Polymorphisms in CYP17, COMT, and ESR1 genes in women after menopause and association with bone mineral density.

In this study, we evaluated genetic factors related to the mineral density during post-menopause. We evaluated 110 women in the first 5 years post-menopause, without previous hormone replacement therapy. Cytochrome P450 17 (CYP17) (rs743572), catechol-O-methyl transferase (COMT) (rs4680), and estrogen receptor 1 (ESR1) (rs9322331) were examined for the presence of polymorphisms. Clinical data were collected by anamnesis; all patients had the osseous densitometry examined using a lunar instrument to determine mineral osseous densitometry in the lumbar column (L2-L4). CYP17, COMT, and ESR1 genotyping was carried out by polymerase chain reaction with DNA collected from buccal swabs. The average age was 51.96 years. The average weights of the patients in control and osteopenia groups were 70.25 ± 12.00 and 62.45 ± 11.64, respectively (P = 0.001) and body mass index (P = 0.006; control: 29.43 ± 5.25; osteopenia: 26.72 ± 4.57). Related to CYP17 polymorphisms, 28.18% of women were TT (wild-type homozygous), 60% were TC (heterozygous), and 11.82% were CC (mutated homozygous). Related to COMT polymorphisms, 53.64% of women were GG (wild-type homozygous), 37.27% were GA (heterozygous), and 9.09% were AA (mutated homozygous). Related to ESR1, 53.64% of women were CC (wild-type homozygous), 40.91% were CT (heterozygous), and 5.45% were TT (mutated homozygous). The ESR1 variant allele was significantly higher in the osteopenia group when compared with women in the normal group (P = 0.02). ESR1 may be associated with low mineral osseous densitometry, while CYP17 and COMT gene polymorphisms were not associated with mineral osseous densitometry.

Do extracts of oral soybean augment the trophic effect of estrogen on the rat uterus?

OBJECTIVE: To evaluate whether soybean extracts and estrogens present additive effects on adult rat uterus. METHODS: Fifty ovariectomized rats were randomly divided into five equal groups of ten animals: Control, treated with vehicle; SE46 and SE120, treated with 46 and 120 mg/kg soybean concentrated extract (SE), respectively; EE, treated with conjugated equine estrogens (CE) 50 µg/kg; SE120 + EE, treated with 50 µg/kg (CE) plus 120 mg/kg SE. The substances were administered daily by gavage for 21 consecutive days. Thereafter the animals were weighed and killed by decapitation; trunk blood was collected for hormone determinations. Uteri were removed immediately and fixed in 10% formaldehyde, followed by dehydration, embedding in paraffin and 6-m sections staining with hematoxylin and eosin for histomorphometric analyses of myometrium and endometrium. After ANOVA analysis of the data, the study was complemented with the Tukey-Kramer test for multiple comparisons. RESULTS: The concentrated extract of soybean at high concentration (SE 120 kg/mg) and estrogens proved to have a trophic effect on the uterus (endometrium and myometrium) of castrated rats. In groups SE120, EE and SE120 + EE, all morphometric parameters examined (number of glands, eosinophils, blood vessels and the glandular area) were increased. No significant addictive effects of soybean extract plus estrogens were detected in the SE120 + EE group. CONCLUSIONS: Our results indicate that soy extract has a trophic effect on rat uterine structures. Treatment of ovariectomized rats with a concentrated soy extract in combination with conjugated estrogens had no addictive effect on the uterine response.

Multidisciplinary approach during menopausal transition and postmenopause in Brazilian women.

OBJECTIVE: To identify clinical, physical, life quality and nutritional aspects of Brazilian women during menopausal transition and postmenopausal periods. METHODS: 115 women agreed to participate in the study. They were divided into two groups: GI--menopausal transition (n = 48) and GII--postmenopause (n = 67). The Kupperman-Blatt Menopausal Index (IMK) and Women's Health Questionnaire (WHQ), Food Frequency Questionnaire and functional capacity were used. All patients were examined and underwent clinical and gynecological examination. RESULTS: There was no significant difference in IMK, WHQ and functional capacity in either group. There was a higher caloric intake, especially in sugars, in postmenopause women than in menopausal transition women. Both groups presented reduced parameters in life quality and functional capacity. CONCLUSION: Our data suggests that there is no significant difference between women in menopausal transition and postmenopause, except in relation to the nutritional parameter. In both groups, the women presented low quality of life and reduced functional capacity.

Effect of estrogen receptor-alpha (ESR1) gene polymorphism on high density lipoprotein levels in response to hormone replacement therapy

Studies have shown that estrogen replacement therapy and estrogen plus progestin replacement therapy alter serum levels of total, LDL and HDL cholesterol levels. However, HDL cholesterol levels in women vary considerably in response to hormone replacement therapy (HRT). A significant portion of the variability of these levels has been attributed to genetic factors. Therefore, we investigated the influence of estrogen receptor-alpha (ESR1) gene polymorphisms on HDL levels in response to postmenopausal HRT. We performed a prospective cohort study on 54 postmenopausal women who had not used HRT before the study and had no significant general medical illness. HRT consisted of conjugated equine estrogen and medroxyprogesterone acetate continuously for 1 year. The lipoprotein levels were measured from blood samples taken before the start of therapy and after 1 year of HRT. ESR1 polymorphism (MspI C>T, HaeIII C>T, PvuII C>T, and XbaI A>G) frequencies were assayed by restriction fragment length polymorphism. A general linear model was used to describe the relationships between HDL levels and genotypes after adjusting for age. A significant increase in HDL levels was observed after HRT (P = 0.029). Women with the ESR1 PvuII TT genotype showed a statistically significant increase in HDL levels after HRT (P = 0.032). No association was found between other ESR1 polymorphisms and HDL levels. According to our results, the ESR1 PvuII TT genotype was associated with increased levels of HDL after 1 year of HRT.

Effect of estrogen receptor-alpha (ESR1) gene polymorphism on high density lipoprotein levels in response to hormone replacement therapy.

Studies have shown that estrogen replacement therapy and estrogen plus progestin replacement therapy alter serum levels of total, LDL and HDL cholesterol levels. However, HDL cholesterol levels in women vary considerably in response to hormone replacement therapy (HRT). A significant portion of the variability of these levels has been attributed to genetic factors. Therefore, we investigated the influence of estrogen receptor-alpha (ESR1) gene polymorphisms on HDL levels in response to postmenopausal HRT. We performed a prospective cohort study on 54 postmenopausal women who had not used HRT before the study and had no significant general medical illness. HRT consisted of conjugated equine estrogen and medroxyprogesterone acetate continuously for 1 year. The lipoprotein levels were measured from blood samples taken before the start of therapy and after 1 year of HRT. ESR1 polymorphism (MspI C>T, HaeIII C>T, PvuII C>T, and XbaI A>G) frequencies were assayed by restriction fragment length polymorphism. A general linear model was used to describe the relationships between HDL levels and genotypes after adjusting for age. A significant increase in HDL levels was observed after HRT (P = 0.029). Women with the ESR1 PvuII TT genotype showed a statistically significant increase in HDL levels after HRT (P = 0.032). No association was found between other ESR1 polymorphisms and HDL levels. According to our results, the ESR1 PvuII TT genotype was associated with increased levels of HDL after 1 year of HRT.

CYP17 polymorphism and hot flushes in postmenopausal women.

OBJECTIVE: To investigate the association between the CYP17alpha gene polymorphism and hot flushes in postmenopausal women. METHODS: Ninety-three non-hysterectomized, postmenopausal women were enrolled in this study. Vasomotor symptoms were assessed at the baseline visit and based on information provided by each participant. The genotypic polymorphism of CYP17alpha gene was analyzed by PCR-RFLP assay using genomic DNA isolated from peripheral blood lymphocytes. RESULTS: Thirty-six women reported hot flushes of mild intensity, 25 reported hot flushes of moderate intensity and 32 of severe intensity. There was no significant difference between the severity of hot flushes and the CYP17 genotype or allele frequencies, 0.58 and 0.67 respectively. No association was found between hot flush severity and the CYP17 allele (odds ratio = 1.17, p = 0.61). CONCLUSION: The results of this study suggest that the CYP17 MspAI polymorphism was not significantly associated with an increased risk of reporting hot flushes.

Estrogen effects on the vaginal pH, flora and cytology in late postmenopause after a long period without hormone therapy.

In this report we evaluated the action of conjugated equine estrogens (CEE) on vaginal symptoms, cytology, pH, and flora in late postmenopausal women without any previous hormone therapy. The study was a randomized, double-blind, placebo-controlled trial with 48 late postmenopausal women who received placebo or unopposed CEE (0.625mg/day of CEE orally) during three months of treatment. Vaginal and sexual complaints were evaluated through daily diary cards. We analyzed vaginal changes through cytology and pH measurements. After three months of treatment, 20% of placebo-treated patients and 80% of the CEE-treated patients reported improvement in vaginal dryness and irritation. In the latter group, the vaginal cells and Lactobacillus increased and the vaginal pH decreased, without other changes in sexual complaints. We concluded that estrogen ameliorated the genital tract of late postmenopausal women without any previous hormone therapy.

The benefits of finasteride for hirsute women with polycystic ovary syndrome or idiopathic hirsutism.

Finasteride has been used frequently in the treatment of prostate hyperplasia, but this drug inhibits 5alpha-reductase and for this reason could be useful for the treatment of hirsutism. The aim of this study was to evaluate the clinical and hormonal effects of finasteride on hirsute women with idiopathic hirsutism or polycystic ovary syndrome. Twenty-four women were randomly divided into two groups: those given placebo and those given finasteride 5 mg/day. The treatment period was 6 months. All patients were evaluated before the beginning of treatment (baseline) and after 3 and 6 months of treatment using clinical examination through Ferriman-Gallwey score, blood pressure, cardiac frequency and body mass index. Also, we collected blood for hormonal determination of levels of prolactin, 17alpha-hydroxyprogesterone, follicle stimulating hormone, luteinizing hormone, total and free testosterone, dehydroepiandrosterone sulfate, androstenedione and dihydrotestosterone. Furthermore, all patients were asked about their concerns and satisfaction with the treatment. The results showed that the Ferriman-Gallwey score in the 6th month of finasteride treatment was significantly lower than at baseline and the 3rd month of this drug treatment. The dihydrotestosterone level in the finasteride group was also significantly reduced compared to that in the placebo group. The other hormones did not show any statistical difference during the study. All the patients treated with finasteride perceived a reduction in hirsutism after 6 months. In conclusion, our data suggest that finasteride may be effective for the treatment of the hirsute woman with idiopathic hirsutism or polycystic ovary syndrome.

A randomized, open-label study of conjugated equine estrogens plus medroxyprogesterone acetate versus tibolone: effects on symptom control, bleeding pattern, lipid profile and tolerability.

OBJECTIVE: To compare the effects of continuous combined conjugated equine estrogens plus medroxyprogesterone acetate (CEE/MPA) with those of tibolone on symptom control, bleeding pattern, lipid profile and tolerability in postmenopausal women. METHODS: This was a randomized, open-label, parallel-group, multicenter study. Generally healthy postmenopausal women with an intact uterus and no contraindications to hormone replacement therapy (HRT) or tibolone were enrolled. Each subject was randomly assigned to receive CEE/MPA 0.625 mg-5.0 mg or tibolone 2.5 mg daily for 13 treatment cycles, each of 28 days. RESULTS: A total of 85 subjects were enrolled and received at least one dose of study medication, of which 76 (89.4%) subjects completed the study (n = 40, CEE/MPA; n = 36, tibolone). The incidence of postmenopausal symptoms decreased significantly over time in both treatment groups, compared with baseline, including significant decreases in the incidence of urogenital and sexual health symptoms. Significant differences in symptom control (other than hot flushes) were observed between treatment groups in a few different cycles for different symptoms, but no consistent or clinically significant trends were observed. No statistically significant differences in the incidence of bleeding were observed between treatment groups after cycle 4. Significant decreases in total cholesterol (5.6%) and low-density lipoprotein (LDL) cholesterol (7.5%) were observed at cycle 13, compared with baseline, in the CEE/MPA group, and significant decreases in high-density lipoprotein (HDL) cholesterol (8.5%) and triglycerides (13.7%) were observed at cycle 13, compared with baseline, in the tibolone group. Significant weight gain was observed at cycle 13 in the tibolone group (3.05 kg), compared with the CEE/MPA group (0.96 kg). The incidences of adverse events were similar in both treatment groups. CONCLUSIONS: Women treated with CEE/MPA or tibolone showed significant improvement of postmenopausal symptoms, including urogenital and sexual health symptoms, and had similar bleeding patterns after four cycles of therapy. CEE/MPA and tibolone each induced a different mix of changes in the lipid profile.

Ultrastructural aspects of the postmenopausal endometrium after oral or transdermal estrogen administration.

In this report we examined the ultrastructural features of the postmenopausal endometrial cells of women treated with different doses of conjugated equine estrogen (CEE), or transdermal 17beta-estradiol. Eight women with uterine prolapse and at least 5 years of menopause were randomly divided into four groups and treated as follows: (I) no hormonal treatment; (II) 0.625mg/day of CEE orally; (III) 1.25mg/day of CEE orally; (IV) 50microg/day of 17beta-estradiol transdermally. Hormones were administered for 28 days followed by vaginal hysterectomy. Fragments of the endometrium were prepared for transmission electron microscopic analysis. We observed that the postmenopausal endometrium of the untreated group was atrophic with lined superficial epithelial cuboidal cells. The presence of gland and stroma cells with clear cytoplasm containing few organelles and heterochromatin nuclei were also observed. On the contrary, the endometrium of the group that received 0.625mg/day of CEE showed signs of proliferative cells such as the presence of numerous organelles in the cytoplasm and euchromatic nuclei. All of the proliferative effects on the endometrium were more pronounced in the groups that received 1.25mg/day of CEE and 50microg/day of transdermal 17beta-estradiol. We concluded that the ultrastructural proliferative changes of the postmenopausal endometrium induced by 1.25mg/day of CEE were similar to 50microg/day of transdermal 17beta-estradiol.

Estrogen, progestogen and tamoxifen increase synaptic density of the hippocampus of ovariectomized rats.

The effects of gonadal steroids or tamoxifen over the synaptic density of the CA1 region of the hippocampus was investigated in ovariectomized (OVX) rats. Chronic oral administration of conjugated equine estrogen, conjugated equine medroxyprogesterone, a combination of both or tamoxifen was performed in ovariectomized (OVX) rats over a period of 60 days. Synaptic density of the stratum radiatum of the CA1 region was evaluated by means of electron microscopy. Significant increases in the range of 34-49% were found for treated animals as compared to OVX controls not subject to hormonal replacement. Our results confirm previously reported effects of estradiol over synaptic density in this region and reports for the first time an effect of medroxyprogesterone (alone or in combination with estrogen) and tamoxifen. Our findings support the notion that hormonal replacement therapy and tamoxifen might have beneficial effects for cognitive function.

ATM gene deletion in patients with adult acute lymphoblastic leukemia.

BACKGROUND: Loss of heterozygosity (LOH) at the ATM gene (mutated in ataxia telangiectasia [AT] patients) and ATM protein deficiency occur in 14% and 34%, respectively, of patients with chronic lymphocytic leukemia (CLL). ATM protein deficiency also is associated with aggressive disease and worse patient survival. Considering the aberrations in the ATM gene in CLL and the high rate of incidence of lymphoid neoplasias in AT patients, the authors investigated its incidence rate and significance in patients with adult acute lymphoblastic leukemia (ALL). METHODS: Samples from 36 adults with ALL were analyzed for LOH and homozygous deletion (HD) using a panel of three microsatellite markers located at the ATM gene (D11S2179), the MLL gene (D11S1356), and the BCL1 gene (D11S987) loci. These markers are located within the 11q13-q23 locus. RESULTS: Of the 36 informative cases, 10 (28%) showed deletions (7 LOH and 3 HDs) at the D11S2179 marker. In two patients, the deletions were extended to the MLL gene locus. These deletions were submicroscopic because only 3% (1 of 36) of patients showed abnormalities involving 11q23 using cytogenetic studies. The authors also estimated the levels of ATM protein in 15 ALL patients and 12 healthy volunteers by radioimmunoassay. The ATM protein levels in cases with LOH at the ATM gene were between 15-50% of those from normal bone marrow. In contrast to CLL patients, patients with LOH or HD at the ATM gene locus showed better survival compared with patients without ATM gene deletions (P = 0.003). CONCLUSIONS: LOH of the ATM gene and protein deficiency are common in adult ALL, are not demonstrated at the cytogenetic level, and are associated with a favorable prognosis. The authors speculate that ATM deficiency may increase the sensitivity of leukemic blasts to the chemotherapy used during induction and after disease remission in patients with adult ALL. The relatively high frequency of deletion of the D11S2179 marker compared with the D11S1356 marker suggests that ATM is the target gene of the deletion at the 11q23 locus, and that such deletions may play a role in the pathogenesis of ALL.

Downregulation of the p53 tumor suppressor gene and upregulation of the bcl-2 gene in retinoic acid receptor alpha-deficient transgenic mice.

We recently demonstrated lymphoma development in transgenic mice deficient in retinoic acid receptor alpha (RARalpha). High incidence of lymphoma development in this transgenic mouse model system was similar to lymphoma development in p53 knockout mice. In an effort to understand the molecular basis of lymphomagenesis in RARalpha-deficient transgenic mice, we compared the levels of RARalpha to the levels of p53 mRNA, and Bcl-2, and Bax proteins in lymphoid and non-lymphoid tissues and in lymphomas derived from the RARalpha-deficient transgenic mice. The p53 mRNA levels were depleted in various tissues including spleen ( approximately 96%), thymus ( approximately 29%) and bone marrow ( approximately 62%) of RARalpha-deficient transgenic mice when compared with the normal littermates, and the reduction in p53 mRNA expression in the various tissues examined was proportional to the reduction in RARalpha expression. Bcl-2 to Bax ratios were highly increased in the lymphoid compartments (spleen >bone marrow >thymus) because of selective overexpression of Bcl-2 protein. In summary, RARalpha downmodulation in this transgenic mouse model system was accompanied by p53 downmodulation and deregulation of Bcl-2 to Bax ratios in the lymphoid compartments.

Ovarian activity before and after gonadal suppression by GnRH-a in patients with polycystic ovary syndrome, hyperandrogenism, hyperinsulinism and acanthosis nigricans.

OBJECTIVE: To investigate the ovarian activity before and after gonadal suppression with GnRH-analog in patients with PCO, hyperandrogenism, hyperinsulinism and acanthosis nigricans. DESIGN: Controlled clinical study. SETTING: Tertiary academic medical center. PATIENTS: Six patients with clinical findings of PCO, hirsutism and acanthosis nigricans. INTERVENTIONS: Morning blood samples in the follicular phase to determine the steroid levels, glucose and insulin curve, comparing to a control group. Administration for 2 consecutive months of a GnRH-analog, comparing, in the study group, the free testosterone levels before and after ovarian suppression. MAIN OUTCOME MEASURE: Determination of insulin levels in PCO, hirsutism and acanthotic patients and the free-testosterone levels before and after gonadal suppression. RESULTS: Insulin levels were significantly higher in the study group when compared to normal women during the glycemic test. We also found a significant decrease in the free-testosterone levels after 2 months of gonadal suppression with GnRH-analog when compared to the initial time. CONCLUSIONS: Patients with PCO, hirsutism and acanthosis nigricans present high levels of insulin, suggesting an ovarian hyperesponsiveness, which is not sustained when gonadotrophic blockage was achieved.

Ovarian activity before and after gonadal suppression by GnRH-a in patients with polycystic ovary syndrome, hyperandrogenism, hyperinsulinism and acanthosis nigricans

Objective. To investigate the ovarian activity before and after gonadal suppression with GnRH-analog in patients with PCO, hyperandrogenism, hyperinsulinism and ancathosis nigricans. Design: Controlled clinical study. Setting: Tertiary academic medical center. Patients: Six patients with clinical findings of PCO, hirsutism and acanthosis nigricans. Interventions. Morning blood samples in the follicular phase to determine the seteroid levels, glucose and insulin curve, comparing to a control group. Administration for 2 consecutive months of a GnRH-analog, comparing, in the study group, the free testosterone levels before and after ovarian suppression. Main Outcome Measure. Determination of insulin levels in PCO, hirsutism and acanthotic patients and the free-testosterone levels before and after gonadal suppression. Results. Insulin levels were significantly higher in the study group when compared to normal women during the glycemic test. We also found a significant decrease in the free-testosterone levels after 2 months of gonadal suppression with GnRH-analog when compared to the initial time. Conclusions. Patients with PCO, hirsutism and acanthosis nicrigans present high levels of in sulin, suggesting an ovarian hyperesponsiveness, which is not sustained when gonadotrophic blockage was achieved.

Morphologic and morphometric aspects of the endometrium of postmenopausal women before and after cyclic oestrogen replacement treatment.

OBJECTIVE: The aim of the study is to observe the morphology and morphometry of the endometrium of postmenopausal women treated with cyclic conjugated oestrogens. STUDY DESIGN: Three groups of nine postmenopausal women received cyclic conjugated oestrogens for 21 days (with a seven-day pause) during six months. The endometrial specimens were obtained using a modified Novak suction curet, in the second or third day of the period of drug washout. The slides were stained with haematoxylin and eosin (H.E.) in order to measure epithelial height and determine the gland/stroma ratio. RESULTS: Morphologic examination showed that single daily doses of 0.3 mg of conjugated oestrogens caused discrete endometrial proliferation after three and six months of treatment. However, a more intense effect was observed in women receiving doses of 0.625 and 1.25 mg/day of the hormone, in the same period. Morphometric study revealed significant increases both in epithelial thickness and in the gland-stroma ratio, specially in women receiving higher doses of the conjugated oestrogen (0.625 and 1.25 mg/day). CONCLUSIONS: We concluded that there were marked proliferative alterations without atypias in the endometrium of women that received 0.625 and 1.25 mg of conjugated oestrogens during six months.

Electron microscopic localization of lacZ expression in the proximal convoluted tubular cells of the kidney in transgenic mice carrying chimeric erythropoietin/lacZ gene constructs.

Regulated expression of the erythropoietin (EPO) gene in the adult kidney plays a key role in the regulation of erythropoiesis. However, uncertainty exists regarding the type of kidney cells involved in EPO gene expression. We previously showed by light microscopy that the lacZ reporter gene is expressed and inducible by hypoxia/anemia in the proximal convoluted tubular (PCT) cells of the kidneys of transgenic mice carrying the 5'-lacZ construct, in which the lacZ gene was placed downstream of a 7.0-kb mouse EPO gene segment containing 6.5 kb of the 5'-flanking sequence. We, report here the light and transmission electron microscopic examination of lacZ expression in the kidneys of transgenic mice carrying the 5'-lacZ construct and two additional constructs carrying the 6.5-kb 5'-flanking sequence with the body of the gene alone, or along with the 1.2-kb 3'-flanking sequence. The electron microscopic analyses unequivocally demonstrated that lacZ under the regulatory control of the 6.5-kb 5'-flanking sequence with or without the body of the gene and the 1.2-kb 3'-flanking sequence was expressed predominantly in the proximal convoluted tubular cells of the kidney following hypoxia induction.

The ACTH test in the diagnosis of hirsutism.

The ACTH test has been used to confirm the diagnosis of adrenal insufficiency and the classic and the non-classic adrenal hyperplasia due to the 3-HSD, 21 OH e 110H deficiencies. This article reviews the historical aspects of the use of ACTH in the diagnosis of hirsutism and points out its mains indications. In spite of new biological molecular advances in the diagnosis of adrenal enzymatic deficiencies, the use of the ACTH test can help the physician to predict both genothipus and fenothipus in populations with hyperandrogenic manifestations due to non-classical or late-onset congenital adrenal hyperplasia.

Expression profile of MDM-2 proteins in chronic lymphocytic leukemia and their clinical relevance.

The MDM-2 oncoprotein exists in an autoregulatory feedback loop with the tumor suppressor protein p53. Therefore, intracellular levels of these two proteins may play important roles in cell proliferation and tumorigenesis. Several MDM-2 proteins (Mr 35-100 Kd) have been demonstrated in human cell lines. We report here the expression profile of MDM-2 and p53 proteins in 87 cases of chronic lymphocytic leukemia (CLL) as detected by immunoblot analysis. The MDM-2 proteins (p57, p59, p67, and p90) were found to be overexpressed in different combinations in 56/87 (64%) of cases of CLL when compared with normal volunteers. The MDM-2 protein p57 was predominantly overexpressed 46/87 (53%) in CLL. In 22/87 (25%) cases of CLL p57 was overexpressed alone, and in 24/87 (28%) cases it was co-overexpressed with other MDM-2 proteins p59/p67/p90. Six of the 87 cases of CLL showed overexpression of the tumor suppressor protein p53 by immunoblot analysis, and five of those cases also co-overexpress MDM-2 protein p57. No statistically significant correlation of MDM-2 protein overexpression to clinical disease stage and history of previous chemotherapy of CLL patients has been found. However, considering the oncogenic potential of overexpressed MDM-2 proteins, a possible role of MDM-2 proteins in the promotion of CLL disease remains to be evaluated.

Differential expression of lacZ in the liver and kidney of transgenic mice carrying chimeric lacZ-erythropoietin gene constructs with or without its 1.2 kb 3'-flanking sequence.

Erythropoietin (EPO) plays a key role in erythropoiesis and is expressed predominantly in the fetal liver and in the adult kidney. The EPO gene is up-regulated at the transcriptional level under hypoxic/anemic conditions. We studied the role of the 5'- and 3'-flanking sequences of the mouse EPO gene in its tissue-specific and hypoxia-induced expression by developing transgenic mouse lines carrying chimeric EPO-lacZ gene constructs. Transgenic mice carrying a 6.5 kb segment of the 5'-sequence and most of the EPO gene in which lacZ was substituted for exon 1 (5'-lacZ-EPO) demonstrated induction of lacZ expression following hypoxia/ anemia induction in both the liver and kidney of adult mice. However, transgenic mice carrying the above construct along with the 1.2 kb 3'-flanking sequence (5'-lacZ-EPO-3') showed a high level of lacZ expression following hypoxia/anemia induction in adult kidney but not in adult liver. With the aim of further understanding the role of the 3'-flanking sequence in tissue-specific expression of the EPO gene, we studied the interactions of protein factors with this 1.2 kb 3' region and demonstrated that multiple sets of protein factors interact tissue specifically with a 10 bp sequence, TCAAAGATGG, located downstream of the previously characterized 3' hypoxia-responsive enhancer element.