Dedicated finite elements for electrode thin films on quartz resonators.

The accuracy of the finite element analysis for thickness shear quartz resonators is a function of the mesh resolution; the finer the mesh resolution, the more accurate the finite element solution. A certain minimum number of elements are required in each direction for the solution to converge. This places a high demand on memory for computation, and often the available memory is insufficient. Typically the thickness of the electrode films is very small compared with the thickness of the resonator itself; as a result, electrode elements have very poor aspect ratios, and this is detrimental to the accuracy of the result. In this paper, we propose special methods to model the electrodes at the crystal interface of an AT cut crystal. This reduces the overall problem size and eliminates electrode elements having poor aspect ratios. First, experimental data are presented to demonstrate the effects of electrode film boundary conditions on the frequency-temperature curves of an AT cut plate. Finite element analysis is performed on a mesh representing the resonator, and the results are compared for testing the accuracy of the analysis itself and thus validating the results of analysis. Approximations such as lumping and Guyan reduction are then used to model the electrode thin films at the electrode interface and their results are studied. In addition, a new approximation called merging is proposed to model electrodes at the electrode interface.

Nuclear expression of S100A4 is associated with aggressive behavior of epithelial ovarian carcinoma: an important autocrine/paracrine factor in tumor progression.

Although S100A4 expression has reportedly been associated with metastasis of various malignancies, little is known about its biological significance in ovarian carcinomas. In this study, we investigated expression and secretion of S100A4 and its extracellular function in ovarian carcinoma cells. We first used immunohistochemistry to examine the expression and localization of S100A4 in 113 epithelial ovarian neoplasms (24 benign, 20 borderline, and 69 malignant tumors) and analyzed its prognostic significance in patients with ovarian carcinoma. Then we investigated the expression, subcellular localization, and secretion of S100A4 in four ovarian carcinoma cell lines. Finally, we examined the effect of S100A4 treatment on the cell proliferation and invasiveness of ovarian carcinoma cells, along with activation of small GTPase, RhoA. Both cytoplasmic and nuclear expressions of S100A4 were significantly stronger in carcinomas than those in benign and borderline tumors. Ovarian carcinoma patients with strong nuclear S100A4 expression showed a significantly shorter survival than those without (P = 0.0045). This was not the case for cytoplasmic S100A4 expression. Ovarian carcinoma cell lines were shown to express S100A4, and secrete S100A4 into the culture media. Treatment with recombinant S100A4 resulted in the upregulation of S100A4 expression, translocation of S100A4 into the nucleus, and enhancement of invasiveness, which was associated with the upregulation of small GTPase, RhoA. These findings suggest that the nuclear expression of S100A4 is involved in the aggressive behavior of ovarian carcinoma and S100A4 is an autocrine/paracrine factor that plays an important role in the aggressiveness of ovarian carcinoma cells.

Elevated expression of E-cadherin and alpha-, beta-, and gamma-catenins in metastatic lesions compared with primary epithelial ovarian carcinomas.

E-cadherin and catenins play key roles in cell adhesion and motility. Little is known about the changes in expression of these molecules in the progression of ovarian carcinomas. In the present study, the immunohistochemical expression of E-cadherin and alpha-, beta-, and gamma-catenins was examined in 77 cases of ovarian carcinoma. In addition, the expression of these molecules was evaluated in 26 matched pairs of primary and metastatic lesions of advanced ovarian carcinomas. Of the 77 primary lesions, positive staining for E-cadherin and alpha-, beta-, and gamma-catenin was observed in 75 (97%), 63 (82%), 71 (92%) and 57 (74%) cases, respectively. Positivity for E-cadherin and alpha-, beta-, and gamma-catenin was significantly decreased in stage III and IV tumors compared with stage I and II tumors, suggesting that expression of the cadherin-catenin complex is reduced with the advancing stages of a tumor. Interestingly, expression of E-cadherin and alpha-, beta-, and gamma-catenin in the lesions of peritoneal dissemination was significantly increased compared with the primary lesions. These findings suggest that expression of the cadherin-catenin complex changes markedly and that reexpression may occur during the peritoneal dissemination of ovarian carcinoma cells.

Expression of BRCA1 protein in benign, borderline, and malignant epithelial ovarian neoplasms and its relationship to methylation and allelic loss of the BRCA1 gene.

BRCA1 is a putative tumour suppressor gene responsible for a hereditary ovarian cancer syndrome. To clarify the possible involvement of BRCA1 in the development of sporadic ovarian neoplasms, this study analysed the immunohistochemical expression of BRCA1 protein in normal ovarian surface epithelium and 119 epithelial ovarian tumours (19 benign, 24 borderline, and 76 malignant tumours). Loss of heterozygosity (LOH) of BRCA1 was examined using three microsatellite markers to analyse the relationship between BRCA1 expression and alterations of the BRCA1 gene. Methylation of the BRCA1 promoter was also analysed by methylation-specific PCR. In ovarian carcinomas showing heterogeneous expression of BRCA1 protein in the same tumour, LOH and methylation status were analysed using microdissection techniques. Finally, the relationship of BRCA1 expression or its genetic alteration to clinicopathological parameters and patient survival was analysed. Ovarian surface epithelial cells expressed BRCA1 protein. Decreased expression of BRCA1 was found in 16% of benign tumours, 38% of borderline tumours, and 72% of carcinomas. LOH of BRCA1 was demonstrated in no benign tumours, 15% of borderline tumours, and 66% of carcinomas. Methylation of BRCA1 was not detected in benign or borderline tumours, but was present in 31% of carcinomas. Reduced expression of BRCA1 correlated with the presence of gene methylation. The frequency of BRCA1 methylation and LOH was higher in serous carcinomas than in other types. In one of the three serous carcinomas that showed heterogeneous expression of BRCA1, BRCA1-positive borderline-like tumour cells were LOH-positive and methylation-negative, whereas adjacent BRCA1-negative carcinoma cells were LOH-positive and methylation-positive. The prognosis of carcinoma patients did not correlate with BRCA1 expression or genetic status. These findings suggest that reduced expression of BRCA1 protein along with genetic and epigenetic changes of the BRCA1 gene play an important role in the development of sporadic ovarian carcinomas, particularly those of serous histology.

Hypoxia attenuates the expression of E-cadherin via up-regulation of SNAIL in ovarian carcinoma cells.

Since ovarian carcinoma cells detach from the primary lesion and metastasize via peritoneal dissemination, we hypothesized that these cells are exposed to hypoxia, which may affect cell attachment and invasiveness. To address this hypothesis, we first examined in vivo the immunohistochemical expression of hypoxia-inducible factor-1alpha (HIF-1alpha) and its topological correlation with E-cadherin expression in ovarian carcinomas. We then examined in vitro the effect of hypoxia on the mRNA and protein expressions of E-cadherin using two ovarian cancer cell lines, SKOV3 and OVCAR3, and normal ovarian surface epithelial (OSE) cells. In addition, hypoxia-induced change in the expression of SNAIL, a transcriptional factor repressing E-cadherin expression, was also analyzed. Finally, we examined the facilitation of invasiveness of ovarian cancer cells under hypoxia using Matrigel invasion assay. Immunohistochemically, nuclear localization of HIF-1alpha was observed in 32 of the 76 (42%) carcinomas studied, and showed a topological correlation with loss of E-cadherin expression. Northern blotting, real-time PCR and Western blotting demonstrated that E-cadherin expression was remarkably decreased under hypoxia in both SKOV3 and OVCAR3 cells, but not in normal OSE cells. mRNA expression of SNAIL was increased under hypoxia in both ovarian cancer cell lines. Invasion assay revealed that hypoxia increases the invasiveness of ovarian cancer cells. Accordingly, the present study demonstrated that hypoxia induces down-regulation of E-cadherin in ovarian carcinoma cells, via up-regulation of the transcriptional repressor SNAIL. These findings suggest that hypoxia plays an important role in the change in intercellular attachment, which may be involved in the initiation of tumor progression of ovarian cancer cells.

Up-regulation of small GTPases, RhoA and RhoC, is associated with tumor progression in ovarian carcinoma.

To clarify the role of small GTPases Rho in the biologic behavior of ovarian carcinoma, we first examined the mRNA expression of RhoA, RhoB, and RhoC in benign, borderline, and malignant ovarian tumors using RT-PCR and real-time RT-PCR. The expression and localization of RhoA protein were also analyzed by Western blotting and immunohistochemistry. Finally, we examined whether up-regulation of Rho enhances the invasiveness of ovarian cancer cells in vitro. Analysis of mRNA levels of the Rho family genes revealed that levels of both RhoA and RhoC were significantly higher in carcinomas than in benign tumors (RhoA, p = 0.0035; RhoC, p = 0.0006). According to histologic subtype, both RhoA and RhoC mRNA levels in serous carcinomas were significantly higher than those in other histologic types. With regard to the International Federation of Gynecological and Obstetrics stage classification, both of RhoA and RhoC mRNA levels were significantly higher in tumors of Stages III+IV than in those of Stages I+II (RhoA, p = 0.0200; RhoC, p = 0.0057). In addition, analysis of matched pairs of primary and disseminated lesions demonstrated that expression of both RhoA and RhoC mRNA was significantly higher in metastatic than in primary tumors. Examination of the protein level showed that expression of RhoA was also increased in advanced ovarian carcinomas, especially those of serous histology. Accordingly, we hypothesized that up-regulation of Rho GTPases plays an important role in the progression of ovarian carcinoma. Matrigel invasion assay using the ovarian cancer cell line, SKOV3, showed that up-regulation and activation after treatment with lysophosphatidic acid was associated with enhanced invasion of the cancer cells. This increase in invasiveness was suppressed by the addition of C3, a specific inhibitor of Rho. These findings suggest that up-regulation of Rho GTPases is important in the tumor progression of ovarian carcinoma and that Rho family proteins could be a molecular target in cancer therapy.

Immunohistochemical expression of cyclins, cyclin-dependent kinases, tumor-suppressor gene products, Ki-67, and sex steroid receptors in endometrial carcinoma: positive staining for cyclin A as a poor prognostic indicator.

Although aberrant expression of several cell-cycle regulators has been reported in endometrial carcinoma, correlations among these factors and their prognostic significance have not fully been elucidated. In the present study, expression of cyclins (D1, E, A, and B1), cyclin-dependent kinases (cdk2, cdk4, and cdc2), and tumor-suppressor gene products (p53, p21, and p27) were systematically examined by immunohistochemistry in 82 cases of endometrial carcinoma and 20 normal endometria. Results were compared with the expression of Ki-67, sex steroid receptor status, clinicopathological parameters, and patient outcomes. Positive staining for cyclin D1, cyclin E, cyclin A, cyclin B1, cdk2, cdk4, cdc2, p53, p21, and p27 was observed in 63%, 66%, 31%, 32%, 51%, 77%, 71%, 43%, 35%, and 60% of the 82 carcinomas, respectively. Among these factors, positive staining for cyclin D1, cdk4, and p53 was significantly frequent in advanced-stage tumors, and that for cyclin D1, cyclin A, cdk4, p21, and p53 was more frequent in higher-grade tumors. High correlation was found between cyclin A and p53 expression, between cyclin D1 and cdk4 expression, between cdk4 and Ki-67 expression, and between p21 and Ki-67 expression. Multivariate analysis showed that the factors for poor prognosis were advanced stage and cyclin A positivity. These findings suggest that various cell-cycle regulators are involved in activated cell growth of endometrial carcinoma, and that positive staining for cyclin A could be a useful marker for unfavorable patient prognosis.

Hypoxia-induced changes in the expression of VEGF, HIF-1 alpha and cell cycle-related molecules in ovarian cancer cells.

BACKGROUND: Hypoxia is important in cancer progression, and at the stage of detachment of the cancer cells from the primary lesion. This study was undertaken to analyze the effect of hypoxia on angiogenesis and cell proliferation in ovarian cancer. MATERIALS AND METHODS: We first used immunohistochemistry to examine the expression of VEGF in 42 cases of ovarian carcinoma, with relevance to the p53 expression. Then, the expression of VEGF, HIF-1 alpha, cell cycle-related molecules and cell numbers were examined in 4 ovarian cancer cell lines with various p53 gene status. RESULTS: Immunohistochemistry showed that there was no significant correlation between VEGF and p53 expression. Moreover, hypoxia increased the expression of VEGF via up-regulation of HIF-1 alpha irrespective of p53 gene status. However hypoxia did not change the cell numbers, but influenced the expression of cell cycle-related molecules (increased p27 and decreased cyclin D1 and Rb). CONCLUSION: Hypoxia increased VEGF expression in ovarian cancer cells irrespective of p53 gene status.