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Elevated Levels of Estrogen Suppress Hepcidin Synthesis and Enhance Serum Iron Availability in Premenopausal Women
Abstract
Clinical and experimental observations have long suggested that elevated levels of estrogen associate with increased serum iron availability. Additionally, recent work has shown that estrogen can downregulate hepcidin synthesis in vitro. This study aims at assessing whether the ability of estrogen to downregulate hepcidin synthesis translates into changes in serum iron status. Hepcidin synthesis was evaluated in MCF-7, Hep-G2 and SKOV-3 cells treated with increasing concentrations of estrogen and cultured for up to 24 h post treatment. The correlation between levels of serum estrogen, hepcidin and iron was assessed using serum samples collected from 153 premenopausal women at random and samples collected from 6 women at days 1, 5, 10, 16, 21 and 28 of the monthly cycle. Estrogen-treated MCF-7 cells showed a significant reduction in hepcidin synthesis, especially at 20 nM/24 h E2 treatment. Hepcidin synthesis was also significantly reduced in Hep-G2 and SKOV-3 cells at 20 nM/24 h E2 treatment. In serum samples collected at random, estrogen (P=0.022; R=−0.213) and iron (P=0.028; R=−0.316) correlated negatively with hepcidin and positively with each other (P=0.033; R=0.319). An overall similar pattern was also observed in monthly cycle-timed samples. These findings suggest that elevated levels of estrogen reduce hepcidin synthesis as means of enhancing serum iron content in menstruating women.
... Several lines of evidence in humans have supported the relationship between high concentrations of sex steroids and Hepc production. Specifically, high estrogen concentrations have been shown to downregulate Hepc synthesis [12][13][14] and reduce Fpn expression at the cell membrane [15]. Progesterone (P4) is associated with the opposite effect [16]. ...
... Experimental evidence in women suggests that estrogens lead to a greater availability of Fe due to the inhibitory effect of this hormone on Hepc synthesis [32]. The decrease of Hepc maintains the integrity of Fpn and increases the release of Fe by duodenal enterocytes (which absorb Fe), and macrophages and hepatocytes (which store Fe) [14]. ...
... Although the exact mechanism by which E2 influences Fe regulation is unknown, some in vitro studies in human breast [14], human ovarian cancer cell lines with epithelial-like morphology [32], human liver cells [21,32], and rodents have suggested that E2 positively regulates the genes involved in Fe metabolism (ferroportin, lactotransferrin, ceruloplasmin ferroxidase, lipocalin 2) [14,15], probably mediated likely by downregulation of Hepc activity. In ovarian epithelial cancer cells, E2-induced downregulation of Hepc gene (HAMP) expression resulting in reduced Hepc concentration [21,32]. ...
Several studies have demonstrated that in woman the sex hormones such as estrogen (E2) and progesterone (P4) influence iron (Fe) regulation, contributing to variations in Fe parameters along the menstrual cycle. These mechanisms based on the regulation of hepcidin (Hepc) which limits Fe availability during the cycle, remain poorly characterized in healthy mares. The objective of this study was to establish the relationship between Hepc, Fe, ferritin (Ferr), and the primary ovarian hormones E2 and P4 in cycling Purebred Spanish mares. Blood samples were taken from 31 Purebred Spanish mares day −5, on day 0, day +5 and day +16 of the cycle. Fe and Ferr significantly increased and Hepc decreased during pre- and ovulatory periods. The secretion peak of estradiol-17β (E2) was reached on day 0 and progesterone (P4) between days +5 and +16. Fe and Ferr were positively correlated (r = 0.57). Fe and Ferr were negatively correlated with Hepc (r = −0.72 and r = −0.02, respectively). E2 and P4 were negatively and positively correlated with Hepc (r = −0.753 and r = 0.54, respectively). In cycling Purebred Spanish mares there is a measurable relationship between steroid hormones and systemic Fe metabolism. Estrogenic dominance in the pre- and ovulatory period allows for a more effective iron status, mediated by hepcidin inhibition. However, P4 during the luteal phase substantially reduces serum Fe and iron stores, possibly related to Hepc stimulation. Future research is required to clarify the relationship between steroid hormones and iron metabolism at the molecular level in equids.
... Although the exact mechanism by which E 2 participates in Fe regulation remains unknown, some in vitro studies in breast (MCF7 and MDA-MB231) [3], ovarian (SKOV3) [37] and liver (HepG2) [38] cancer cell lines of women and other experimental models in rodents suggest that E 2 can positively regulate genes involved in Fe metabolism [1]. In this context, it has been reported that the human E 2 downregulates Hepc synthesis and upregulates Fpn expression, as a means of enhancing intracellular iron efflux [3]. ...
... Although the exact mechanism by which E 2 participates in Fe regulation remains unknown, some in vitro studies in breast (MCF7 and MDA-MB231) [3], ovarian (SKOV3) [37] and liver (HepG2) [38] cancer cell lines of women and other experimental models in rodents suggest that E 2 can positively regulate genes involved in Fe metabolism [1]. In this context, it has been reported that the human E 2 downregulates Hepc synthesis and upregulates Fpn expression, as a means of enhancing intracellular iron efflux [3]. In ovarian cells, the positive regulation of the transcription factor HIF-1a, that regulates a variety of proteins induced by E 2 , reduces the expression of the Hepcidin Antimicrobial Peptide gene (HAMP), decreasing the concentration of Hepc [38]. ...
In woman and in animal models, estrogens are involved in iron (Fe) homeostasis supporting the hypothesis of the existence of an "estrogen-iron axis". Since advancing age leads to a decrease in estrogen levels, the mechanisms of Fe regulation could be compromised. In cyclic and pregnant mares, to date, there is evidence linking the iron state with estrogens pattern. Then, the objective of this study was to determine the relationship among Fe, ferritin (Ferr), hepcidin (Hepc) and estradiol-17β (E2) in cyclic mares with advancing age. A total of 40 Spanish Purebred mares of different ranges of age was analyzed: 4-6 years (n = 10), 7-9 years (n = 10), 10-12 years (n = 10), and >12 years (n = 10). Blood samples were obtained on days -5, 0, +5 and + 16 of the cycle. Compared to mares of 4-6 years, serum Ferr was significantly higher (P < 0.01) and Fe significantly lower (P < 0.01) in mares >12 years of age. Hepc was significantly higher in mares >12 years (P < 0.01) than in those 7-9 years of age. E2 levels were higher in mares of 7-9 years (P < 0.01) than in 4-6 and >12 years of age. Fe and Ferr were negatively correlated with Hepc (r = -0.71 and r = -0.02, respectively). E2 was negatively correlated with Ferr and Hepc (r = -0.28 and r = -0.50, respectively), and positively with Fe (r = 0.31). There is a direct relationship between E2 and Fe metabolism, mediated by the inhibition of Hepc in Spanish Purebred mares. The reduction of E2 decreases the inhibitory effects on Hepc, increasing the levels of stored Fe and mobilizing less the free Fe in circulation. Based on the fact that ovarian estrogens participate in changes in the parameters indicative of iron status with age, the existence of an "estrogen-iron axis" in the mares'estrous cycle could be considered. Future studies are required to clarify these hormonal and metabolic interrelationships in the mare.
... Estrogen is considered the primary female sex steroid, and elevated concentrations of E2 have been positively correlated with iron demand and the release of iron into the systemic circulation, and negatively correlated with hepcidin concentration (Hamad et al., 2020), however, the exact mechanism by which E2 influences iron regulation is still to be determined. In vitro studies in breast (Bajbouj et al., 2018), ovarian SKOV3 (Yang et al., 2012), liver cells (HUH7 and Hep-G2) (Hou et al., 2012) and rodent models suggest E2 may support the upregulation of genes involved in iron metabolism (e.g., ferroportin, lactotransferrin, ferroxidase ceruloplasmin, lipocalin 2) (Stuckey et al., 2006;Bajbouj et al., 2018;Hamad et al., 2020), likely by downregulating hepcidin activity. In ovarian cells (E2-S and SKOV-3), E2-induced upregulation of HIF-1α has also been shown to downregulate hepcidin gene (HAMP) expression, subsequently reducing hepcidin concentration (Hou et al., 2012). ...
... Estrogen is considered the primary female sex steroid, and elevated concentrations of E2 have been positively correlated with iron demand and the release of iron into the systemic circulation, and negatively correlated with hepcidin concentration (Hamad et al., 2020), however, the exact mechanism by which E2 influences iron regulation is still to be determined. In vitro studies in breast (Bajbouj et al., 2018), ovarian SKOV3 (Yang et al., 2012), liver cells (HUH7 and Hep-G2) (Hou et al., 2012) and rodent models suggest E2 may support the upregulation of genes involved in iron metabolism (e.g., ferroportin, lactotransferrin, ferroxidase ceruloplasmin, lipocalin 2) (Stuckey et al., 2006;Bajbouj et al., 2018;Hamad et al., 2020), likely by downregulating hepcidin activity. In ovarian cells (E2-S and SKOV-3), E2-induced upregulation of HIF-1α has also been shown to downregulate hepcidin gene (HAMP) expression, subsequently reducing hepcidin concentration (Hou et al., 2012). ...
Iron metabolism research in the past decade has identified menstrual blood loss as a key contributor to the prevalence of iron deficiency in premenopausal females. The reproductive hormones estrogen and progesterone influence iron regulation and contribute to variations in iron parameters throughout the menstrual cycle. Despite the high prevalence of iron deficiency in premenopausal females, scant research has investigated female-specific causes and treatments for iron deficiency. In this review, we provide a comprehensive discussion of factors that influence iron status in active premenopausal females, with a focus on the menstrual cycle. We also outline several practical guidelines for monitoring, diagnosing, and treating iron deficiency in premenopausal females. Finally, we highlight several areas for further research to enhance the understanding of iron metabolism in this at-risk population.
... Moreover, gender has also influence over iron losses and stores due to menstrual menses 1,12 . Along with this, several studies have already suggested changes over hepcidin production by high concentrations of sex hormones; specifically, high oestrogen concentrations are supposed to downregulate hepcidin synthesis [13][14][15][16] and reduce ferroportin expression in cells membrane 17 . Interestingly, progesterone seems to produce an opposite effect 18 . ...
... Only Peeling et al. 21 tested men and women´s response, but the results were presented as a whole, ignoring the possible influence of female sex hormones1 [3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18] . It is unknown the hepcidin response to endurance exercise of the active females in different menstrual cycle phases, depending on their pre-exercise ferritin concentrations. ...
Serum ferritin has been proposed as a predictor of hepcidin concentrations in response to exercise. However, this fact has not been studied in physically-active women. Therefore, the main objective of this study was to analyse the hepcidin response at different ferritin status before and after running exercise in physically active females. Fifteen eumenorrheic women performed a 40-min running protocol at 75% of VO2peak speed in different menstrual cycle phases (early-follicular phase, mid-follicular phase and luteal phase). Blood samples were collected pre-exercise, 0h post-exercise and 3h post-exercise. For statistics, participants were divided into two groups according to their pre-exercise ferritin levels (<20 and ≥20 μg/L). Through menstrual cycle, hepcidin was lower in both early follicular phase (p=0.024; 64.81±22.48 ng/ml) and mid-follicular phase (p=0.007; 64.68±23.91 ng/ml) for <20 μg/L ferritin group, in comparison with ≥20 μg/L group (81.17±27.89 and 79.54±22.72 ng/ml, respectively). Hepcidin showed no differences between both ferritin groups in either pre-exercise, 0h post-exercise and 3h post-exercise. Additionally, no association between pre-exercise ferritin and hepcidin levels 3h post-exercise (r=-0.091; p=0.554) was found. Menstrual cycle phase appears to influence hepcidin levels depending on ferritin reserves. In particular, physically-active females with depleted ferritin reserves seems to present lower hepcidin levels during the early-follicular phase and mid-follicular phase. However, no association between ferritin and hepcidin levels was found in this study. Hence, ferritin levels alone may not be a good predictor of hepcidin response to exercise in this population. Multiple factors such as sexual hormones, training loads and menstrual bleeding must be taken into account.
... 29 In this context, it has been reported that the human sex hormone estrogen (17β-estradiol; E 2 ) downregulates hepcidin synthesis and upregulates Fpn expression as a means of enhancing intracellular iron efflux. 31 E 2 has also been reported to induce oxidative stress, membrane damage, and cell-cycle arrest in MCF7 cells 32 in a manner related to disrupted intracellular iron status. 33 Given that disrupted iron metabolism is a common finding in cancer and that E 2 and Dox disrupt intracellular iron metabolism and cause DNA damage, we hypothesized that treating cancer cells with E 2 plus Dox could limit the ability of cancer cells to maintain high iron content, making them more susceptible to Dox-induced apoptosis. ...
... 47 The findings also demonstrate that E 2 +Dox treatment results in a significant disruption of intracellular iron metabolism. Previous work has shown that E 2 disrupts intracellular iron metabolism 31 and that this is associated with increased oxidative stress, DNA damage, and cell-cycle arrest in MCF7 and SKOV3. 32,33 The role of E 2 in iron metabolism stems mainly from its ability to reduce hepcidin synthesis through upregulated HIF1α expression 48,49 or direct interaction with E 2 -responsive elements in the hepcidin gene. ...
Introduction
Increased iron content in cancer cells is associated with resistance to chemotherapy. Recent studies have demonstrated that estrogen (E2) suppresses hepcidin synthesis and enhances intracellular iron efflux. Herein, we investigated whether E2-driven intracellular iron efflux renders cancer cells more susceptible to doxorubicin (Dox)-induced cytotoxicity.
Methods
Breast, ovarian, and liver cancer cell lines treated with E2, Dox, or a combination of both were assessed for intracellular iron status, mitochondrial function, cell cycle, and apoptosis.
Results
E2+Dox treatment in MCF7, SKOV3 and MDA-MB231 cells resulted in enhanced apoptosis compared with Dox-treated cells. Expression of γH2AX was significantly higher and that of survivin significantly lower in E2+Dox-treated cells than Dox-treated cells. At 48 hours, E2+Dox had induced a significant increase in the percentage of sub-G1 apoptotic cells, increased CHK1 expression, and decreased cyclin D1, CDK4, and CDK6 expression. Ferroportin and ferritin expression was significantly higher and that of TfR1 significantly lower in E2+Dox-treated cells than Dox-treated cells. Intracellular iron content was significantly reduced in E2+Dox-treated cells at 48 hours posttreatment. Lastly, E2+Dox-treated cells showed higher levels of mitochondrial membrane hyperpolarization than Dox-treated cells.
Conclusion
These findings suggest that E2 disrupts intracellular iron metabolism in such a way that increases cell susceptibility to Dox-induced cytotoxicity.
... 29 In this context, it has been reported that the human sex hormone estrogen (17β-estradiol; E 2 ) downregulates hepcidin synthesis and upregulates Fpn expression as a means of enhancing intracellular iron efflux. 31 E 2 has also been reported to induce oxidative stress, membrane damage, and cell-cycle arrest in MCF7 cells 32 in a manner related to disrupted intracellular iron status. 33 Given that disrupted iron metabolism is a common finding in cancer and that E 2 and Dox disrupt intracellular iron metabolism and cause DNA damage, we hypothesized that treating cancer cells with E 2 plus Dox could limit the ability of cancer cells to maintain high iron content, making them more susceptible to Dox-induced apoptosis. ...
... 47 The findings also demonstrate that E 2 +Dox treatment results in a significant disruption of intracellular iron metabolism. Previous work has shown that E 2 disrupts intracellular iron metabolism 31 and that this is associated with increased oxidative stress, DNA damage, and cell-cycle arrest in MCF7 and SKOV3. 32,33 The role of E 2 in iron metabolism stems mainly from its ability to reduce hepcidin synthesis through upregulated HIF1α expression 48,49 or direct interaction with E 2 -responsive elements in the hepcidin gene. ...
... This disparity may be attributed to hormonal differences, with testosterone promoting erythropoiesis by stimulating erythropoietin production and suppressing hepcidin, thus enhancing iron utilization [82]. Conversely, elevated estrogen levels in females can increase hepcidin expression, leading to reduced iron absorption and utilization [83,84]. Trace metals such as Ni, Co, and Mn are involved in hormone regulation and have been linked to variations in serum testosterone and estrogen levels [85][86][87]. ...
Background: Anemia significantly contributes to the global disease burden, with its incidence potentially influenced by the trace metal content within the body. Objective: This study aims to examine the associations between trace metals and anemia risk, with a particular focus on investigating the potential mediating roles of iron status and inflammation in these associations. Methods: Five trace metals (Ni, Co, Mn, Se, and Mo) were examined in 1274 US adults, utilizing data from the National Health and Nutrition Examination Survey (NHANES) 2017–2020. The individual and combined effects of these metals on anemia were assessed using logistic regression, quantile g-computation (QGC), and Bayesian kernel machine regression (BKMR). A sex-stratified analysis was conducted to discern any gender-specific susceptibilities. Additionally, mediation analysis was employed to explore the potential mediating roles of iron status and inflammation in the associations between these metals and anemia. Results: Increased risks of anemia were positively associated with Co and Ni levels but negatively correlated with Se and Mn levels (all with p < 0.05). The trace metal mixture was negatively associated with anemia, with the highest weights of Co and Se in different directions in both the QGC and BKMR models. In the sex-specific analysis, we observed less pronounced protective effects from trace metals in females. Moreover, the mediating proportion of the iron status and inflammation in these relationships ranged from 10.29% to 58.18%. Conclusion: Our findings suggest that the trace element mixture was associated with decreased anemia risk, among which Se was a protective factor while Co was a risk factor, and females were more susceptible. The effects of these trace metals on anemia may be mediated by the iron status and inflammation.
... Glucocorticoid receptor activation stimulates erythroid progenitor self-renewal but inhibits terminal differentiation [40][41][42], which would be compatible with the shift towards immature cells that we have described. Furthermore, oestrogens, which are derived in obesity via peripheral aromatisation in subcutaneous adipose tissue [43], increase iron availability and facilitate erythropoiesis by suppressing hepcidin production [44,45], and increase haematopoietic stem-cell renewal [46]. Nevertheless, oestrogens inhibit hypoxia induced EPO production in animal models [47] and circulating oestradiol was Table 1 Anthropometric characteristics and erythrocyte and reticulocyte parameters of obesity and body shape phenotypes ABSI a body shape index (cut-offs ≥ 73 for women, ≥ 80 for men); Apple large-ABSI-small-HI, BMI body mass index, HI hip index (cut-offs ≥ 64 for women, ≥ 49 for men), Imm. ...
Abstract Background Obesity is associated with type 2 diabetes mellitus and chronic low-grade inflammation. Although chronic inflammatory conditions and diabetes are associated with anaemia, less is known about associations of obesity and body shape, independent of each other, with erythrocyte and reticulocyte parameters. Methods We investigated the associations of body mass index (BMI) and the allometric body shape index (ABSI) and hip index (HI), which are uncorrelated with BMI, with erythrocyte and reticulocyte parameters (all continuous, on a standard deviation (SD) scale) in UK Biobank participants without known metabolic, endocrine, or major inflammatory conditions (glycated haemoglobin HbA1c
... The increasing in hematocrit and hemoglobin due to testosterone is associated with erythropoietin stimulation and with decreasing in hepcidin and ferritin levels, it was suggested that testosterone induces erythropoiesis by stimulating erythropoietin and also by consumption of iron in the erythropoiesis process (17). High estrogen is associated with high iron, as estrogen lowers the level of hepcidin to improve the level of iron in the blood of menstruating women (18). Hepcidin decreases iron absorption by binding to ferroportin causing iron breakdown and administrate the patient a dose of growth hormone or testosterone and reduces the level of hepcidin, also it was recorded gonadotropin-stimulated estrogen decreases the concentration of hepcidin-25 (19). ...
Polycystic ovary syndrome (PCOS) is one of the most common gynecological diseases that affect the fertility in women in Basra governorate. The current study was designed in order to assess iron aberrations in PCOS patients by measuring the related parameters and their relationship with sex hormones in patients with PCOS. Serum samples were collected from 45 PCOS patients and 45 controls from a private women's clinic and were measured by ELISA in a private medical laboratory. The results showed a significant decrease in the level of hepcidin, transferrin and estradiol versus a significant increase in iron, ferritin, progesterone and testosterone. The current study showed a clear imbalance in the level of iron and its serum regulating parameters in in PCOS women, and there is an effective correlation between iron status and sex hormones.
... The increasing in hematocrit and hemoglobin due to testosterone is associated with erythropoietin stimulation and with decreasing in hepcidin and ferritin levels, it was suggested that testosterone induces erythropoiesis by stimulating erythropoietin and also by consumption of iron in the erythropoiesis process (17). High estrogen is associated with high iron, as estrogen lowers the level of hepcidin to improve the level of iron in the blood of menstruating women (18). Hepcidin decreases iron absorption by binding to ferroportin causing iron breakdown and administrate the patient a dose of growth hormone or testosterone and reduces the level of hepcidin, also it was recorded gonadotropin-stimulated estrogen decreases the concentration of hepcidin-25 (19). ...
Polycystic ovary syndrome (PCOS) is one of the most common gynecological diseases that affect the fertility in women in Basra governorate. The current study was designed in order to assess iron aberrations in PCOS patients by measuring the related parameters and their relationship with sex hormones in patients with PCOS. Serum samples were collected from 45 PCOS patients and 45 controls from a private women’s clinic and were measured by ELISA in a private medical laboratory. The results showed a significant decrease in the level of hepcidin, transferrin and estradiol versus a significant increase in iron, ferritin, progesterone and testosterone. The current study showed a clear imbalance in the level of iron and its serum regulating parameters in in PCOS women, and there is an effective correlation between iron status and sex hormones.
... Subsequently, Yang et al. observed an inhibition of hepcidin synthesis induced by 17b-estradiol, and an increased Fe uptake, a mechanism to compensate Fe loss during menstruation (50). Recent studies have confirmed that estrogen activity contributes to the changes in serum Fe status; a negative correlation between hepcidin and estrogen or Fe was demonstrated in menstruating women which is a mechanism increasing serum Fe content (51). The latest reports indicated that testosterone suppressed hepcidin (44), either through direct or indirect mechanisms, resulting in the increased Fe turnover and maintaining erythropoiesis during severe energy deficit (52). ...
Hepcidin is a protein responsible for maintaining iron (Fe) homeostasis. Data regarding the role of hepcidin in the pathomechanism of Fe balance disturbances associated with acromegaly (AG) are scarce. The aim of the study was to assess the impact of alterations in complete blood count parameters, Fe homeostasis, gonadal status and GH/IGF-1 on the level of hepcidin in AG patients. The study evaluated the differences in hepcidin concentration and iron homeostasis between patients newly diagnosed with AG in comparison to healthy control subjects (CS). We prospectively enrolled 25 adult patients newly diagnosed with AG and 25 healthy volunteers who served as CS. The level of hepcidin was measured using the Hepcidin 25 (bioactive) hs ELISA, which is a highly sensitive enzyme immunoassay for the quantitative in vitro diagnostic measurement (DRG Instruments GmbH, Germany). The median of hepcidin concentration in the serum of patients with AG was significantly lower 9.8 (6.2–18.2) ng/ml as compared to CS 21.3 (14.3–34.0) ng/ml (p = 0.003). In the AG group, a statistically significant negative correlation between hepcidin and IGF-1 (rho = −0.441) was observed. Our study demonstrated a decreased hepcidin level in AG patients in comparison to CS what may have a potentially protective effect against anemia through an increased bioavailability of Fe. Additionally, GH may have a positive direct or indirect effect on erythropoiesis. Further studies on larger patient groups are necessary in order to clarify the exact role of hepcidin in the regulation of erythropoiesis in the excess of GH/IGF-1.
... Furthermore, hepcidin may enhance hepatic lipid accumulation, as seen in male and female haloperidol and male clozapine treated rats. It seems to play a role in the regulation of hepatic metabolic pathways involved in the pathogenesis of NAFLD [50], but the underlying mechanism is unclear as well as the effect of estrogen, which is known to modulate hepcidin synthesis and serum iron availability [51]. HO-1 and ALAS1 are the rate limiting enzymes of heme degradation and synthesis, respectively. ...
Background
Patients with liver diseases often have some form of anemia. Hematological dyscrasias are known side effects of antipsychotic drug medication and the occurrence of agranulocytosis under clozapine is well described. However, the sex-dependent impact of clozapine and haloperidol on erythrocytes and symptoms like anemia, and its association with hepatic iron metabolism has not yet been completely clarified. Therefore, in the present study, we investigated the effect of both antipsychotic drugs on blood parameters and iron metabolism in the liver of male and female Sprague Dawley rats.
Methods
After puberty, rats were treated orally with haloperidol or clozapine for 12 weeks. Blood count parameters, serum ferritin, and liver transferrin bound iron were determined by automated counter. Hemosiderin (Fe³⁺) was detected in liver sections by Perl’s Prussian blue staining. Liver hemoxygenase (HO-1), 5’aminolevulinate synthase (ALAS1), hepcidin, heme-regulated inhibitor (HRI), cytochrome P4501A1 (CYP1A1) and 1A2 (CYP1A2) were determined by Western blotting.
Results
We found anemia with decreased erythrocyte counts, associated with lower hemoglobin and hematocrit, in females with haloperidol treatment. Males with clozapine medication showed reduced hemoglobin and increased red cell distribution width (RDW) without changes in erythrocyte numbers. High levels of hepatic hemosiderin were found in the female clozapine and haloperidol medicated groups. Liver HRI was significantly elevated in male clozapine medicated rats. CYP1A2 was significantly reduced in clozapine medicated females.
Conclusions
The characteristics of anemia under haloperidol and clozapine medication depend on the administered antipsychotic drug and on sex. We suggest that anemia in rats under antipsychotic drug medication is a sign of an underlying liver injury induced by the drugs. Changing hepatic iron metabolism under clozapine and haloperidol may help to reduce these effects of liver diseases.
... Subsequently, Yang et al. observed an inhibition of hepcidin synthesis induced by 17b-estradiol, and an increased Fe uptake, a mechanism to compensate Fe loss during menstruation (50). Recent studies have confirmed that estrogen activity contributes to the changes in serum Fe status; a negative correlation between hepcidin and estrogen or Fe was demonstrated in menstruating women which is a mechanism increasing serum Fe content (51). The latest reports indicated that testosterone suppressed hepcidin (44), either through direct or indirect mechanisms, resulting in the increased Fe turnover and maintaining erythropoiesis during severe energy deficit (52). ...
... During the sugar or placebo pill week, a rebound in endogenous hormones occurs, raising luteinizing hormone, follicle-stimulating hormone and particularly oestradiol levels to those that are equivalent to the early follicular phase in naturally menstruating women (Reape et al., 2008;Rechichi et al., 2009;Schlaff et al., 2004;Spona et al., 1996;Sullivan et al., 1999). With previous research demonstrating that oestrogen is positively correlated with serum iron levels in females (Bajbouj et al., 2018), the elevations of serum iron with contraceptive use during the withdrawal bleed period is likely to be associated with the increase in endogenous oestrogen levels. Angeli et al. (2016) noted that female participants taking the contraceptive pill maintained higher serum iron concentrations throughout the cycle (or during the active pill phase), a result that may be due to several undocumented factors including; dose of ethinylestradiol based on the brand of pill ingested and hepatic metabolism of ethinylestradiol by the individual. ...
Iron deficiency is a common nutrient deficiency within athletes, with sport scientists and medical professionals recognizing that athletes require regular monitoring of their iron status during intense training periods. Revised considerations for athlete iron screening and monitoring have suggested that males get screened biannually during heavy training periods and females require screening biannually or quarterly, depending on their previous history of iron deficiency. The prevalence of iron deficiency in female athletes is higher than their male counterparts and is often cited as being a result of the presence of a menstrual cycle in the premenopausal years. This review has sought to revise our current understanding of female physiology and the interaction between primary reproductive hormones (oestrogen and progesterone) and iron homoeostasis in females. The review highlights an apparent symbiotic relationship between iron metabolism and the menstrual cycle that requires additional research as well as identifying areas of the menstrual cycle that may be primed for nutritional iron supplementation. ARTICLE HISTORY
... During low levels of iron in females, potentially induced by regular menstruation, DMT1 expression increases to promote ferrous iron and other divalent iron absorption [102]. Clinical studies reported elevated levels of estrogen are associated with enhanced serum iron due to reduction in hepcidin synthesis, which improves serum iron level in menstruating females [103]. In addition, there is a positive association between iron and zinc status in premenopausal women [104], which supports the notion that iron and zinc absorption kinetics may be enhanced in young women. ...
Customization of mental health therapies needs to consider the differences in degree of brain maturity between young (18–29 years) and mature (30 years or older) adults as well as brain morphology among men and women. The aim of this study was to identify the significant dietary and lifestyle contributors to mental distress in these sub-populations. Independent repeated cross-sectional sampling was performed for over a 5-year period (2014–2019) to collect data from different populations at different time-points and seasons. A backward stepwise regression analysis was used on 2628 records. Mental distress in young women was associated with high consumption of caffeine and fast-food, and it was negatively correlated with moderate-high levels of exercise as well as frequent breakfast consumption. Mature women shared several common factors with young women; however, high fruit consumption was negatively associated with mental distress. For young men, high exercise, moderate consumption of dairy, and moderate-high intake of meat were negatively associated with mental distress. In addition, high fast-food and caffeine consumption were positively associated with mental distress in young men. For mature men, strong negative associations between higher education, moderate intake of nuts and mental distress surfaced. Our results support the need to customize dietary and lifestyle recommendations to improve mental wellbeing.
... CMS patients 24,[27][28][29]47 , but the effect of estrogen on RBC production is still unclear in CMS patients at high altitude. It is possible that all sex hormones have a direct effect on EPO and hepcidin levels and iron metabolism, which directly contribute to the increased hematocrit under hypoxia [48][49][50][51][52][53][54][55][56][57] . Physiological factors, such as ventilation, cardiopulmonary responses, and oxygenation, also depend on sex hormones and can also produce differences in hematocrit levels between the two sexes 25,[58][59][60][61][62][63][64][65] . ...
Monge’s disease (Chronic mountain sickness), is a maladaptive condition caused by
chronic exposure to high altitude hypoxia. One defining feature of chronic
mountain sickness (CMS) is excessive erythrocytosis with extremely high hematocrit
levels. In the Andean population, CMS prevalence is vastly different between men and
women, being rare in the latter. Furthermore, there is a sharp increase in CMS incidence
in women after menopause. In this study, we assess the role of sex hormones
(testosterone, progesterone and estrogen) in CMS and non-CMS cells using well
characterized in-vitro erythroid platform. While we found that there was a mild (and
insignificant) increase in RBC production with testosterone; estrogen, at physiological
concentrations, reduced sharply CD235a-positive cells (Glycophorin A-marker of RBC),
from 56% in the untreated CMS sample to 10% in the treated CMS cells in a stage-specific
and dose responsiveness manner. At the molecular level, we determined that estrogen has direct effect on GATA1, remarkably decreasing mRNA and protein levels (p<0.01) and its target genes (Alas2, BclxL and EPOR, p<0.001). These changes result in
significant increase in apoptosis of erythroid cells. We also demonstrate that estrogen
regulates erythropoiesis in CMS patients through estrogen beta signaling and its
inhibition can diminish the effects of estrogen by significantly increasing HIF1, VEGF
and GATA1 mRNA levels. Taken altogether, our results indicate that estrogen has a
major impact on the regulation of erythropoiesis, particularly under chronic hypoxic
conditions, and has the potential of treating blood diseases, such as high altitude severe
erythrocytosis.
... Finally, it has been suggested that estrogen decreases hepcidin synthesis, 34,35 and hereby possibly causing a gender influence on hepcidin concentrations. We found a HbA1c, glycated hemoglobin A1c; hsCRP, high-sensitivity C-reactive protein. ...
Chronic low-grade inflammation in type 1 diabetes (T1D) might increase hepcidin synthesis, possibly resulting in functional iron deficiency (FID). We hypothesized that in T1D children with FID, hepcidin concentrations are increased compared to those with normal iron status and those with absolute iron deficiency (AID). We evaluated hepcidin concentrations in T1D children in relation to iron status, and investigated whether hepcidin is useful in assessing FID. A cross-sectional study was conducted. FID was defined as elevated zinc protoporphyrin/heme ratio and/or red blood cell distribution width, and AID as low serum ferritin concentration. Post-hoc analyses with different definitions of FID were performed, using transferrin saturation and reticulocyte hemoglobin content. Serum hepcidin concentrations were measured using mass-spectrometry. The IRODIAB-study is registered at www.trialregister.nl (NTR4642). This study included 215 T1D children with a median age of 13.7 years (Q1-Q3: 10.1-16.3). The median (Q1-Q3) hepcidin concentration in patients with normal iron status was 1.8 nmol/l (0.9-3.3), in AID-patients, 0.4 nmol/l (0.4-0.4) and in FID-patients, 1.6 nmol/l (0.7-3.5). Hepcidin concentrations in FID-patients were significantly higher than in AID-patients (p < 0.001). Irrespective of FID-definition used, hepcidin concentrations did not differ between FID-patients and patients with normal iron status. This might be explained by the influence of various factors on hepcidin concentrations, and/or by differences in response of iron parameters over time. Single hepcidin measurements do not seem useful in assessing FID in T1D children. Multiple hepcidin measurements over time in future studies, however, might prove to be more useful in assessing FID in children with T1D.
... Before the onset of menopause hepcidin levels in women are nearly 50% lower than in males of corresponding ages. After the menopause, hepcidin levels tend to be similar in both sexes [212,213] or slightly increased in men [214]. These results demonstrate a negative correlation between ferritin and estrogen levels during the menopausal transition period [215]. ...
Iron is an essential element that participates in numerous cellular processes. Any disruption of iron homeostasis leads to either iron deficiency or iron overload, which can be detrimental for humans' health, especially in elderly. Each of these changes contributes to the faster development of many neurological disorders or stimulates progression of already present diseases. Age-related cellular and molecular alterations in iron metabolism can also lead to iron dyshomeostasis and deposition. Iron deposits can contribute to the development of inflammation, abnormal protein aggregation, and degeneration in the central nervous system (CNS), leading to the progressive decline in cognitive processes, contributing to pathophysiology of stroke and dysfunctions of body metabolism. Besides, since iron plays an important role in both neuroprotection and neurodegeneration, dietary iron homeostasis should be considered with caution. Recently, there has been increased interest in sex-related differences in iron metabolism and iron homeostasis. These differences have not yet been fully elucidated. In this review we will discuss the latest discoveries in iron metabolism, age-related changes, along with the sex differences in iron content in serum and brain, within the healthy aging population and in neurological disorders such as multiple sclerosis, Parkinson's disease, Alzheimer's disease, and stroke.
... Nausea, a common side effect of excessive iron consumption, has also been postulated as an evolved mechanism to keep individuals from consuming too much iron, particularly during pregnancy [37]. Finally, there is a growing but inconclusive body of literature that demonstrates that estrogen is implicated in the regulation of iron homeostasis, and could be another factor that regulates reproductive-aged women's iron levels to function within a certain range [38][39][40]. Assessing the relative contribution of these proximate mechanisms of iron control in humans-menstrual blood loss, iron-related nausea, and estrogen-is not an easy task. Overt menstruation is a derived characteristic within the Primate order [41], while the role of estrogen in internal gestation is a primitive characteristic among placental mammals [42]. ...
Background and objectives:
Iron is critical for women's reproduction, and iron-deficiency anemia is a global health problem for mothers. While public health programs have aimed to correct iron deficiency in reproductive-aged women with supplementation, a small group of studies have shown that too much iron also has negative effects on birth outcomes. We hypothesize that women's iron levels evolved within a narrow optimum, and predict that hemoglobin (Hb) levels would be associated with women's fecundity.
Methodology:
We used the publicly available, longitudinal Tsimane' Amazonian Panel Study to test the association between -Hb levels and hazard of having a next birth (a measure of fecundity) among 116 parous, reproductive-aged Tsimane' women of Bolivia. A Cox proportional hazards model was used to model Hb level and other predictors against the event of next birth across the observation period, which began at each woman's previous birth.
Results:
The higher the Hb level, the lower the hazard of a woman giving birth within the study observation period (hazard ratio=0.82, P = 0.03). However, there was no evidence that low Hb reduced women's fecundity.
Conclusions and implications:
These results demonstrate that high Hb influences women's fecundity. These results supports the growing body of literature showing that iron metabolism is critical for understanding the evolution of women's reproduction. More work is needed to determine the evolved optimal range of iron levels for reproductive-aged women.
Lay summary:
Lower chance of pregnancy among Tsimane' women with high Hb levels, suggesting evolved optimal Hb levels in women.
... Lastly, the organic pollutant polychlorinated biphenyls (PCBs) inhibits HAMP expression through an E2-like effect that associates with disrupted iron homeostasis [22]. In humans, several reports have suggested that elevated levels of serum E2 correlate positively with serum iron status [18][19][20][21][22] and negatively with serum HAMP concertation [23,24]. Mean serum iron levels significantly increase in women on contraceptives [25][26][27][28]. ...
Clinical and experimental evidence suggest that estrogen manipulates intracellular iron metabolism and that elevated levels of estrogen associate with increased systemic iron availability. This has been attributed to the ability of estrogen to suppress hepcidin synthesis, maintain ferroportin integrity and enhance iron release from iron-absorbing duodenal enterocytes and iron-storing macrophages and hepatocytes. These observations speak of a potential “estrogen-iron” axis that manipulates iron metabolism in response to hematologic (erythropoiesis) and non-hematologic (uterine growth, pregnancy, lactation) needs for iron. Such an axis could contribute to minimizing iron deficiency in premenopausal women and iron overload in postmenopausal women. It could also exacerbate iron overload and related clinical consequences including cancer, osteoporosis, cardiovascular complications and neurodegenerative symptoms, especially in postmenopausal women on hormonal replacement therapy. Understanding the role of estrogen in iron metabolism may shed some light on the pleotropic, but often paradoxical, roles of estrogen in human health and disease.
... These findings are in line with a study demonstrating a significant reduction in serum hepcidin without correlation with serum iron in female patients with elevated estrogens, due to fertility treatments [157]. On the other hand, in a study with premenopausal women during various phases of their monthly cycle, serum 17β-estradiol was negatively correlated with hepcidin and positively correlated with iron [158]. Definitely, more work is required to clarify the mechanism and physiological relevance of hepcidin regulation by estrogens before considering any therapeutic applications. ...
Hepcidin is a key hormonal regulator of systemic iron homeostasis and its expression is induced by iron or inflammatory stimuli. Genetic defects in iron signaling to hepcidin lead to “hepcidinopathies” ranging from hereditary hemochromatosis to iron-refractory iron deficiency anemia, which are disorders caused by hepcidin deficiency or excess, respectively. Moreover, dysregulation of hepcidin is a pathogenic cofactor in iron-loading anemias with ineffective erythropoiesis and in anemia of inflammation. Experiments with preclinical animal models provided evidence that restoration of appropriate hepcidin levels can be used for the treatment of these conditions. This fueled the rapidly growing field of hepcidin therapeutics. Several hepcidin agonists and antagonists, as well as inducers and inhibitors of hepcidin expression have been identified to date. Some of them were further developed and are currently being evaluated in clinical trials. This review summarizes the state of the art.
The mammalian dive reflex, characterized by bradycardia and peripheral vasoconstriction, occurs in all mammals, including humans, in response to apnea. However, the dive reflex to a single, maximal, dry, dynamic apnea (DYN), and how it compares to a time-matched exercise control trial (EX) or dry static apnea (SA), has not been studied. We examined the hypotheses that, compared to EX and SA, the magnitude of the (a) cardiovascular response and (b) hematological response to DYN would be greater. Cardiovascular parameters (heart rate [HR], systolic [SBP], diastolic [DBP], and mean arterial [MAP] blood pressure) were continuously collected in twenty-three (F=6) moderate and elite freedivers, first during a maximal DYN, then during a time-matched SA and EX on a swimming ergometer in randomized order. Venous blood draws were made prior to and following each trial. The change in calculated oxygen saturation (DYN:-17±13%, EX:-2±1%, ΔSA:-2±1%;P<0.05, all comparisons) was greater during DYN compared to EX and SA. During DYN, ΔSBP (DYN:104±31mmHg, EX:38±23mmHg, SA:20±11mmHg), ΔDBP (DYN:45±12mmHg, EX:14±10mmHg, SA:15±8mmHg) and ΔMAP (DYN:65±17mmHg EX:22±13mmHg, SA:16±9mmHg) were increased compared to EX and SA, while ΔHR was greater during EX (DYN:-24±23bpm, EX:33±13bpm, SA:-1±10bpm) than either DYN or SA (P<0.0001, all comparisons). Females had greater pressor response to EX (ΔSBP:59±30mmHg, ΔDBP:24±14mmHg, ΔMAP:35±8mmHg) than males (ΔSBP:31±15mmHg, ΔDBP:11±6mmHg, ΔMAP:18±8mmHg; P<0.01, all comparisons). Together, these data indicate that DYN elicits a distinct, exaggerated cardiovascular response compared to EX or SA alone.
The maintenance of iron (Fe) homeostasis is vital for the physiological function along life. In sexually mature humans and experimental animals, estrogens downregulate hepcidin (Hpc) expression, in order to improve the intestinal absorption and to mobilize Fe stores for maternal erythropoietic expansion and placental development. However, changes of these mechanisms related to regulation of Hpc on the availability of Fe during gestation with advancing age in mares, remain unknown. The objective of this study was to evaluate the interrelationships between serum Fe, Ferritin (Ferr) and Hpc with estrone (E1) and estradiol-17β (E2) concentrations in pregnant mares of different ages. Blood samples were taken from 40 pregnant Spanish Purebred mares belonging to 4 different age groups, 10 subjects for each group: 4-6 years, 7-9 years, 10-12 years, and > 12 years were used in this study. Fe concentrations of 4-6 and 7-9 years groups were higher (P < .01) than 10-12 and >12 years groups. Ferr concentrations of 4-6 years group were higher (P < .01) than other groups. Hpc concentrations increased and E1 decreased (P < .01) in > 12 years group compared to other age groups. E2 concentrations of 7-9, 10-12 and >12 years groups were higher (P < .01) than those of 4-6 years group; 7-9 years group had higher E2 concentrations (P < .01) than > 12 years group. Fe and Ferr were negatively correlated with Hpc (r = -0.81 and r = -0.67, respectively). E1 and E2 were negatively correlated with Fe (r = -0.23 and r = -0.11, respectively). E2 was positively correlated with Hpc (r = 0.78). In pregnant Spanish Purebred mare, the increase of estrogens, according to the more efficient iron status in response to Hpc inhibition and consequent mobilization of circulating and iron reserve, shows the existence of "estrogen-iron axis" in young mares. Nevertheless, these mechanisms are reversed in old mares, suggesting a less efficient iron metabolism with advancing age. It is hoped that new investigations are needed to understand in depth and clarify further the complex metabolic and hormonal mechanisms involved also in equine species.
Several lines of evidence suggest that estrogen (17-β estradiol; E2) protects against diabetes mellitus and plays important roles in pancreatic β-cell survival and function. Mounting clinical and experimental evidence also suggest that E2 modulates cellular iron metabolism by regulating the expression of several iron regulatory genes, including hepcidin (HAMP), hypoxia-inducible factor 1-α, ferroportin (SLC40A1), and lipocalin (LCN2). However, whether E2 regulates cellular iron metabolism in pancreatic β cells and whether the antidiabetic effects of E2 can be, at least partially, attributed to its role in iron metabolism is not known. In this context, pancreatic β cells express considerable levels of conventional E2 receptors (ERs; mainly ER-α) and nonconventional G protein-coupled estrogen receptors and hence responsive to E2 signals. Moreover, pancreatic islet cells require significant amounts of iron for proper functioning, replication and survival and, hence, well equipped to manage cellular iron metabolism (acquisition, utilization, storage, and release). In this review, we examine the link between E2 and cellular iron metabolism in pancreatic β cells and discuss the bearing of such a link on β-cell survival and function.
There are many biochemical tools available to assess the state of iron metabolism, but the diagnosis of iron pathology remains problematic. As molecular mechanisms responsible for iron uptake, transport, storage, and utilization become more precisely defined, efforts to exploit this new knowledge for diagnosis and therapy have begun to enter pediatric practice environments. This chapter will review the current understanding of iron (patho)physiology, current tools for biochemical assessment of iron status, and future prospects for the molecular manipulation of pathologic iron metabolism and storage.
Increased expression of Yes-associated protein-1 (YAP1) was shown to correlate with reduced survival in breast cancer (BC) patients. However, the exact mechanism of YAP1 regulation in BC cells remains ambiguous. Genomic sequence search showed that the promoter region of the YAP1 gene contains CpG Islands, hence the likelihood of epigenetic regulation by DNA methylation. To address this possibility, the effect of estrogen (17β estradiol; E2) on YAP1 gene expression and YAP1 promoter methylation status was evaluated in BC cells. The functional consequences of E2 treatment in control and YAP1-silenced BC cells were also investigated. Our data showed that E2 modulates YAP1 expression by hypomethylation of its promoter region via downregulation of DNA methyltransferase 3B (DNMT3B); an effect that seems to facilitate tumor progression in BC cells. Although the effect of E2 on YAP1 expression was estrogen receptor (ER) dependent, E2 treatment also upregulated YAP1 expression in MDA-MB231 and SKBR3 cells, which are known ER-negative BC cell lines but expresses ERα. Functionally, E2 treatment resulted in increased cell proliferation, decreased apoptosis, cell cycle arrest, and autophagic flux in MCF7 cells. The knockdown of the YAP1 gene reversed these carcinogenic effects of E2 and inhibited E2-induced autophagy. Lastly, we showed that YAP1 is highly expressed and hypomethylated in human BC tissues and that increased YAP1 expression correlates negatively with DNMT3B expression but strongly associated with ER expression. Our data provide the basis for considering screening of YAP1 expression and its promoter methylation status in the diagnosis and prognosis of BC.
The total iron level in the brain increases with age, and excess iron is associated with neurodegenerative diseases; however, the mechanism of brain iron deposition is unknown. In peripheral cells, the expression of hepcidin, a master regulator of iron homeostasis, is regulated by estrogen. This study aimed to determine whether hepcidin was involved in iron deposition in the brain and brain endothelial cells of estrogen-deficient aged female mice. Aged mice showed increased levels of hepcidin and ferritin in the brain and brain microvessels compared to young mice, and these levels were reduced by estrogen replacement in ovariectomized aged mice. In the brain endothelial cell line bEnd.3, the lipopolysaccharide (10 ng/ml)-induced increases of hepcidin mRNA and protein levels, number of Prussian blue-positive cells, and free radicals were reduced after estrogen treatment. These results suggest that estrogen deficiency with an increase of hepcidin is partly responsible for iron deposition in the brain and brain endothelial cells, and that hepcidin can be a target to prevent brain aging and neurodegeneration in postmenopausal women.
The ability of monocytes to release or sequester iron affects their role in cancer and inflammation. Previous work has shown that while IL-6 upregulates hepcidin synthesis and enhances iron sequestration, E2 reduces hepcidin synthesis and increases iron release. Given that E2 upregulates IL-6 production in monocytes, it is likely that the exact effect of E2 on iron metabolism in monocytes is shaped by its effect on IL-6 expression. To address this issue, the expression of key iron regulatory proteins was assessed in E2-treated U937, HuT-78, THP-1 and Hep-G2 cells. Iron status was also evaluated in U937 cells treated with the ERα agonist PPT, the ER antagonist ICI-182780, dexamethasone + E2, IL-6 + E2 and in IL-6-silenced U937 cells. E2 treatment reduced hepcidin synthesis in HuT-78, HTP-1 and Hep-G2 cells but increased hepcidin synthesis and reduced FPN expression in U937 cells. E2-treated U937 cells also showed reduced HIF-1α and FTH expression and increased TFR1 expression, which associated with increased labile iron content as compared with similarly treated Hep-G2 cells. While treatment of U937 cells with interleukin 6 (IL-6) resulted in increased expression of hepcidin, dexamethasone treatment resulted in reduced hepcidin synthesis relative to E2- or dexamethasone + E2-treated cells; IL-6 silencing also resulted in reduced hepcidin synthesis in U937 cells. Lastly, while iron depletion resulted in increased cell death in U937 cells, E2 treatment resulted in enhanced cell survival and reduced apoptosis. These findings suggest that E2 differentially alters iron metabolism in monocytes in an IL-6 dependent manner.
It is well accepted that intracellular iron overload that associate with various forms of cancer fuels tumor mutagenesis and growth. Hence, iron chelation therapy is being increasingly used to minimize iron overload in cancer patients despite significant safety and efficacy concerns. Mounting evidence suggests that estrogen (E2) downregulates hepcidin synthesis and increases serum iron concentration. It is postulated therefore that, by downregulating hepcidin synthesis, E2 may maintain ferroportin integrity and enhance intracellular iron efflux. Here, MCF-7 and SKOV-3 cancer cells treated with increasing concentrations (5, 10 and 20 nM) of E2 were assessed for intracellular labile iron content, the expression of hepcidin, ferroportin, and transferrin receptors 1 and 2 along with cell viability at different time points post treatment. In MCF-7 cells, E2 treatment resulted in a significant reduction in hepcidin synthesis, most noticeably at the 20 nM/24 h dose, a significant increase in ferroportin expression and a marked decrease in transferrin receptors 1 and 2 expression. E2-treated cells also showed reduced intracellular labile iron content most evidently at 20 nM/48 h dose and reduced viability especially at 20 nM/72 h dose. E2-treated SKOV-3 showed slightly reduced intracellular labile iron content, reduced expression of hepcidin and significantly increased expression of TFR1 but not TFR2; FPN expression was overall similar to that of controls. The effects of E2 on intracellular iron metabolism in SKOV-3 were most evident at 5 nM/24 h dose. These findings suggest that E2 treatment induces intracellular iron efflux, which may minimize intracellular iron overload in cancer cells; disrupted expression of transferrin receptor 1 and/or 2 may help sustain a low intracellular iron environment.
Objective
Hepcidin reduces iron absorption by binding to the intestinal iron transporter ferroportin, thereby causing its degradation. Although short-term administration of testosterone or growth hormone (GH) has been reported to decrease circulating hepcidin levels, little is known about how hepcidin is influenced in human endocrine conditions associated with anemia.
Research design and methods
We used a sensitive and specific dual–monoclonal antibody sandwich immunoassay to measure hepcidin-25 in patients (a) during initiation of in vitro fertilization when endogenous estrogens were elevated vs. suppressed, (b) with GH deficiency before and after 12 months substitution treatment, (c) with hyperthyroidism before and after normalization, and (d) with hyperprolactinemia before and after six months of treatment with a dopamine agonist.
Results
In response to a marked stimulation of endogenous estrogen production, median hepcidin levels decreased from 4.85 to 1.43 ng/mL (p < 0.01). Hyperthyroidism, hyperprolactinemia, or GH substitution to GH-deficient patients did not influence serum hepcidin-25 levels.
Conclusions
In humans, gonadotropin-stimulated endogenous estrogen markedly decreases circulating hepcidin-25 levels. No clear and stable correlation between iron biomarkers and hepcidin-25 was seen before or after treatment of hyperthyroidism, hyperprolactinemia or growth hormone deficiency.
Naturally occurring CD4+CD25+ regulatory T cells (Treg) exert an important role in mediating maternal tolerance to the fetus during pregnancy, and this effect might be regulated via maternal estrogen secretion. Although estrogen concentration in the pharmaceutical range has been shown to drive expansion of CD4+CD25+ Treg cells, little is known about how and through what mechanisms E2 within the physiological concentration range of pregnancy affects this expansion. Using in vivo and in vitro mouse models in these experiments, we observed that E2 at physiological doses not only expanded Treg cell in different tissues but also increased expression of the Foxp3 gene, a hallmark for CD4+CD25+ Treg cell function, and the IL-10 gene as well. Importantly, our results demonstrate that E2, at physiological doses, stimulated the conversion of CD4+CD25− T cells into CD4+CD25+ T cells which exhibited enhanced Foxp3 and IL-10 expression in vitro. Such converted CD4+CD25+ T cells had similar regulatory function as naturally occurring Treg cells, as demonstrated by their ability to suppress naïve T cell proliferation in a mixed lymphocyte reaction. We also found that the estrogen receptor (ER) exist in the CD4+CD25− T cells and the conversion of CD4+CD25− T cells into CD4+CD25+ T cells stimulated by E2 could be inhibited by ICI182,780, a specific inhibitor of ER(s). This supports that E2 may directly act on CD4+CD25− T cells via ER(s). We conclude that E2 is a potential physiological regulatory factor for the peripheral development of CD4+CD25+ Treg cells during the implantation period in mice.
The emergence of antiestrogen resistance in breast cancer is an important clinical phenomenon affecting long-term survival in this disease. Identifying factors that convey cell survival in this setting may guide improvements in treatment. Estrogen (E2) can induce apoptosis in breast cancer cells that have been selected for survival after E2 deprivation for long periods (MCF-7:5C cells), but the mechanisms underlying E2-induced stress in this setting have not been elucidated. Here, we report that the c-Src kinase functions as a key adapter protein for the estrogen receptor (ER, ESR1) in its activation of stress responses induced by E2 in MCF-7:5C cells. E2 elevated phosphorylation of c-Src which was blocked by 4-hydroxytamoxifen (4-OHT), suggesting that E2 activated c-Src through the ER. We found that E2 activated the sensors of the unfolded protein response (UPR), IRE1α (ERN1) and PERK kinase (EIF2AK3), the latter of which phosphorylates eukaryotic translation initiation factor-2α (eIF2α). E2 also dramatically increased reactive oxygen species (ROS) production and up-regulated expression of heme oxygenase HO-1 (HMOX1), an indicator of oxidative stress, along with the central energy sensor kinase AMPK (PRKAA2). Pharmacological or RNAi-mediated inhibition of c-Src abolished the phosphorylation of eIF2α and AMPK, blocked E2-induced ROS production, and inhibited E2-induced apoptosis. Together, our results establish that c-Src kinase mediates stresses generated by E2 in long-term E2-deprived cells that trigger apoptosis. This work offers a mechanistic rationale for a new approach in the treatment of endocrine-resistant breast cancer.
Hepcidin, a liver-derived iron regulatory protein, plays a crucial role in iron metabolism. It is known that gender differences exist with respect to iron storage in the body; however, the effects of sex steroid hormones on iron metabolism are not completely understood. We focused on the effects of the female sex hormone estrogen on hepcidin expression. First, ovariectomized (OVX) and sham-operated mice were employed to investigate the effects of estrogen on hepcidin expression in an in vivo study. Hepcidin expression was decreased in the livers of OVX mice compared to the sham-operated mice. In OVX mice, bone morphologic protein-6 (BMP6), a regulator of hepcidin, was also found to be downregulated in the liver, whereas ferroportin (FPN), an iron export protein, was upregulated in the duodenum. Both serum and liver iron concentrations were elevated in OVX mice relative to their concentrations in sham-operated mice. In in vitro studies, 17β-estradiol (E(2)) increased the mRNA expression of hepcidin in HepG2 cells in a concentration-dependent manner. E(2)-induced hepatic hepcidin upregulation was not inhibited by ICI 182720, an inhibitor of the estrogen receptor; instead, hepcidin expression was increased by ICI 182720. E(2) and ICI 182720 exhibit agonist actions with G-protein coupled receptor 30 (GPR30), the 7-transmembrane estrogen receptor. G1, a GPR30 agonist, upregulated hepcidin expression, and GPR30 siRNA treatment abolished E(2)-induced hepcidin expression. BMP6 expression induced by E(2) was abolished by GPR30 silencing. Finally, both E(2) and G1 supplementation restored reduced hepatic hepcidin and BMP6 expression and reversed the augmentation of duodenal FPN expression in the OVX mice. In contrast, serum hepcidin was elevated in OVX mice, which was reversed in these mice with E(2) and G1. Thus, estrogen is involved in hepcidin expression via a GPR30-BMP6-dependent mechanism, providing new insight into the role of estrogen in iron metabolism.
Interaction of estrogen with iron at the systemic level is long suspected, but direct evidence linking the two is limited. In the present study, we examined the effects of 17β-estradiol (E2) on hepcidin, a key negative regulator of iron absorption from the liver. We found that transcription of hepcidin was suppressed by E2 treatment in human liver HuH7 and HepG2 cells, and this down-regulation was blocked by E2 antagonist ICI 182780. Chromatin immunoprecipitation, deletion, and EMSA detected a functional estrogen responsive element half-site that is located between -2474 and -2462 upstream from the start of transcription of the hepcidin gene. After cloning the human hepcidin promoter into the pGL3Luc-Reporter vector, luciferase activity was also down-regulated by E2 treatment in HepG2 cells. E2 reduced hepcidin mRNA in wild-type mice as well as in hemochromatosis Fe gene knockout mice. In summary, our data suggest that hepcidin inhibition by E2 is to increase iron uptake, a mechanism to compensate iron loss during menstruation. This mechanism may also contribute to increased iron stores in oral contraceptive users.
Fertile women may be encouraged to use contraception during clinical trials to avoid potential drug effects on fetuses. However, hormonal contraception interferes with pharmacokinetics and pharmacodynamics and modifies internal milieus. Macrophages depend on the milieu to which they are exposed. Therefore, we assessed whether macrophage function would be affected by the use of combined oral contraceptives (OCs) and if this influence depended on the androgenic or non-androgenic properties of progestin.
Healthy adult women were enrolled and stratified into two groups: women who did not use OCs (Fs) and women treated with OCs (FOCs). FOCs were further stratified as a function of androgenic (FOCA+) and non-androgenic (FOCA-) properties of progestins. Routine hematological, biochemical, inflammatory and endothelial dysfunction parameters were measured. Monocyte-derived macrophages (MDMs) were evaluated for the expression and activity of estrogen receptors and androgen receptors, and release of tumor necrosis factor α (TNFα) was measured from unstimulated and lipopolysaccharide-stimulated cells.
As is already known, the use of OCs changed numerous parameters: the number of lymphocytes, iron levels, total iron-binding capacity of transferrin, triglycerides, high-density lipoprotein, total cholesterol, and C-reactive protein increased, while prothrombin time and alkaline phosphatase decreased. Hormonal levels also varied: cortisol was higher in FOCs, while luteinizing hormone, follicle-stimulating hormone, and testosterone were lower in FOCs. Asymmetric dimethylarginine, an index of endothelial function, was lower in FOC than in Fs, as were cysteine and bilirubin. The androgenic properties of progestins affected the activity of OCs: in particular, white blood cell count, hemoglobin, high-density lipoprotein and calcium were higher in FOCA- than in FOCA+, whereas percentage oxygen saturation and γ-glutamyl transpeptidase were lower in FOCA- than in FOCA+. Importantly, FOCs had a lower global DNA methylation, indicating that OC may have epigenetic effects on gene expression. OC did not modify the expression of androgen receptor but increased estrogen receptor α expression, more considerably in FOCA+, and decreased estrogen receptor β, more considerably in FOCA-. Importantly, the activation state of estrogen receptor β in FOCs was decreased, while estrogen receptor α was not active in either Fs or FOCs. Unstimulated MDMs obtained from FOCs showed higher release of TNFα in comparison with Fs. After lipopolysaccharide stimulation, the release of TNFα was significantly higher in Fs than in FOCs.
OC use induced many changes in hematological and plasmatic markers, modifying hormonal levels, endothelial function, inflammation index and some redox state parameters, producing a perturbation of the internal milieu that impacted macrophagic function. In fact, different levels of estrogen receptor expression and release of TNFα were observed in macrophages derived from OC users. Some of the above activities were linked to the androgenic properties of progestin. Even though it is not known whether these effects are reversible, the results indicate that to avoid potential skewing of results only a single type of OC should be used during a single clinical trial.
The regulation of iron metabolism involves multiple organs including the duodenum, liver and bone marrow. The recent discoveries of novel iron-regulatory proteins have brought the liver to the forefront of iron homeostasis. The iron overload disorder, genetic hemochromatosis, is one of the most prevalent genetic diseases in individuals of Caucasian origin. Furthermore, patients with non-hemochromatotic liver diseases, such as alcoholic liver disease, chronic hepatitis C or nonalcoholic steatohepatitis, often exhibit elevated serum iron indices (ferritin, transferrin saturation) and mild to moderate hepatic iron overload. Clinical data indicate significant differences between men and women regarding liver injury in patients with alcoholic liver disease, chronic hepatitis C or nonalcoholic steatohepatitis. The penetrance of genetic hemochromatosis also varies between men and women. Hepcidin has been suggested to act as a modifier gene in genetic hemochromatosis. Hepcidin is a circulatory antimicrobial peptide synthesized by the liver. It plays a pivotal role in the regulation of iron homeostasis. Hepcidin has been shown to be regulated by iron, inflammation, oxidative stress, hypoxia, alcohol, hepatitis C and obesity. Sex and genetic background have also been shown to modulate hepcidin expression in mice. The role of gender in the regulation of human hepcidin gene expression in the liver is unknown. However, hepcidin may play a role in gender-based differences in iron metabolism and liver diseases. Better understanding of the mechanisms associated with gender-related differences in iron metabolism and chronic liver diseases may enable the development of new treatment strategies.
Ferroportin and hepcidin are critical proteins for the regulation of systemic iron homeostasis. Ferroportin is the only known mechanism for export of intracellular non-heme-associated iron; its stability is regulated by the hormone hepcidin. Although ferroportin profoundly affects concentrations of intracellular iron in tissues important for systemic iron absorption and trafficking, ferroportin concentrations in breast cancer and their influence on growth and prognosis have not been examined. We demonstrate here that both ferroportin and hepcidin are expressed in cultured human breast epithelial cells and that hepcidin regulates ferroportin in these cells. Further, ferroportin protein is substantially reduced in breast cancer cells compared to nonmalignant breast epithelial cells; ferroportin protein abundance correlates with metabolically available iron. Ferroportin protein is also present in normal human mammary tissue and markedly decreased in breast cancer tissue, with the highest degree of anaplasia associated with lowest ferroportin expression. Transfection of breast cancer cells with ferroportin significantly reduces their growth after orthotopic implantation in the mouse mammary fat pad. Gene expression profiles in breast cancers from >800 women reveal that decreased ferroportin gene expression is associated with a significant reduction in metastasis-free and disease-specific survival that is independent of other breast cancer risk factors. High ferroportin and low hepcidin gene expression identifies an extremely favorable cohort of breast cancer patients who have a 10-year survival of >90%. Ferroportin is a pivotal protein in breast biology and a strong and independent predictor of prognosis in breast cancer.
High dose oestrogen therapy was used as a treatment for postmenopausal patients with breast cancer from the 1950s until the introduction of the safer antioestrogen, tamoxifen in the 1970s. The anti-tumour mechanism of high dose oestrogen therapy remained unknown. There was no enthusiasm to study these signal transduction pathways as oestrogen therapy has almost completely been eliminated from the treatment paradigm. Current use of tamoxifen and the aromatase inhibitors seek to create oestrogen deprivation that prevents the growth of oestrogen stimulated oestrogen receptor (ER) positive breast cancer cells. However, acquired resistance to antihormonal therapy does occur, but it is through investigation of laboratory models that a vulnerability of the cancer cell has been discovered and is being investigated to provide new opportunities in therapy with the potential for discovering new cancer-specific apoptotic drugs. Laboratory models of resistance to raloxifene and tamoxifen, the selective oestrogen receptor modulators (SERMs) and aromatase inhibitors demonstrate an evolution of drug resistance so that after many years of oestrogen deprivation, the ER positive cancer cell reconfigures the survival signal transduction pathways so oestrogen now becomes an apoptotic trigger rather than a survival signal. Current efforts are evaluating the mechanisms of oestrogen-induced apoptosis and how this new biology of oestrogen action can be amplified and enhanced, thereby increasing the value of this therapeutic opportunity for the treatment of breast cancer. Several synergistic approaches to therapeutic enhancement are being advanced which involve drug combinations to impair survival signaling with the use of specific agents and to impair bcl-2 that protects the cancer cell from apoptosis. We highlight the historical understanding of oestrogen's role in cell survival and death and specifically illustrate the progress that has been made in the last five years to understand the mechanisms of oestrogen-induced apoptosis. There are opportunities to harness knowledge from this new signal transduction pathway to discover the precise mechanism of this oestrogen-induced apoptotic trigger. Indeed, the new biology of oestrogen action also has significance for understanding the physiology of bone remodeling. Thus, the pathway has a broad appeal in both physiology and cancer research.
Estrogen deficiency has been regarded as the main causative factor in menopausal symptoms and diseases. Here, we show that although estrogen decreases by 90%, a concurrent but inverse change occurs in iron levels during menopausal transition. For example, levels of serum ferritin are increased by two- to threefold from before menopause to after menopause. This observation has led us to hypothesize that, in addition to estrogen deficiency, increased iron as a result of menopause could be a risk factor affecting the health of postmenopausal women. Further studies on iron and menopause are clinically relevant and may provide novel therapeutic treatments.
Hepcidin is a hormone secreted in response to iron loading and inflammation. Hepcidin binds to the iron exporter ferroportin, inducing its degradation and thus preventing iron entry into plasma. We determined that hepcidin binding to ferroportin leads to the binding and activation of the protein Janus Kinase2 (Jak2), which is required for phosphorylation of ferroportin. Ferroportin is a dimer and both monomers must be capable of binding hepcidin for Jak2 to bind to ferroportin. Once Jak2 is bound to the ferroportin dimer, both ferroportin monomers must be functionally competent to activate Jak2 and for ferroportin to be phosphorylated. These results show that cooperativity between the ferroportin monomers is required for hepcidin-mediated Jak2 activation and ferroportin down-regulation. These results provide a molecular explanation for the dominant inheritance of hepcidin resistant iron overload disease.
Mucormycosis is an increasingly common fungal infection with an unacceptably high mortality despite first-line antifungal therapy. Iron acquisition is a critical step in the causative organisms' pathogenetic mechanism. Therefore, abrogation of fungal iron acquisition is a promising therapeutic strategy to impact clinical outcomes for this deadly disease.
The increased risk of mucormycosis in patients with renal failure receiving deferoxamine iron chelation therapy is explained by the fact that deferoxamine actually acts as a siderophore for the agents of mucormycosis, supplying previously unavailable iron to the fungi. The iron liberated from deferoxamine is likely transported into the fungus by the high-affinity iron permease. In contrast, two other iron chelators, deferiprone and deferasirox, do not supply iron to the fungus and were shown to be cidal against Zygomycetes in vitro. Further, both iron chelators were shown to effectively treat mucormycosis in animal models, and one has been successfully used as salvage therapy for a patient with rhinocerebral mucormycosis.
Further investigation and development of iron chelators as adjunctive therapy for mucormycosis is warranted.
Ferroportin (Fpn) is the only known iron exporter in vertebrates. Hepcidin, a peptide secreted by the liver in response to iron or inflammation, binds to Fpn, inducing its internalization and degradation. We show that after binding of hepcidin, Fpn is tyrosine phosphorylated at the plasma membrane. Mutants of human Fpn that do not get internalized or that are internalized slowly show either absent or impaired phosphorylation. We identify adjacent tyrosines as the phosphorylation sites and show that mutation of both tyrosines prevents hepcidin-mediated Fpn internalization. Once internalized, Fpn is dephosphorylated and subsequently ubiquitinated. An inability to ubiquitinate Fpn does not prevent hepcidin-induced internalization, but it inhibits the degradation of Fpn. Ubiquitinated Fpn is trafficked through the multivesicular body pathway en route to degradation in the late endosome/lysosome. Depletion of proteins involved in multivesicular body trafficking (Endosome Sorting Complex Required for Transport proteins), by small-interfering RNA, reduces the trafficking of Fpn-green fluorescent to the lysosome.
Iron is essential for many biological processes, including oxygen delivery, and its supply is tightly regulated. Hepcidin, a small peptide synthesized in the liver, is a key regulator of iron absorption and homeostasis in mammals. Hepcidin production is increased by iron overload and decreased by anemia and hypoxia; but the molecular mechanisms that govern the hepcidin response to these stimuli are not known. Here we establish that the von Hippel-Lindau/hypoxia-inducible transcription factor (VHL/HIF) pathway is an essential link between iron homeostasis and hepcidin regulation in vivo. Through coordinate downregulation of hepcidin and upregulation of erythropoietin and ferroportin, the VHL-HIF pathway mobilizes iron to support erythrocyte production.
The increasing prevalence of multiple comorbidities among anemic patients with chronic kidney disease has made the use of serum ferritin and transferrin saturation more challenging in diagnosing iron deficiency. Because serum ferritin is an acute-phase reactant and because the inflammatory state may inhibit the mobilization of iron from reticuloendothelial stores, the scenario of patients with serum ferritin >800 ng/ml, suggesting iron overload, and transferrin saturation <20%, suggesting iron deficiency, has become more common. This article revisits the basis for the Kidney Disease Outcomes Quality Initiative recommendations regarding the use of serum ferritin and transferrin saturation in guiding iron therapy, then explores some of the newer alternative markers for iron status that may be useful when serum ferritin and transferrin saturation are insufficient. These newer tests include reticulocyte hemoglobin content, percentage of hypochromic red cells, and soluble transferrin receptor, all of which have shown some promise in limited studies. Finally, the role of hepcidin, a hepatic polypeptide, in the pathophysiology of iron mobilization is reviewed briefly.
In thalassemia, deficient globin-chain production during erythropoiesis results in anemia. Thalassemia may be further complicated by iron overload (frequently exacerbated by blood transfusion), which induces numerous endocrine diseases, hepatic cirrhosis, cardiac failure and even death. Accumulation of iron in the absence of blood transfusions may result from inappropriate suppression of the iron-regulating peptide hepcidin by an erythropoietic mechanism. To test this hypothesis, we examined erythroblast transcriptome profiles from 15 healthy, nonthalassemic donors. Growth differentiation factor 15 (GDF15), a member of the transforming growth factor-beta superfamily, showed increased expression and secretion during erythroblast maturation. Healthy volunteers had mean GDF15 serum concentrations of 450 +/- 50 pg/ml. In comparison, individuals with beta-thalassemia syndromes had elevated GDF15 serum levels (mean 66,000 +/- 9,600 pg/ml; range 4,800-248,000 pg/ml; P < 0.05) that were positively correlated with the levels of soluble transferrin receptor, erythropoietin and ferritin. Serum from thalassemia patients suppressed hepcidin mRNA expression in primary human hepatocytes, and depletion of GDF15 reversed hepcidin suppression. These results suggest that GDF15 overexpression arising from an expanded erythroid compartment contributes to iron overload in thalassemia syndromes by inhibiting hepcidin expression.
Iron is an essential dietary element. However, the ability of iron to cycle between oxidized and reduced forms also renders it capable of contributing to free radical formation, which can have deleterious effects, including promutagenic effects that can potentiate tumor formation. Dysregulation of iron metabolism can increase cancer risk and promote tumor growth. Cancer cells exhibit an enhanced dependence on iron relative to their normal counterparts, a phenomenon we have termed iron addiction. Work conducted in the past few years has revealed new cellular processes and mechanisms that deepen our understanding of the link between iron and cancer. Control of iron efflux through the combined action of ferroportin, an iron efflux pump, and its regulator hepcidin appears to play an important role in tumorigenesis. Ferroptosis is a form of iron-dependent cell death involving the production of reactive oxygen species. Specific mechanisms involved in ferroptosis, including depletion of glutathione and inhibition of glutathione peroxidase 4, have been uncovered. Ferritinophagy is a newly identified mechanism for degradation of the iron storage protein ferritin. Perturbations of mechanisms that control transcripts encoding proteins that regulate iron have been observed in cancer cells, including differences in miRNA, methylation, and acetylation. These new insights may ultimately provide new therapeutic opportunities for treating cancer.
Elevated levels of estrogen often associate with increased susceptibility to infection. This has been attributed to the ability of estrogen to concomitantly enhance the growth and virulence of pathogens and suppress host immunity. But the exact mechanism of how estrogen mediates such effects, especially in cases where the pathogen and/or the immune components in question do not express estrogen receptors, has yet to be elucidated. Here we propose that translating the adverse effects of estrogen during infection is dependent to a significant degree upon its ability to manipulate iron homeostasis. For elevated levels of estrogen alter the synthesis and/or activity of several factors involved in iron metabolism including hypoxia inducible factor 1α (HIF-1α) and hepcidin among others. This leads to the inhibition of hepcidin synthesis in hepatocytes and the maintenance of ferroportin (FPN) integrity on the surface of iron-releasing duodenal enterocytes, hepatocytes, and macrophages. Intact FPN permits the continuous efflux of dietary and stored iron into the circulation, which further enhances pathogen growth and virulence on the one hand and suppresses host immunity on the other. This new conceptual framework may help explain a multitude of disparate clinical and experimental observations pertinent to the relationship between estrogen and infection.
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Gender-related disparities in the regulation of iron metabolism may contribute to the differences exhibited by men and women in the progression of chronic liver diseases associated with reduced hepcidin expression, e.g., chronic hepatitis C, alcoholic liver disease, or hereditary hemochromatosis. However, their mechanisms remain poorly understood. In this study we took advantage of the major differences in hepcidin expression and tissue iron loading observed between Bmp6-deficient male and female mice to investigate the mechanisms underlying this sexual dimorphism. We found that testosterone robustly represses hepcidin transcription by enhancing Egfr signaling in the liver and that selective epidermal growth factor receptor (Egfr) inhibition by gefitinib (Iressa) in males markedly increases hepcidin expression. In males, where the suppressive effects of testosterone and Bmp6-deficiency on hepcidin expression are combined, hepcidin is more strongly repressed than in females and iron accumulates massively not only in the liver but also in the pancreas, heart, and kidneys.
Conclusion:
Testosterone-induced repression of hepcidin expression becomes functionally important during homeostatic stress from disorders that result in iron loading and/or reduced capacity for hepcidin synthesis. These findings suggest that novel therapeutic strategies targeting the testosterone/EGF/EGFR axis may be useful for inducing hepcidin expression in patients with iron overload and/or chronic liver diseases.
Testosterone administration increases hemoglobin levels and has been used to treat anemia of chronic disease. Erythrocytosis is the most frequent adverse event associated with testosterone therapy of hypogonadal men, especially older men. However, the mechanisms by which testosterone increases hemoglobin remain unknown. Testosterone administration in male and female mice was associated with a greater increase in hemoglobin and hematocrit, reticulocyte count, reticulocyte hemoglobin concentration, and serum iron and transferring saturation than placebo. Testosterone downregulated hepatic hepcidin mRNA expression, upregulated renal erythropoietin mRNA expression, and increased erythropoietin levels. Testosterone-induced suppression of hepcidin expression was independent of its effects on erythropoietin or hypoxia-sensing mechanisms. Transgenic mice with liver-specific constitutive hepcidin over-expression failed to exhibit the expected increase in hemoglobin in response to testosterone administration. Testosterone upregulated splenic ferroportin expression and reduced iron retention in spleen. After intravenous administration of transferrin-bound (58) Fe, the amount of (58) Fe incorporated into red blood cells was significantly greater in testosterone-treated mice than in placebo-treated mice. Serum from testosterone-treated mice stimulated hemoglobin synthesis in K562 erythroleukemia cells more than that from vehicle-treated mice. Testosterone administration promoted the association of androgen receptor (AR) with Smad1 and Smad4 to reduce their binding to BMP-response elements in hepcidin promoter in the liver. Ectopic expression of AR in hepatocytes suppressed hepcidin transcription; this effect was blocked dose-dependently by AR antagonist flutamide. Testosterone did not affect hepcidin mRNA stability. Conclusion: Testosterone inhibits hepcidin transcription through its interaction with BMP-Smad signaling. Testosterone administration is associated with increased iron incorporation into red blood cells. © 2013 The Authors Aging Cell © 2013 Blackwell Publishing Ltd/Anatomical Society of Great Britain and Ireland.
Estrogen replacement is an effective therapy of postmenopausal symptoms such as hot flushes, bone loss, and vaginal dryness. Undesired estrogen effects are the stimulation of uterine and mammary gland epithelial cell proliferation as well as hepatic estrogenicity. In this study, we examined the influence of different estradiol release kinetics on tissue responsivity in ovariectomized (OVX) rats. Pulsed release kinetics was achieved by ip or sc administration of estradiol dissolved in physiological saline containing 10% ethanol (EtOH/NaCl) whereas continuous release kinetics was achieved by sc injection of estradiol dissolved in benzylbenzoate/ricinus oil (1+4, vol/vol). Initial 3-d experiments in OVX rats showed that pulsed ip estradiol administration had profoundly reduced stimulatory effects on the uterus and the liver compared with continuous release kinetics. On the other hand, both administration forms prevented severe vaginal atrophy. Based on these results, we compared the effects of pulsed (sc in EtOH/NaCl) vs. continuous (sc in benzylbenzoate/ricinus oil) estradiol release kinetics on bone, uterus, mammary gland, and liver in a 4-month study in OVX rats. Ovariectomy-induced bone loss was prevented by both administration regimes. However, pulsed estradiol resulted in lower uterine weight, reduced induction of hepatic gene expression, and reduced mammary epithelial hyperplasia relative to continuous estradiol exposure. We conclude that organ responsivity is influenced by different hormone release kinetics, a fact that might be exploited to reduce undesired estradiol effects in postmenopausal women.
Hepcidin is a peptide hormone secreted by the liver in response to iron loading and inflammation. Decreased hepcidin leads
to tissue iron overload, whereas hepcidin overproduction leads to hypoferremia and the anemia of inflammation. Ferroportin
is an iron exporter present on the surface of absorptive enterocytes, macrophages, hepatocytes, and placental cells. Here
we report that hepcidin bound to ferroportin in tissue culture cells. After binding, ferroportin was internalized and degraded,
leading to decreased export of cellular iron. The posttranslational regulation of ferroportin by hepcidin may thus complete
a homeostatic loop: Iron regulates the secretion of hepcidin, which in turn controls the concentration of ferroportin on the
cell surface.
We investigated the effect of 17β-estradiol (E(2)) alone and separately vitamin E treatment on trace element status of rats following an ovariectomic operation. Forty rats were equally divided into four groups: Group 1, control, non-ovariectomized rats; Group 2, (OVX) rats, ovariectomized under general anesthesia; Group 3, (OVX+E(2)) rats, the group received a 40 µg kg(-1) subcutan dose of E(2) per day after ovariectomy; and Group 4, (OVX + E(2) + vitamin E) rats, received the same E(2) treatment, but with an additional 100 mg kg(-1) intraperitoneal dose of vitamin E per day after ovariectomy. At the end of the 30-day experiment, the rats were sacrificed and their blood was collected for the measurement of zinc, copper, iron, phosphorus, selenium, magnesium, calcium, manganese, and chromium; copper-zinc superoxide dismutase (SOD); manganese-superoxide dismutase (Mn-SOD); glutathione peroxidase (Se-GSH-Px); and catalase (CAT). The levels of zinc, copper, iron, phosphorus, selenium, calcium, chromium, and manganese and activities of SOD, Mn-SOD, Se-GSH-Px, and CAT were lower in the OVX than in the control group, but magnesium level was unaffected. However, zinc, copper, iron, phosphorus, selenium, calcium, chromium, and manganese levels and SOD, Mn-SOD, Se-GSH-Px, and CAT activities were higher under separate E(2) and E(2) + vitamin E treatments. The level of magnesium in the treated-OVX groups was not different than in the OVX group. In conclusion, E(2) treatment has an ameliorating effect on the trace element status in OVX, and this effect may be enhanced with the addition of vitamin E.
Iron is essential for many biological processes and its deficiency or excess is involved in pathological conditions. At cellular level, the maintenance of iron homeostasis is largely accomplished by the transferrin receptor (TfR-1) and by ferritin, whose expression is mainly regulated post-transcriptionally by iron regulatory proteins (IRPs). This study examines the hypothesis that modification of serum estrogen levels by ovariectomy and 17beta-estradiol (E(2)) treatment in rats modulate serum iron-status parameters and iron metabolism in adipose tissue. In particular, we evaluated the RNA binding of IRP1 by electrophoretic mobility-shift assay and IRP1, ferritin, and TfR-1 expression in adipose tissue by Western blot analysis. Ovariectomy, besides a lowered serum iron and transferrin iron binding capacity, remarkably decreased the binding activity of IRP1 in peritoneal and subcutaneous adipose tissues, and these effects were reversed by E(2) treatment. Moreover, ovariectomy determined a decrease of IRP1 expression, which was significant in subcutaneous adipose tissue. Consistent with IRP1 regulation, an increase of ferritin and a decrease of TfR-1 expression were observed in peritoneal adipose tissue from ovariectomized animals, while the treatment with E(2) reconstituted TfR-1 level. A similar expression profile of TfR-1 was observed in subcutaneous adipose tissue, where ferritin level did not change in ovariectomized animals, and was increased after E(2) treatment. Our results indicate that estrogen level changes can regulate the binding activity of the IRP1, and consequently ferritin and TfR-1 expression in adipose tissue, suggesting a relationship among serum and tissue iron parameters, estrogen status and adiposity.
Interferon-gamma (IFNgamma)-mediated indoleamine 2,3-dioxygenase (IDO) expression, important in innate immunity, immune suppression, and tolerance, can be counteracted by ferrous iron (FeSO(4)). Elevation of intracellular iron levels during stimulation with IFNgamma impeded IFNgamma-induced IDO mRNA and protein expression in HEp-2 cells. Decreased IDO expression was accompanied by decreased tryptophan degradation. Accordingly, IFNgamma-mediated suppressing effects on Chlamydia trachomatis (CT) infection were reduced or even abolished in the presence of FeSO(4). Conversely, lowering intracellular iron levels by deferoxamine (DFO) did not increase IFNgamma-induced IDO expression but potentiated Chlamydia-suppressing effects by lowering intracellular iron availability. Additionally, DFO led to a CT-induced IDO expression in HEp-2 cells not treated with IFNgamma. In summary, this study demonstrates that iron acts as a regulatory element for modulating IDO expression, in addition to its function as an essential element for chlamydial growth. This may represent an important control mechanism of IDO expression at the transcriptional level.
Our previous study revealed that estrogen regulates nm23-H1 expression thus promoting cell migration-invasion via activating PIK3/Akt pathway. In this study, we explored the effect of hormone on hypoxia-inducible factor-1 (HIF-1alpha), a key factor in cancer invasion and metastasis, via activation of Akt signaling transduction pathway. We treated two ovarian cancer cell lines ES-2 and SKOV3 with 17beta-estradiol, methoxyprogesterone acetate (MPA) only, or hormone combined with and Akt, MAPK pathway inhibitor, or transefected with siRNA targeting Akt sequenced with hormone. Expression of HIF-1alpha was measured by Western blotting. We observed the effect of hormone on nm23-H1 expression after the cells were transfected by siRNA targeting HIF-1alpha or treated with CoCl2 to induce HIF-1alpha overexpression. The 17beta-estradiol increased HIF-1alpha expression in ovarian cancer cells, and this upregulatory effect was abrogated by Akt inhibitor LY294002 (P<0.05) and Akt siRNA interference (P<0.05), but not affected by MAPK inhibitor PD980059 (P>0.05). MPA had the opposite effect. Nm23-H1 protein expression in ES-2 and SKOV3 cells were decreased after treatment with 17beta-estradiol (P<0.05), whereas MPA had the opposite effect. The effect was attenuated by HIF-1alpha siRNA (P<0.05) and enhanced by HIF-1alpha overexpression after CoCl2 treatment (P<0.05). Our data suggest that estrogen and progestin regulate HIF-1alpha expression via Akt signaling pathway, affecting nm23-H1 expression in influencing cell metastasis.
Ferritin values for 250 selected sera were compared with values for iron, total iron-binding capacity (TIBC), and transferrin saturation, to assess the potential of the ferritin assay for the detection of latent iron deficiency. The specimens were grouped (50 in each group) according to their values for iron and TIBC. In Group 1 (low iron, high TIBC) the saturation and ferritin values both indicated iron deficiency in all but one. In the 100 specimens of Groups 2 (normal iron, high TIBC) and 4 (normal iron, high normal TIBC), the saturation values revealed 16 iron-deficient cases, the ferritin test 55. For Groups 3 (low iron, normal TIBC) and 5 (low iron, low TIBC), the ferritin test revealed fewer cases of iron deficiency than did the saturation values (37 cases vs 51 cases, in the 100 specimens). Evidently the ferritin test detects iron deficiency in many cases for whom the serum iron and TIBC tests are not positively indicative. The correlation of serum ferritin with iron, TIBC, and transferrin saturation in the five groups was good only in the case of specimens for which the TIBC was normal; if it was abnormal the correlation was very poor.
The effect of iron overload on susceptibility of mice to Candida albicans infection and on the type of T helper (Th) immunity elicited was investigated. Iron overload greatly increased susceptibility
to disseminated infection with low-virulence C. albicans cells of exogenous origin. The candidacidal activity and the ability to release nitric oxide and bioactive interleukin (IL)-12
were greatly impaired in neutrophils and macrophages from infected mice. CD4 T cells from spleens of iron-overloaded mice
were found to produce high levels of IL-4 and IL-10 and low levels of interferon-γ. Treatment of iron-overloaded mice with
the iron chelator, deferoxamine, resulted in the cure of mice from infection, restored the antifungal effector and immunomodulatory
functions of the phagocytic cells, and allowed the occurrence of CD4 Th1 protective antifungal responses. These data indicate
that iron overload may negatively affect CD4 Th1 development in mice with candidiasis, a function efficiently restored by
therapy with deferoxamine.
During infection microbes attack host tissues, causing damage to specific organs, sepsis or even death. For proliferation microbes desperately need iron for which they have to compete with the host. Micro-organisms have developed an abundant number of strategies to acquire iron from their specific environment and to transport the element to sites of incorporation into biologically important molecules. As part of the non-specific defence mechanisms against infection, the body modifies iron metabolism in order to make iron less available for micro-organisms. Such processes have a profound effect on the immune system and are also expressed in other forms of inflammation. Microbial iron transport systems are explored as targets for antibiotic treatment and vaccines. In particular, iron chelators, used for the treatment of iron overload may become important drugs for fighting bacterial and viral infections.
Hypoferremia is a common response to systemic infections or generalized inflammatory disorders. In mouse models, the development of hypoferremia during inflammation requires hepcidin, an iron regulatory peptide hormone produced in the liver, but the inflammatory signals that regulate hepcidin are largely unknown. Our studies in human liver cell cultures, mice, and human volunteers indicate that IL-6 is the necessary and sufficient cytokine for the induction of hepcidin during inflammation and that the IL-6-hepcidin axis is responsible for the hypoferremia of inflammation.
We have previously used genome-wide transcript profiling to investigate the relationships between changes in gene expression and physiological alterations during the response of the immature mouse uterus to estrogens. Here we describe the identification of a functionally inter-related group of estrogen-responsive genes associated with iron homeostasis, including the iron-binding protein lactotransferrin, the ferroxidase ceruloplasmin, the iron delivery protein lipocalin 2 and the iron-exporter ferroportin. Quantitative real-time PCR revealed that the expression of these genes increases with time during the uterotrophic response, reaching maximal levels in the post-proliferative phase (between 48 and 72 h). In contrast, the heme biosynthesis genes aminolevulinic acid synthase 1 and 2 were maximally induced by estrogen at 2 and 4 h, respectively, prior to increased cell proliferation. Together, these data reveal that estrogen induces the temporally coordinated expression of iron homeostasis genes in the mouse uterus, and suggest an important role for iron metabolism during sex steroid hormone-induced uterine cell growth and differentiation.
Hepcidin is a cationic amphipathic peptide made in the liver, released into plasma and excreted in urine. Hepcidin is the homeostatic regulator of intestinal iron absorption, iron recycling by macrophages, and iron mobilization from hepatic stores, but it is also markedly induced during infections and inflammation. Under the influence of hepcidin, macrophages, hepatocytes, and enterocytes retain iron that would otherwise be released into plasma. Hepcidin acts by inhibiting the efflux of iron through ferroportin, the sole known iron exporter that is expressed in the small intestine, and in hepatocytes and macrophages. As befits an iron-regulatory hormone, hepcidin synthesis is increased by iron loading, and decreased by anemia and hypoxia. Hepcidin is also rapidly induced by cytokines, including IL-6. The resulting decrease in plasma iron levels eventually limits iron availability to erythropoiesis and contributes to the anemia associated with infection and inflammation. The decrease in extracellular iron concentrations due to hepcidin probably limits iron availability to invading microorganisms, thus contributing to host defense.