Assessment of micronucleus frequency in the peripheral blood of female rats in persistent estrus treated with selective estrogen receptor modulators

The aim of this study was to evaluate micronucleus (MN) frequency in polychromatic erythrocytes (PCE) of female rats in persistent estrus (a model developed to mimic polycystic ovary syndrome) treated with selective estrogen receptor modulators (SERMs, tamoxifen, and raloxifene). Forty female Wistar-Hannover rats were divided into four groups of 10 animals each: Group I (normally cycling rats) and Group II (persistent estrus) both received only vehicle, while Group III (persistent estrus) was treated with tamoxifen (250 mu g/animal/day) and Group IV (persistent estrus) was treated with raloxifene (750 mu g/animal/day). Tamoxifen and raloxifene were given by oral gavage beginning on postnatal day 90 and continuing for 30 consecutive days. Peripheral blood samples were collected from tails 1 day following the last exposure. Blood smears were made on glass slides and stained with 10% Giemsa solution. ANOVA and a Tukey post-hoc test were used for data analysis. Mean percentages of MN were 1.82 +/- 0.13, 5.20 +/- 0.24, 3.32 +/- 0.13, and 3.04 +/- 0.12 in Groups I, II, III, and IV, respectively. The results indicate that tamoxifen and raloxifene similarly reduced the formation of MNPCE of female rats in persistent estrus (P < 0.0001 for Groups III and IV vs. Group II), using the dosages and time periods applied in the present study. The data suggest possibly antimutagenic effects of SERMs under high levels of estrogens. The findings also suggest that this is an interesting animal model for studying the genotoxicity of estrogens. Environ. Mol. Mutagen. 2012. (C) 2011 Wiley Periodicals, Inc.

High-Fat Diet Obesity Associated With Insulin Resistance Increases Cell Proliferation, Estrogen Receptor, and PI3K Proteins in Rat Ventral Prostate

In this study, we evaluated the effects of obesity and insulin resistance induced by a high-fat diet on prostate morphophysiology, focusing on cell proliferation, expression of androgen (AR) and estrogen receptors (ER) and proteins of the insulin signaling pathway. Adult male Wistar rats were fed a high-fat diet (20% fat) for 15 weeks, whereas control animals received a balanced diet (4% fat). Both groups were then divided and treated for 2 weeks with 1 mg/kg body weight/day of the aromatase inhibitor letrozole or vehicle only. The ventral prostate was analyzed with immunohistochemical, histopathological, stereological, and Western blotting methods. Obese rats showed insulin resistance, hyperinsulinemia, and reduced plasma testosterone levels. The incidence of prostatic intraepithelial neoplasia (PIN) was 2.7 times higher in obese rats and affected 0.4% of the gland compared with 0.1% PIN areas found in control rats. Obesity doubled cell proliferation in both prostate epithelium and stroma. AR content decreased in the prostate of obese rats and estrogen receptor beta (ER beta) increased in this group. Protein levels of insulin receptor substrate 1 and protein kinase B diminished in the obese group, whereas phosphatidylinositol 3-kinase (PI3K) increased significantly. Most structural changes observed in the prostate of obese rats normalized after letrozole treatment, except for increased stromal cell proliferation and ER beta expression, which might be associated with insulin resistance. This experimental model of obesity and insulin resistance induced by a high-fat diet increases cell proliferation in rat prostate. Such alterations are associated with decreased levels of AR and increased ER beta and PI3K proteins. This change can facilitate the establishment of proliferative lesions in rat prostate.

VIP expression in vessels of the vaginal wall: relationship with estrogen status

Objective Vasoactive intestinal peptide (VIP) is a neuropeptide with elevated expression in regions that control urogenital functions. Estrogen appears to modulate VIP expression in various organs, but this effect has not been demonstrated in the vaginal wall. The aim of this study was to evaluate the influence of estrogen status on VIP expression in vessels of the vaginal wall. Methods Surgical specimens were removed from the vaginal walls of 18 premenopausal women and 12 postmenopausal women who were given surgery for genital prolapse grade I or II. Vaginal specimens were stained with estrogen receptor-alpha (ER-alpha) and VIP antibodies. Levels of follicle stimulating hormone (FSH), estradiol, prolactin, fasting glucose and serum thyroxine stimulating hormone were also measured. Estrogen status was assessed on the basis of FSH and ER-alpha scores. Results The vaginal walls of premenopausal women had significantly higher ER-alpha scores than those of menopausal women (premenopausal group, 3.6 +/- 2.2; menopausal group, 1.4 +/- 1.8; p = 0.01). Premenopausal women also had significantly higher levels of VIP in the vaginal wall than menopausal women (p = 0.02). Increasing age was associated with lower level of VIP staining (odds ratio 0.88; 95% confidence interval 0.78-0.99). Conclusion Levels of ER-alpha and VIP expression in the posterior vaginal wall were higher in premenopausal than in menopausal women, but VIP expression was not associated with estrogen status. Age was an independent predictor of VIP staining in vaginal wall biopsies.

Dose-dependent change in biomarkers during neoadjuvant endocrine therapy with fulvestrant: results from NEWEST, a randomized Phase II study

NEWEST (Neoadjuvant Endocrine Therapy for Women with Estrogen-Sensitive Tumors) is the first study to compare biological and clinical activity of fulvestrant 500 versus 250 mg in the neoadjuvant breast cancer setting. We hypothesized that fulvestrant 500 mg may be superior to 250 mg in blocking estrogen receptor (ER) signaling and growth. A multicenter, randomized, open-label, Phase II study was performed to compare fulvestrant 500 mg (500 mg/month plus 500 mg on day 14 of month 1) versus fulvestrant 250 mg/month for 16 weeks prior to surgery in postmenopausal women with ER+ locally advanced breast cancer. Core biopsies at baseline, week 4, and surgery were assessed for biomarker changes. Primary endpoint: change in Ki67 labeling index (LI) from baseline to week 4 determined by automated computer imaging system (ACIS). Secondary endpoints: ER protein expression and function; progesterone receptor (PgR) expression; tumor response; tolerability. ER and PgR were examined retrospectively using the H score method. A total of 211 patients were randomized (fulvestrant 500 mg: n = 109; 250 mg: n = 102). At week 4, fulvestrant 500 mg resulted in greater reduction of Ki67 LI and ER expression versus 250 mg (-78.8 vs. -47.4% [p < 0.0001] and -25.0 vs. -13.5% [p = 0.0002], respectively [ACIS]); PgR suppression was not significantly different (-22.7 vs. -17.6; p = 0.5677). However, H score detected even greater suppression of ER (-50.3 vs. -13.7%; p < 0.0001) and greater PgR suppression (-80.5 vs. -46.3%; p = 0.0018) for fulvestrant 500 versus 250 mg. At week 16, tumor response rates were 22.9 and 20.6% for fulvestrant 500 and 250 mg, respectively, with considerable decline in all markers by both ACIS and H score. No detrimental effects on endometrial thickness or bone markers and no new safety concerns were identified. This provides the first evidence of greater biological activity for fulvestrant 500 versus 250 mg in depleting ER expression, function, and growth.

Bone mineral density and body composition in girls with idiopathic central precocious puberty before and after treatment with a gonadotropin-releasing hormone agonist

OBJECTIVES: Idiopathic central precocious puberty and its postponement with a (gonadotropin-releasing hormone) GnRH agonist are complex conditions, the final effects of which on bone mass are difficult to define. We evaluated bone mass, body composition, and bone remodeling in two groups of girls with idiopathic central precocious puberty, namely one group that was assessed at diagnosis and a second group that was assessed three years after GnRH agonist treatment. METHODS: The precocious puberty diagnosis and precocious puberty treatment groups consisted of 12 girls matched for age and weight to corresponding control groups of 12 (CD) and 14 (CT) girls, respectively. Bone mineral density and body composition were assessed by dual X-ray absorptiometry. Lumbar spine bone mineral density was estimated after correction for bone age and the mathematical calculation of volumetric bone mineral density. CONEP: CAAE-0311.0.004.000-06. RESULTS: Lumbar spine bone mineral density was slightly increased in individuals diagnosed with precocious puberty compared with controls; however, after correction for bone age, this tendency disappeared (CD = -0.74 +/- 0.9 vs. precocious puberty diagnosis = -1.73 +/- 1.2). The bone mineral density values of girls in the precocious puberty treatment group did not differ from those observed in the CT group. CONCLUSION: There is an increase in bone mineral density in girls diagnosed with idiopathic central precocious puberty. Our data indicate that the increase in bone mineral density in girls with idiopathic central precocious puberty is insufficient to compensate for the marked advancement in bone age observed at diagnosis. GnRH agonist treatment seems to have no detrimental effect on bone mineral density.

Antifibrotic Effect of Tamoxifen in a Model of Progressive Renal Disease

Tamoxifen, a selective estrogen receptor modulator, has antifibrotic properties; however, whether it can attenuate renal fibrosis is unknown. In this study, we tested the effects of tamoxifen in a model of hypertensive nephrosclerosis (chronic inhibition of nitric oxide synthesis with L-NAME). After 30 days, treated rats had significantly lower levels of albuminuria as well as lower histologic scores for glomerulosclerosis and interstitial fibrosis than untreated controls. Tamoxifen was renoprotective despite having no effect on the sustained, severe hypertension induced by L-NAME. Tamoxifen prevented the accumulation of extracellular matrix by decreasing the expression of collagen I, collagen III, and fibronectin mRNA and protein. These renoprotective effects associated with inhibition of TGF-beta 1 and plasminogen activator inhibitor-1, and with a significant reduction in a-smooth muscle actin-positive cells in the renal interstitium. Furthermore, tamoxifen abrogated IL-1 beta- and angiotensin-II-induced proliferation of fibroblasts from both kidney explants and from the NRK-49F cell line. Tamoxifen also inhibited the expression of extracellular matrix components and the production and release of TGF-beta 1 into the supernatant of these cells. In summary, tamoxifen exhibits antifibrotic effects in the L-NAME model of hypertensive nephrosclerosis, likely through the inhibition of TGF-beta 1, suggesting that it may have therapeutic use in CKD treatment.

Tamoxifen Subcellular Localization; Observation of Cell-Specific Cytotoxicity Enhancement by Inhibition of Mitochondrial ETC Complexes I and III

Recently, a nongenomic cytotoxic component of the chemotherapeutic agent tamoxifen (TAM) has been identified that predominantly triggers mitochondrial events. The present study delineates the intracellular fate of TAM and studies its interaction with a spectrum of cell homeostasis modulators primarily relevant to mitochondria. The subcellular localization of TAM was assessed by confocal fluorescence microscopy. The effect of the modulators on TAM cytotoxicity was assessed by standard MTT assays. Our findings show that in estrogen receptor positive MCF7 breast adenocarcinoma cells and DU145 human prostate cancer cells, TAM largely accumulates in the mitochondria and endoplasmic reticulum, but not lysosomes. Our results further demonstrate that in MCF7, but not in DU145 cells, mitochondrial electron transport chain complex I and III inhibitors exacerbate TAM toxicity with an order of potency of myxothiazol = stigmatellin > rotenone > antimycin A, suggesting a cell-specific cytotoxic interplay between mitochondrial complex I and III function and TAM action.

17 beta-Estradiol replacement in young, adult and middle-aged female ovariectomized rats promotes improvement of spatial reference memory and an antidepressant effect and alters monoamines and BDNF levels in memory- and depression-related brain areas

Clinical and experimental evidence suggest that estrogens have a major impact on cognition, presenting neurotrophic and neuroprotective actions in regions involved in such function. In opposite, some studies indicate that certain hormone therapy regimens may provoke detrimental effects over female cognitive and neurological function. Therefore, we decided to investigate how estrogen treatment would influence cognition and depression in different ages. For that matter, this study assessed the effects of chronic 17 beta-estradiol treatment over cognition and depressive-like behaviors of young (3 months old), adult (7 months old) and middle-aged (12 months old) reproductive female Wistar rats. These functions were also correlated with alterations in the serotonergic system, as well as hippocampal BDNF. 17 beta-Estradiol treatment did not influence animals' locomotor activity and exploratory behavior, but it was able to improve the performance of adult and middle-aged rats in the Morris water maze, the latter being more responsive to the treatment. Young and adult rats displayed decreased immobility time in the forced swimming test, suggesting an effect of 17 beta-estradiol also over such depressive-like behavior. This same test revealed increased swimming behavior, triggered by serotonergic pathway, in adult rats. Neurochemical evaluations indicated that 17 beta-estradiol treatment was able to increase serotonin turnover rate in the hippocampus of adult rats. Interestingly, estrogen treatment increased BDNF levels from animals of all ages. These findings support the notion that the beneficial effects of 17 beta-estradiol over spatial reference memory and depressive-like behavior are evident only when hormone therapy occurs at early ages and early stages of hormonal decline. (C) 2011 Elsevier B.V. All rights reserved.

Prognostic significance of CD24 and claudin-7 immunoexpression in ductal invasive breast cancer

This study aimed to identify the CD24 and CD44 immunophenotypes within invasive ductal breast carcinoma (I DC) subgroups defined by immunohistochesmistry markers and to determine its influence on prognosis as well as its association with the expression of Ki-67, cytokeratins (CK5 and CK 18) and claudin-7. Immunohistochemical expression of CD44 and CD24 alone or in combination was investigated in 95 IDC cases arranged in a tissue microarray (TMA). The association with subgroups defined as luminal A and B; HER2 rich and triple negative, or with the other markers and prognosis was analyzed. CD44(+)/CD24(-) and CD44(-)/CD24(+) were respectively present in 8.4% and 16.8% of the tumors, a lack of both proteins was detected in 6.3%, while CD441(-)/CD24(+) was observed in 45.3% of the tumors. Although there was no significant correlation between subgroups and different phenotypes, the CD44(+)/CD24(-) phenotype was more common in the basal subgroups but absent in HER2 tumors, whereas luminal tumors are enriched in CD44(-)/CD24(+) and CD44(+)/CD24(+) cells. The frequency of CD44(+)/CD24(-) or CD44(-)/CD24(+) was not associated with clinical characteristics or biological markers. There was also no significant association of these phenotypes with the event free (DFS) and overall survival (OS). Single CD44(+) was evident in 57.9% of the tumors and was marginally associated to grading and not to any other tumor characteristics as well as OS and DFS. CD24(+) was positive in 74.7% of the tumors, showing a significant association with estrogen receptor, progesterone receptor and Ki-67 and a marginal association with CKI8 and claudin-7. Expression of claudin-7 and Ki-67 did not associate with the cancer subgroups, while a positive association between CK18 and the luminal subgroups was found (P=0.03). CK5, CK18 and Ki-67 expression had no influence in OS or DFS. Single CD24(+) (P=0.07) and claudin-7 positivity (P=0.05) were associated with reduced time of recurrence, suggesting a contribution of these markers to aggressiveness of breast cancer.

Analysis of the concordance rates between core needle biopsy and surgical excision in patients with breast cancer

Objective: To evaluate whether immunohistochemical marker studies performed on core needle biopsy (CNB) specimens accurately reflect the marker status of the tumor obtained from final surgical specimen. Methods: This was a retrospective study that used the database of the Division of Mastology of the Hospital das Clinicas, Sao Paulo, Brazil. Sixty-nine patients submitted to ultrasound-guided CNB diagnosed with breast cancer were retrospectively analyzed. Immunohistochemistry (IHC) on core biopsy specimens was compared to that of excisional biopsy regarding estrogen receptor (ER), progesterone receptor (PR), human epidermal gowth factor receptor 2 gene (HER2), p53, and Ki67. The analysis of the concordance between CNB and surgical biopsy was performed using the kappa (k) coefficient (95% CI). Results: A perfect concordance between the labeling in the surgical specimens and the preoperative biopsies in p53 (k = 1.0; 95% CI: 0.76-1.0) was identified. There was an almost perfect concordance for ER (k = 0.89; 95% CI: 0.65-1.0) and a substantial concordance for PR (k = 0.70; 95% CI: 0.46-0.93). HER2 (k = 0.61; 95% CI: 0.38-0.84) and Ki-67 (k = 0.74; 95% CI: 0.58-0.98) obtained a substantial concordance this analysis. Conclusion: The results of this study indicate that the immunohistochemical analysis of ER, PR, Ki-67, and p53 from core biopsy specimens provided results that accurately reflect the marker status of the tumor. The concordance rate of HER2 was less consistent; although it produced substantial concordance, values were very close to moderate concordance.

Cu(GlyGlyHis) effects on MCF7 cells: Copper uptake, reactive oxygen species generation and membrane topography changes

Elevated levels of copper have been detected in various types of human cancer cells, such as breast cancer cells, and a number of mechanisms have been proposed to explain the action and influence of copper on tumor progress. In this work, we found that stimulating the proliferation of mammary epithelial MCF7 cells with the high-redox-potential copper complex Cu (GlyGlyHis) is associated with the copper-induced intracellular generation of reactive oxygen species (ROS) that induces lipid peroxidation and causes increased roughness of external cell membranes, which leads to the formation of larger cell domes. The results presented herein provide new insights into the molecular link between copper and the proliferation of breast cancer cells and, consequently, into the mechanism by which changes in redox balance and ROS accumulation regulates cell membrane roughness. (C) 2012 Elsevier Inc. All rights reserved.

PTTG expression in different experimental and human prolactinomas in relation to dopaminergic control of lactotropes

Abstract Background Pituitary tumor transforming gene (pttg) is a novel oncogene that is expressed at higher level in most of the tumors analyzed to date compared to normal tissues. Nevertheless, its expression in prolactinomas and its relation with the pituitary dopamine receptor 2 (D2R) are not well defined. We sought to determine the pituitary level of pttg in three different experimental models of prolactinomas with altered dopaminergic control of the pituitary: the dopaminergic D2R knockout female mouse, the estrogen-treated rat, and the senescent female rat. These three models shared the characteristics of increased pituitary weight, hyperprolactinemia, lactotrope hyperplasia and reduced or absent dopaminergic action at the pituitary level. We also studied samples from human macroprolactinomas, which were characterized as responsive or resistant to dopamine agonist therapy. Results When compared to female wild-type mice, pituitaries from female D2R knockout mice had decreased PTTG concentration, while no difference in pttg mRNA level was found. In senescent rats no difference in pituitary PTTG protein expression was found when compared to young rats. But, in young female rats treated with a synthetic estrogen (Diethylstylbestrol, 20 mg) PTTG protein expression was enhanced (P = 0.029). Therefore, in the three experimental models of prolactinomas, pituitary size was increased and there was hyperprolactinemia, but PTTG levels followed different patterns. Patients with macroprolactinomas were divided in those in which dopaminergic therapy normalized or failed to normalize prolactin levels (responsive and resistant, respectively). When pituitary pttg mRNA level was analyzed in these macroprolactinomas, no differences were found. We next analyzed estrogen action at the pituitary by measuring pituitary estrogen receptor α levels. The D2R knockout female mice have low estrogen levels and in accordance, pituitary estrogen receptors were increased (P = 0.047). On the other hand, in senescent rats estrogen levels were slightly though not significantly higher, and estrogen receptors were similar between groups. The estrogen-treated rats had high pharmacological levels of the synthetic estrogen, and estrogen receptors were markedly lower than in controls (P < 0.0001). Finally, in patients with dopamine resistant or responsive prolactinomas no significant differences in estrogen receptor α levels were found. Therefore, pituitary PTTG was increased only if estrogen action was increased, which correlated with a decrease in pituitary estrogen receptor level. Conclusion We conclude that PTTG does not correlate with prolactin levels or tumor size in animal models of prolactinoma, and its pituitary content is not related to a decrease in dopaminergic control of the lactotrope, but may be influenced by estrogen action at the pituitary level. Therefore it is increased only in prolactinomas generated by estrogen treatment, and not in prolactinomas arising from deficient dopamine control, or in dopamine resistant compared with dopamine responsive human prolactinomas. These results are important in the search for reliable prognostic indicators for patients with pituitary adenomas which will make tumor-specific therapy possible, and help to elucidate the poorly understood phenomenon of pituitary tumorigenesis.

Melatonin reduces LH, 17 beta-estradiol and induces differential regulation of sex steroid receptors in reproductive tissues during rat ovulation

Abstract Background Melatonin is associated with direct or indirect actions upon female reproductive function. However, its effects on sex hormones and steroid receptors during ovulation are not clearly defined. This study aimed to verify whether exposure to long-term melatonin is able to cause reproductive hormonal disturbances as well as their role on sex steroid receptors in the rat ovary, oviduct and uterus during ovulation. Methods Twenty-four adult Wistar rats, 60 days old (+/- 250 g) were randomly divided into two groups. Control group (Co): received 0.9% NaCl 0.3 mL + 95% ethanol 0.04 mL as vehicle; Melatonin-treated group (MEL): received vehicle + melatonin [100 μg/100 g BW/day] both intraperitoneally during 60 days. All animals were euthanized by decapitation during the morning estrus at 4 a.m. Results Melatonin significantly reduced the plasma levels of LH and 17 beta-estradiol, while urinary 6-sulfatoximelatonin (STM) was increased at the morning estrus. In addition, melatonin promoted differential regulation of the estrogen receptor (ER), progesterone receptor (PR), androgen receptor (AR) and melatonin receptor (MTR) along the reproductive tissues. In ovary, melatonin induced a down-regulation of ER-alpha and PRB levels. Conversely, it was observed that PRA and MT1R were up-regulated. In oviduct, AR and ER-alpha levels were down-regulated, in contrast to high expression of both PRA and PRB. Finally, the ER-beta and PRB levels were down-regulated in uterus tissue and only MT1R was up-regulated. Conclusions We suggest that melatonin partially suppress the hypothalamus-pituitary-ovarian axis, in addition, it induces differential regulation of sex steroid receptors in the ovary, oviduct and uterus during ovulation.

Manipulation of the periovulatory sex steroidal milieu affects endometrial but not luteal gene expression in early diestrus Nelore cows

In beef cattle, the ability to conceive has been associated positively with size of the preovulatory follicle (POF). Proestrus estradiol and subsequent progesterone concentrations can regulate the endometrium to affect receptivity and fertility. The aim of the present study was to verify the effect of the size of the POF on luteal and endometrial gene expression during subsequent early diestrus in beef cattle. Eighty-three multiparous, nonlactating, presynchronized Nelore cows received a progesterone-releasing device and estradiol benzoate on Day–10 (D 10). Animals received cloprostenol (large follicle-large CL group; LF-LCL; N ¼ 42) or not (small follicle-small CL group; SF-SCL; N ¼ 41) on D 10. Progesterone devices were withdrawn and cloprostenol administered 42 to 60 hours (LF-LCL) or 30 to 36 hours (SF-SCL) before GnRH treatment (D0). Tissues were collected at slaughter on D7. The LF-LCL group had larger (P < 0.0001) POF (13.24 0.33 mm vs. 10.76 0.29 mm), greater (P < 0.0007) estradiol concentrations on D0 (2.94 0.28 pg/mL vs. 1.27 0.20 pg/mL), and greater (P < 0.01) progesterone concentrations on D7 (3.71 0.25 ng/mL vs. 2.62 0.26 ng/mL) compared with the SF-SCL group. Luteal gene expression of vascular endothelial growth factor A, kinase insert domain receptor, fms-related tyrosine kinase 1, steroidogenic acute regulatory protein, cytochrome P450, family 11, subfamily A, polypeptide 1, and hydroxy-delta-5-steroid dehydrogenase, 3 beta- and steroid deltaisomerase 7 was similar between groups. Endometrial gene expression of oxytocin receptor and peptidase inhibitor 3, skin-derived was reduced, and estrogen receptor alpha 2, aldo-keto reductase family 1, member C4, and lipoprotein lipase expression was increased in LF-LCL versus SF-SCL. Results support the hypothesis that the size of the POF alters the periovulatory endocrine milieu (i.e., proestrus estradiol and diestrus progesterone concentrations) and acts on the uterus to alter endometrial gene expression. It is proposed that the uterine environment and receptivity might also be modulated. Additionally, it is suggested that increased progesterone secretion of cows ovulating larger follicles is likely due to increased CL size rather than increased luteal expression of steroidogenic genes.

Zearalenone, an estrogenic mycotoxin, is an immunotoxic compound

The aim of this study was to assess the toxic effects of zearalenone (ZEA) on the immune function. Ovariectomised rats were treated daily by gavage with 3.0 mg/kg of ZEA for 28 days. Body weight gain, food consumption, haemotological parameters, lymphoid organs, and their cellularities were evaluated. Moreover, acquired immune responses and macrophage activity were also assessed. ZEA promoted reduction in body weight gain, which is not fully explained by diminished food consumption. Despite no effect on haematological parameters, ZEA caused thymic atrophy with histological and thymocyte phenotype changes and decrease in the B cell percentage in the spleen. With respect to acquired and innate immune responses, no statistically significant differences in delayed-type hypersensitivity were noticed; however, in the ZEA-treated rats, antibody production and peroxide release by macrophages were impaired. The observed results could be related to ZEA activity on ERs; thus, ZEA is an immunotoxic compound similar to estrogen and some endocrine disruptors.