Selenium-Binding Protein 1 (SBP1) autoantibodies in ovarian disorders and ovarian cancer.

Infertility is a risk factor for ovarian cancer (OvCa). The goal was to determine if antibodies to selenium-binding protein 1 (SBP1), an autoantibody we identified in patients with premature ovarian failure (POF), occurs in both infertility and OvCa patients, and thus could be associated with preneoplasia. Anti-SBP1 was measured by immunoassay against recombinant SBP1, in sera from OvCa (n = 41), infertility (n = 92) and control (n = 87) patients. Infertility causes were POF, unexplained, irregular ovulation or endometriosis. The percent of anti-SBP1-positive sera was higher in POF (P = 0.02), irregular ovulation (P = 0.001), unexplained causes (P = 0.02), late (III-IV)-stage OvCa (P = 0.02) but was not significant in endometriosis, benign ovarian tumors/cysts, early stage (I-II) OvCa or uterine cancer compared to healthy controls. Anti-SBP1 was significantly higher in women with serous (P = 0.04) but not non-serous (P = 0.33) OvCa compared to controls. Also, we determined if anti-SBP1 was associated with CA125 or anti-TP53, markers often studied in OvCa. Anti-TP53 and CA125 were measured by established immunoassays. The ability of anti-SBP1 alone to discriminate infertility or OvCa from controls or when combined with anti-TP53 and CA125, to identify OvCa was evaluated by comparing the area under the curve (AUC) in ROC analysis. Anti-SBP1 alone discriminated infertility (AUC = 0.7; P = 0.001) or OvCa (AUC = 0.67; P = 0.03) from controls. The sensitivity and specificity of OvCa identification was increased by combining CA125, anti-TP53 and anti-SBP1 (AUC = 0.96). Therefore, anti-SBP1 occurs in infertile women with POF, ovulatory disturbances or unexplained infertility and in serous OvCa. This suggests an autoimmune process is associated with the development of serous OvCa.

Interpersonal violence, PTSD, and inflammation: potential psychogenic pathways to higher C-reactive protein levels.

Interpersonal violence (IPV) is major public health concern with wide-ranging sequelae including depression, posttraumatic stress disorder (PTSD), and possible alterations of immune and inflammation processes. There is a need to identify the psycho-biological pathways through which IPV may translate to altered inflammatory processes since both PTSD and inflammation are associated with serious physical health conditions such as obesity, diabetes, and cardiovascular disease. This study investigated the relationships between IPV, psychological distress, and the inflammatory marker C-reactive protein (CRP), in a sample of 139 urban women who have a high likelihood for having experienced IPV. Participants were recruited from an outpatient gynecology clinic to complete self-report measures about their IPV histories and psychological symptoms, as well as to have their blood sampled using a finger stick. Results indicated that exposure to IPV predicted the presence of probable depression and PTSD diagnoses. Individuals who experience clinical levels of PTSD exhibited higher CRP levels, and this relationship held after adjusting for comorbid depression. Correlational analyses suggested that reexperiencing symptoms may explain the link between PTSD diagnosis and higher levels of CRP. Follow-up path analytic models provided good fit to the overall data, and indicated that the relationship between probable PTSD status and CRP is not explained by higher BMI. Overall, these findings call for increased attention to the role of PTSD in explaining links between trauma and diminished health.

Anti-HE4 antibodies in infertile women and women with ovarian cancer.

OBJECTIVES: To develop an assay for anti-HE4 antibodies and assess such antibodies in sera from women with increased epidemiologic risk for ovarian cancer (infertility) and patients with ovarian cancer in comparison to controls. METHODS: An ELISA was developed to measure antibodies to recombinant full length HE4 and cut-off values were determined for different levels of specificity (up to 99%). RESULTS: Infertile women more frequently had anti-HE4 antibodies than controls (23% at 98% specificity, p < 0.001) with antibodies most frequent in women with POF (31%) and ovulatory dysfunction (47%). There was also an increased frequency of anti-HE4 antibodies in patients with ovarian cancer (14% at 97% specificity, p < 0.01), but more women with certain types of infertility have anti-HE4 antibodies than women with ovarian cancer. Most patients with ovarian cancer have circulating HE4 antigen, which may interfere with detection of antibodies, while the level of HE4 antigen in sera from infertile women was not higher than in normal controls. There was a statistically significant correlation between antibodies to HE4 and antibodies to mesothelin in the same patients. CONCLUSIONS: Women with certain types of infertility, which have increased risk to develop ovarian cancer, and women with ovarian cancer more frequently than controls have antibodies to HE4, a biomarker for ovarian cancer. The antibodies may reflect a tumor-promoting Th2 type of inflammation.

Contrast-enhanced sonography depicts spontaneous ovarian cancer at early stages in a preclinical animal model.

OBJECTIVE: Our goal was to examine the feasibility of using laying hens, a preclinical model of human spontaneous ovarian cancer, in determining the kinetics of an ultrasound contrast agent indicative of ovarian tumor-associated neoangiogenesis in early-stage ovarian cancer. METHODS: Three-year-old White Leghorn laying hens with decreased ovarian function were scanned before and after intravenous injection of a human serum albumin-perflutren contrast agent at a dose of 5 µL/kg body weight. Gray scale morphologic characteristics, Doppler indices, the arrival time, peak intensity, and wash-out of the contrast agent were recorded and archived on still images and video clips. Hens were euthanized thereafter; sonographic predictions were compared at gross examination; and ovarian tissues were collected. Archived clips were analyzed to determine contrast parameters and Doppler intensities of vessels. A time-intensity curve per hen was drawn, and the area under the curve was derived. Tumor types and the density of ovarian microvessels were determined by histologic examination and immunohistochemistry and compared to sonographic predictions. RESULTS: The contrast agent significantly (P < .05) enhanced the visualization of microvessels, which was confirmed by immunohistochemistry. Contrast parameters, including the time of wash-out and area under the curve, were significantly different (P < .05) between ovaries of normal hens and hens with ovarian cancer and correctly detected cancer at earlier stages than the time of peak intensity. CONCLUSIONS: The laying hen may be a useful animal model for determining ovarian tumor-associated vascular kinetics diagnostic of early-stage ovarian cancer using a contrast agent. This model may also be useful for testing the efficacy of different contrast agents in a preclinical setting.

Decreased severity of ovarian cancer and increased survival in hens fed a flaxseed-enriched diet for 1 year.

OBJECTIVE: With the exception of the laying hen, no other animal model of spontaneous ovarian surface epithelial cancer replicates the human disease. Flaxseed is the richest vegetable source of omega-3 fatty acids, which are chemopreventive in breast cancer and may be important in other cancers. The objective of this study was to determine if a flaxseed-enriched diet had a chemopreventive effect on ovarian cancer in the laying hen. METHODS: White Leghorn hens were fed with 10% flaxseed-enriched or standard diet for 1 year. The incidence and severity of ovarian cancer were determined by gross pathology and histology in the two groups. General health markers were also measured. Eggs were collected and analyzed by gas chromatography to determine omega-3 fatty acid levels. RESULTS: A significant reduction in late stage ovarian tumors was detected in the flaxseed-fed hens. Incidence rates of ovarian cancer were not significantly different between the two groups. The results indicate that a flaxseed diet increases overall survival in the laying hen. Flaxseed-fed hens' eggs incorporated significantly more omega-3 fatty acids compared to control hens. CONCLUSIONS: These findings show that 10% flaxseed supplementation for 1 year in the laying hen results in a significant reduction in the severity of ovarian cancer, but no change in the incidence of the disease. Hens fed flaxseed had overall better health and reduced mortality. These findings may provide the basis for a clinical trial that evaluates the efficacy of flaxseed as a chemosuppressant of ovarian cancer in women.

Detection of tumor-associated neoangiogenesis by Doppler ultrasonography during early-stage ovarian cancer in laying hens: a preclinical model of human spontaneous ovarian cancer.

OBJECTIVE: Tumor-associated neoangiogenesis (TAN) is one of the earliest events in ovarian tumor growth and represents a potential target for early detection of ovarian cancer (OVCA). Because it is difficult to identify patients with early-stage OVCA, the goal of this study was to explore a spontaneous animal model of in vivo ovarian TAN associated with early-stage OVCA detectable by Doppler ultrasonography (DUS). METHODS: White Leghorn laying hens were scanned transvaginally at 15-week intervals up to 45 weeks. Gray scale ovarian morphologic characteristics and Doppler indices were recorded. Hens were euthanized at diagnosis for ultrasonographic morphologic/vascular abnormalities or at the end of the study (those that remained normal). Ovarian morphologic and histologic characteristics were evaluated. Vascular endothelial growth factor (VEGF) and alpha(v)beta(3)-integrin expression was assessed by immunohistochemical analysis. Doppler ultrasonographic observations were compared with histologic and immunohisto-chemical findings to determine the ability of DUS to detect ovarian TAN. RESULTS: Significant changes in ovarian blood flow parameters were observed during transformation from normal to tumor development in the ovary (P < .05). Tumor-related changes in ovarian vascularity were identified by DUS before the tumor became detectable by gray scale imaging. Increased expression of VEGF and alpha(v)beta(3)-integrins was associated with tumor development. Ovarian TAN preceded tumor progression in hens. CONCLUSIONS: The results suggest that ovarian TAN may be an effective target for the detection of early-stage OVCA. The laying hen may also be useful for studying the detection and inhibition of ovarian TAN using various means, including the efficacy of contrast agents, targeted molecular imaging, and antiangiogenic therapies.

Prevalence of antitumor antibodies in laying hen model of human ovarian cancer.

Antitumor antibodies are associated with tumors in human cancers. There is relatively little information on the timing and progression of antibody response to tumors. The objective of the study was to determine if spontaneous ovarian cancer in the egg-laying hen is associated with antitumor antibodies. Antibodies were detected by immunoassay and immunoblotting using proteins from normal ovary and ovarian tumors. Candidate antigens were identified by mass spectrometry of immunoreactive spots cut from 2-dimensional gels and Western blot. Antitumor (serum reacting against tumor ovarian extract) and antiovarian (serum reacting against normal ovarian extract) antibodies were significantly associated with ovarian cancer (67%; P

Histopathology of ovarian tumors in laying hens: a preclinical model of human ovarian cancer.

The high mortality rate due to ovarian cancer (OVCA) is attributed to the lack of an effective early detection method. Because of the nonspecificity of symptoms at early stage, most of the OVCA cases are detected at late stages. This makes the access to women with early-stage disease problematic and presents a barrier to development and validation of tests for detection of early stage of OVCA in humans. Animal models are used to elucidate disease etiologies and pathogenesis that are difficult to study in humans. Laying hen is the only available animal that develops OVCA spontaneously; however, detailed information on ovarian tumor histology is not available. The goal of this study was to determine the histological features of malignant ovarian tumors in laying hens. A total of 155 young and old (1-5 years of age) laying hens (Gallus domesticus) were selected randomly and evaluated grossly and microscopically for the presence of ovarian tumors. Histological classification of tumors with their stages and grades was determined with reference to those for humans. Similar to humans, all 4 types including serous, endometrioid, mucinous, and clear cell or mixed carcinomas were observed in hen ovarian tumors. Some early neoplastic as well as putative ovarian lesions were also observed. Similarities in histology, metastasis, and stages of hen OVCA to those of humans demonstrate the feasibility of the hen model for additional delineation of the mechanism underlying ovarian carcinogenesis, preclinical testing of new agents for the prevention, and therapy of this disease.

Selenium-Binding Protein 1 expression in ovaries and ovarian tumors in the laying hen, a spontaneous model of human ovarian cancer.

OBJECTIVE: Reduced Selenium-Binding Protein 1 (SELENBP1) expression was recently shown in multiple cancers. There is little information on the expression and function of SELENBP1 in cancer progression. In order to develop a better understanding of the role of SELENBP1 in ovarian cancer, our objective was to determine if SELENBP1 is expressed in the normal ovaries and ovarian tumors in the egg-laying hen, a spontaneous model of human ovarian cancer. METHODS: SPB1 mRNA expression in normal ovary (n=20) and ovarian tumors (n=23) was evaluated by RT-PCR. Relative levels of mRNA were compared by quantitative RT-PCR (qRT-PCR) in selected samples. SELENBP1 protein expression was evaluated by 1D Western blot and immunohistochemistry with a commercial anti-human SELENBP1 antibody. RESULTS: SELENBP1 mRNA and protein was expressed in 100% of normal and ovarian tumors and qRT-PCR confirmed decreased mRNA expression in 80% of ovarian tumors. SELENBP1 was primarily localized in surface epithelial cells of normal ovaries. In ovaries containing early tumor lesions, SELENBP1 expression was reduced in the surface epithelium near the tumor and was expressed in tumor cells, while more distant regions with normal histology retained SELENBP1 expression in the surface epithelium. CONCLUSIONS: We have shown for the first time that SELENBP1 is expressed in both normal ovaries and ovarian tumors in the hen and that SELENBP1 expression is altered in the vicinity of the tumor. Furthermore, SELENBP1 expression in normal ovarian surface epithelium and in ovarian tumors parallels that previously reported for ovarian cancer in women.

Correlates of circulating androgens in mid-life women: the study of women's health across the nation.

CONTEXT: Androgens influence sexual differentiation and behavior, body composition, and physical functioning in men, but their role in women is less well understood. Because circulating androgens decline with age, the use of androgen supplementation for women to improve health and well-being has been increasing. OBJECTIVE: The aim of this study was to assess the association between androgens and a variety of end points thought to be affected by androgens. DESIGN: In a community-based baseline cohort of women aged 42-52 yr from the Study of Women's Health Across the Nation, we measured circulating testosterone (T), dehydroepiandrosterone sulfate, and SHBG, and calculated a free androgen index (FAI) in 2961 women. MAIN OUTCOME MEASURES: Correlations of androgen measures with each other and with body mass index, waist circumference, and waist-hip ratio were computed, and odds ratios (OR) were estimated for the categorical outcomes of functional limitations, functional status, self-reported health, scores indicative of depressed mood, quality of life, sexual desire and arousal, and the presence of the metabolic syndrome. RESULTS: Androgens, and particularly SHBG, were associated most strongly with body mass index, waist circumference, and waist-hip ratio. SHBG was associated prominently inversely with the metabolic syndrome (OR = 0.32; 95% confidence interval = 0.26-0.39), which was present in 17% of women at baseline. Dehydroepiandrosterone sulfate was associated modestly with functional status and self-reported health. T was associated minimally with increased sexual desire (OR = 1.09; 95% confidence interval = 1.00-1.18). The association of FAI with self-reported health and depressive symptomatology based on the Center for Epidemiologic Studies Depression Scale score was explained more by T than by SHBG, whereas the association of FAI with sexual arousal and metabolic syndrome was due more to SHBG than to T. CONCLUSIONS: Circulating SHBG and androgens are most strongly associated with physical characteristics and the metabolic syndrome in women in this community-based cohort. Androgens are related weakly to physical functioning and other symptoms to which they commonly are attributed, such as sexual desire, sexual arousal, and well-being.

Change in estradiol and follicle-stimulating hormone across the early menopausal transition: effects of ethnicity and age.

Serum reproductive hormone concentrations were measured longitudinally in a community-based, multiethnic population of midlife women to assess whether ethnic differences exist in the patterns of change in estradiol (E2) and FSH and, if so, whether these differences are explained by host characteristics. We studied 3257 participants from seven clinical sites in the Study of Women's Health Across the Nation (SWAN) who were aged 42-52 yr at baseline and self-identified as African American (28.2%), Caucasian (47.1%), Chinese (7.7%), Hispanic (8.4%), or Japanese (8.6%). E2 and FSH were assayed in serum collected primarily in the early follicular phase of a spontaneous menstrual cycle in three consecutive annual visits. The primary explanatory variables included in repeated-measures regression analyses were race/ethnicity, menopausal status, age, body mass index (BMI), day of the cycle, smoking, parity, socioeconomic status, study site, and the self-report of diabetes at baseline. At the baseline visit, 46.2% of the women were classified as being early perimenopausal, with the remaining being premenopausal. By the second follow-up visit, 5.5% of the women in that cohort were postmenopausal, 66.8% were early perimenopausal, 8.3% were late perimenopausal, and 19.4% remained premenopausal. Serum E2 concentrations decreased significantly with age, with a steeper decline at higher ages. FSH concentrations increased significantly with age, with a steeper increase at higher ages. Similar patterns in the decline of E2 and the increase in FSH with age were found across ethnic groups, but the levels of these hormones differed by race/ethnicity. Specifically, over time, Chinese and Japanese women had lower E2 concentrations but similar FSH levels, compared with Caucasian women, and African American women had higher FSH concentrations but comparable E2 levels with those of Caucasian women. These ethnic differences in E2 and FSH were independent of menopausal status. The effect of BMI on serum E2 and FSH levels varied by menopausal status. Increasing BMI was associated with decreasing concentrations of E2 among premenopausal and early perimenopausal women but was associated with increasing concentrations of E2 among late perimenopausal and postmenopausal women. Increasing BMI was associated with decreasing concentrations of FSH, with the effect of BMI becoming larger as women transitioned through menopause. We conclude that serum E2 levels decrease and FSH concentrations increase with increasing age in midlife women, that ethnic differences in E2 over time differ from ethnic differences in FSH and suggest ethnic differences in the pituitary-ovarian relationship, and that the effect of BMI on E2 and FSH concentrations varies by menopausal status.

Body size and ethnicity are associated with menstrual cycle alterations in women in the early menopausal transition: The Study of Women's Health across the Nation (SWAN) Daily Hormone Study.

The dynamics of reproductive hormones that characterize the menopausal transition (perimenopause) are incompletely understood, particularly in non-Caucasian women. The Study of Women's Health across the Nation (SWAN) is a multiethnic cohort study of 3302 women at seven sites who were aged 42-52 yr at baseline. All participants are seen annually to assess a variety of endpoints. A subcohort of 848 women undergoes further investigation of their daily patterns of reproductive hormones in the Daily Hormone Study (DHS). DHS enrollees annually complete a daily collection of first morning voided urine for an entire menstrual cycle or up to 50 d (whichever comes first). Chemiluminescent assays measured urinary LH and FSH, as well as metabolites of estradiol [estrone conjugates (E1c)] and progesterone [pregnanediol glucuronide (Pdg)]. Cycles were assessed for evidence of luteal activity and day of luteal transition using previously developed algorithms. Midreproductive-aged women who underwent similar daily urinary analyses served as historical controls. Correlates of cycle features were identified. Eight hundred thirty-three cycles were evaluable and had complete data on covariates. Six hundred seventy-four (80.9%) cycles had evidence of luteal activity, and 159 (19.1%) did not. Women who were at least 49 yr old were less likely to have cycles with luteal activity and had more variable cycle length, higher total-cycle FSH, and lower total-cycle Pdg. Compared with heavier women, those with body mass index less than 25 kg/m2 had shorter cycles and higher total-cycle LH, FSH, and Pdg but not E1c. Chinese- and Japanese-American women had overall lower adjusted total-cycle E1c excretion. Smoking was not significantly associated with cycle length or hormones. When compared with cycles of younger control women, the cycles of the SWAN DHS participants had higher gonadotropins, lower total integrated Pdg, and E1c levels that were not different, which suggests that the ovary retains sensitivity to elevated FSH in the early menopausal transition. In this cross-sectional study of women over age 42 who are premenopausal or in the early menopausal transition, there were important differences in the characteristics of cycles related to age, body mass index, and ethnicity. Comparisons to younger women indirectly support the inhibin hypothesis, which proposes that the initiating event in the menopausal transition is the loss of inhibin negative feedback on FSH secondary to a diminished follicular reserve.

Immune cells in the normal ovary and spontaneous ovarian tumors in the laying hen (Gallus domesticus) model of human ovarian cancer.

BACKGROUND: Spontaneous ovarian cancer in chickens resembles human tumors both histologically and biochemically. The goal was to determine if there are differences in lymphocyte content between normal ovaries and ovarian tumors in chickens as a basis for further studies to understand the role of immunity in human ovarian cancer progression. METHODS: Hens were selected using grey scale and color Doppler ultrasound to determine if they had normal or tumor morphology. Cells were isolated from ovaries (n = 6 hens) and lymphocyte numbers were determined by flow cytometry using antibodies to avian CD4 and CD8 T and B (Bu1a) cells. Ovarian sections from another set of hens (n = 26) were assessed to verify tumor type and stage and to count CD4, CD8 and Bu1a immunostained cells by morphometric analysis. RESULTS: T and B cells were more numerous in ovarian tumors than in normal ovaries by flow cytometry and immunohistochemistry. There were less CD4+ cells than CD8+ and Bu1a+ cells in normal ovaries or ovarian tumors. CD8+ cells were the dominant T cell sub-type in both ovarian stroma and in ovarian follicles compared to CD4+ cells. Bu1a+ cells were consistently found in the stroma of normal ovaries and ovarian tumors but were not associated with follicles. The number of immune cells was highest in late stage serous tumors compared to endometrioid and mucinous tumors. CONCLUSIONS: The results suggest that similar to human ovarian cancer there are comparatively more immune cells in chicken ovarian tumors than in normal ovaries, and the highest immune cell content occurs in serous tumors. Thus, this study establishes a foundation for further study of tumor immune responses in a spontaneous model of ovarian cancer which will facilitate studies of the role of immunity in early ovarian cancer progression and use of the hen in pre-clinical vaccine trials.

Autoantibodies to mesothelin in infertility.

BACKGROUND: According to extensive epidemiologic data, infertility is associated with increased ovarian cancer risk. Previous studies showed that both women with infertility and those with ovarian cancer have autoantibodies to ovarian antigens. The objective was to determine if women with infertility have antibodies to mesothelin, a well-characterized ovarian cancer antigen. METHODS: Sera were obtained from women with infertility (n = 109), ovarian cancer (n = 28), benign ovarian tumors or cysts (n = 24), and from healthy women (n = 152). Infertility included those with a risk for ovarian cancer; endometriosis (n = 23), ovulatory dysfunction (n = 17), premature ovarian failure (POF; n = 25) and unexplained infertility (n = 44). Sera were assayed for mesothelin antibodies and for circulating mesothelin antigen by immunoassay and compared with assay control sera (n = 16) to determine a positive result. RESULTS: Mesothelin antibodies were significantly more frequent in women with prematurely reduced ovarian function including ovulatory dysfunction (59%), ovarian failure (44%) and unexplained infertility (25%) compared with controls. In contrast, women with endometriosis, who also have a high risk for ovarian cancer, did not have mesothelin antibodies. Serum levels of mesothelin were rarely elevated in women with infertility but were high in most patients with ovarian cancer. CONCLUSIONS AND IMPACT: We show for the first time that antibodies to mesothelin, a well-characterized ovarian cancer antigen, occur in some women with epidemiologic risk for ovarian cancer. The results suggest it may be possible to identify which women with infertility have ovarian cancer risk.

The hen model of human ovarian cancer develops anti-mesothelin autoantibodies in response to mesothelin expressing tumors.

OBJECTIVE: Study of the hen immune system led to seminal contributions to basic immunological principles. Recent studies of spontaneous ovarian cancer in the laying hen show strikingly similar tumor types and antigen expression compared to human ovarian cancer, suggesting hens would be valuable for studies of tumor immunology and pre-clinical vaccine development. Circulating mesothelin is a relatively specific marker for human ovarian cancer and autoantibodies to mesothelin were reported. We hypothesized that hen tumors express mesothelin and that circulating anti-mesothelin antibodies occur in response to tumors. METHODS: Mesothelin mRNA expression was analyzed by RT-PCR in hen ovarian tumors and normal ovaries. Mesothelin protein expression was evaluated by immunohistochemistry (IHC) and two-dimensional SDS-PAGE Western blots. Anti-mesothelin antibodies were assessed by immunoassay of sera from hens with normal ovaries and with ovarian tumors. RESULTS: Significant mesothelin mRNA expression was observed in 57% (12/21) of hen ovarian tumors but not in normal ovaries and was found predominantly in serous tumors as in humans. Mesothelin protein was detected in tumors with mesothelin mRNA by IHC and 2D Western blots, but not in normal ovaries or tumors without mesothelin mRNA. Circulating anti-mesothelin antibodies occurred in 44% (n = 4/9) of hens with ovarian tumors which express mesothelin mRNA and were not found in hens with tumors that did not express mesothelin (n = 0/5) or normal ovaries (n = 0/5). CONCLUSION: The results support the utility of the hen as a novel model for preclinical studies of mesothelin as a biomarker and a target for immunotherapy.

Sphingosine-1 phosphate receptor (S1p1), a critical receptor controlling human lymphocyte trafficking, is expressed in hen and human ovaries and ovarian tumors.

BACKGROUND: Sphingosine-1 receptor 1 (S1P1) plays a major role in regulating lymphocyte egress from peripheral lymph tissue. Lymphocyte trafficking is potentially a critical response to tumors and to tumor vaccines. Also, the receptor has been shown to influence metastasis. However, there is little information on its expression in the aged ovary or ovarian tumors. As a basis for further studies in the laying hen model of spontaneous ovarian cancer, the objective of this study was to determine if S1P1 is expressed in hens, and if the morphological distribution of S1P1 is similar in hen and human ovary and ovarian tumors. METHODS: S1P1 mRNA was ascertained in hen tissue by RT-PCR using hen specific primers. S1P1 protein expression and localization was evaluated in hen and human tissue with a human S1P1 antibody by Western blot and immunohistochemistry. RESULTS: S1P1 mRNA was expressed in all hen tissues examined. Protein was detected in human and hen ovary and ovarian tumors at 47, 72 and 108 kDa in Western blots. S1P1 was similarly expressed on endothelial cells, lymphocytes and surface epithelial cells in normal ovaries and tumor-containing ovaries of the hen. In addition, S1P1 distribution was heterogeneous in both hen and human ovarian tumors by immunohistochemistry. CONCLUSION: The results show that S1P1 is expressed in the hen and human ovary as well as in ovarian tumors. These findings support the use of the hen in further studies of the role of S1P1 in metastasis and immune cell trafficking in ovarian tumor development.

Differential expression of aldehyde dehydrogenase 1a1 (ALDH1) in normal ovary and serous ovarian tumors.

BACKGROUND: We showed there are specific ALDH1 autoantibodies in ovarian autoimmune disease and ovarian cancer, suggesting a role for ALDH1 in ovarian pathology. However, there is little information on the ovarian expression of ALDH1. Therefore, we compared ALDH1 expression in normal ovary and benign and malignant ovarian tumors to determine if ALDH1 expression is altered in ovarian cancer. Since there is also recent interest in ALDH1 as a cancer stem cell (CSC) marker, we assessed co-expression of ALDH1 with CSC markers in order to determine if ALDH1 is a potential CSC marker in ovarian cancer. METHODS: mRNA and protein expression were compared in normal human ovary and serous ovarian tumors using quantitative Reverse-Transcriptase PCR, Western blot (WB) and semi-quantitative immunohistochemistry (IHC). ALDH1 enzyme activity was confirmed in primary ovarian cells by flow cytometry (FC) using ALDEFLUOR assay. RESULTS: ALDH1 mRNA expression was significantly reduced (p < 0.01; n = 5) in malignant tumors compared to normal ovaries and benign tumors. The proportion of ALDH1+ cells was significantly lower in malignant tumors (17.1 ± 7.61%; n = 5) compared to normal ovaries (37.4 ± 5.4%; p < 0.01; n = 5) and benign tumors (31.03 ± 6.68%; p < 0.05; n = 5). ALDH1+ cells occurred in the stroma and surface epithelium in normal ovary and benign tumors, although surface epithelial expression varied more in benign tumors. Localization of ALDH1 was heterogeneous in malignant tumor cells and little ALDH1 expression occurred in poorly differentiated malignant tumors. In benign tumors the distribution of ALDH1 had features of both normal ovary and malignant tumors. ALDH1 protein expression assessed by IHC, WB and FC was positively correlated (p < 0.01). ALDH1 did not appear to be co-expressed with the CSC markers CD44, CD117 and CD133 by IHC. CONCLUSIONS: Total ALDH1 expression is significantly reduced in malignant ovarian tumors while it is relatively unchanged in benign tumors compared to normal ovary. Thus, ALDH1 expression in the ovary does not appear to be similar to breast, lung or colon cancer suggesting possible functional differences in these cancers. SIGNIFICANCE: These observations suggest that reduced ALDH1 expression is associated with malignant transformation in ovarian cancer and provides a basis for further study of the mechanism of ALDH1 in this process.

Autoantigens in ovarian autoimmunity associated with unexplained infertility and premature ovarian failure.

OBJECTIVE: To identify ovarian autoantigens associated with ovarian autoantibodies. DESIGN: Hypothesis-generating prospective study. SETTING: Urban infertility referral centers and academic research institution. PATIENT(S): Seventy-four patients with infertility, 19 patients with premature ovarian failure (POF), and 16 healthy control women. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Identification of autoantigens. RESULT(S): To identify major antigens for ovarian autoimmunity, sera from 74 women with unexplained infertility were screened for ovarian autoantibodies (AOAs) by immunoassay and one-dimensional Western blot. The majority of sera had immunoreactions at 50-56 kDa. Six representative positive infertility sera were used to identify antigens between 40 and 60 kD by two-dimensional Western blot and mass spectrometry. Antigens included aldehyde (retinal) dehydrogenases (ALDH1A1, ALDH1A2, and ALDH7A1), protein disulfide isomerase A3, vimentin, α-enolase, phosphoglycerate dehydrogenase, and selenium-binding protein 1 (SBP1). Sixty percent (24 out of 40) of infertility and POF sera were positive for recombinant ALDH1A1, SBP1, or enolase; 80.7% (21 out of 26) of AOA-positive sera had antibodies to one or more of the three antigens, and only 7% (1 out of 14) of AOA-negative sera had antibodies to recombinant proteins. CONCLUSION(S): ALDH1A1 and SBP1 are unique to ovarian autoimmunity associated with infertility and POF, and may provide the basis for specific tests to identify patients with ovarian autoimmunity.

Cyclooxygenases expression and distribution in the normal ovary and their role in ovarian cancer in the domestic hen (Gallus domesticus).

Cyclooxygenase (COX) (PTGS) is the rate-limiting enzyme in the biosynthesis of prostaglandins. Two COX isoforms have been identified, COX-1 and COX-2, which show distinct cell-specific expression and regulation. Ovarian cancer is the most lethal gynecological malignancy and the disease is poorly understood due to the lack of suitable animal models. The laying hen spontaneously develops epithelial ovarian cancer with few or no symptoms until the cancer has progresses to a late stage, similar to the human disease. The purpose of this study was to examine the relative expression and distribution of COX-1 and COX-2 in the ovaries of normal hens and in hens with ovarian cancer. The results demonstrate that COX-1 was localized to the granulosa cell layer and cortical interstitium, ovarian surface epithelium (OSE) and postovulatory follicle (POF) of the normal ovary. In ovarian cancer, COX-1 mRNA was significantly increased and COX-1 protein was broadly distributed throughout the tumor stroma. COX-2 protein was localized to the granulosa cell layer in the follicle and the ovarian stroma. COX-2 mRNA expression did not change as a function of age or in ovarian cancer. There was significantly higher expression of COX-1 mRNA in the first POF (POF-1) compared to POF-2 and POF-3. COX-2 mRNA expression was not significantly different among POFs. There was no difference in COX-1 or COX-2 mRNA in the OSE isolated from individual follicles in the follicular hierarchy. The results confirm previous findings of the high expression of COX-1 in ovarian tumors further supporting the laying hen as a model for ovarian cancer, and demonstrate for the first time the high expression of COX-1 in POF-1 which is the source of prostaglandins needed for oviposition.

Anti-tumor and anti-ovarian autoantibodies in women with ovarian cancer.

PROBLEM: There is a lack of validated marker(s) for the diagnosis of early-stage ovarian cancer (OVCA). The objective was to determine if women with OVCA had antibodies, to assess their potential as markers of ovarian cancer. The secondary objective was to compare the prevalence of antibodies to proteins from normal ovary and ovarian tumors to determine if antibodies primarily recognize tumor antigens, as many antigens are common to tumor and normal ovary. METHOD OF STUDY: Serum samples from patients with OVCA, borderline or benign ovarian tumors, endometrial cancer and healthy women were examined for anti-ovarian and anti-tumor antibodies by immunoassay. Immunoreactive proteins were characterized by one- and two-dimensional Western blot. RESULTS: Ovarian (81%, P < or = 0.001) and anti-tumor (69%, P < or = 0.001) autoantibodies in OVCA were significantly different from those of control sera. A majority of OVCA serum samples reacted with proteins at about 50 kDa from normal ovary or ovarian tumors in one-dimensional Western blot. While there were similar reactions in two-dimensional Western blots, there are differences between reactions to normal and tumor antigens and between reactions to autologous and allogeneic tumors. CONCLUSION: Serum autoantibodies are significantly associated with OVCA. Anti-tumor antibodies may provide a useful marker for the detection of ovarian cancer.