Prognostic Significance of Tumor Tissue NeuGcGM3 Ganglioside Expression in Patients Receiving Racotumomab Immunotherapy.

Expression of N-glycolyl GM3 (NeuGcGM3) ganglioside was detected in the tumor specimens of patients who were on Racotumomab anti-idiotype vaccine maintenance treatment, and prognostic significance as a biomarker was investigated. No statistically significant association was observed in the multivariate analysis between overall survival and tissue NeuGcGM3 IHC levels. Although numerically there was a difference favoring less intense IHC for better prognosis, this did not reach statistical power. However, there was a strong correlation between Racotumomab doses and overall survival (OS). Mean OS of the patient with more than 10 Racotumomab application was significantly longer than the patient who had less than 10 injections (70.7 months vs. 31.1 months, p < 0.001). We propose that, regardless of staining intensity, the presence of NeuGcGM3 in patient tissues might be an indicator of benefit in Racotumomab treatment.

Role of gamma probe-assisted intraoperative sentinel lymph node evaluation in predicting axillary breast cancer metastasis after neoadjuvant chemotherapy.

OBJECTIVE: Intraoperative sentinel lymph node biopsy is a universally accepted technique to identify patients who are candidates for axillary lymph node dissection during breast cancer surgery. However, there is controversy over its use in patients who underwent preoperative neoadjuvant chemotherapy. This study aimed to examine the diagnostic value of gamma probe-assisted intraoperative sentinel lymph node examination with frozen section in breast cancer patients who had undergone preoperative neoadjuvant chemotherapy. METHODS: This retrospective study included 94 tumors diagnosed with stage IIA, IIB or IIIA invasive breast cancer with locoregional lymph node metastasis who underwent surgical treatment after neoadjuvant chemotherapy. Intraoperatively, axillary sentinel lymph node sampling was done using radioactive colloid and gamma probe and materials were examined with frozen section method. Patients with positive sentinel nodes underwent axillary resection. Histopathological examination of all surgical samples was done postoperatively. RESULTS: In 87 of 94 tumors (92.6%), a sentinel lymph node could be identified using the method. The sensitivity, specificity and accuracy of the method for predicting axillary macro metastasis were 85.7, 86.5 and 86.2%, respectively, with 5.7% false negative rate. CONCLUSIONS: Sentinel lymph node identification using preoperative scintigraphy and intraoperative use of gamma probe seems to be a feasible and efficient method in terms of differentiating patients that require axillary lymph node dissection during breast cancer surgery, even when they have received neoadjuvant chemotherapy. Further large prospective studies allowing subgroup analyses are warranted.

Plexin C1 Marks Liver Cancer Cells with Epithelial Phenotype and Is Overexpressed in Hepatocellular Carcinoma.

Background and Aims: Hepatocellular carcinoma is an aggressive malignancy of the liver and is ranked as the sixth most common cancer worldwide. There is still room for novel markers to improve the diagnosis and monitoring of HCC. Our observations in cancer databases that PLXNC1 is upregulated in HCC led us to investigate the expression profile of Plexin C1 mRNA and protein in HCC cell lines and tissues. Methods: A recombinant protein encompassing part of the extracellular domain of Plexin C1 was used as an antigen for monoclonal antibody development. Transcript and protein levels of Plexin C1 in HCC cell lines were determined by RT-qPCR and Western blotting, respectively. In vivo evaluation of Plexin C1 expression in HCC tissues was accomplished by immunohistochemistry studies in tissue microarrays. Results: A monoclonal antibody, clone PE4, specific to Plexin C1, was generated. In silico and in vitro analyses revealed a Plexin C1-based clustering of well-differentiated HCC cell lines. Staining of HCC and nontumoral liver tissues with PE4 showed a membrane-localized overexpression of Plexin C1 in tumors (p=0.0118). In addition, this expression was correlated with the histological grades of HCC cases. Conclusions: Plexin C1 distinguishes HCC cells of epithelial characteristics from those with the mesenchymal phenotype. Compared to the nontumoral liver, HCC tissues significantly overexpress Plexin C1. The newly generated PE4 antibody can be evaluated in larger HCC cohorts and might be exploited for the examination of Plexin C1 expression pattern in other epithelial malignancies.

Histologic Evidence of New Collagen Formulation Using Platelet Rich Plasma in Skin Rejuvenation: A Prospective Controlled Clinical Study.

BACKGROUND: Platelet-rich plasma (PRP) is an autologous concentration of human platelets contained in a small volume of plasma and has recently been shown to accelerate rejuvenate aging skin by various growth factors and cell adhesion molecules. OBJECTIVE: This study was conducted to evaluate the efficacy and safety of intradermal injection of PRP in the human facial rejuvenation. METHODS: This study was a prospective, single-center, single-dose, open-label, non-randomized controlled clinical study. PRP injected to the upper site of this right infra-auricular area and all face. Saline was injected to the left infra-auricular area. Histopathological examinations were performed before PRP treatment, 28 days after the PRP, and saline (control) treatments. RESULTS: Twenty women ranging in age from 40 to 49 years (mean age, 43.65±2.43 years) were enrolled in the study. The mean optical densities (MODs) of collagen in the pre-treatment, control, and PRP-treated area were measured. They were 539±93.2, 787±134.15, 1,019±178, respectively. In the MOD of PRP, 89.05 percent improvement was found when MOD of PRP was compared with MOD of pre-treatment. The mean MOD of collagen fibers was clearly highest on the PRP side (p<0.001). The PRP-to-saline improvement ratio (89.05% to 46.01%) was 1.93:1. No serious side effects were detected. CONCLUSION: PRP increases dermal collagen levels not only by growth factors, but also by skin needling (the mesotherapy technique 'point by point'). PRP application could be considered as an effective (even a single application) and safety procedure for facial skin rejuvenation.

A rare cause of mechanical intestinal obstruction: Pharmacobezoar.

A bezoar is a hard, and solid, foreign body located in the gastrointestinal tract that may recur. Bezoar is classified according to its origin. Pharmacobezoars develop in the gastrointestinal tract due to alterations in anatomical structure and/or intestinal motility. In this paper, a case, not yet defined in the literature, of a pharmacobezoar causing a mechanical obstruction that is accompanied by a malignancy in the colon is reported, with the aim of contributing to the literature.

Clinical significance of miR-140-5p and miR-193b expression in patients with breast cancer and relationship to IGFBP5.

The functional role of IGFBP5 in breast cancer is complicated. Experimental and bioinformatics studies have shown that IGFBP5 is targeted by miR-140-5p and miR-193b, although this has not yet been proven in clinical samples. The aim of this study was to evaluate the expression of miR-140-5p and miR-193b in breast cancer and adjacent normal tissue and assess its correlation with IGFBP5 and the clinicopathological characteristics of the tumors. IGFBP5 protein expression was analyzed immunohistochemically and IGFBP5, miR-140 and miR-193b mRNA expression levels were analyzed with real-time RT-PCR. Tumor tissue had higher miR-140-5p expression than adjacent normal tissue (p = 0.015). Samples with no immunohistochemical staining for IGFBP5 showed increased miR-140-5p expression (p = 0.009). miR-140-5p expression was elevated in invasive ductal carcinomas (p = 0.002), whereas basal-like tumors had decreased expression of miR-140-5p compared to other tumors (p = 0.008). Lymph node-positive samples showed an approximately 13-fold increase in miR-140-5p expression compared to lymph node-negative tissue (p = 0.049). These findings suggest that miR-140-5p, but not miR-193b, could be an important determinant of IGFBP5 expression and clinical phenotype in breast cancer patients. Further studies are needed to clarify the expressional regulation of IGFBP5 by miR-140-5p.

EGFR and KRAS mutations in Turkish non-small cell lung cancer patients: a pilot study.

EGFR and KRAS mutation profile in non-small cell lung cancers (NSCLCs) shows wide variations due to geographic and ethnic background. We aimed to determine the frequency and types of EGFR and KRAS mutations in a sample group of Turkish NSCLC cases. The study included 14 adenocarcinomas (ACs), 11 squamous cell carcinoma (SCC) patients selected from archival material including small biopsy or surgical specimens. Their formalin fixed paraffin-embedded tumor tissues were used for genomic DNA extraction for EGFR exon 19 and 21, and KRAS exon 2 mutations. Eleven NSCLCs (44 %) had EGFR mutations. Exon 19 and 21 mutations were found in 8 (32 %) and 5 (20 %) cases. Two cases showed double EGFR mutations. In ACs, 5 (35.7 %) patients had EGFR gene mutation, 3 in exon 19 and 3 in exon 21. In SCCs, 6 (54.5 %) cases had EGFR mutation, 5 in exon 19 and 2 in exon 21. All exon 19 mutations were deletion-type mutations. For exon 21, 3 cases had L858R point mutation (CTG>CGG) and two cases showed deletion-type mutations. Six (24 %) NSCLCs showed KRAS mutations (three ACC, three SCC), 5 codon 12 mutations (G>T, T>C, G>A) and one codon 13 mutation (G>T). Three NSCLC cases showed both EGFR and KRAS mutations together. The profile of KRAS mutation in our AC cases was quite similar to those seen in the Western countries; however, frequency and clustering of EGFR mutations were similar to those seen in the Eastern countries.

MicroRNA-21 as an indicator of aggressive phenotype in breast cancer.

OBJECTIVE: The recently discovered microRNAs (miRNAs) are non-protein-coding, endogenous small RNAs. The aim of this study was to investigate the relationship between microRNA-21 (miR-21) and the clinicopathological features of breast cancer. MATERIALS AND METHODS: Formalin-fixed, paraffin-embedded (FFPE) tissue samples were collected from 15 patients who had undergone surgery for primary breast cancer. The miR-21 expression levels in normal and cancer tissues were analyzed using real-time quantitative reverse transcriptase polymerase chain reaction (real-time qRT-PCR). The correlations between the miR-21 expression level and the stage of disease, the tumor size, lymph node involvement, hormone receptor status, human epidermal growth factor receptor 2 (HER2) status, and disease-free survival (DFS) were investigated. RESULTS: The miR-21 expression levels were significantly higher in patients with stage III disease, patients with more than 3 axillary lymph node metastases and HER2-positive patients than in patients with stage I-II disease, patients with 1-3 axillary lymph node metastases and HER2-negative patients (p = 0.005, p = 0.037, and p = 0.014, respectively). Patients with high miR-21 expression level had a significantly shorter DFS than patients with low miR-21 expression level (median DFS: 18 and 56 months, respectively; p = 0.004). CONCLUSION: These results show that miR-21 is an indicator of an aggressive breast cancer phenotype and that it may be a new therapeutic target in the treatment of breast cancer in the future.

[A new remedy in pathology practice: molecular solution to sample mix-up]. / Patoloji Pratiginde Yeni Bir Çare Uygulamasi: Örnek Karisikligina Moleküler Çözüm.

OBJECTIVE: Although very rare, suspicious situations about the identity of diagnostic tissue material have been encountering in pathology practice. Such situations undoubtedly have the potential to create undesirable results. In the present study, an application targeting getting rid of any doubts about the identity of the diagnostic tissue samples is described. MATERIAL AND METHOD: A combination of short tandem repeats (STR) of the human genome consisting of CSF1PO, TH01, TPOX, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539 and Penta E were selected on the basis of ease of application and bioinformatic discrimination power. Possible forms of diagnostic tissue mix up were set in 3 different models with 3 diagnostic tissue samples of 2 different cases. Of the tissue samples selected, A (salivary gland) and B (striated muscle) belonged to the same case and C (uterus wall) belonged to another case. In the first model, there was no problem about tissue identity (M1: A/B). In the second model, two different diagnostic material were mixed up (M2: B/C). In the last model, there were 3 diagnostic material obtained from 2 different cases (M3: A/B/C). DNA was extracted from all tissue samples and all of the selected 10 STR were amplified with specially designed primers by PCR. After chemical denaturation, amplicons were submitted to polyacrylamide gel electrophoresis for discrimination of single DNA strands according to their conformation polymorphism (SSCP). Special patterns of each STR in the gel matrix obtained from M1, M2 and M3 models, were evaluated on the principle of being 'same or different' to determine the diagnostic material identity. RESULTS: Each of the salivary gland, striated muscle and uterus wall samples were correctly identified (matched with the right source cases) after evaluating 10 different STR SSCP patterns designed under M1, M2 and M3 models. CONCLUSION: This application targeting to solve diagnostic tissue identity problems is a simple and cheap application of SSCP and its efficacy was proven on the designed models.

Stromal cells do not share KRAS mutations that the epithelial component harbors in colorectal carcinomas.

Epithelia and stroma are the main components of carcinomas that have impact on carcinogenesis. Many of the genetic changes harboring in the epithelia may possibly be seen in stromal cells during neoplastic transformation. We intended to investigate weather KRAS mutations are shared by the stromal cells in colorectal adenocarcinomas. Forty cases with KRAS mutation were studied. Glandular/epithelial and the stromal components of each primary tumor were collected and KRAS mutation analysis was performed. None of the cases revealed KRAS mutations in stromal integral. We concluded that stromal cells in colorectal carcinoma do not share KRAS mutations that the epithelial component harbors.

Investigation of human papillomavirus DNA in colorectal carcinomas and adenomas.

Human papillomavirus (HPV) has been considered to be an etiological agent for anogenital cancers, such as cervical cancer and possibly a subset of cancers of the aerodigestive tract. The aim of the study was to evaluate the presence of human papillomavirus DNA in colorectal carcinomas and adenomas. Formalin-fixed and paraffin-embedded archival tissue samples were used for DNA extraction. One hundred and six colorectal carcinomas and 62 adenomas were screened by nested polymerase chain reaction (PCR) for HPV DNA with a control group of 49 cervical tissues with invasive cervical carcinoma and cervical intraepithelial neoplasia (CIN). In the study group, we did not find HPV DNA positivity in any of all the colorectal carcinomas and adenomas. In the control group with cervical lesions, 34 out of 49 (69.4%) samples were positive for the HPV DNA. These results indicated that there was no correlation between HPV infection and colorectal carcinomas and adenomas.

Role of human papillomavirus in the development of urothelial carcinoma.

It has been estimated that almost 10% of the worldwide cancer burden is linked to human papillomavirus (HPV) infection. Although the association between HPV and bladder carcinoma has been extensively investigated, data on the role of HPV in bladder carcinogenesis are controversial. The aim of the study was to assess the possible role of human papillomavirus in the development of urothelial bladder carcinomas. Formalin-fixed and paraffin-embedded archival tissue samples were used for DNA extraction. Seventy urothelial bladder carcinoma tissues were screened by nested-polymerase chain reaction (PCR) for HPV DNA with a control group of total 18 cervical tissues with invasive cervical carcinoma and cervical intraepithelial neoplasia III (CIN III). In the study group, we did not find HPV DNA positivity in any of the urothelial carcinomas. In the control group, 15 out of 18 (83.3%) samples were positive for the HPV DNA. These results indicated that there was no association between HPV infection and urothelial carcinomas.

A valuable adjunct to FNA diagnosis of papillary thyroid carcinoma: in-house PCR assay for BRAF T1799A (V600E).

The diagnostic approach to thyroid nodules generally starts with FNA cytology. However, approximately one-fifth of cytologic evaluations yield indeterminate cytological findings but only 20% of cases with indeterminate thyroid nodule cytology have a cancer diagnosis, emphasizing the need for an effective ancillary test based on FNA material to help prevent unnecessary surgery. Detection of BRAFV600E mutation, the genetic signature of papillary thyroid carcinoma (PTC) in FNA material provides an invaluable diagnostic adjunct to overcome the limitations of FNA cytology. There are many ways to detect V600E, such as direct DNA sequencing, allele-specific PCR and hybridization-based colorimetric methods. In this study, a newer simple PCR method is presented that removes requirements for sequencing special equipment and commercial kits. Two forward primers including the mutant sequence specific (F2), and one common reverse (R) primer were optimized to generate a 241 bp fragment (F1R), an internal PCR control, and a 141 bp fragment (F2R) denoting the presence of V600E. Sensitivity studies revealed that the assay is capable of detecting V600E even in 1 ng of DNA. Direct sequencing data of 241 bp F1R fragment proved the specificity of the assay. For validation studies of the sequence specific multiplex PCR assay, archival FNA slides were used in a group of thyroid lesions including PTC, follicular carcinoma, follicular adenoma, Hashimoto thyroiditis, and benign thyroid nodules. The newer PCR-based method presented in this study is a practical, inexpensive one-step assay to detect the BRAF T1796A mutation on FNA samples.

Mitochondrial DNA common deletion is not associated with thyroid, breast and colorectal tumors in Turkish patients.

Recently, efforts have been focused on mitochondrial DNA changes and their relation to human cancers. Among them, a 4977 bp deletion of mitochondrial DNA, named "common deletion", has been investigated in several types of tumors, with inconsistent results. In this study, we investigated the presence of the common deletion in tissues from 25 breast, 25 colorectal and 50 thyroid tumors and in the adjacent healthy tissues from Turkish patients. Samples from healthy volunteers were also evaluated for comparison. Two PCR-based methods were used for the detection of the common deletion. First, two pairs of primers were used to amplify wild-type and deleted mtDNA. Then, a highly sensitive nested-PCR was performed, to determine low amounts of deleted genomes. By the first method, wild-type mtDNAs were observed in all samples, but a deletion was observed in only six thyroid samples, by using the nested-PCR method. In conclusion, the mitochondrial common deletion was very rare in our study group and did not appear to be not related with cancer.

Mitochondrial DNA common deletion is not associated with thyroid, breast and colorectal tumors in Turkish patients

Recently, efforts have been focused on mitochondrial DNA changes and their relation to human cancers. Among them, a 4977 bp deletion of mitochondrial DNA, named "common deletion", has been investigated in several types of tumors, with inconsistent results. In this study, we investigated the presence of the common deletion in tissues from 25 breast, 25 colorectal and 50 thyroid tumors and in the adjacent healthy tissues from Turkish patients. Samples from healthy volunteers were also evaluated for comparison. Two PCR-based methods were used for the detection of the common deletion. First, two pairs of primers were used to amplify wild-type and deleted mtDNA. Then, a highly sensitive nested-PCR was performed, to determine low amounts of deleted genomes. By the first method, wild-type mtDNAs were observed in all samples, but a deletion was observed in only six thyroid samples, by using the nested-PCR method. In conclusion, the mitochondrial common deletion was very rare in our study group and did not appear to be not related with cancer.

Computerized image analysis in differentiation of skin lesions caused by electrocution, flame burns, and abrasion.

In the practice of forensic science, sometimes, it is not easy to understand whether skin lesion is due to electrocution and to differentiate the thermal burns and abrasion-type lesions, especially when electricity source cannot be revealed by death science investigation. Based on the causes of the lesions, cases were classified into three groups. Group 1 included 30 deaths from electrocution. Group 2 included 30 deaths with flame burns. Group 3 included 30 deaths from traffic accident cases, which had abrasions. In this study, epidermal nuclear area, perimeter, nuclear form factor, nuclear minimum axes, nuclear maximum axes, and minimum axes/maximum axes ratio were measured. As a result, we think that computerized image analysis beside light microscopic examination can be useful in the differentiation of the electrocution, flame burn, and abrasion type lesions.

The effects of tissue fixation alternatives on DNA content: a study on normal colon tissue.

The aim of this study is to investigate the effects of different fixatives on DNA, and to evaluate the fixation options for molecular studies including polymerase chain reaction (PCR) and fluorescence in-situ hybridization (FISH). Three normal-looking colonic segments from surgical resections were used for tissue sampling. The full thickness of the colonic tissues (3 mm diameter) was sampled. Tissues were fixed in 70% ethanol, 10% neutral-buffered formalin, Hollande, B5, Bouin, and Zenker solutions for 1, 2, 5, 12, 24, and 48 hours, and processed and embedded in paraffin in a standard protocol. Quantitative measurements of the extracted DNA were carried out. DNA quality was tested by PCR for the human beta globin gene. Tissue sections were also tested for the availability of FISH, by using a Her-2/neu protocol. All fixation alternatives were found to be reasonable sources of DNA for molecular studies, and they enabled the successful PCR amplification of a housekeeping gene. DNA yields were predominantly over 1000 bp in 70% ethanol and 24-hour 10% neutral-buffered formalin fixations. As for B5 and Hollande, the DNA molecules obtained were almost all smaller than 100 bp. All tissues fixed in formalin, ethanol, and Hollande were found suitable for Her-2/neu visualization after standard FISH applications, but tissues fixed in Zenker, B5, and Bouin were not found suitable.