Decreased expression of autophagy-related proteins in malignant epithelial ovarian cancer

Some studies have shown that the change of autophagic capacity may correlate with malignant transformation. Our study was designed to investigate the expression and significance of two autophagy-related proteins, microtubule-associated protein 1 light chain 3 (LC3) and Beclin 1 in the tumorigenesis and development of epithelial ovarian carcinoma. We observed that the positive expression of LC3 and Beclin 1 was significantly higher in the samples of benign and borderline ovarian tumors than those in malignant epithelial ovarian cancers. The expression of LC3 and Beclin 1 was associated with FIGO stage and histological grade. No significant relationship was observed between age and histological grade. However, we observed that the expression of LC3 was not related to Beclin 1. Therefore, the decrease of autophagic capacity may be related to tumorigenesis and development of epithelial ovarian cancer.     

Decreased expression of autophagy-related proteins in malignant epithelial ovarian cancer

Some studies have shown that the change of autophagic capacity may correlate with malignant transformation. Our study was designed to investigate the expression and significance of two autophagy-related proteins, microtubule-associated protein 1 light chain 3 (LC3) and Beclin 1 in the tumorigenesis and development of epithelial ovarian carcinoma. We observed that the positive expression of LC3 and Beclin 1 was significantly higher in the samples of benign and borderline ovarian tumors than those in malignant epithelial ovarian cancers. The expression of LC3 and Beclin 1 was associated with FIGO stage and histological grade. No significant relationship was observed between age and histological grade. However, we observed that the expression of LC3 was not related to Beclin 1. Therefore, the decrease of autophagic capacity may be related to tumorigenesis and development of epithelial ovarian cancer.

Addendum: Shen Y, Liang LZ, Hong MH, Xiong Y, Wei M, Zhu XF. Expression and clinical significance of microtubule-associated protein 1 light chain 3 (LC3) and Beclin1 in epithelial ovarian cancer. Ai Zheng 2008; 27:595-9.     

Cell surface proteins and malignant transformation

Many of the altered properties of malignant cells are thought to involve alterations in cell surface functions. In order to understand these alterations it is necessary to know more about the molecular structure of the surface. Methods for analyzing surface proteins are discussed and their application to normal and transformed tissue culture cells are reviewed. A number of surface proteins are observed to be altered by transformation. Most of the alterations are reductions in amounts of particular species, although a few proteins do increase. Evidence concerning the reasons for these alterations and the possible functions of some of the molecules is reviewed. Working hypotheses arising from these data are presented and prospects for understanding the physiological changes in terms of molecular effects are discussed. Particular emphasis is placed on the idea that surface molecules are associated in specific-non-covalent complexes which are important for their functions.     

Localization of a carbohydrate antigen associated with growing oocytes and ovarian surface epithelium.

We used a monoclonal antibody (PS1) to a carbohydrate antigen to study the development of the oocyte and follicle during early stages of differentiation in several mammalian species. This antigen has been shown to localize within the cytoplasm of oocytes in primordial follicles as well as in growing oocytes. It is also localized within distinct layers of the zona pellucida (ZP) of developing follicles. Although this antibody was made against a specific ZP glycoprotein, the antigen also appears to be abundant in cells of the ovarian surface epithelium (OSE). The localization of this carbohydrate moiety has been observed in ovaries of rabbits of different ages as well as in the ovarian surface epithelium of other mammalian species including cat, cynomolgus monkey, baboon, and human. These studies demonstrate that there is an abundant carbohydrate antigenic determinant which is associated with both the mammalian oocyte and the ovarian surface epithelium but which is not apparent in other ovarian cell types or in non-ovarian secretory epithelium. This antibody probe should provide a valuable tool for studying the development and differentiation of the ovary, since this antigen is associated with two highly differentiated but distinct cell types.     

Localization of MCT2 at peroxisomes is associated with malignant transformation in prostate cancer

Previous studies on monocarboxylate transporters expression in prostate cancer (PCa) have shown that monocarboxylate transporter 2 (MCT2) was clearly overexpressed in prostate malignant glands, pointing it out as a putative biomarker for PCa. However, its localization and possible role in PCa cells remained unclear. In this study, we demonstrate that MCT2 localizes mainly at peroxisomes in PCa cells and is able to take advantage of the peroxisomal transport machinery by interacting with Pex19. We have also shown an increase in MCT2 expression from non‐malignant to malignant cells that was directly correlated with its peroxisomal localization. Upon analysis of the expression of several peroxisomal β‐oxidation proteins in PIN lesions and PCa cells from a large variety of human prostate samples, we suggest that MCT2 presence at peroxisomes is related to an increase in β ‐oxidation levels which may be crucial for malignant transformation. Our results present novel evidence that may not only contribute to the study of PCa development mechanisms but also pinpoint novel targets for cancer therapy.     

Progressive changes in Met-dependent signaling in a human ovarian surface epithelial model of malignant transformation

We used an experimental in vitro model of human ovarian surface epithelium (OSE), the tissue of origin of >90% of ovarian cancers, to more precisely define the contribution of hepatocyte growth factor (HGF) to various OSE phenotypes at different stages of neoplastic progression. Neoplastic transformation of OSE in cultures was achieved by multiple genetic manipulations, resulting in the nontumorigenic line IOSE-29, the tumorigenic IOSE-Ov29, and the tumor-derived, more highly malignant IOSE-Ov29/T4. We demonstrate here that, compared to IOSE-29, IOSE-Ov29 and IOSE-Ov29/T4 exhibited higher levels of the HGF receptor Met and an increasing duration of ERK1/2 activation with malignant progression, in conjunction with other neoplastic properties. HGF activated Met signaling in all lines but elicited different responses: HGF induced cell dispersion (scattering) and collagen gel invasion in IOSE-Ov29 and IOSE-Ov29/T4 but did not alter the growth pattern of IOSE-29. Inhibition with PD98059 and LY294002 independently prevented HGF-induced invasive growth. Furthermore, our results show that HGF-induced invasion can be mediated through a rapamycin-sensitive p70 S6K cascade, which demonstrates that p70S6K can regulate cell motility in addition to its well-established role in protein synthesis. Taken together, our data correlate specific responses to HGF-mediated signaling with specific signaling pathways and with progressive neoplastic changes.     

Decreased expression of the ARID1A gene is associated with poor prognosis in primary gastric cancer

Background

The ARID1A gene encodes adenine-thymine (AT)-rich interactive domain-containing protein 1A, which participates in chromatin remodeling. ARID1A has been showed to function as a tumor suppressor in various cancer types. In the current study, we investigated the expression and prognosis value of ARID1A in primary gastric cancer. Meanwhile, the biological role of ARID1A was further investigated using cell model in vitro.

Methodology/Principal Findings

To investigate the role of ARID1A gene in primary gastric cancer pathogenesis, real-time quantitative PCR and western blotting were used to examine the ARID1A expression in paired cancerous and noncancerous tissues. Results revealed decreased ARID1A mRNA (P = 0.0029) and protein (P = 0.0015) expression in most tumor-bearing tissues compared with the matched adjacent non-tumor tissues, and in gastric cancer cell lines. To further investigate the clinicopathological and prognostic roles of ARID1A expression, we performed immunohistochemical analyses of the 224 paraffin-embedded gastric cancer tissue blocks. Data revealed that the loss of ARID1A expression was significantly correlated with T stage (P = 0.001) and grade (P = 0.006). Consistent with these results, we found that loss of ARID1A expression was significantly correlated with poor survival in gastric cancer patients (P = 0.003). Cox regression analyses showed that ARID1A expression was an independent predictor of overall survival (P = 0.029). Furthermore, the functions of ARID1A in the proliferation and colony formation of gastric cell lines were analyzed by transfecting cells with full-length ARID1A expression vector or siRNA targeting ARID1A. Restoring ARID1A expression in gastric cancer cells significantly inhibited cell proliferation and colony formation. Silencing ARID1A expression in gastric epithelial cell line significantly enhanced cell growth rate.

Conclusions/Significance

Our data suggest that ARID1A may play an important role in gastric cancer and may serve as a valuable prognostic marker and potential target for gene therapy in the treatment of gastric cancer.     

Cell surface expression of v-fms-coded glycoproteins is required for transformation.

The viral oncogene v-fms encodes a transforming glycoprotein with in vitro tyrosine-specific protein kinase activity. Although most v-fms-coded molecules remain internally sequestered in transformed cells, a minor population of molecules is transported to the cell surface. An engineered deletion mutant lacking 348 base pairs of the 3.0-kilobase-pair v-fms gene encoded a polypeptide that was 15 kilodaltons smaller than the wild-type v-fms gene product. The in-frame deletion of 116 amino acids was adjacent to the transmembrane anchor peptide located near the middle of the predicted protein sequence and 432 amino acids from the carboxyl terminus. The mutant polypeptide acquired N-linked oligosaccharide chains, was proteolytically processed in a manner similar to the wild-type glycoprotein, and exhibited an associated tyrosine-specific protein kinase activity in vitro. However, the N-linked oligosaccharides of the mutant glycoprotein were not processed to complex carbohydrate chains, and the glycoprotein was not detected at the cell surface. Cells expressing high levels of the mutant glycoprotein did not undergo morphological transformation and did not form colonies in semisolid medium. The transforming activity of the v-fms gene product therefore appears to be mediated through target molecules on the plasma membrane.     

High expression of LASS2 is associated with unfavorable prognosis in patients with ovarian cancer

Homo sapiens longevity assurance homolog 2 of yeast LAG1 (LASS2), is a gene isolated from a human liver complementary DNA library. In this study, we found that LASS2 protein level was positively related to International Federation of Gynecology and Obstetrics (FIGO) stage and LASS2-negative tumors showed significant association with longer disease-free survival (DFS) and overall survival (OS) in ovarian cancer patients. The heterogeneous expression of LASS2 had been exhibited in diverse ovarian cancer cells. A significantly lower messenger RNA (mRNA) and protein level of LASS2 was seen in 3AO cell compared with those in other types of ovarian cancer cells. Meanwhile, the mRNA and protein levels of LASS2 in ES-2 and NIH:OVCAR-3 cells were obviously higher. LASS2 overexpression in 3AO cell could promote migration, invasion, and metastasis abilities in vitro and in vivo, while LASS2 knockdown in ES-2 and NIH:OVCAR-3 cells had the opposite effects. The oncogenic capacity of LASS2 in ovarian cancer may be mediated by increased expression of YAP/TAZ. It is indicated that lowering the expression of LASS2 is likely to serve as an unprecedented approach for the treatment of ovarian cancer.     

Cytoskeletal control of the apical surface transformation of rat uterine epithelium

Summary— During early pregnancy, in the lead up to blastocyst implantation, the apical cell surface of luminal epithelial cells of the rat uterus undergo a dramatic shape transformation. This study aims to investigate the role of the cytoskeleton in this apical transformation by considering the effects of the drugs cytochalasin D and colchicine on the uterine luminal cell surface. The results are determined using transmission and scanning electron microscopy. In vivo exposure to cytochalasin D during oestrus, as well as on day 1 of pregnancy, did not affect the long, regular surface microvilli. This drug, however, did disrupt the terminal web within the apical cytoplasm of these cells. Disruption of microfilament (MF) polymerization by cytochalasin D on day 4 of pregnancy induced a cell surface transformation, resulting in the appearance of numerous irregular projections normally present during blastocyst implantation on day 6 of pregnancy. Colchicine did not alter the uterine microvilli of oestrus or day 1 pregnant tissue. Unlike the effect of cytochalasin D, colchicine-induced microtubule (MT) disruption on day 4 of pregnancy did not increase irregular projections and hence this treatment did not result in the cell surface appearance associated with blastocyst implantation. These results indicate that the disruption of MF, rather than MT, contributes to the transformation of the uterine luminal cell surface during the lead up to blastocyst attachment.     

Decreased 5-hydroxymethylcytosine is associated with neural progenitor phenotype in normal brain and shorter survival in malignant glioma

Epigenetic modification of DNA by cytosine methylation to produce 5-methylcytosine (5mC) has become well-recognized as an important epigenetic process in human health and disease. Recently, further modification of 5mC by the ten eleven translocated (TET) family of enzymes to produce 5-hydroxymethylcytosine (5hmC) has been described. In the present study, we used immunohistochemistry to evaluate the distribution of 5hmC in human brain during different periods of development and in a large series of gliomas (n=225). We found that during development, 5hmC levels are high in more differentiated compartments like the fetal cortex, but low in the periventricular progenitor cell regions. In adults, we found 5hmC levels to be highest in the cortex, but present in all intrinsic cell types in the brain including stromal elements. In brain tumors, 5hmC levels were high in low grade tumors and reduced in malignant glioma, but did not exhibit any correlation with IDH1 mutation status. Additionally, we identified a significant relationship between low levels of 5hmC and reduced survival in malignant glioma. This observation was further supported by in silico analysis showing differential expression of genes involved in 5hmC homeostasis in aggressive subsets of glioblastoma. Finally, we show that several genes involved in regulating the levels of 5hmC are also prognostic in malignant glioma. These findings suggest that 5hmC regulation in malignant glioma may represent an important determinant of tumor differentiation and aggressive behavior, as well as a potential therapeutic target.     

Notch mediated epithelial to mesenchymal transformation is associated with increased expression of the Snail transcription factor

Notch signalling pathway has been implicated as an important contributor to epithelial to myofibroblast transformation (EMT) in tumourigenesis. However, its role in kidney tubular cells undergoing EMT is not defined. This study assessed Notch signalling and the downstream effects on Snail in cultured proximal tubular epithelial cells. EMT was induced by exposure to transforming growth factor beta-1 (TGFβ₁) and angiotensin II (AngII). The expressions of Notch1, Snail, E-cadherin and α-smooth muscle actin (α-SMA) were determined by Western blot. Matrix Metalloproteinase (MMP)-2 and -9 production were determined by zymography. The specific roles of Notch1-ICD and Snail were determined by gene expression or siRNA technique respectively. TGFβ₁ and AngII resulted in EMT as characterized by the expected decrease in E-cadherin expression, an increase in α-SMA, MMP-2 and MMP-9 expression and associated increase of Notch1 and Snail. Over-expression of Notch1-ICD similarly resulted in increased Snail expression, loss of E-cadherin and increasedα-SMA. Inhibiting Snail degradation by pre-treatment with lithium chloride (LiCl) led to a further decrease in E-cadherin expression in cells concurrently exposed to TGFβ₁ + AngII, confirming that Snail is a repressor of E-cadherin. Silencing of Snail blocked TGFβ₁ + AngII induced EMT. Inhibition of Notch activation, by concurrent exposure to DAPT during the induction of EMT attenuated the decrease in E-cadherin expression, limited the increase in α-SMA and MMP-2 and -9 expression and decreased Snail expression. These results suggest a direct role for Notch signalling via the Snail pathway in the development of EMT and renal fibrosis.     

Transport changes associated with growth control and malignant transformation.

We can distinguish two classes of membrane transport changes in cultured cells: (a) growth-rate contingent changes are those which occur in coordination with the onset of density-dependent inhibition of growth; (b) transformation-specific changes are those which occur when cells become transformed, and which can be detected even when normal and transformed cells are growing at the same rate. Growth-rate contingent changes include the density-dependent changes in phosphate, nucleoside, glucose, amino acid, and potassium transport. Only one transformation-specific transport change has been found in Rous-transformed chicken embryo fibroblasts: an increased rate of hexose transport. The variation in potassium transport are associated with variations in the number of ouabain binding sites in the membrane. The molecular basis for changes in the rate of hexose transport is unknown, although gross changes in membrane bilayer composition and "fluidity" seem not to be involved. In analyzing the regulation of hexose transport activity, we find that decreased cAMP may play a role in the transformation-specific increase in hexose transport, but that fibrinolytic activity is not necessary.     

Transdifferentiation of murine squamous vaginal epithelium in proestrus is associated with changes in the expression of keratin polypeptides

The superficial layers of the stratified squamous epithelium of the murine vagina undergo transdifferentiation into cuboidal mucinous cells during the proestrus phase of the normal estrous cycle. In contrast to their squamous progenitor cells which have the cytoskeletal characteristics of squamous epithelium, mucinous cells express keratin polypeptides typical of simple nonstratified epithelia. Accordingly, the transdifferentiation of squamous cell into mucinous cells involves not only a change in cell morphology but also a switch in the expression of keratin polypeptides. These data indicate that the stratified squamous cells of the vagina are not terminally differentiated and their phenotype can be hormonally modulated.     

Threshold of pre-T-cell-receptor surface expression is associated with alphabeta T-cell lineage commitment.

BACKGROUND: The development of immature thymocytes is regulated by the pre-T-cell receptor (pre-TCR). The pre-TCR is involved in several developmental processes including rescuing cells from programmed cell death, allelic exclusion and alphabeta versus gammadelta T-cell lineage commitment. A major issue is how the pre-TCR functions to integrate these processes in developing thymocytes. RESULTS: We have used a sensitive immunofluorescence technique to reveal the surface-expression profile of the pre-TCR on immature thymocyte subsets. We show that early pre-T cells (CD25(+)CD44(-)) can be subdivided on the basis of the level of surface pre-TCR expression. Detectable surface pre-TCR expression identified a rapidly cycling population of early pre-T cells which had successfully undergone beta-selection and been rescued from programmed cell death. Late pre-T cells (CD25(-)CD44(-)), which had traversed the beta-selection checkpoint, expressed surprisingly heterogeneous surface levels of the pre-TCR: high levels of surface pre-TCR expression were associated with commitment to the alphabeta T-cell lineage, whereas late pre-T cells with lower levels of surface pre-TCR could develop along both the alphabeta or gammadelta T-cell lineages. CONCLUSIONS: These data demonstrate that the surface expression of the pre-TCR can be used to reveal newly identified stages of T-cell development and to provide insights into alphabeta T-cell lineage commitment. They show that, although pre-TCR expression does not act as a developmental switch per se, its level of surface expression on late pre-T cells predicts their developmental potential.     

Decreased tumstatin-mRNA is associated with poor outcome in patients with NSCLC

Tumstatin is a candidate tumor suppressor that plays an important role in tumor growth and angiogenesis. The purpose of this study was to evaluate the correlation between tumstatin-mRNA expression and the clinicopathologic characteristics, tumor angiogenesis, outcome of patients with non-small cell lung cancer (NSCLC). Specimens from 68 patients with NSCLC were recruited in this study. Tumstatin-mRNA expression and protein level in tumor tissues were quantified respectively by quantitative RT-PCR and ELISA. Microvessel density (MVD) was determined by CD34 immunostaining. The correlation of tumstatin-mRNA expression levels with clinicopathologic variables, tumor angiogenesis, and prognosis was analyzed. Tumstatin-mRNA expression levels were decreased in tumor. Tumstatin-mRNA expression level was significantly correlated with its protein level in tumor (r = 0.562; P = 0.001). Tumstatin-mRNA expression levels in poorly differentiated tumor tissues were significantly lower than in well-differentiated tumor tissues (P < 0.004). Furthermore, tumor tumstatin-mRNA expression were also significantly related to tumor pathologic stage (P = 0.032) and MVD (r = -0.77, P < 0.0001). Overall survival (OS) and disease-free survival (DFS) analysis indicated that NSCLC patients with low tumstatin-mRNA expression had poorer OS and DFS than those with high expression (P = 0.015 and 0.037; respectively). Multivariable Cox regression analysis revealed that the tumstatin-mRNA expression could be an independent prognostic indicators in both DFS and OS. Tumstatin-mRNA expression levels are down-regulated in NSCLC tissues. Tumstatin-mRNA expression level correlates with prognosis, which suggests that tumstatin-mRNA is a new potential independent marker of favorable prognosis in NSCLC.     

Expression of cellCAM-105 in the apical surface of rat uterine epithelium is controlled by ovarian steroid hormones

Affinity-purified antibodies to cellCAM-105, an adhesive cell surface glycoprotein, were used in immunohistochemical investigations of rat uteri at various functional stages: (i) the oestrous, pro-oestrous, metoestrous, and dioestrous stages of the oestrous cycle, (ii) Days 1-8 of normal pregnancy, (iii) delayed implantation, (iv) 18 h after oestrogen reactivation from delay of implantation, and (v) juvenile rats, and normal ovariectomized adults, respectively, before and after experimental injection of progesterone and/or oestrogen. CellCAM-105 was present in the apical zones of the luminal and glandular epithelium cells in a stage-specific and hormone-dependent manner. The results indicate that: (1) steroid hormones are essential for the expression of cellCAM-105 in the uterine epithelial cells; (2) progesterone induces cellCAM-105 expression in the glandular epithelium, and oestrogen induces cellCAM-105 expression in the luminal epithelium; (3) progesterone induces down-regulation of cellCAM-105 from the surface of the uterine luminal epithelium of juvenile rats; (4) cellCAM-105 is absent in the luminal epithelial cells but present in the glandular epithelial cells of the rat uterus at the time of blastocyst implantation.