Iron chelation with oral deferiprone in patients with thalassemia.
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Deferiprone↗
Biomedical subjects
M B Martin
Publications and source records attributed to M B Martin.
Upregulation of estrogen receptor-alpha expression in rabbit cardiac allograft.
Estrogen receptor (ER) expression has been detected in different tissues, and estradiol-17beta treatment protects against experimental transplant arteriosclerosis. In this study, ER-alpha expression in the rabbit hearts and attached aortas before and after cardiac-aorta allograft transplantation was examined. Ten male New Zealand White rabbits were transplanted with cardiac-aorta allografts from male Dutch Belted rabbits. This transplant arteriosclerosis model uses a 0.5% cholesterol diet and immunosuppression with cyclosporin A (10 mg . kg-1 . d-1) until euthanatization 42 days later. The cardiac grafts with the attached aorta were harvested. Strong staining of ER-alpha protein was shown in the coronary arteries of the cardiac allografts by immunohistochemistry with the use of a mouse anti-human ER-alpha monoclonal antibody (ID5). In contrast, both the nongrafted hearts of the recipients and donor hearts expressed only weak staining. RNase protection assay with the use of a 32P-labeled ER-alpha antisense riboprobe (pOR 300) proved that the basal expression of ER-alpha mRNA is similar in the nongrafted aorta of both recipients and donors. A marked increase of ER-alpha mRNA was observed in the allograft aorta compared with the nongrafted aorta (289+/-69%, P<0. 02) by reverse transcription and polymerase chain reaction. The DNA sequence analysis confirmed that the polymerase chain reaction-amplified fragment corresponded to ER-alpha. This is the first observation of ER-alpha upregulation in the allograft vasculature and may relate to the allograft cardiovascular protective effects of estrogen.
Animals↗
Consumption of a high-fat diet alters estrogen receptor content, protein kinase C activity, and mammary gland morphology in virgin and pregnant mice and female offspring.
Previous studies have shown that a diet high in polyunsaturated fatty acids increases mammary tumor incidence in adult and pregnant mice and rats and in the female offspring. The present study investigated whether a high-fat diet alters the number of estrogen receptor (ER) binding sites and protein kinase C (PKC) activity in the mammary gland of these animals. In the female offspring, the effects of maternal exposure to a high-fat diet during pregnancy on development of the mammary epithelial tree were studied also. BALB/c mice were kept on a diet containing either 43% (high-fat) or 16% (low-fat) calories from corn oil, which consists mostly of n-6 polyunsaturated fatty acids, for 1 month. In adult female mice, a 6-fold increase in the number of ER binding sites and 2-fold increase in PKC activity were found in the mammary glands of the high-fat mice when compared with the low-fat mice. In pregnant mice, a high-fat diet increased ER binding sites by 61% and PKC activity by 51%. In contrast to adult and pregnant mice, females exposed to a high-fat diet only in utero through their pregnant mother exhibited a significantly reduced number of mammary ER binding sites by age 45 days (78% decrease) and a reduction in PKC activity by ages 30 and 100 days (44 and 20% decrease, respectively). The mammary epithelial tree of the high-fat offspring contained more terminal end buds and was less differentiated than that of the low-fat offspring. These findings show that consumption of a high-fat diet increases ER and PKC in the adult and pregnant mouse mammary gland, perhaps contributing to the fat-induced promotion of mammary tumorigenesis. In contrast, reduced ER and PKC following a high-fat exposure in utero may be associated with increased susceptibility to carcinogenesis, possibly due to an increased number of terminal end buds that are the sites of neoplastic transformation in the mammary gland.
Animals↗
Estradiol regulates estrogen receptor mRNA stability.
Previous studies suggest that post-transcriptional events play an important role in estrogen-induced loss of estrogen receptor expression. The present study shows that treatment of MCF-7 cells with estradiol resulted in a six-fold decrease in estrogen receptor mRNA half-life from 4 h in control cells to 40 min in estradiol treated cells. To determine the role of protein synthesis in the regulation of estrogen receptor mRNA stability, several translational inhibitors were utilized. Pactamycin and puromycin, which prevent ribosome association with mRNA, inhibited the effect of estradiol on receptor mRNA stability, whereas cycloheximide, which has no effect on ribosome association with mRNA, had no effect on estradiol regulation of estrogen receptor mRNA stability. In control cells, the total cellular content of estrogen receptor mRNA was associated with high molecular weight polyribosomes. Treatment with estradiol resulted in a 70% decrease in estrogen receptor mRNA associated with polyribosomes but had no effect on the polyribosome distribution of estrogen receptor mRNA. In an in vitro degradation assay, polyribosomes isolated from estradiol-treated cells degraded ER mRNA faster than polyribosomes isolated from control cells. The nuclease activity associated with the polysome fraction appeared to be Mg2+ independent and inhibited by RNasin. Freeze-thawing and heating at 90 degrees C for 10 min resulted in the loss of nuclease activity. These studies suggest that an estrogen-regulated nuclease activity associated with ribosomes alters the stability of estrogen receptor mRNA.
Breast Neoplasms↗
Structure-activity relationships of N-hydroxyurea 5-lipoxygenase inhibitors.
The discovery of second generation N-hydroxyurea 5-lipoxygenase inhibitors was accomplished through the development of a broad structure-activity relationship (SAR) study. This study identified requirements for improving potency and also extending duration by limiting metabolism. Potency could be maintained by the incorporation of heterocyclic templates substituted with selected lipophilic substituents. Duration of inhibition after oral administration was optimized by identification of structural features in the proximity of the N-hydroxyurea which correlated to low in vitro glucuronidation rates. Furthermore, the rate of in vitro glucuronidation was shown to be stereoselective for certain analogs. (R)-N-[3-[5-(4-Fluorophenoxy)-2-furyl]-1-methyl-2-propynyl]-N-hydroxyure a (17c) was identified and selected for clinical development.
Animals↗
The role of transforming growth factor-beta in the regulation of estrogen receptor expression in the MCF-7 breast cancer cell line.
The role of transforming growth factor-beta1 (TGFbeta1) in the regulation of estrogen receptor (ER) expression in MCF-7 cells was investigated. After treatment of the cells with 100 pM TGFbeta1, ER protein declined by about 30% at 6 h from a concentration of 413.5 fmol/mg protein in control cells to 289.5 fmol/mg protein in treated cells. The concentration of receptor remained suppressed for 24 h. Scatchard analysis demonstrated that the decrease in ER protein corresponded to a decrease in estradiol-binding sites, with no effect on the binding affinity of the ER. The dissociation constant of the estradiol-ER complex was 0.117 nM in TGFbeta1-treated cells compared to 0.155 nM in control cells. Treatment with TGFbeta1 did not influence the half-life of the ER. In TGFbeta1-treated cells, as well as in control cells, the half-life of the receptor was approximately 4 h. In contrast to the effect on ER concentration, TGFbeta1 treatment resulted in a greater decrease in the steady state level of ER messenger RNA (approximately 75%) at 6 h. By 24 h, a small recovery in the amount of messenger RNA was observed. Transcription run-on experiments demonstrated a decrease of approximately 70% in the level of ER gene transcription at 3 h. Transient transfections using an ER promoter-chloramphenicol acetyltransferase construct demonstrated that after TGFbeta1 treatment, chloramphenicol acetyltransferase activity decreased by 50%, suggesting that TGFbeta1 inhibition of the ER gene transcription is mediated through the ER promoter. Although treatment with TGFbeta1 decreased the ER concentration, the growth factor had no effect on the activity of ER, as measured by its effects on estradiol induction of progesterone receptor and pS2, suggesting that TGFbeta1 does not inhibit proliferation of MCF-7 cells by blocking ER activity.
Breast Neoplasms↗
Expression of an estrogen receptor by human coronary artery and umbilical vein endothelial cells.
BACKGROUND: Premenopausal women have much lower susceptibility to coronary artery disease than do men or postmenopausal women. It has been proposed that estrogen plays a role in cardioprotection, but little information is available regarding the mechanism by which estrogen may help to protect the vasculature. Here, we describe an estrogen receptor (ER) in human coronary artery and umbilical vein endothelial cells. METHODS AND RESULTS: Human umbilical vein endothelial cells and human coronary artery endothelial cells were cultured in hormone-free medium for 48 hours before experiments. Estradiol (3.7 nmol/L) added to cultures promoted proliferation by a mechanism that is inhibited by the specific ER antagonist ICI182,780. Estradiol-treated cells incorporated twice the [3H]thymidine of hormone-free cells; this increase was prevented by ICI182,780. Endothelial cells from both sources stained in a nuclear pattern with an ER-specific antibody. Ribonuclease protection assay detected mRNA for the ER. Ligand-binding studies estimated 2 x 10(4) to 8 x 10(4) receptors per cell and a Kd of approximately 5 nmol/L. Interaction of ERs with a consensus estrogen response element was shown by an electrophoretic mobility shift assay. In addition, an antibody against the ER supershifted the protein-DNA complex. CONCLUSIONS: These studies define the presence of an ER in human coronary artery and umbilical vein endothelial cells. They support the hypothesis that cardioprotective effects of estrogen are mediated, at least in part, through a classic steroid hormone receptor mechanism.
Arteries↗
Effect of divergent self-monitoring strategies on motor performance and emotion as a function of alternating task complexity.
The aim of this study was to determine the effectiveness of implementing different self-monitoring (SM) strategies as a function of varying task complexity on a computer game, 'Shufflepuck'. Self-monitoring theory suggests that positive self-monitoring (PSM) results in better performance for difficult tasks, whereas negative self-monitoring (NSM) is preferable for easy tasks. When SM was tested under easy and complex task conditions, it was found that performance effectiveness was a function of matching appropriate task difficulty with the SM strategy. Unpaired t-tests compared the first set of 100 trials (pre-task) with the second set of 100 trials (post-task) on a computer skills game requiring hand-eye coordination. The results showed that subjects who changed from the difficult task/PSM condition to the easy task/PSM condition (different task, same SM strategy) performed significantly poorer (P < 0.001) than subjects who changed from the difficult task/PSM condition to the easy task/NSM condition (different task, different SM strategy). A second unpaired t-test comparing the two sets of difference performance scores indicated that performance was significantly superior for subjects using the PSM than the NSM strategy while performing the difficult task (P < 0.05). That is, changing from the easy task/NSM condition to the difficult task/NSM condition produced poorer performance than changing from the easy task/NSM to the difficult task/PSM condition. In terms of subjects' emotions, the use of NSM and PSM when performing easy and difficult tasks, respectively, resulted in markedly less negative affect (P < 0.01), but did not influence positive affect, on pre- and post-task comparisons. Finally, correlations between affect and performance, based on pre- and post-performance and affect scores, were moderate to high. Implications of the present results for slumps in sport performance are offered.
Exercise↗
Regulation of estrogen receptor concentration and activity by an erbB/HER ligand in breast carcinoma cell lines.
Expression of the erbB-2 oncogene in breast cancer patients correlates with poor prognosis and failure of hormonal therapy. In this study, the effects of a putative erbB/HER ligand, gp30, on estrogen receptor (ER) concentration and activity was investigated in the estrogen receptor positive human breast cancer cells, BT474 and MCF-7, which express either high or low levels of erbB-2 and erbB-4, respectively. Treatment of cells with gp30 resulted in a decrease in the steady-state level of estrogen receptor protein by approximately 70-80%. The effect of gp30 on the concentration of ER was independent of serum in the media and was not inhibited by an epidermal growth factor receptor blocking antibody. In addition to the effect on ER protein, gp30 decreased the steady-state level of ER messenger RNA. Transcription run on experiments demonstrated that the decrease in ER expression was mediated by a decrease in ER gene transcription. The effect of gp30 on estrogen receptor activity was also investigated in this study. Treatment of cells with gp30 blocked estradiol induction of progesterone receptor. Inhibition was observed at the level of progesterone receptor protein, messenger RNA, and gene transcription. gp30 also blocked estradiol induction of pS2 gene transcription. In addition to its effects on progesterone receptor and pS2, gp30 blocked activation of an estrogen response element in a transient transfection assay and inhibited ER binding to its response element in a DNA mobility shift assay, suggesting a direct effect on the estrogen receptor. The effects of gp30 on estrogen receptor concentration and activity were independent of the level of erbB-2 and erbB-4 in the cell. These data show that gp30 regulates the concentration of ER and modulates ER activity.
Breast Neoplasms↗
Bidirectional interactions between the estrogen receptor and the cerbB-2 signaling pathways: heregulin inhibits estrogenic effects in breast cancer cells.
The responsiveness of estrogen receptor (ER)-positive breast cancer to endocrine therapy is frequently reduced in cells over-expressing c-erbB-2. Stimulation of ER suppresses c-erbB-2, indicating that estrogen controls the activity of c-erbB-2. Heregulin (HRG) has been described to bind to c-erbB-3/c-erbB-4 and to stimulate c-erbB-2. Here we describe the effects of HRG on cell growth and on ER and c-erbB-2 expression in breast cancer cell lines containing distinct levels of c-erbB-2 and ER (BT-474: c-erbB-2 , ER+; MDA-MB-361: c-erbB-2++, ER++; MCF-7: c-erbB-2+, ER ). Proliferation of estrogen-stimulated, c-erbB-2 and ER-positive cells is inhibited by HRG in a dose-dependent manner. In addition, HRG dose-dependently inhibits ER expression. Estrogen, however, inhibits c-erbB-2. Estrogen-mediated down-regulation of c-erbB-2 is most pronounced in MCF-7 but weaker in BT-474. In the latter cells HRG efficiently blocks the estrogenic effect on c-erbB-2. In MCF-7 cells, however, the inhibition of c-erbB-2 cannot be completely reverted by HRG. This modulation occurs in all 3 cell lines at protein, RNA and transcriptional levels, suggesting that the activity of the c-erbB-2 promoter, which contains an estrogen-responsive region, is affected by HRG. The intensity of the mutual inhibition between the HRG/c-erbB-2 and the estrogen/ER system depends on the relative levels of ER and c-erbB-2 expression in the respective cell lines.
Antineoplastic Agents↗
Effects of 12-O-tetradecanoylphorbol-13-acetate on estrogen receptor activity in MCF-7 cells.
The effects of long term treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA) on estrogen receptor (ER) expression in the human breast cancer cell line, MCF-7, were studied. This study demonstrates that treatment of cells with the phorbol ester blocked estrogen receptor activity. Treatment of cells with 100 nM TPA resulted in an 80% decrease in the level of ER protein and a parallel decrease in ER mRNA and binding capacity. Following removal of TPA from the medium, the level of ER protein and mRNA returned to control values; however, the receptor failed to bind estradiol. These cells also failed to induce progesterone receptor in response to estradiol. In addition, TPA treatment blocked transcription from an estrogen response element in transient transfection assays and inhibited ER binding to its response element in a DNA mobility shift assay. The estrogen receptor in treated cells was recognized by two monoclonal anti-ER antibodies and was not quantitatively different from ER in control cells. RNase protection analysis failed to detect any qualitative changes in the ER mRNA transcript. Mixing experiments suggest that TPA induces/activates a factor which interacts with the ER to block binding of estradiol. The effects of TPA on ER levels and binding capacity were concentration-dependent. Low concentrations of TPA inhibited estradiol binding without a decrease in the level of protein, whereas higher concentrations were required to decrease the level of ER protein. The effects of TPA appear to be mediated by activation of protein kinase C since the protein kinase C inhibitors, H-7 and bryostatin, block the effects of TPA on estradiol induction of progesterone receptor. TPA treatment had no effect on the level or binding capacity of the glucocorticoid receptor, indicating that the effects are not universal for steroid receptors. These data demonstrate that activation of the protein kinase C signal transduction pathway modulates the estrogen receptor pathway. The long term effect of protein kinase C activation is to inhibit estrogen receptor function through induction/activation of a factor which interacts with the receptor.
1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
Genetic alterations of microsatellites on chromosome 18 in human breast carcinoma.
Allelic alterations of chromosome 18 microsatellites were determined using normal and tumor DNA pairs from 29 patients with infiltrating ductal carcinoma of the breast. Loss of heterozygosity was detected in 62% (18 of 29 patients) of the tumors at one or more of these microsatellites. Eight of the 18 patients exhibited deletions in the region at 18q21.1. This chromosomal band is known to contain a tumor suppressor gene (DCC) whose expression is frequently inactivated in several types of cancer. Ten other patients had deletions in regions not included in the DCC locus. Five of these patients revealed a common deletion at the D18S50 locus (18q23), and the other five patients had deletions in various other regions of the chromosome. No apparent correlation between loss of heterozygosity of chromosome 18 microsatellites and the clinical stage was found in this series. The results indicate that, in addition to the DCC locus, the 18q23 region is likely to contain a second tumor suppressor gene relevant to breast carcinogenesis. Four percent of all microsatellites tested in these patients showed allelic differences in the sizes of repeat units between tumor and the corresponding constitutional DNAs. The pattern of allele instability observed in breast carcinoma differed from that originally reported in a hereditary type of colorectal carcinoma. The observation suggests that this phenomenon is not a mechanism specific to neoplastic processes in breast carcinoma.
Alleles↗
Regulation of retinoblastoma gene expression in hormone-dependent breast cancer.
Studies have shown an increased risk for breast cancer in the mothers of children suffering from retinoblastoma and osteosarcoma, suggesting a role for the retinoblastoma susceptibility (Rb) gene product in breast cancer. We now show that estradiol decreases the expression of Rb at the level of protein and messenger RNA (mRNA) in estrogen-dependent breast cancer cell lines. Treatment of MCF-7 cells with 10(-9) M estradiol for 48 h resulted in a 70% decrease in the level of Rb protein. Ribonuclease protection assays showed a 50% decrease in the steady state levels of Rb mRNA by 12 h and a 70% decrease in Rb mRNA by 24 h. Treatment with estradiol had no effect on the rate of Rb gene transcription or on Rb mRNA stability, but resulted in an increase in the steady state level of Rb mRNA in the nucleus. The effect of estradiol was inhibited by 10(-7) M 4-hydroxytamoxifen. In the absence of estradiol, the antiestrogens 4-hydroxytamoxifen and ICI 164,384 increased Rb mRNA by 50% over that in estrogen-depleted conditions. Estradiol regulation of Rb mRNA also occurred in other estrogen-dependent breast cancer cell lines. Insulin-like growth factor I, insulin, progestins, and epidermal growth factor had no effect on Rb expression. In summary, these results show that estradiol specifically regulates the expression of the Rb susceptibility gene product in hormone-dependent breast cancer by a posttranscriptional mechanism that occurs in the nucleus. The results from this study suggest that the negative regulation of Rb expression by estradiol, rather than Rb loss or mutation, may play an important role in breast carcinogenesis.
Breast Neoplasms↗
Effect of cadmium on estrogen receptor levels and estrogen-induced responses in human breast cancer cells.
The effects of cadmium on estrogen receptor and other estrogen-regulated genes in the human breast cancer cell line MCF-7 were studied. Treatment of MCF-7 cells with 1 microM cadmium decreased the level of estrogen receptor 58%. Cadmium induced a parallel decrease in estrogen receptor mRNA (62%). Progesterone receptor levels increased 3.2-fold after cadmium treatment. This induction was blocked by the anti-estrogen ICI-164,384. Progesterone receptor mRNA was also increased by cadmium, as well as cathepsin D mRNA. An in vitro nuclear transcription run-on assay showed that cadmium increased the transcription of the progesterone receptor and pS2 genes and decreased transcription of the estrogen receptor gene. These are not general effects of heavy metals, as zinc, 25 and 100 microM, did not affect progesterone receptor protein and mRNA levels. Cadmium stimulated pS2 and progesterone receptor mRNAs in a clone of MDA-MB-231 cells transfected with the human estrogen receptor, but had no effect in MDA-MB-231 cells transfected with antisense estrogen receptor. Cadmium also stimulated an estrogen response element in transient transfection experiments. These data suggest that the effects of cadmium are mediated by the estrogen receptor independent of estradiol. In addition to its effect on gene expression, cadmium induced the growth of MCF-7 cells 5.6-fold.
Breast Neoplasms↗
Erythrocytapheresis therapy to reduce iron overload in chronically transfused patients with sickle cell disease.
Chelation therapy with deferoxamine is effective in preventing the risk of transfusional iron overload, but treatment failure is common because of noncompliance. To reduce the transfusional iron load, we have evaluated longterm erythrocytapheresis in 14 subjects with sickle cell disease and stroke (11) or other complications (3) as an alternative to simple transfusion. Subjects were treated with erythrocytapheresis using the Haemonetics V50 (Haemonetics Corp, Braintree, MA) to maintain the target pretransfusion hemoglobin S (Hb S) level less than 50% for 6 to 71 months. The transfusional iron load and the donor blood usage were analyzed for a 6- to 36-month study period and were compared with similar data from a subset of 7 subjects previously treated with conventional (target Hb S < 30%) and modified (target Hb S < 50%) simple transfusion protocols. The effect of erythrocytapheresis on iron accumulation was determined by assessment of serum ferritin levels in the absence of iron chelation. The mean transfusional iron load and donor blood usage with erythrocytapheresis were 19 +/- 14 mg iron/kg/yr (range, 6 to 50) and 188.4 +/- 55.2 mL packed-red blood cells (RBC)/kg/yr (range, 107 to 281), respectively. Of 6 subjects receiving no iron chelation therapy, 5 maintained normal or nearly normal serum ferritin levels during 11 to 36 months of erythrocytapheresis. In comparison with conventional simple transfusion and modified simple transfusion, erythrocytapheresis reduced iron loading by 87% (P < .01) and 82% (P < .01), respectively, but increased donor blood usage by 23% and 73%, respectively. Subjects with pre-erythrocytapheresis Hb levels > or = 8.0 g/dL had lower iron accumulation (P < .001) and less donor blood usage (P < .005) than subjects with Hb levels < or = 8.0 g/dL. Although donor blood usage is increased in comparison with simple transfusion, long-term erythrocytapheresis markedly reduces or prevents iron accumulation. This form of transfusion therapy allows the cessation of iron chelation in well-chelated subjects and, if used as the initial form of transfusion therapy, may prevent long-term complications of sickle cell disease without risk of iron overload and the need for chelation therapy.
Adolescent↗
Regulation of estrogen receptor expression.
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Breast Neoplasms↗
Familial translocation t(10;14) (q26.1;q32.3): report of three offspring with 10q deletion and 14q duplication.
We describe two brothers and a cousin with common clinical features, including mild mental retardation, motor delays, hypotonia with truncal ataxia, esotropia, and mild facial and hand dysmorphia. The initial routine chromosome study failed to detect any abnormality in the proband. Based on a high index of clinical suspicion, high-resolution chromosome studies were performed on the proband's parents. A small reciprocal translocation t(10;14) (q26.1;q32.3) was detected in the father. The breakpoint on the derivative chromosome 14 was further placed telomeric to the immunoglobulin heavy-chain gene cluster at the band q32.33 by fluorescence in situ hybridization. Studies of the proband and two affected paternal cousins revealed that each had inherited the same derivative chromosome 10 from their carrier parents. This unbalanced karyotype resulted from an adjacent-1 segregation of the 10;14 translocation.
Abnormalities, Multiple↗
Hepatic iron stores and plasma ferritin concentration in patients with sickle cell anemia and thalassemia major.
To examine the relationship between hepatic iron stores and plasma ferritin concentration in individuals treated with red cell transfusion and iron chelation therapy, 37 patients with sickle cell anemia and 74 patients with thalassemia major were studied. In each patient, hepatic iron stores were measured by an independently validated noninvasive magnetic method, and plasma ferritin was determined by immunoassay. The correlation between hepatic iron and plasma ferritin was significant both in patients with sickle cell anemia (R = 0.75, P < 0.0001) and in those with thalassemia major (R = 0.76, P < 0.0001). Regression analysis showed no significant difference between the two groups in the linear relationships between hepatic iron stores and plasma ferritin. Considering all 111 transfused patients as a group, the coefficient of correlation between hepatic iron stores and plasma ferritin was highly significant (R = 0.76, P < 0.0001). Regression analysis found that variation in body iron stores, as assessed by magnetic determinations of hepatic iron, accounted for only approximately 57% of the variation in plasma ferritin, suggesting that the remainder was the result of other factors, such as hemolysis, ineffective erythropoiesis, ascorbate deficiency, inflammation, and liver disease. The 95% prediction intervals for hepatic iron concentration, given the plasma ferritin, were so broad as to make a single determination of plasma ferritin an unreliable predictor of body iron stores. Variability resulting from factors other than iron status limits the clinical usefulness of the plasma ferritin concentration as a predictor of body iron stores.
Adolescent↗