder cancer treated with BCG, demonstrating that about 30% of patients are at risk of progression and that late progressions even more than 3 years after the initial resection and BCG treatment are rare but not exceptional.

OBJECTIVE

To report the long‐term results of bacille Calmette‐Guérin (BCG) intravesical therapy in relation to disease progression and recurrence in primary T1 high‐grade (HG) bladder cancer (BC) confirmed by central pathological review.

PATIENTS AND METHODS

In all, 136 patients from two university centres (Rotterdam, n= 49; Toronto, n= 87) were diagnosed with primary T1HG BC. One experienced uro‐pathologist reviewed all slides, ensuring all cases were indeed HG and that muscle was present in all specimens. Patients were treated with BCG induction (six instillations) after transurethral resection (TUR) of the tumour and followed with cystoscopy and urinary cytology. Predictors for recurrence, progression and survival were assessed with multivariable Cox regression models.

RESULTS

Mean (range) follow‐up was 6.5 (0.3–21.6) years. There were no significant differences for recurrence (P= 0.52), progression (P= 0.35) and disease‐specific survival (DSS) (P= 0.69) between the two centres. Among the cohort, 47 patients (35%) recurred and 42 (30.9%) progressed with a median time to progression of 2.1 years; 16 (38%) of these progressions occurred ≥3 years after the initial BCG course; 22 (16%) patients who progressed died from BC. Overall, 96 (71%) patients had no evidence of disease at the last follow‐up. Carcinoma in situ was the only independent predictor for recurrence in multivariate analysis (P= 0.011). No independent predictors were found for progression.

CONCLUSIONS

Conservative treatment with BCG is a valid option in primary T1HG BC. Nevertheless, the aggressive nature of T1HG BC is evident in the fact that 30% progressed, with a high proportion of these progression events occurring ≥3 years after BCG. Caution should be exercised when relying on the long‐term effects of BCG, and close follow‐up of these patients should not be neglected.     

Continent urinary diversion and bladder augmentation in children: the Mainz pouch procedure

The formation of a bowel reservoir of large capacity at low pressure by using small and large bowel (ileocaecal segment) has proved reliable for achieving continent urinary diversion (n=80), for bladder augmentation (n=42) as well as for total bladder replacement (n=24). Encouraged by the results we obtained in our adult patients, we have used this technique during the last 3.5 years in 29 children. Indications for urinary diversions in children have been: neurogenic bladder with diplegia (n=8), bladder exstrophy (n=2), traumatic loss of the bladder (n=1), urogenital sinus (n=1) and rhabdomyosarcoma of the prostate or bladder (n=2). Bladder augmentation was indicated in 6 children with iatrogenic bladder loss, in 5 children with neurogenic bladder without diplegia and in 4 boys with incontinent epispadias or exstrophy. In children with bladder exstrophy or incontinent epispadias, continence was achieved using a modified Young-Dees technique with formation of a long intra-abdominal muscular tube made out of the bladder plate or the low-capacity bladder. The capacity of the urinary reservoir was guaranteed by bladder augmentation or bladder replacement with an ileocaecal pouch. During a mean follow-up period of 26 months (bladder augmentation) and 21 months (continent diversion) there was only one postoperative complication (intussusception ileus) which required operative revision. Two children had to undergo reoperation because of nipple problems. Follow-up, with monitoring of biochemical and metabolic parameters, is necessary to show whether this technique will provide a long-term successful solution for these problems.     

Thymidylate synthase was associated with patient prognosis and the response to adjuvant therapy in bladder cancer

OBJECTIVE

To investigate the expression of thymidylate synthase (TS), a key enzyme in DNA synthesis that is over‐expressed in several cancer cells, in bladder cancer and its association with patient prognosis and the response to adjuvant therapy.

PATIENTS AND METHODS

In all, 67 bladder tissue specimens were obtained from patients who had undergone transurethral resection (TUR). TS expression in bladder cancer and normal bladder tissue was analysed by immunohistochemistry.

RESULTS

Of the 67 bladder tissue specimens, 47 (70%) and 10 (15%) had positive expression for TS in cancer and normal tissues, respectively. TS expression was greater in patients with Grade 3 (16/17, 94%) than in Grade 1 and 2 (31/50, 64%; P = 0.002). It was also greater in Stage T1 (14/14) than in Stage Ta (33/53, 62%; P = 0.001). Furthermore, patients with negative TS expression had a longer postoperative recurrence‐free survival (RFS) than those with positive expression during the 5 year follow‐up (P = 0.028). In the patients with positive TS‐expressing tumours, adjuvant therapy significantly improved RFS (P < 0.001).

CONCLUSIONS

High TS expression might be a marker of poor prognosis for patients with bladder cancer. In addition, patients with high TS expression might also be benefit from adjuvant therapy.     

Use of the novel marker BLCA-1 for the detection of bladder cancer

PURPOSE: Bladder cancer specific nuclear structural alterations have been identified. We examined the expression pattern of one of these proteins, BLCA-1, in tissue and urine samples from individuals with bladder cancer as well as in samples from normal controls. MATERIALS AND METHODS: BLCA-1 sequence data were used to produce antibodies to this protein, which were used in immunoblot and enzyme-linked immunosorbent assays. RESULTS: BLCA-1 was detectable in tissue from patients with bladder cancer but not in normal adjacent areas of the bladder or in normal donor bladder tissue. This protein was also detectable in the urine of patients with bladder cancer by immunoblot and immunoassay. Using a cutoff of 0.025 optical density units (absorbance value) BLCA-1 was detected in 20 of 25 urine samples from patients with bladder cancer but in only 6 of 46 normal, high risk, prostate or renal cancer samples tested, resulting in a test with 80% sensitivity and 87% specificity. Expression of this protein did not appear to correlate with tumor grade. CONCLUSIONS: This research indicates that BLCA-1 is a urine based marker of bladder cancer which may be useful for the detection of this disease.     

Presence of PSA auto‐antibodies in men with prostate abnormalities (prostate cancer/benign prostatic hyperplasia/prostatitis)

Prostate‐specific antigen (PSA), produced by the prostate, liquefies post‐ejaculate semen. PSA is detected in semen and blood. Increased circulating PSA levels indicate prostate abnormality [prostate cancer (PC), benign prostatic hyperplasia (BPH), prostatitis (PTIS)], with variance among individuals. As the prostate has been proposed as an immune organ, we hypothesise that variation in PSA levels among men may be due to presence of auto‐antibodies against PSA. Sera from healthy men (n = 28) and men having prostatitis (n = 25), BPH (n = 30) or PC (n = 29) were tested for PSA antibody presence using enzyme‐linked immunosorbent assay (ELISA) values converted to standard deviation (SD) units, and Western blotting. Taking ≥2 SD units as cut‐off for positive immunoreactivity, 0% of normal men, 0% with prostatitis, 33% with BPH and 3.45% with PC demonstrated PSA antibodies. One‐way analysis of variance (anova ) performed on the mean absorbance values and SD units of each group showed BPH as significantly different (P < 0.01) compared with PC and prostatitis. All others were nonsignificant (P < 0.05). Men (33%) with BPH had PSA antibodies by ELISA and Western blot. These discoveries may find clinical application in differential diagnosis among prostate abnormalities, especially differentiating BPH from prostate cancer and prostatitis.     

Endothelin-1 in children with chronic renal failure

Endothelin-1 (ET-1) was meansured after extraction from plasma of normal adults (5.9±1.9 pg/ml,n=22), normal children (7.1±1.86 pg/ml,n=29), nonhaemodialysed children with chronic renal failure (CRF) (11.1±1.8 pg/ml),n=10), renal graft recipients (9.5±3.4 pg/ml,n=37), haemodialysed children 24 h after a haemodialysis session (20.02±10.9 pg/ml,n=26) and haemodialysed children before and after a haemodialysis session (15.31±10.6 and 13.8±8.5 respectively,n=14). A sensitive and specific radioimmunoassay was used. ET-1 was significantly higher in non-haemodialysed CRF children and in renal graft recipients than in normal children (P<0.001 andP<0.01, respectively) and significantly higher in haemodialysed children when compared with normal children, non-haemodialysed CRF children and renal graft recipients (P<0.001). ET-1 concentrations were similar in normal children and normal adults. ET-1 was inversely correlated with glomerular filtration rate in non-haemodialysed CRF children (r=–0.39,P<0.01) and positively correlated with extracellular volume in haemodialysed children (r=0.435,P<0.03). After haemodialysis, ET-1 increased in 6 and decreased in 8 of the 14 children studied before and after a haemodialysis session.     

Involvement of the insulin-like growth factor I receptor and its downstream antiapoptotic signaling pathway is revealed by dysregulated microRNAs in bladder carcinoma

ObjectiveUrothelial carcinoma is one of the most common pathological types of bladder cancer. Several studies have shown that dysregulated microRNAs (miRNAs) play an important role in bladder cancer progression. We performed the present miRNA microarray analysis in samples of urothelial carcinoma of the bladder and adjacent normal bladder tissue from Taiwanese patients to investigate dysregulated miRNAs.Materials and methodsTo detect dysregulated miRNAs in urothelial carcinoma of the bladder, samples of tumor and adjacent normal tissues were collected from 10 patients. Tissue samples from three patients were subjected to miRNA microarray analysis, and the remaining tissue samples from the other seven patients were used to validate the results obtained from the microarray data. Potential targets of these dysregulated miRNAs were identified using online databases, including MicroCosm and TargetScan.ResultsA panel of 30 differentially expressed miRNAs with at least fourfold differences in expression compared with normal controls, including 19 upregulated and 11 downregulated miRNAs, was generated. The expression levels of miR-30a-5p, miR-30a-3p, miR-99a, miR-130b, miR-133b, miR-135b, miR-145, miR-195, miR-204, and miR-214 were experimentally verified using real-time RT-PCR analysis. Using an online miRNA target database, we discovered that these dysregulated miRNAs potentially control components of the insulin-like growth factor 1 receptor (IGF1R) signaling pathway.ConclusionOur results indicate that dysregulated miRNAs may be involved in bladder cancer pathogenesis and are potential biomarkers.     

Pituitary‐tumour‐transforming‐gene 1 expression in testicular cancer

Genomic instability is a feature of germ cell tumours. The pituitary‐tumour‐transforming‐gene 1 (PTTG1) is the major effector of chromosome segregation during mitosis, protecting the cell from aneuploidy. The protein expression of this gene has been evaluated in testicular tumours by immunohistochemistry. Formalin‐fixed and paraffin‐embedded specimens of testicular tissues from 83 patients undergoing therapeutic orchidectomy for seminomas (n = 53), embryonal carcinoma (n = 10), yolk sac tumour (n = 10) and teratoma (n = 10) were examined. Seminoma was associated with in situ carcinoma (CIS) in 23 samples. PTTG1 immunostaining was performed using rabbit anti‐PTTG1 as a primary antibody. In CIS, only isolated cells showed nuclear staining for PTTG1. In the peripheral area of seminoma, PTTG1 was mostly detected as localised in the nucleus; in the central area of seminoma, PTTG1 staining was more intense in cytoplasm. PTTG1‐positive cells were also present in the areas of seminoma infiltration. On the other hand, in embryonal carcinoma, cells had a diffuse positive immunostaining, mainly cytoplasmatic, while we did not observe an expression of PTTG1 in yolk sac tumour and mature teratoma. We firstly identified the PTTG1 expression pattern in normal testis, CIS and testicular cancer. Further investigation is needed to clarify the functional activity of PTTG1 in testicular oncogenesis.     

Human urinary bladder-carcinoma cells are non-osteoinductive

Summary Small pieces (ca. 2–3×3–5 mm) of the urinary-tract mucosa from noninvasive papillary transitional-cell carcinomas of the bladder (ca. urotheliale papillare, n=33), invasive transitional-cell carcinomas of the bladder (ca. urotheliale papillare infiltrans, n=6, papillary transitional-cell carcinomas of the bladder with squamous metaplasia (ca. urotheliale papillare cum metaplasia planoepitheliale, n=4), transitional-cell carcinomas in situ (ca. urotheliale in situ, n=2), and squamous-cell carcinomas of the bladder (ca. planoepitheliale, n=2) were grafted intramuscularly into cortisone-immunosuppressed mice to test the ability of transformed transitional epithelium to induce heterotopic osteogenesis. Altogether, 156 implants from 47 cases of urinary bladder carcinoma were performed. Histological examination of implants, excised 10–17 days later, revealed relatively good survival of the grafted epithelium, which had proliferated and, in some cases, formed cysts and islands but failed to induce heterotopic osteogenesis in the surrounding host tissues. In nine implants prepared from four cases (noninvasive papillary transitional-cell carcinoma of the bladder and invasive papillary transitional-cell carcinoma of the bladder, two cases each) a small amount of cartilage and/or bone was found in the stroma of grafted tissue. The rarity of this phenomenon — together with the observation that implants of normal human urinary-tract mucosa have never induced the formation of cartilage/bone, whereas in a similar system, dog or guineapig grafts are osteogenic — suggests that the cartilage/bone present in the stroma of implanted cancers is the result of metaplasia of the stroma of the neoplasm and not the product of any osteoinductive potency of human urothelium.     

Immunohistochemical evidence of canine morbillivirus (canine distemper) infection in coatis (Nasua nasua) from Southern Brazil

The pathological and immunohistochemical (IHC) findings associated with infection due to canine morbilivírus (canine distemper virus, CDV) are described in coatis (Nasua nasua). Tissue fragments of coatis (n = 13) that died at the Bela Vista Sanctuary, Paraná, Southern Brazil, were routinely processed for histopathology to identify the main histopathologic patterns as compared to that of the domestic dog. Selected formalin‐fixed paraffin‐embedded (FFPE) tissue fragments of the lungs, liver, urinary bladder and small intestine were used in IHC assays designed to identify the antigens of CDV, canine adenovirus (CAdV‐1 and CAdV‐2) and canine parvovirus type 2 (CPV‐2). The main histopathologic patterns identified were interstitial pneumonia (n = 9), interstitial nephritis (n = 6), atrophic enteritis (n = 4) and ballooning degeneration of the uroepithelium (n = 3). Positive immunolabelling for intralesional antigens of CDV was identified in the lung with interstitial pneumonia (n = 3), in the intestine (n = 2) and in the degenerated epithelium of the urinary bladder (n = 2). Antigens of CPV‐2, CAdV‐1 and CAdV‐2 were not identified in any FFPE tissue sections evaluated. These findings indicate that these wild carnivores were infected by a viral disease pathogen common to the domestic dog and develop similar histopathologic findings. Collectively, these findings suggest that these coatis were infected by CDV and can serve as a potential host for this infectious disease pathogen.