Anthropometric Measures and Risk of Epithelial Ovarian Cancer: Results From the Nurses' Health Study

Epidemiologic evaluations of the relationship between anthropometry and ovarian cancer risk have not been conclusive. Using data collected from two large cohorts, the Nurses' Health Study (NHS) and NHSII, we prospectively evaluated the association between waist and hip circumference, the waist‐to‐hip ratio (WHR), and BMI with risk of epithelial ovarian cancer. Women completed biennial questionnaires assessing ovarian cancer risk factors beginning in 1976 (NHS) and 1989 (NHSII). For the WHR and BMI analyses, 333 and 862 confirmed cases were identified, respectively, through 1 June 2006 (NHS) and 1 June 2005 (NHSII). WHR and waist circumference were not associated with risk (P‐trend = 0.63 and 0.65, respectively). There was evidence for a decreased risk with increasing hip circumference among postmenopausal women (P‐trend = 0.03), but a suggestive positive association among premenopausal women (P‐trend = 0.04) (P‐interaction = 0.01). The hazard ratios (HRs) comparing the highest vs. lowest quintile of hip circumference among pre‐ and postmenopausal women were 1.54 (95% confidence interval (CI) = 0.45–5.23) and 0.66 (95%CI = 0.37–1.16), respectively. BMI was not clearly associated with risk in pre‐ or postmenopausal women. Results from this large prospective study suggest that hip circumference could be a possible risk factor for premenopausal ovarian cancer, but may reduce risk of postmenopausal ovarian cancer. The differential effect of hip circumference based on menopausal status requires further confirmation.     

Expression of c-Src and phospho-Src in epithelial ovarian carcinoma

Abnormal c-Src expression and activation has been observed in a number of tumors. To determine the therapeutic potential of Src inhibitors for ovarian cancer patients, this study aimed to explore the expression patterns of c-Src and phospho-Src in epithelial ovarian cancer. A total of 82 patients with epithelial ovarian cancer treated at Sun Yat-sen University Cancer Center from January 1999 to December 2005 were enrolled along with 25 patients with benign ovarian lesions; 20 normal ovarian tissues served as controls. Expression of c-Src and phospho-Src (Tyr416) was examined using immunohistochemistry. Survival analyses were performed using Kaplan–Meier curves. As compared to the control group, a significantly greater proportion of ovarian cancer tissues were positive for c-Src and phospho-Src expression (P < 0.001). c-Src expression was associated with age, while phospho-Src expression was significantly associated with age, FIGO stage, histology grade, and residual tumor size after surgery (all P < 0.05). The mean survival time was associated with phospho-Src expression, but not with c-Src expression. The mean survival times of patients with phospho-Src-positive tumors were significantly greater than those with phospho-Src-negative tumors (87.4 months, 95 % CI = 74.3–100.5 months and 91.5 months, 95 % CI = 84.7–98.2 months, respectively; P = 0.013). The increased c-Src expression and activation in epithelial ovarian cancer suggests that ovarian cancer patients may benefit from tyrosine kinase inhibitors such as Dasatinib. Activation of c-Src through phosphorylation at Tyr416 may play a role in the early stages of ovarian cancer development, and evaluation of its expression may be a useful prognostic marker of epithelial ovarian cancer.     

p27Kip1 as a prognostic factor in breast cancer: a systematic review and meta‐analysis

The aim of this study was to comprehensively evaluate via a meta‐analysis the association between p27 expression and clinical outcome in breast cancer patients. We conducted a meta‐analysis of 20 studies (n= 6463 patients) that evaluated the correlation between p27 expression and indicators of breast cancer clinical outcome, including overall survival (OS), disease‐free survival (DFS) and relapse‐free survival (RFS). Data pooling was performed by RevMan 4.2. A total of 60% (9 of 15) of the studies showed a significant association between p27 high expression and OS, whereas 25% (2 of 8) and 60% (3 of 5) studies demonstrated a correlation between p27 high expression and DFS and RFS, respectively. The relative risks (RRs) were 1.34 (1.26–1.42) for OS (P < 0.00001), 1.27 (1.10–1.47) for DFS (P= 0.001) and 1.49 (0.92–2.42) for RFS (P= 0.10). In lymph node‐negative breast cancer patients, the RRs for OS and RFS were 1.84 (1.30–2.59; P= 0.0005) and 1.30 (0.20–8.50; P= 0.78), respectively. In lymph node‐positive breast cancer patients, the RRs for OS and RFS were 2.99 (1.77–5.07; P < 0.0001) and 1.49 (0.80–2.77; P= 0.21), respectively. This meta‐analysis indicates that reduced p27 is an independent prognostic factor for poor overall and disease‐free cancer survival.     

PIWIL4在人卵巢癌中的表达及其功能研究

采用半定量RT-PCR方法检测人卵巢透明细胞癌ES-2细胞中PIWIL1、PIWIL2、PIWIL3、PIWIL4 mRNA表达水平,以及PIWIL4在卵巢癌组织和癌旁正常组织中的表达情况．设计并化学合成针对PIWIL4的siRNA,用脂质体转染法将其转入ES-2细胞内,通过MTT和克隆形成实验,观察PIWIL4-siRNA对ES-2细胞生长活性和增殖能力的影响．半定量RT-PCR实验结果发现,ES-2细胞中,PIWIL4相对PIWIL1、PIWIL2、PIWIL3,其表达水平最高(P < 0.05),而且PIWIL4在卵巢癌组织中的表达明显高于癌旁正常组织(P < 0.01)．MTT实验和克隆形成实验结果显示,转染PIWIL4-siRNA的ES-2细胞生长活性和克隆形成率明显低于对照组(P < 0.05)．由此得出结论：PIWIL4在卵巢癌细胞系ES-2细胞表达较高,且它在卵巢癌组织中的表达明显上调,同时PIWIL4-siRNA可有效抑制ES-2细胞的生长活性和增殖能力,提示PIWIL4可能与卵巢癌发生、发展相关．     

RETRACTION: C-reactive Protein and Metabolic Syndrome in Youth: A Strong Relationship?

Retraction: Note from the Editor‐in‐Chief: This paper is retracted Objective: Metabolic syndrome (MS) is on the rise in youth. As high‐sensitivity C‐reactive protein (hs‐CRP) is associated with cardiovascular/metabolic disorders, we evaluated the association between MS and its components and hs‐CRP in a sample of Brazilian overweight and obese youth. Methods and Procedures: A total of 407 students (229 girls, 273 with excessive weight, 11.3 ± 3.2 years) were evaluated. Measurement included BMI, waist circumference (WC), blood pressure, lipids, insulin, and hs‐CRP. Excessive weight was defined using BMI z ‐score; MS by the modified National Cholesterol Education Program—Adult Treatment Panel III. Results: Subjects were classified into two groups: with MS (n = 72) and without (n = 335). hs‐CRP means and medians were higher in MS group (1.41 mg/l vs. 1.06 mg/l, P < 0.001; 2.21 mg/l vs. 1.23 mg/l, P < 0.001). Associations between hs‐CRP quartiles and insulin resistance (IR) (P < 0.001), MS (P < 0.001), WC (P < 0.000), BMI z‐score (P < 0.001), hypertension (P < 0.001), hypertriglyceridemia (P < 0.001), and low HDL‐c (P = 0.023) were significant; adjustment of hs‐CRP for BMI z‐score eliminated the previous association, except for the number of MS components (nMSc) (P < 0.001). Adjusting for homeostasis model assessment method of IR (HOMA‐IR) did not eliminate the relation between hs‐CRP and MS components. Furthermore, increases in BMI z ‐score and nMSc were associated with an increased hs‐CRP. Excessive weight (odds ratio (OR), 7.9; confidence interval (CI), 4.7–13.4; P = 0.000), hypertension (OR, 2.3; CI, 1.3–4.2; P = 0.003), and hypertriglyceridemia (OR, 2.3; CI, 1.5–3.7; P < 0.001) were independently associated with hs‐CRP. Discussion: In youth, hs‐CRP is strongly related with MS and its components, and is also determined by the body composition. This association indicates a precocious proinflammatory state.     

Aromatase expression in ovarian epithelial cancers

Our study focused on aromatase cytochrome P450 (CYP19) expression in ovarian epithelial normal and cancer cells and tissues. Aromatase mRNA expression was analyzed by real-time PCR in ovarian epithelial cancer cell lines, in human ovarian surface epithelial (HOSE) cell primary cultures, and in ovarian tissue specimens (n=94), including normal ovaries, ovarian cysts and cancers. Aromatase mRNA was found to be expressed in HOSE cells, in BG1, PEO4 and PEO14, but not in SKOV3 and NIH:OVCAR-3 ovarian cancer cell lines. Correlation analysis of aromatase expression was performed according to clinical, histological and biological parameters. Aromatase expression in ovarian tissue specimens was higher in normal ovaries and cysts than in cancers (P<0.0001). Using laser capture microdissection in normal postmenopausal ovaries, aromatase was found to be predominantly expressed in epithelial cells as compared to stromal component. Using immunohistochemistry (IHC), aromatase was also detected in the epithelium component. There was an inverse correlation between aromatase and ERalpha expression in ovarian tissues (P<0.001, r=-0.34). In the cancer group, no significant differences in aromatase expression were observed according to tumor histotype, grade, stage and survival. Aromatase activity was evaluated in ovarian epithelial cancer (OEC) cell lines by the tritiated water assay and the effects of third-generation aromatase inhibitors (AIs) on aromatase activity and growth were studied. Letrozole and exemestane were able to completely inhibit aromatase activity in BG1 and PEO14 cell lines. Interestingly, both AI showed an antiproliferative effect on the estrogen responsive BG1 cell line co-expressing aromatase and ERalpha. Aromatase expression was found in ovarian epithelial normal tissues and in some ovarian epithelial cancer cells and tissues. This finding raises the possibility that some tumors may respond to estrogen and provides a basis for ascertaining an antimitogenic effect of AI in a subgroup of ovarian epithelial cancers.     

High density of IL-17-producing cells is associated with improved prognosis for advanced epithelial ovarian cancer

Interleukin (IL)-17 is the signature cytokine of T helper 17 cells. The role of IL-17 in the tumor microenvironment is still controversial. Few studies describing IL-17 expression in ovarian cancer have been reported. We have therefore analyzed the in situ tumor expression of IL-17 in advanced ovarian cancer and the possible correlation of IL-17 expression with tumor-associated macrophages (TAMs) and with survival in advanced ovarian cancer. Clinical data of 104 patients with stage III–IV epithelial ovarian cancer at the Sun Yat-sen University Cancer Center between 2000 and 2008 were retrospectively reviewed. Immunohistochemical staining of IL-17 and CD163 (marker for TAMs) was performed. Our data showed that levels of IL-17 were significantly increased in ovarian cancer compared with normal ovarian tissues (P<0.001). The high IL-17 expression group included more patients with grade 1 tumors than the low IL-17 expression group (P=0.042). High IL-17 expression correlated with improved progression-free survival (PFS) in advanced ovarian cancer (P<0.001). However, no significant difference was observed in overall survival between the high and low IL-17 expression groups. Multivariate analysis revealed that the density of IL-17-producing cells was a positive prognostic factor for PFS (P=0.001). Moreover, a positive correlation between the density of IL-17-producing cells and TAMs was identified (r=0.354, P<0.001). Our results indicate that the infiltration of IL-17-producing cells might contribute to improved PFS in advanced ovarian cancer. Our findings provide a new insight into the complex role of IL-17 in the tumor microenvironment of ovarian cancer.     

Genetic variants in telomere‐maintenance genes are associated with ovarian cancer risk and outcome

Most ovarian cancer patients present at an advanced stage with poor prognosis. Telomeres play a critical role in protecting chromosomes stability. The associations of genetic variants in telomere maintenance genes and ovarian cancer risk and outcome are unclear. We genotyped 137 single nucleotide polymorphisms (SNPs) in telomere‐maintenance genes in 417 ovarian cancer cases and 417 matched healthy controls to evaluate their associations with cancer risk, survival and therapeutic response. False discovery rate Q‐value was calculated to account for multiple testing. Eleven SNPs from two genes showed nominally significant associations with the risks of ovarian cancer. The most significant SNP was TEP1: rs2228026 with participants carrying at least one variant allele exhibiting a 3.28‐fold (95% CI: 1.72‐6.29; P < 0.001, Q = 0.028) increased ovarian cancer risk, which remained significant after multiple testing adjusting. There was also suggested evidence for the associations of SNPs with outcome, although none of the associations had a Q < 0.05. Seven SNPs from two genes showed associations with ovarian cancer survival (P < 0.05). The strongest association was found in TNKS gene (rs10093972, hazard ratio = 1.88; 95% CI: 1.20‐2.92; P = 0.006, Q = 0.076). Five SNPs from four genes showed suggestive associations with therapeutic response (P < 0.05). In a survival tree analysis, TEP1:rs10143407 was the primary factor contributing to overall survival. Unfavourable genotype analysis showed a cumulative effect of significant SNPs on ovarian cancer risk, survival and therapeutic response. Genetic variations in telomere‐maintenance genes may be associated with ovarian cancer risk and outcome.     

Detection of Autoantibodies to Annexin A11 in Different Types of Human Cancer

Introduction  Annexin A11 was previously identified as an autoantigen in 4.1–10.1% of patients with various systemic autoimmune diseases. In this study, an enzyme-linked immunosorbent assay (ELISA) was developed to investigate the occurrence and features of anti-annexin A11 autoantibodies in sera from patients with different types of cancer. Methods  The recombinant protein of GST fused to the N-terminal domain (1–175 residues) of human annexin A11 was expressed and used as antigen in ELISA. A total of 246 serum specimens were analyzed, which includes sera from healthy women (77), patients with ovarian cancer (72), breast cancer (18), colon cancer (19), pancreatic cancer (20), prostate cancer (20), and diabetes (20). Results  The overall titer of anti-annexin A11 autoantibodies in ovarian cancer patients (or primary tumors only) was found much higher than that in healthy controls (P < 0.05). At the cut-off value designating positive reaction, anti-annexin A11 autoantibodies were detected in 12.5% (5/40) of primary ovarian cancer patients with a significant difference from 2.6% (2/77) of the healthy controls (P < 0.05), but only in 6.25% (2/32) of recurrent tumors. ROC curve demonstrated the potential diagnostic value of anti-annexin A11 autoantibodies in primary ovarian cancer patients with an AUC of 0.62 (0.52–0.73). Anti-annexin A11 autoantibodies were also detected in 5.26% (1/19) of colon cancer and 10% (2/20) of diabetes patients but without significant difference from the healthy controls. Conclusion  A convenient assay to detect anti-annexin A11 autoantibodies in patients was developed, and the experimental data are promising but need to be expanded to address their biological/clinical relevance.     

Eg5 high expression predicts dismal prognosis in epithelial ovarian cancer

The expression of kinesin spindle protein (Eg5) and its significance of clinical prognosis of patients with epithelial ovarian cancer (EOC) were evaluated in this study. Forty-five fresh frozen tissue samples for quantitative real-time polymerase chain reaction (qPCR) and 196 samples for immunohistochemistry (IHC) analysis with tissue microarray (TMA) were applied to characterize Eg5 mRNA and protein expressions in EOC. The correlation between clinical parameters and Eg5 protein expression was investigated using statistical analysis. The expression of Eg5 protein was significantly higher in EOC tissues compared with that in corresponding non-cancerous tissues (P < 0.05). The high Eg5 expression was significantly correlated with older age (P = 0.003), higher stage (P = 0.001), presence of metastasis (P = 0.041) and higher CA125 serum level (P = 0.013). For univariate analysis, associated prognostic markers in patients with ovarian cancer were analyzed for correlations with poor overall survival, including Eg5 (P = 0.011), age (P = 0.001), FIGO stage (P = 0.011), CA125 serum level (P = 0.001), lymph nodes (P = 0.012), and metastasis (P = 0.001). For multivariate analysis, Eg5 expression, FIGO stage and age were independent factors found contributing to a largely unfavorable prognosis in patients with ovarian cancer. In conclusion, the high expression of Eg5 is correlated with an unfavorable prognosis in EOC.     

hMSH2、PCNA蛋白和金葡菌L型感染在卵巢肿瘤中的临床意义

目的 探讨hMSH2(human mut shomolog 2)基因、增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)和金葡菌L型感染在卵巢肿瘤中的表达及临床意义.方法 用免疫组化法检测97例卵巢乳头状癌及23例卵巢乳头状瘤组织中hMSH2、PCNA蛋白以及金葡菌L型抗原的表达,用革兰染色法检测这些组织中有无L型细菌的存在,并用x2检验进行统计学处理.结果 hMSH2蛋白在卵巢恶性肿瘤组织中的阳性表达率明显低于良性肿瘤的(P＜0.01),在卵巢乳头状癌中临床分期Ⅰ、Ⅱ期表达率明显高于Ⅲ、Ⅳ期(P＜0.01),随着病理分级增高而显著降低(P＜0.05),腹腔淋巴结无转移者、无腹水者比有转移、有腹水者明显增高(P ＜0.01 ～0.05).而PCNA蛋白在卵巢恶性肿瘤组织中的阳性表达率明显高于良性肿瘤的(P＜0.01),在卵巢乳头状癌中临床分期Ⅲ、Ⅳ期的表达明显高于Ⅰ、Ⅱ期(P＜0.05),随着病理分级增高而显著增加(P＜0.05),腹腔淋巴结有转移者比无转移者明显增高(P＜0.05),有无腹水者之间差异无统计学意义(P＞0.05).金葡菌L型抗原阳性表达和L型菌的检出率与病理分级、临床分期有明显相关性(P＜0.05),有腹腔淋巴结转移、有腹水者均高于无转移和无腹水者(P＜0.01).结论 hMSH2、PCNA蛋白在卵巢肿瘤中有不同程度的异常表达,均可作为判断卵巢肿瘤生物学行为及患者预后的参考指标,金葡菌L型的感染极有可能导致基因的突变或过表达,因此L型感染可能成为诱发肿瘤形成的原因之一,它们相互协同在卵巢肿瘤发生和发展过程中起重要作用.     

Ceramide metabolism is affected by obesity and diabetes in human adipose tissue

Ceramide is involved in development of insulin resistance. However, there are no data on ceramide metabolism in human adipose tissue. The aim of our study was to examine sphingolipid metabolism in fat tissue from obese nondiabetic (n = 11), obese diabetic (n = 11), and lean nondiabetic (n = 8) subjects. The content of ceramide (Cer), dihydroceramide (dhCer), sphingosine (SPH), sphinganine (SPA), sphingosine‐1‐phosphate (S1P; pmol/mg of protein), the expression (mRNA) and activity of key enzymes responsible for Cer metabolism: serine palmitoyltransferase (SPT), neutral and acidic sphingomyelinase (nSMase and aSMase, respectively), and neutral and acidic ceramidase (nCDase and aCDase, respectively) were examined in human adipose tissue. The contents of SPA and Cer were significantly lower whereas the content of dhCer was higher in both obese groups than the respective values in the lean subjects. The expression of examined enzymes was elevated in both obese groups. The SPT and CDases activity increased whereas aSMase activity deceased in both obese groups. We have found correlation between adipose tissue Cer content and plasma adiponectin concentration (r = 0.69, P < 0.001) and negative correlation between total Cer content and HOMA‐IR index (homeostasis model of insulin resistance) (r = ?0.67, P < 0.001). We have found that both obesity and diabetes affected pathways of sphingolipid metabolism in the adipose tissue. J. Cell. Physiol. 227: 550–557, 2012. © 2011 Wiley Periodicals, Inc.     

EGFR和KRAS在上皮性卵巢癌组织中的表达及临床意义

目的:探讨表皮生长因子受体(EGFR)和鼠Kirsten肉瘤病毒致癌基因(KRAS)蛋白在上皮性卵巢癌组织中的表达水平及临床意义。方法:利用免疫组化SP法对上皮性卵巢癌组织(病例组,n=57)、卵巢良性肿瘤组织(良性组,n=50)以及正常卵巢组织(对照组,n=50)中的EGFR、KRAS蛋白水平进行检测,并分析其在上皮性卵巢癌发生发展中的作用。结果:病例组的EGFR、KRAS蛋白阳性率高于良性组和对照组(P0.05),良性组和对照组的EGFR、KRAS蛋白阳性率差异无统计学意义(P0.05)。浆液性腺癌组EGFR、KRAS蛋白的阳性表达率高于非浆液性腺癌组,Ⅲ~Ⅳ期的上皮性卵巢癌组织中EGFR、KRAS蛋白阳性表达率高于Ⅰ~Ⅱ期,中、低分化组中EGFR、KRAS蛋白阳性表达率高于高分化组,差异均具有统计学意义(P0.05)。上皮性卵巢癌组织中EGFR与KRAS的蛋白的阳性表达率呈正相关关系(r=0.469,P0.05)。结论:EGFR及KRAS蛋白在上皮性卵巢癌组织中的表达水平明显升高,可能参与了上皮性卵巢癌的发生发展过程,且两者之间呈正相关关系,联合检测可作为早期诊断上皮性卵巢癌的重要指标。     

A novel treatment strategy for ovarian cancer based on immunization against zona pellucida protein (ZP) 3

We tested the principle of treating malignant ovarian tumors by vaccination against their ectopically expressed protein, zona pellucida glycoprotein (ZP) 3, using as the experimental model the granulosa cell tumors that develop in transgenic mice expressing the simian virus 40 T-antigen under the inhibin-α promoter (inhα/Tag). We found high ZP3 expression in granulosa cell tumors of the transgenic mice, in human surface ovarian cancer and granulosa cell lines, and in human granulosa cell tumors and their metastases. Early preventive immunization (between 2 and 5.5 mo of age) of transgenic mice with recombinant human (rh) ZP3 prevented ovarian tumorigenesis, and delayed therapeutic immunization (between 4.5 and 7 mo) reduced weights of existing tumors by 86 and 75%, respectively (P<0.001), compared to vehicle-treated control mice. No objective side effects of the immunizations were observed. Liver metastases were found in nontreated/vehicle-treated controls (n=7/39), but none following active rhZP3 immunizations (n=0/36; P<0.05). Immunization with rhZP3 was highly effective, as demonstrated by the induction of anti-ZP3 antibodies, as well as proliferative responses to the ZP3 antigen. These results signal rhZP3 immunization as a novel strategy to be developed for the immunotherapy of ovarian granulosa cell tumors, as well as for that of other malignancies that may express ZP3.     

Characterization of reproductive cycles and adrenal activity in the black‐footed ferret (Mustela nigripes) by fecal hormone analysis

The reproductive cycle of the black‐footed ferret (Mustela nigripes) was characterized by enzyme immunoassay (EIA) analysis of ovarian fecal steroids (estradiol, progestins) in 29 females over two consecutive breeding seasons. Estrous status was determined by measuring the vulva size and examining the percentage of superficial cells in vaginal lavages. Mean fecal estradiol concentrations were correlated with vulval area (r = 0.370, P < 0.0001) and the percentage of superficial cells (r = 0.380, P < 0.0001). Ovulation resulted in a rise in fecal progestin concentrations 5 days after breeding that differed (P < 0.05) between pregnant (n = 14) and pseudopregnant (n = 12) females during the late luteal phase (days 12–40), with concentrations remaining higher in pregnant animals. Gestation length was 41.3 ± 0.7 days with 3.6 ± 0.4 kits produced per female. Litter size correlated significantly (P < 0.05) with fecal estradiol, but not progestins during the 12 to 40 days after breeding. Females failing to breed (n = 3) remained in estrus for 31 ± 6.2 days before ovulation induction with human chorionic gonadotropin. Adrenal activity in male (n = 4) and female (n = 6) black‐footed ferrets was monitored by quantifying fecal corticoid metabolites after a series of manipulations (physical restraint, intramuscular saline, intramuscular gel adrenocorticotropic hormone (ACTH), intramuscular liquid ACTH). A significant (P < 0.0001) increase in fecal corticoids above the pre‐treatment baseline occurred 20 to 44 hours after restraint (five of 10 animals), saline (six of nine), gel ACTH (seven of 10), and liquid ACTH (nine of 10) treatments. Immunoreactivity of high‐performance liquid chromatography–separated fecal elutes was compared using antibodies against cortisol and corticosterone. The cortisol EIA demonstrated immunoreactivity that co‐eluted with ³H‐cortisol, whereas a corticosterone radioimmunoassay detected a metabolite peak that co‐eluted with ³H‐corticosterone in addition to a slightly less polar and one considerably more polar peak. Despite recognizing different metabolites, both assays produced similar temporal profiles of corticoid excretion after manipulation. This study provides new information on the black‐footed ferret regarding differences in fecal steroid excretion patterns between pregnancy and pseudopregnancy and the potential application of fecal corticoid metabolite monitoring for evaluating responses to stressors associated with practices used in breeding management. Zoo Biol 20:517–536, 2001. © 2002 Wiley‐Liss, Inc.     

<Emphasis Type="Italic">RASSF1A</Emphasis> expression level in primary epithelial tumors of various locations

Tumor suppressor activity of RASSF1A in vitro and in vivo was established, in particular, in studies of knockout mice cells. Data on methylation of the promoter region and a lower expression of RASSF1A were mostly obtained with cancer cell lines. Here, the RASSF1A mRNA was quantified the first time in primary epithelial malignant tumors of five various locations from 130 patients by semi-quantitative RT-PCR. Representative samples of kidney, lung, and breast carcinomas were examined. Preliminary data were obtained for RASSF1A expression in ovarian and colorectal carcinomas. System studies showed unexpected expression profiles, namely, the mRNA level increased (two- to sevenfold) more frequently than decreased in renal, breast, ovarian, and colorectal carcinomas. A higher RASSF1A mRNA level was significantly more frequent in renal cell carcinomas (24/38, 63% vs 8/38, 21%, P = 0.0004 by Fisher’s exact test) and ovarian carcinomas (8/13, 62% vs 2/13, 15%, P = 0.0114). Equal frequencies of lower and higher RASSF1A expression levels were only observed in non-small cell lung cancer (16/38, 42%). Noteworthy, an increase in expression was more common at early clinical stages of squamous cell lung cancer and adenocarcinoma, while a decrease in RASSF1A expression was more frequent at advanced clinical stages. In clear cell renal cell carcinoma, an increase in RASSF1A expression occurred more often at both early and advanced stages and was significant at advanced stages (P = 0.0094). The findings suggested tumor specificity for changes in RASSF1A expression. The observed regularities may also indicate that RASSF1A has dual functions in tumors, acting as a tumor suppressor and as a protooncogene.     

Proteomic tracking of serum protein isoforms as screening biomarkers of ovarian cancer

Epithelial ovarian cancer is the fourth leading cause of cancer death among women. Due to the asymptomatic nature and poor survival characteristic of the disease, screening for specific biomarkers for ovarian cancer is a major health priority. Differentially expressed proteins in the serum of ovarian cancer patients have the potential to be used as cancer-specific biomarkers. In this study, proteomic methods were used to screen 24 serum samples from women with high-grade ovarian cancer and compared to a control group of 11 healthy women. Affigel-Blue treated serum samples were processed either by linear (pH 4-7) or narrow range (pH 5.5-6.7) IEF strips for the first dimension. Proteins separated in first dimension were resolved by 8-16% gradient SDS-PAGE. Protein spots were visualized by SYPRO Ruby staining, imaged by FX-imager and compared and analyzed by PDQuest software. Twenty-two protein spots were consistently differentially expressed between normal and ovarian cancer patients by resolving proteins in a linear pH strip of 4-7 for the first dimension. Six of the protein spots, significantly up-regulated in grade 3 ovarian cancer patients (p < 0.05), were identified by MALDI-TOF MS and Western blotting as the isoforms of haptoglobin precursor. When serum proteins were resolved on narrow pH range strips (5.5-6.7), 23 spots were consistently differentially expressed between normal and grade 3 ovarian cancer patients. Of these, 4 protein spots significantly down regulated in grade 3 ovarian cancer patients (p < 0.05) were identified by MALDI-TOF MS and Western blotting, as isoforms of transferrin precursor. Increased expression of serum haptoglobin and transferrin was also identified in peritoneal tumor fluid obtained from women diagnosed with grade 2/3 ovarian cancer (n = 7). Changes in the expression of haptoglobin and transferrin in the serum of women with different pathological grades of ovarian cancer was examined by one-dimensional Western blotting method. Serum samples collected from women suffering from benign, borderline, grade 1, grade 2 and grade 3 cancer (n = 4 for haptoglobin and n = 5 for transferrin in each group) were analyzed and compared to the serum of normal healthy women. The mean serum haptoglobin expression in grade 3 ovarian cancer patients was fourfold higher than in the control subjects (p < 0.05). On the other hand, transferrin expression in grade 3 ovarian cancer patients was decreased by twofold than in normal healthy women (p < 0.05). Haptoglobin expression in the serum of cancer patients (n = 7) decreased following chemotherapy (six cycles of taxol/carboplatin). Concomitant with the decrease of haptoglobin, transferrin expression remained constant in four patients, but increased in three out of seven patients included in the study. Changes in serum expression of haptoglobin correlated with the change of CA 125 levels before and after chemotherapy. In conclusion, proteomic profiling of differentially expressed proteins in the sera of normal women compared to women with ovarian cancer can greatly facilitate the discovery of a panel of biomarkers that may aid in the detection of ovarian cancer with greater specificity.     

DPP4 Gene DNA Methylation in the Omentum is Associated With Its Gene Expression and Plasma Lipid Profile in Severe Obesity

Severely obese subjects with the metabolic syndrome (MS) have higher dipeptidyl peptidase‐4 (DPP4) expression in their visceral adipose tissue (VAT) compared to obese individuals without MS. We tested the hypothesis that methylation level of CpG sites in the DPP4 promoter CpG island in VAT was genotype‐dependent and associated with DPP4 mRNA abundance and MS‐related phenotypes. The VAT DNA was extracted in 92 severely obese premenopausal women undergoing biliopancreatic derivation for the treatment of obesity. Women were nondiabetic and none of them used medication to treat MS features. Cytosine methylation rates (%) of 102 CpG sites in the DPP4 CpG island were assessed by pyrosequencing of sodium bisulfite‐treated DNA. Methylation rates were >10% for CpG sites 94–102. Their mean methylation rate (%Meth_94–102) was different between genotypes for DPP4 polymorphisms rs13015258 (P = 0.001), rs17848915 (P = 0.0004), and c.1926 G>A (P = 0.001). The %Meth_94–102 correlated negatively with DPP4 mRNA abundance (r = ?0.25, P < 0.05) and positively with plasma high‐density lipoprotein (HDL) cholesterol concentrations (r = 0.22, P < 0.05), whereas DPP4 mRNA abundance correlated positively with plasma total‐/HDL‐cholesterol ratio (r = 0.25; P < 0.05). In the VAT of nondiabetic severely obese women, genotype‐dependent methylation levels of specific CpG sites in the DPP4 promoter CpG island were associated with DPP4 gene expression and variability in the plasma lipid profile. Higher DPP4 gene expression in VAT and its relationship with the plasma lipid profile may be explained by actually unknown DPP4 biological effect or, to another extent, may also be a marker of VAT inflammation known to be associated with metabolic disturbances.     

TRPM3表达上调通过调控Beclin1的表达促进卵巢癌细胞自噬

目的:探讨瞬时受体电位离子通道3(TRPM3)和Beclin1在卵巢癌中的表达和对卵巢癌细胞自噬的影响。方法:收集6例正常卵巢组织标本和20例卵巢癌组织标本,应用免疫组织化学染色法检测TRPM3和Beclin1在卵巢癌组织中的表达,采用western blotting法检测TRPM3和Beclin1在正常卵巢上皮细胞Moody和卵巢癌细胞Hey、ES-2中的表达差异。用TRPM3-siRNA瞬时转染细胞Hey和ES-2,通过western blotting法检测TRPM3基因沉默情况及Beclin1、p62和LC3的蛋白表达变化。结果:在正常卵巢组织和卵巢癌组织中,TRPM3的阳性表达率分别为33.3%、80.0%(P0.05),而Beclin1的阳性表达率分别为33.3%、65.0%(P0.05)。与正常卵巢上皮细胞Moody相比,卵巢癌细胞Hey和ES-2中TRPM3、Beclin1蛋白表达水平明显较高(P0.05)。沉默TRPM3基因表达的卵巢癌细胞中Beclin1和LC3蛋白表达与对照组相比明显降低,而p62表达升高(P0.05)。结论:卵巢癌组织和细胞中TRPM3蛋白呈高表达,可能通过调控Beclin1促进卵巢癌细胞的自噬。