Significance of topoisomerase III beta expression in breast ductal carcinomas: strong associations with disease-specific survival and metastasis

Topoisomerases are ubiquitous nuclear enzymes that regulate DNA structure in eukaryotic cells. The role of topoisomerase III beta, the newest member of the topoisomerase family, in the clinical outcome of breast cancer is still poorly understood. This study aims to investigate the immunoexpression of topoisomerase III beta in breast cancer and its relationships with clinicopathologic features and immunohistochemical markers of prognostic significance in breast pathology. Using tissue microarrays containing 171 cases of primary invasive breast cancer, we analyzed the immunoexpression of topoisomerase III beta, estrogen receptor, progesterone receptor, HER-2, BRCA-1, p53, and Ki67. Immunostaining for topoisomerase III beta was found in 33.9% of breast carcinomas, and immunopositivity was correlated with distant metastasis (P = .036) and death (P = .006). Decreased expression of topoisomerase III beta correlated with low expression of Ki67 (P < .001) and negativity for HER-2 (P < .001), BRCA-1 (P = .001), and p53 (P < .001). In the multivariate analysis, topoisomerase Hip expression was a significant predictor of survival (hazard ratio, 3.006 [95% confidence interval, 1.582-5.715]; P = .001). In conclusion, topoisomerase III beta expression can be a useful marker in assessing the prognosis of patients with breast cancer and is an independent predictor of survival. (C) 2010 Elsevier Inc. All rights reserved.

beta 1 Integrin and VEGF expression in an experimental model of brain tissue heterotopia in the lung

Integrins and vascular endothelial growth factor (VEGF) are crucially involved in interaction, proliferation, migration, and survival of the cells. However, there is no report in the literature about beta 1 integrin and VEGF expression in heterotopic brain tissue. The aim of this study was to assess beta 1 integrin and VEGF expression in experimental brain tissue heterotopia in the lung during both fetal and neonatal periods. Twenty-four pregnant female Swiss mice were used to induce brain tissue heterotopia on the 15th gestational day. Briefly, the brain of one fetus of each dam was extracted, disaggregated, and injected into the right hemithorax of siblings. Six of these fetuses with pulmonary brain tissue implantation were collected on the 18th gestational day (group E18) and six other on the eighth postnatal day (group P8). Immunohistochemistry of the fetal trunks showed implantation of glial fibrillary acidic protein- and neuronal nuclei-positive heterotopic brain tissue, which were also positive for beta 1 integrin and VEGF in both groups E18 and P8. These results indicate that brain tissue heterotopia during fetal and postnatal period is able to complete integration with the lung tissue as well as to induce vascular proliferation which are the necessary steps for a successful implantation.

Determination of P-glycoprotein, MDR-related protein 1, breast cancer resistance protein, and lung-resistance protein expression in leukemic stem cells of acute myeloid leukemia

Background: The most primitive leukemic precursor in acute myeloid leukemia (AML) is thought to be the leukemic stem cell (LSC), which retains the properties of self-renewal and high proliferative capacity and quiescence of the hematopoietic stem cell. LSC seems to be immunophenotypically distinct and more resistant to chemotherapy than the more committed blasts. Considering that the multidrug resistance (MDR) constitutive expression may be a barrier to therapy in AML, we have investigated whether various MDR transporters were differentially expressed at the protein level by different leukemic subsets. Methods: The relative expression of the drug-efflux pumps P-gp, MRP, LRP, and BCRP was evaluated by mean fluorescence index (MFI) and the Kolmogorov-Smirnov analysis (D values) in five leukemic subpopulations: CD34(+)CD38(-)CD123(+) (LSCs), CD34(+)CD38(+)CD123(-), CD34(+)CD38(+)CD123(+), CD34(+)CD38(+)CD123(-), and CD34(-) mature cells in 26 bone marrow samples of CD34(+) AML cases. Results: The comparison between the two more immature subsets (LSC versus CD34(+)CD38(-)CD123(-) cells) revealed a higher P-gp, MRP, and LRP expression in LSCs. The comparative analysis between LSCs and subsets of intermediate maturation (CD34(+)CD38(+)) demonstrated the higher BCRP expression in the LSCs. In addition, P-gp expression was also significantly higher in the LSC compared to CD34(+)CD38(+)CD123(-) subpopulation. Finally, the comparative analysis between LSC and the most mature subset (CD34(-)) revealed higher MRP and LRP and lower P-gp expression in the LSCs. Conclusions: Considering the cellular heterogeneity of AML, the higher MDR transporters expression at the most immature, self-renewable, and quiescent LSC population reinforces that MDR is one of the mechanisms responsible for treatment failure. (C) 2008 Clinical Cytometry Society.

Oestrogen and Progesterone Receptor Gene Expression in Canine Oocytes and Cumulus Cells Throughout the Oestrous Cycle

The aim of this research was to analyze oestrogen receptor-alpha (ER alpha), ER beta and progesterone receptor (PR) gene expression in the canine oocyte and cumulus cells throughout the oestrous cycle. Ovaries from 38 bitches were recovered after ovariohysterectomy and sliced. The phase of the oestrous cycle was determined by vaginal cytology, vaginoscopy and serum hormonal measurements. Oocytes were mechanically denuded by repeated pipetting. For each phase of the cycle, a sample was composed by a pool of 50 oocytes (sample number: prooestrus = 3, oestrus = 8, dioestrus = 5 and anoestrus = 5) or a pool of cumulus cells (prooestrus = 4, oestrus = 7, dioestrus = 4 and anoestrus = 6). Oocyte and cumulus cells` total RNA was isolated and reverse transcription was conducted to perform real-time PCR. Oestrogen receptor-alpha was expressed throughout the cycle in the oocyte (33.33%, 25.0%, 20.0% and 60.0% for prooestrus, oestrus, dioestrus and anoestrus, respectively) and cumulus cells (50.0%, 47.14%, 25.0% and 66.67% for prooestrus, oestrus, dioestrus and anoestrus, respectively). In the oocyte, the ER beta was also expressed in all phases of the cycle (33.33%, 50.0%, 20.0% and 60.0% for prooestrus, oestrus, dioestrus and anoestrus, respectively), whereas in cumulus cells, ER beta was only expressed during prooestrus (50%) and oestrus (14.29%). Interestingly, while the oocyte PR was not detected in any phase of the cycle, this receptor was expressed during prooestrus (50%), oestrus (42.86%) and anoestrus (16.67%) in cumulus cells. In conclusion, canine oocytes express ER alpha and ER beta throughout the oestrous cycle, however, there is a lack of PR expression in all these phases. Moreover, in cumulus cells, only ER alpha was expressed throughout the oestrous cycle.

Prognostic impact of adenomatous polyposis coil gene expression in osteosarcoma of the extremities

Purpose: To evaluate the impact of adjuvant chemotherapy on the outcome of osteosarcoma of the extremities, and to identify prognostic factors using the expression of adenomatous polyposis coli (APC), cadherin and beta-catenin Wnt-signalling markers. Methods: The clinical, demographic, anatomic and pathological factors including a detailed analysis of the immunohistochemical expression of cadherin, B-catenin and APC were retrospectively examined in 97 patients with osteosarcoma of the extremities (metastatic and non-metastatic at diagnosis), treated with surgery and/or chemotherapy from 1985 to 2000. Results: APC immunoreactivity showed a statistically significant association with age and serum alkaline phosphatase levels (p = 0.025 and p = 0.038). When survival was the end-point of multivariate analysis, race segregated patients with poor survival as did lack of cadherin expression. For overall survival, cadherin immunoreactivity and the interaction between APC expression and response to adjuvant chemotherapy were significant (p = 0.012 and p < 0.001). No significant clinical association was evident with immunohistochemical expression of cadherin, beta-catenin. Conclusion: Lack of expression of cadherin was a significant variable to overall and disease-free survival. Significantly, positive APC immunoreactivity and adjuvant chemotherapy were associated with a favourable treatment response. Studies using newer immunohistochemical markers within the Wnt-signalling pathway may guide the development of more appropriate therapeutic targets for future individualised treatment. (C) 2010 Elsevier Ltd. All rights reserved.

Cigarette smoke inhalation modulates gene expression in sites of bone healing: a study in rats

Objective. The aim of this study was to assess the effect of cigarette smoke inhalation (CSI) on gene expression in alveolar bone healing sites. Study design. Wistar rats were randomly assigned to the groups: control [animals not exposed to CSI (n = 20)] and test [animals exposed to CSI, starting 3 days before teeth extraction and maintained until killing them (n = 20)]. First mandibular molars were bilaterally extracted, and the expression of alkaline phosphatase, bone morphogenetic protein (BMP) 2 and 7, receptor activator of nuclear factor kappa B ligand, osteoprotegerin, and d2 isoform of vacuolar adenosine triphosphatase V(o) domain were assessed by quantitative polymerase chain reaction in the newly formed tissue in the sockets. Results. Overall, data analysis demonstrated that CSI significantly affected the expression pattern of all of the studied genes except BMP-7. Conclusion. The expression of key genes for bone healing may be affected by CSI in tooth extraction sites. (Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2010;110:447-452)

Glucose transporter protein 1 expression in mucoepidermoid carcinoma of salivary gland: correlation with grade of malignancy

P>Mucoepidermoid carcinoma (MEC), the most common primary salivary malignancy, shows great variability in clinical behaviour, thus demanding investigation to identify of prognostic markers. Since Warburg`s studies, unrestricted cell growth during tumorigenesis has been linked to altered metabolism, implying hypoxic stimulation of glycolysis and diminished contribution of mitochondrial oxidative phosphorylation to cellular ATP supply. Hypothesizing that the study of MEC metabolic status could lead to the discovery of prognostic markers, we investigated by immunohistochemistry the expression of glucose transporter 1 (Glut-1), mitochondrial antigen and peroxiredoxin I (Prx I) in samples of MEC from different histological grades. Our results showed that mitochondrial antigen and Prx I were expressed in the majority of the MEC cases independent of the histological grade. In contrast Glut-1 expression increased significantly as the tumours became more aggressive. These results suggested that oxidative phosphorylation may contribute to ATP supply in all stages of MEC progression, and that the relative contribution of glycolysis over mitochondria for cellular ATP supply increases during MEC progression, favouring growth under low oxygen concentration. In addition, the observed high Prx I protein levels could provide protection to tumour cells against reactive oxygen species generated as a consequence of mitochondrial function and hypoxia-reoxygenation cycling. Altogether our findings suggest that upregulation of Glut-1 and Prx I constitute successful adaptive strategies of MEC cells conferring a growth advantage over normal salivary gland cells in the unstable oxygenation tumour environment.

The severity of epithelial dysplasia is associated with loss of maspin expression in actinic cheilitis

Background Prolonged exposure of the lip to sunlight may cause actinic cheilitis (AC) and squamous cell carcinoma (SCC). Maspin is a serpin with tumor suppressor functions. This work analyzed the presence and distribution of maspin in AC and lip SCC. Methods Sections from 36 cases diagnosed as AC (18 cases with mild epithelial dysplasia, 11 with moderate and 7 with severe), 18 cases diagnosed as lip SCC and 7 specimens containing normal lip vermillion epithelium were submitted for immunohistochemical analysis to detect maspin. Results All AC cases with mild and two cases with moderate dysplasia were scored 3. The remaining nine cases with moderate dysplasia were identified as score 2, whereas all cases with severe dysplasia were scored 1. Positive staining for maspin decreased from the basal layer to the surface. Among the 18 lip SCCs studied, 15 cases showed abundant staining for maspin. Epithelium adjacent to the SCCs also showed intense positive staining in all cells. Conclusions Our results suggest that the loss of maspin expression occurs from the basal layer to the surface. Lip SCCs related to solar radiation show an intense presence of maspin protein in almost all tumor cells as well as the neighboring epithelium. Fontes A, Sousa SM, Santos E, Martins MT. The severity of epithelial dysplasia is associated with loss of maspin expression in actinic cheilitis.

CASPASE EXPRESSION IN ORAL SQUAMOUS CELL CARCINOMA

Background. Apoptosis is a genetically programmed form of cell death, of which caspases are the central components. Methods. By tissue microarray of 229 cases of oral squamous cell carcinoma (OSCC), we analyzed the immunoexpression of caspases 3, 6, 7, 8, 9, and 10. Results. All proteins that we examined were expressed in primary OSCC samples. Caspases 8 and 9 were prominently expressed, and caspases 3, 6, 7, and 10 were occasionally expressed. Disease-free survival differed significantly between caspase 7 high-expressing and low-expressing patients, and our multivariate analysis suggested that expression of caspase 7 is an independent prognostic factor for patients with OSCC. Conclusion. This study suggests that caspases regulate the tumorigenesis of OSCC and that caspase 7 expression is a predictor of locoregional recurrence of OSCC. (C) 2010 Wiley Periodicals, Inc. Head Neck 33: 1191-1198, 2011

STAT3 expression in salivary gland tumours

The aim of the present study was to evaluate the signal transducer and activator of transcription (STAT3) expression, which is constitutively active in different types of malignant tumours, in salivary gland tumours. Fifty biopsies of salivary gland tumours (9 pleomorphic adenomas, 12 adenoid cystic carcinomas, 7 epithelial-myoepithelial carcinomas, 10 polymorphous low-grade adenocarcinomas and 12 mucoepidermoid carcinomas) and 10 normal. salivary glands were immunohistochemically labeled for STAT3 and Phospho-STAT3 (STAT3P). The labeled sections were quatitatively and quantitatively evaluated. The results showed that, in normal salivary gland, STAT3 was expressed in cytoplasm and STAT3P in nuclei of all tissue cells, except in large mucous acinar cells for which both antibodies were negative. In pleomorphic adenoma, the expression was the same as in normal glands. In malignant tumours, there were variations in the expression of these antibodies. The most important one was the presence of STAT3 in the nuclei of the malignant tumour cells, most evident in the cribriform-type of adenoid cystic carcinoma. Both toss and variation of STAT3P expression were also observed. The presence of STAT3 in the nuclei of malignant salivary gland tumours may represent an important event in oncogenesis probably contributing to tumour cell proliferation white blocking apoptosis. However, further investigation will. be necessary to support this hypothesis. (c) 2007 Pubtished by Elsevier Ltd.

Enhanced programmed death 1 (PD-1) and PD-1 ligand (PD-L1) expression in patients with actinic cheilitis and oral squamous cell carcinoma

PD-1 and PD-L1 can be involved in tumor escape, and little is known about the role of these molecules in oral tumors or pre-malignant lesions. In the present study, we investigated the expression of PD-1 and PD-L1 in the blood and lesion samples of patients with actinic cheilitis (AC) and oral squamous cell carcinoma (OSCC). Our results showed that lymphocytes from peripheral blood and tissue samples exhibited high expression of PD-1 in both groups analyzed. Patients with AC presented higher percentage as well as the absolute numbers of CD4(+)PD-1(+) and CD8(+)PD-1(+) lymphocytes in peripheral blood mononuclear cells (PBMC) than healthy individuals, while patients with OSCC presented an increased frequency of CD8(+)PD1(+) in PBMC when compared with controls. On the other hand, increased frequency of CD4(+) and CD8(+) T cells expressing PD-1(+) accumulate in samples from OSCC, and the expression of PD-L1 was intense in OSCC and moderate in AC lesion sites. Lower levels of IFN-gamma and higher levels of TGF-beta were detected in OSCC samples. Our data demonstrate that PD-1 and PD-L1 molecules are present in blood and samples of AC and OSCC patients. Further studies are required to understand the significance of PD-1 and PD-L1 in oral tumors microenvironment.

Substance P and NK-1R expression in oral precancerous epithelium

The objectives of this study were to investigate the presence and distribution of substance P and neurokinin 1 receptor in oral premalignant epithelium and their relation with the presence of dysplasia, and to analyze whether the expression of substance P can be considered an early oncogenic event in oral carcinogenesis. Substance P and neurokinin I receptor expression was immunohistochemically studied in 83 oral carcinomas and adjacent nontumor epithelia. The presence and degree of epithelial dysplasia was assessed according to WHO criteria. The nuclear, cytoplasmic, and membrane expression of substance P and the cytoplasmic and membrane expression of neurokinin 1 receptor were assessed in tumor and adjacent non-tumor epithelium. Nuclear and cytoplasmic expression of substance P in non-tumor epithelium was significantly associated with the presence of epithelial dysplasia (p<0.001) and carcinoma in. situ (p=0.021). Nuclear, cytoplasmic, and membrane expressions of substance P in non-tumor epithelium were significantly (p<0.001) associated with its expression in the corresponding tumor. These findings suggest that substance P plays a role in early oral carcinogenesis by promoting the proliferation and growth of premalignant fields.

Ki-67 expression in non-tumour epithelium adjacent to oral cancer as risk marker for multiple oral tumours

Objective: The aim of this study was to determine whether the differential assessment of epithelial proliferation is useful to diagnose premalignant fields and assess the risk of multiple tumours. Material and methods: We analysed 83 oral carcinomas with associated non-tumour epithelium classified as distant or close according to its distance (> or < 1 cm) from the invasion point, and as squamous hyperplasia, mild, moderate, severe dysplasia or carcinoma in situ. Twenty-five healthy oral mucosa samples were used as controls. An immunohistochemical technique was applied using Mib-1. Ki-67 in premalignant epithelium was assessed in basal layer, parabasal layer, medium and upper third. Results: Parabasal expression was significantly higher or showed a tendency to be higher in close and distant epithelia with any histological grade than in the controls. Parabasal Ki-67 significantly differed between distant epithelia associated with multiple vs single tumours (P < 0.001) and between distant epithelia associated with multiple tumours vs controls (P < 0.001). This difference was not observed between distant epithelia associated with single tumours and controls (P = 0.175). The cut-off point that differentiated epithelia associated with multiple tumours was > 50% of Ki-67 + parabasal cells in distant epithelia, which yielded 0.88 sensitivity and 0.79 specificity. Conclusions: The concept of a precancerous field may be linked to an increase in the proliferative activity of parabasal cells.

Altered mucin expression in the gastrointestinal tract: a review

Early studies of changes in mucin expression in disorders of the gastrointestinal tract focused on alterations in the carbohydrate chain. This review briefly considers the various mechanisms by which such alterations may come about: (a) normal variation, (b) sialic acid alterations, (c) defective assembly of carbohydrate side-chains, (d) changed expression of core proteins and (e) epithelial metaplasia. The availability of monoclonal antibodies to mucin core proteins adds a new dimension to mucin histochemistry. It is now possible to offer explanations for traditional mucin histochemical findings on the basis of lineage-specific patterns of mucin core protein expression. Changes in core protein expression are described in inflammatory, metaplastic and neoplastic disorders of the gastrointestinal tract. The possibility that mucin change could be important in the aetiology of some diseases such as ulcerative colitis and H. pylori gastritis is considered. It is more probable, however, that changes in mucin expression are secondary to reprogramming of cellular differentiation and altered cell turnover. As such they may serve as markers to explain pathogenesis and provide novel diagnostic and prognostic information.