Nilay Sen Turk

No Strong Association Between HER-2/neu Protein Overexpression and Gene Amplification in High-grade Invasive Urothelial Carcinomas

The generation of urothelial carcinoma is caused by the accumulation of various molecular changes, as in most malignancies. There are conflicting data about the status of HER-2/neu oncogene in urothelial carcinomas. The aim of this study was to determine the status of HER-2/neu oncogene in high-grade invasive urothelial carcinoma of urinary bladder both in protein and DNA level. We evaluated HER-2/neu protein overexpression by immunohistochemistry (IHC) and gene amplification by fluorescent in situ hybridization (FISH) and real-time quantitative PCR in paraffin-embedded samples of high-grade invasive urothelial carcinoma obtained from 36 patients. Polysomy 17 was also assessed by FISH. Immunohistochemically, HER-2/neu protein overexpression was observed in 22 (61.1%) tumors (ten tumors with score 3+ and 12 with score 2+). Fourteen of 36 tumors (38.9%) were evaluated as negative (score 0 or 1+). Complete concordance between FISH and the PCR was seen in all of the samples scored as 0 and 1+ by IHC. HER-2/neu gene amplification was observed in three of 27 (11.1%) tumors by FISH (nine samples were non-informative) and in eight of 36 (22.2%) tumors by the PCR. The complete concordance between HER2-2/neu protein overexpression and gene amplification was seen only in three of 27 tumors. Polysomy 17 was seen in nine tumors (33.3%). The results indicated that, in contrast to breast cancer, there was no strong association between HER-2/neu overexpression and gene amplification in invasive urothelial carcinomas, and polysomy 17 was higher in tumors showing HER-2/neu overexpression.

Effects of Tempol on Experimental Acute Necrotizing Pancreatitis Model in Rats

ABSTRACT Aim: We aimed to investigate the effects of Tempol on local organ damage in an experimental acute pancreatitis model. Methods: This experimental study was conducted on 40 male Wistar- albino rats. The animals were randomly allocated into four groups: (i) Sham-operated group, laparotomies and cannulations of the pancreatic duct without acute necrotizing pancreatitis (ANP) (n=10); (ii) Sham + Tempol group, identical to group 1 except for intravenous tempol treatment for 4 hours (n = 10); (iii) ANP group, glycodeoxycholic acid was infused into the pancreatic duct and cerulein was infused intravenously for 6 hours for development of ANP (n=10); and (iv) ANP + Tempol treated group, in addition to the procedure in group 3, rats were administered tempol intravenously for 4 hours (n = 10). Injury of the pancreas was evaluated histopathologically. Malondialdehyde and myeloperoxidase levels of the pancreatic tissue, blood gas analysis, leukocyte and hematocrit levels were measured. Wet/dry weight of pancreatic tissue was also measured. Results: Serum amylase levels, pancreatic tissue malondialdehyde and myeloperoxidase levels, wet/dry weight ratio, pancreatic edema, acinar necrosis, fat necrosis and hemorrhage, inflammation and perivascular infiltration were significantly lower in the ANP + Tempol group compared with the ANP group. Conclusion: Tempol infusion reduced local organ damage due to acute necrotizing pancreatitis in this experimental study. These findings demonstrate that tempol has protective effects on local organ damage due to acute necrotizing pancreatitis in rats.

Expression and Amplification of Topoisomerase-2α in Type 1 and Type 2 Papillary Renal Cell Carcinomas and Its Correlation with HER2/neu Amplification

The current study was undertaken to investigate chromosomal and genetical aberrations leading to overexpression of Topoisomerase-2α (TOP2α) and to reveal the possible association of these aberrations with HER2/neu overexpression and gene amplification, and to search for the relationship between TOP2α and HER2/neu status with prognostical biomarkers in papillary renal cell carcinoma (RCC), a group of tumors with diverse molecular, chromosomal and clinical features. Archival cases of papillary RCC obtained from Departments of Pathology of Pamukkale, Ege and Dokuz Eylul Universities were studied in two groups (type 1 and type 2) each containing 20 cases. The level of TOP2α and HER2/neu expression by tumor cells were determined immunohistochemically. A multicolor FISH probe was used to define both amplification of HER2/neu and TOP2α genes, and polysomy 17. The ratio of cells expressing TOP2α in type 1 and type 2 papillary RCC were 24.29% and 6.89%, respectively. The difference was statistically significant comparing the average or median values of groups separately (p = 0.002). The expression levels of TOP2α and HER2/neu were also correlated. TOP2α and HER2/neu were co-amplified in both groups. Immunohistochemical expression was not observed in 15 of 23 cases with HER2/neu amplification. The most frequent finding detected by FISH method was polysomy of chromosome 17. We had contradictory results compared with the findings reported in the limited numbers of literature. It shows us that papillary RCC constitute a heterogenous group of tumors with various cytogenetic features and morphological classification of these tumors may not be compatible with their molecular characteristics.

Determination of apoptosis, proliferation status and O6-methylguanine DNA methyltransferase methylation profiles in different immunophenotypic profiles of diffuse large B-cell lymphoma

OBJECTIVE Our aim was to investigate the expression of apoptosis-associated proteins (bcl-2, bcl-xl, bax, bak, bid), apoptotic index (AI) and proliferation index (PI) in germinal center B-cell-like immunophenotypic profile (GCB) and non-GCB of diffuse large B-cell lymphoma (DLBCL). METHODS The methylation status of the promoter region of O6-methylguanine-DNA yerine O6-methylguanine-DNA methyltransferase (MGMT) gene and its relation with immunophenotypic differentiation of DLBCLs were also investigated. 101 cases were classified as GCB (29 cases) or non-GCB (72 cases). Apoptosis-associated proteins and PI were determined by IHC, and TUNEL method was used to determine AI. MGMT methylation analysis was performed by real-time PCR. RESULTS The PI was significantly higher in GCB compared with non-GCB (p=0.011). Percentage of cells stained with bcl-6 was positively correlated with the percentage of cells expressing bcl-2 (p=0.023), AI (p=0.006) and PI (p<0.001), while a significant negative correlation was observed with the percentage of cells expressing bax (p=0.027). The percentage of cells stained with MUM1 showed a significantly positive correlation with the percentage of cells expressing bcl-xl (p=0.003), bid (p=0.002), AI (p<0.001), and PI (p=0.001). MGMT methylation analysis was performed in 95 samples, and methylated profile was found in 31 cases (32.6%). GCB was found in 6 cases (22.2%) and non-GCB was determined in 25 cases (36.8%) out of 31 with MGMT methylated samples. There was no significant association between MGMT methylation status and immunophenotypic profiles (p=0.173). CONCLUSION These results suggest that bcl-6 protein expression may be responsible for the high PI in GCB. Additionally, we found that apoptosis-associated proteins were not significantly associated with immunophenotypic profiles.

Fine‐needle aspiration biopsy of an invasive ductal carcinoma with medullary features presented with abscess formation

Dear Dr. Bedrossian: We point out that breast carcinomas infrequently appear as cystic masses, which may mimic an abscess formation clinically and cytologically. A 38-year-old woman was presented with a breast mass for 2 weeks. In her physical examination, a palpable, tender breast mass was found within the upper outer quadrant of the right breast without axillary lymphadenopathy. Mammography showed a suspicious microcalcified mass with slightly irregular margins (3 cm in diameter). An ultrasound of the breast revealed a cystic lesion with rough papillary projections. Fine-needle aspiration biopsies (FNABs) from solid and cystic areas of the mass were performed. The smears showed syncytial groups of atypical cells (nuclear pleomorphism, hyperchromasia, nuclear contour irregularity but no macronucleoli) in a background composed of prominent neutrophils and sparse plasma cells (Fig. 1). Tubular formation was also observed in some groups of tumor cells. A diagnosis of malignant cytology consistent with high-grade ductal carcinoma was rendered. The patient, then, underwent breast conservative surgery with sentinel lymph node dissection and was diagnosed as invasive ductal carcinoma with medullary features. No axillary lymph node metastasis was identified. Breast carcinomas are rarely seen clinically as cystic masses. Cystic breast carcinomas comprise intracystic papillary carcinoma, solid tumors infiltrating benign cysts, or cystic degeneration of solid tumors. FNABs from these lesions may reveal varying amounts of neutrophils, which may obscure the tumor cells and mimic abscess formation easily. Therefore, smears with abundant neutrophils in the background should be considered with caution. The differential diagnosis of a breast carcinoma from an abscess is important. Both breast carcinomas and abscess may cause skin retraction, erythema of breast skin, and tenderness. A carcinoma can occur in any site of the breast, whereas breast abscess mostly is confined to subareolar region with a previous history of multiple abscesses in the same area. Breast abscess may have sparse epithelial cells with regenerative atypia on FNAB. However, ultrasound-directed FNAB from breast mass may better reveal groups of many carcinoma cells. High-grade invasive ductal carcinomas with or without medullary features, medullary carcinomas, or pure squamous cell carcinomas have been reported as cystic degenerative masses with extensive neutrophils on their smears. Akbulut et al. reported cytologic features in FNAB from 20 histologically verified medullary breast carcinomas. Only 1 of these 20 cases appeared radiologically as a partly cystic, solid mass (as in our case) and the rest as solid circumscribed masses. However, the presence of neutrophils was observed in 11 of the 20 cases, 3 of which strongly suggested abscess formation. This may show that abundant neutrophils may exist in the background of smears even if the tumors include no cystic components. Moreover, not only a prominent lymphoplasmacytic background but also a neutrophilic infiltrate may be seen in medullary carcinomas. Kleer and Michael reported cytological features of breast carcinomas with prominent lymphocytic infiltrate in 18 cases. Of these 18 cases, 9 were invasive ductal carcinomas and 9 were medullary carcinomas (6 typical and 3 atypical). A prominent neutrophilic infiltrate was observed in one of typical medullary carcinoma cases. *Correspondence to: Ender Duzcan, M.D., Department of Pathology, Pamukkale University School of Medicine, Morfoloji Binasi, Kınıkli 20070, Denizli, Turkey. E-mail: eduzcan@pau.edu.tr Received 26 August 2009; Accepted 6 October 2009 DOI 10.1002/dc.21266 Published online 15 January 2010 in Wiley InterScience (www. interscience.wiley.com).

Characterizing the Association Between Toll-like Receptor Subtypes and Nephrolithiasis With Renal Inflammation in an Animal Model

OBJECTIVE To show experimentally induced renal stone disease and to evaluate secondary inflammatory responses in vivo, and to characterize changes in the expression of Toll-like receptor (TLR) subtypes in this model. METHODS Twenty 5- to 6-week-old male Wistar rats were divided into control and hyperoxaluria groups (n = 10 per group) and were supplied with normal water or 1% ethylene glycol, respectively, for 16 weeks. The animals were then placed in metabolic cages, and urine was collected for a 24-hour urine oxalate level evaluation. Following sacrifice, rats were subjected to bilateral nephrectomy and both kidneys were histopathologically evaluated. A 1-mm3 biopsy section from the right kidney of each rat was subjected to real-time polymerase chain reaction of the TLR expression. RESULTS At the end of week 16, the hyperoxaluria group had a higher mean 24-hour urine oxalate level (1.91) than the control group (0.29) (P <.05) and a remarkably increased deposition of renal CaOx crystals (15/20) than the control group (0/20) (P <.05), which was universally accompanied by inflammation (15/15). Twelve and no rats in the hyperoxaluria and control groups, respectively, had macroscopically visible renal pelvic stones (P <.05). Quantitative real-time polymerase chain reaction revealed significant decreases in the expression of several TLRs, particularly TLR11 and TLR7. Decreases in TLR1, TLR3, and TLR6 expressions and an increase in the TLR2 expression did not differ significantly between the groups. CONCLUSION We believe that is the first evaluation of TLR expression associated with renal stone formation in an animal model of inflammation. These results might lead to novel TLR-based treatments for nephrolithiasis and related inflammatory renal damage.

Human TLR gene family members are differentially expressed in patients with urothelial carcinoma of the bladder

PURPOSE Toll-like receptors (TLRs) have an important role in the activation of both innate and adaptive immunity in response to pathogens and endogenous danger signals from damaged or dying cells. The aim of this study was to determine the relationship between urothelial carcinoma (UC) and TLR expression. BASIC PROCEDURES Real-time polymerase chain reaction evaluation was made of the messenger RNA expression of TLRs 1-10 in 24 UC samples and 46 nontumoral bladder tissue samples. The levels of proinflammatory cytokines (IL-1β, IL-6, and IL-8) in the urine samples were also determined with enzyme-linked immunosorbent assay. MAIN FINDINGS TLR2-7 and TLR10 expressions were significantly higher in UC than in the control group (P<0.05 for all comparisons). No concordance was found between matched tumor tissue and urine samples in terms of TLR expression. IL-1β, IL-6, and IL-8 levels were significantly higher in urine specimens of patients with UC (P = 0.033, P = 0.001, and P = 0.008, respectively). PRINCIPAL CONCLUSIONS The results of this study demonstrated that the TLR gene expression profiles reflect the heterogeneity within UC. These results might also prompt further investigation to better understand the role of the TLR gene family expression in the tumor progression of UC.