Decreased efficacy of twice-weekly intravaginal dehydroepiandrosterone on vulvovaginal atrophy.

OBJECTIVE: While daily intravaginal administration of 0.50% (6.5 mg) dehydroepiandrosterone (DHEA, prasterone) for 12 weeks has shown clinically and statistically significant effects on moderate to severe (MS) dyspareunia as the most bothersome symptom (MBS), the present study analyzes the effect of a reduced dosing regimen on MBS vaginal dryness. METHOD: Daily intravaginal 0.50% prasterone for 2 weeks followed by twice weekly for 10 weeks versus placebo. RESULTS: Maximal beneficial changes in vaginal parabasal and superficial cells and pH were observed at 2 weeks as observed for intravaginal 10 µg estradiol (E2). This was followed by a decrease or lack of efficacy improvement after switching to twice-weekly dosing. The decrease in percentage of parabasal cells, increase in percentage of superficial cells and decrease in vaginal pH were all highly significant (p < 0.0001 to 0.0002 over placebo) at 12 weeks. In parallel, the statistical significance over placebo (p value) on MBS vaginal dryness at 6 weeks was 0.09 followed by an increase to 0.198 at 12 weeks. For MBS dyspareunia, the p value of 0.008 at 6 weeks was followed by a p value of 0.077 at 12 weeks, thus illustrating a decrease of efficacy at the lower dosing regimen. The improvements of vaginal secretions, color, epithelial integrity and epithelial surface thickness were observed at a p value < 0.01 or 0.05 over placebo at 2 weeks, with a similar or loss of statistical difference compared to placebo at later time intervals. No significant adverse event was observed. Vaginal discharge related to the melting of Witepsol was reported in 1.8% of subjects. CONCLUSION: The present data show that daily dosing with 0.50% DHEA for 2 weeks followed by twice-weekly dosing is a suboptimal treatment of the symptoms/signs of vulvovaginal atrophy resulting from a substantial loss of the efficacy achieved at daily dosing.

DHEA and intracrinology at menopause, a positive choice for evolution of the human species.

Menopause has been chosen by evolution as the convergence of three factors, namely cessation of ovarian function (reproduction and estrogen secretion), high circulating dehydroepiandrosterone (DHEA), and intracrine enzymes able to convert DHEA into active sex steroids in peripheral tissues. The arrest of estrogen secretion by the ovaries at menopause causes a decrease of circulating estradiol below the threshold of biological activity, thus eliminating stimulation of the endometrium and risk of endometrial cancer. As much as the arrest of secretion of estradiol by the ovaries is essential to protect the uterus, it is of major importance that sex steroids continue to be made available in most other tissues which need estrogens and/or androgens for their normal functioning. Evolution, through 500 million years, has progressively provided the peripheral tissues with the enzymes able to make androgens and estrogens while high levels of DHEA, the precursor of all sex steroids, have appeared much later with the primates approximately 20 million years ago. All elements were thus in place for the functioning of intracrinology or the cell-specific formation of estrogens and androgens in peripheral tissues from the inactive precursor DHEA, with no significant release of active sex steroids in the circulation, thus eliminating the risks of adverse effects in the other tissues, especially the uterus. The presence of subthreshold levels of circulating estradiol combined with the formation of sex steroids from DHEA in specific peripheral tissues (intracrinology) makes menopause a positive characteristic supporting many years of good-quality postmenopausal life, useful for taking care of children and grandchildren. DHEA, however, decreases with age and is present at very different concentrations between different women, with the consequence that approximately 75% of postmenopausal women have too low circulating DHEA levels and suffer from symptoms/signs of hormone deficiency.

Intravaginal dehydroepiandrosterone (prasterone), a highly efficient treatment of dyspareunia.

OBJECTIVE: To examine the effect of intravaginal dehydroepiandrosterone (DHEA) on pain at sexual activity (dyspareunia) identified as the most bothersome symptom of vaginal atrophy in postmenopausal women at both screening and day 1. METHODS: This prospective, randomized, double-blind and placebo-controlled phase III clinical trial studied the effect of prasterone (DHEA) applied locally in the vagina on the severity of dyspareunia in 114 postmenopausal women who had identified dyspareunia as their most bothersome symptom of vaginal atrophy, while meeting the criteria for superficial cells ≤ 5% and pH > 5.0 at both screening and day 1. RESULTS: At the standard duration of 12 weeks of treatment, increasing doses of 0.25%, 0.5% and 1.0% DHEA decreased the percentage of parabasal cells by 48.6  ±â€Š 6.78%, 42.4  ±  7.36% and 54.9  ±â€Š 6.60% (p < 0.0001 vs. placebo for all) with no change with placebo (p = 0.769). The effects on superficial cells and pH were also highly significant compared to placebo at all DHEA doses. The severity score of pain at sexual activity decreased by 0.5, 1.4, 1.6 and 1.4 units in the placebo and 0.25%, 0.5% and 1.0% DHEA groups, respectively, with the p value of differences from placebo ranging from 0.0017 to < 0.0001. CONCLUSIONS: Intravaginal DHEA, through local estrogen and androgen formation, causes a rapid and highly efficient effect on pain at sexual activity without systemic exposure of the other tissues, thus avoiding the recently reported systemic effects of estrogens.

Chronic administration of dehydroepiandrosterone (DHEA) to female monkey and rat has no effect on mammary gland histology.

Dehydroepiandrosterone (DHEA), the major steroid precursor of androgens and estrogens produced in peripheral tissues in primates, has been shown to exert chemopreventive effect on the development of carcinogen-induced rat mammary tumors. Since little is known on the effect of DHEA administration on mammary gland physiology and histology, we have studied the effect of long-term administration of DHEA to normal female monkey and rat on mammary gland histology as well as on serum DHEA, DHEA sulphate (DHEA-S), testosterone and estradiol levels. In monkeys, DHEA treatment (2 or 10 mg/(kg b.w.day)) induced a dose-related increase in serum DHEA and DHEA-S (above 20-fold) levels. At the highest dose of DHEA, serum testosterone levels were significantly increased (three- to fourfold), while serum estradiol concentration was not modified. DHEA treatment did not modify the histological characteristics of monkey mammary glands. In the rat, following DHEA administration (10 or 100 mg/(kg b.w.day)), a dose-related marked increase in serum DHEA and DHEA-S was observed. Serum testosterone was also increased in DHEA-treated animals, while no significant changes in serum estradiol levels were detected. As in the monkey, the histology of the female rat mammary gland remained unchanged following long-term treatment with any of the two doses of DHEA.

Pangenomic changes induced by DHEA in the skin of postmenopausal women.

The objective of this study was to explore, for the first time, the changes in the pangenomic profile induced in human skin in women treated with dehydroepiandrosterone (DHEA) applied locally. Sixty postmenopausal women participated in this phase II prospective, randomized, double-blind and placebo-controlled study. Women were randomized to the twice daily local application of 0% (placebo), 0.3%, 1% or 2% DHEA cream. Changes in the pangenomic expression profile were studied using Affymetrix Genechips. Significant changes (p<0.05) in sixty-six DHEA-responsive probe sets corresponding to 52 well-characterized genes and 9 unknown gene sequences were identified. A dose-dependent increase in the expression of several members of the collagen family was observed, namely COL1, COL3 and COL5 as well as the concomitant modulation of SPARC, a gene required for the normal deposition and maturation of collagen fibrils in the dermis. Several genes involved in the proliferation and differentiation of keratinocytes were also modulated. In addition, topical DHEA reduced the expression of genes associated with the terminal differentiation and cornification of keratinocytes. Our results strongly suggest the possibility that DHEA could exert an anti-aging effect in the skin through stimulation of collagen biosynthesis, improved structural organization of the dermis while modulating keratinocyte metabolism.

Bioavailability and metabolism of oral and percutaneous dehydroepiandrosterone in postmenopausal women.

To study the bioavailability of dehydroepiandrosterone (DHEA) administered by the oral and percutaneous routes, three groups of 12 postmenopausal women aged 60-70 years received two capsules of 50mg of DHEA orally before breakfast daily for 14 days or applied 4 g of a 10% DHEA cream or gel at the same time of the day on a 30 cm x 30 cm surface area on the thighs. Detailed serial blood sampling over 24h was performed following 1st and 14th DHEA administration for measurement of DHEA and nine of its metabolites by liquid chromatography tandem mass spectrometry (LC-MS/MS) or gas chromatography mass spectrometry (GC-MS). Serum levels of estrone (E1) and estradiol (E2) did not change following DHEA administration by any of the three formulations, while serum androstenedione (4-dione), testosterone, DHEA sulfate (DHEA-S), E(1)-S, androsterone glucuronide (ADT-G) and 3alpha-androstanediol-G (3alpha-diol-G), increased in all cases, the effect on these parameters being more important after oral than percutaneous administration due to the metabolism of DHEA into these metabolites in the gastrointestinal tract and liver. No qualitative differences in DHEA metabolism are observed between the oral and percutaneous routes of DHEA administration while the levels of all steroids remain on a plateau during the 24h period during chronic percutaneous DHEA administration. The present data show that DHEA is transformed into active androgens and estrogens in peripheral intracrine tissues with no or minimal release of the active steroids E(1), E(2) or testosterone in the circulation. Moreover, DHEA is preferentially transformed into androgens rather than into estrogens. Most importantly, the present data show that changes in serum DHEA following oral or percutaneous DHEA administration are not a valid parameter of DHEA action since the increase in serum DHEA is at least 100% greater than the increase in the formation of active androgens and estrogens and thus much higher than the potential physiological effects.

A complex promoter element mediates transactivation of the human proliferating cell nuclear antigen promoter by the 243-residue adenovirus E1A oncoprotein.

The adenovirus E1A oncoproteins interfere with the normal regulation of cellular proliferation through interactions with cell cycle regulatory proteins. In view of the essential role of proliferating-cell nuclear antigen (PCNA) in DNA replication, we performed a mutational analysis of the minimal human PCNA promoter (nucleotides -87 to +62) to define sequence elements which mediate transactivation by the 243-residue E1A protein (E1A 243R). Linker-scanning and site-directed mutants were examined for basal and E1A-induced expression of chloramphenicol acetyltransferase (CAT) from PCNA promoter-CAT reporter constructs transiently expressed in HeLa cells. The results define the cis-acting element required for induction of PCNA by E1A 243R as a region between -59 and -45 relative to the transcription initiation site. This PCNA E1A-responsive element (PERE), which is protected from DNase I digestion by nuclear extracts from 293 cells, includes the sequence AGCGTGG immediately upstream of the ATF binding site previously shown to be important for activation of PCNA by E1A 243R (G. F. Morris and M. B. Mathews, J. Virol. 65:6397-6406, 1991). Mutation of either the upstream component or the ATF site within the PERE diminishes basal promoter activity and abrogates transactivation by E1A 243R. This novel cis-acting element is also essential for both basal and E1A-induced expression in the context of the full-length PCNA promoter.

Modulation of transcriptional activation of the proliferating cell nuclear antigen promoter by the adenovirus E1A 243-residue oncoprotein depends on proximal activators.

Previous analyses defined a proliferating cell nuclear antigen (PCNA) E1A-responsive element (PERE) in the PCNA promoter that is essential for transactivation by the 243-residue product of the adenovirus type 2 E1A 12S mRNA (E1A 243R). In this report, we show that the PERE activates a heterologous basal promoter and confers susceptibility to transactivation by E1A 243R, indicating that the PERE is both necessary and sufficient for the response of the PCNA promoter to this oncoprotein. Insertion of linker sequences between the PERE and the site of transcription initiation in the PCNA promoter severely impairs the promoter's response to E1A 243R transactivation. GAL4 sites can replace the function of the PERE in the E1A 243R response of the PCNA basal promoter if transcriptional activators of suitable strength are supplied as GAL4 fusion proteins. Weak transcriptional activators render the PCNA basal promoter subject to transactivation by E1A 243R but do not endow the adenovirus E1B basal promoter with a similar response. Strong transcriptional activators do not support transactivation by E1A 243R, however; instead, E1A reduces the ability of the strong activators to activate both the PCNA and E1B basal promoters. Although other mechanistic differences might determine the response, the data imply a relationship between the activation strength of promoter-proximal effectors and the response of the PCNA basal promoter to E1A 243R. These experiments indicate that the PERE can function autonomously in mediating transactivation by E1A 243R and that the PCNA basal promoter is configured in a manner that permits modulation by E1A 243R of transcriptional activation by promoter-proximal effectors.

Effect of dehydroepiandrosterone and the antiestrogen EM-800 on growth of human ZR-75-1 breast cancer xenografts.

BACKGROUND: In the mammary gland, androgens are formed from the precursor steroid dehydroepiandrosterone (DHEA). Clinical evidence indicates that androgens have inhibitory effects on breast cancer. Estrogens, on the other hand, stimulate the development and growth of breast cancer. We studied the effect of DHEA alone or in combination with the newly described pure antiestrogen EM-800 on the growth of subcutaneous tumor xenografts formed by the human breast cancer cell line ZR-75-1 in ovariectomized nude mice. METHODS: Immediately after ovariectomy, mice received daily subcutaneous injections of 0.5 microg estrone (E1) (an estrogenic hormone). EM-800 (15, 50, or 100 microg) was given orally once daily. DHEA was administered percutaneously twice daily (total dose of 0.3, 1.0, or 3.0 mg) to the dorsal skin either alone or in combination with a 15-microg daily oral dose of EM-800. Changes in tumor size in response to the treatments (in relation to measurements made on the first day of treatment) were assessed periodically. At the end of the experiments, tumors were dissected and weighed. RESULTS: A 9.4-fold increase in tumor size in 9.5 months was observed in ovariectomized mice receiving E1 alone. Administration of 15, 50, or 100 microg EM-800 in E1-supplemented mice led to inhibitions of 87.5%, 93.5%, and 94.0% in tumor size, respectively. DHEA, on the other hand, at doses of 0.3, 1.0, or 3.0 mg inhibited terminal tumor size by 50.4%, 76.8%, and 80.0%, respectively. Comparable inhibitions in tumor size were obtained with a daily 15-microg oral dose of EM-800 with or without different doses of percutaneous DHEA. CONCLUSIONS: DHEA and EM-800 independently suppressed the growth of E1-stimulated ZR-75-1 xenograft tumors in nude mice. Administration of DHEA at the defined doses did not alter the inhibitory effect of EM-800.

A p18 mutant defective in CDK6 binding in human breast cancer cells.

Progression from G1 to the S-phase of the cell cycle is controlled by a family of low molecular weight cyclin-dependent kinase (CDK) inhibitors. The importance of these proteins in cell growth control is underscored by the observation that some members of this family are deleted or mutated in human cancers. For example, the gene encoding the CDK inhibitor p18 is located on a segment of chromosome 1 that is often abnormal in human breast tumors. We have identified an alanine to proline substitution at codon 72 of the p18 gene in BT-20 human breast cancer cells. This mutation abrogates the ability of p18 to interact with CDK6 and renders p18 deficient in suppressing cell growth in a colony formation assay. Our results suggest that p18 inactivation by point mutations may contribute to deregulated growth control in certain cell lines and/or tumors.

Comparison of the effects of the antiestrogens EM-800 and tamoxifen on the growth of human breast ZR-75-1 cancer xenografts in nude mice.

Although estrone supplementation in ovariectomized (OVX) nude mice bearing ZR-75-1 xenografts caused a 365% increase in average tumor size during the 4-month treatment period, administration of the antiestrogen EM-800 at the daily oral doses of 50, 150, or 400 microg completely prevented estrogen-stimulated tumor growth. At the same doses of tamoxifen, tumor size was inhibited to 189, 117, and 120% above pretreatment values. However, when EM-800 (150 microg/day) was added to the daily 150- and 400-microg doses of tamoxifen, final tumor size was decreased further to 12 and 38% above pretreatment values, respectively. EM-800 (400 microg daily) administered to estrone-supplemented OVX mice caused complete, partial, and stable responses in 11, 22, and 49% of estrone-stimulated tumors, respectively, whereas 19% (7 of 37) progressed. At the same dose of tamoxifen, the corresponding responses were 3% (complete response), 3% (partial response), and 25% (no change), whereas 69% (22 of 32) of tumors progressed. In the absence of estrone supplementation, tamoxifen (400 microg) alone administered to OVX mice stimulated tumor growth to 161% compared with initial size whereas the same dose of EM-800 reduced tumor size by 55%, a value superimposable to that observed in OVX control animals. The agonistic effect of tamoxifen is thus illustrated by the observation that 73% of tumors progressed when tamoxifen was administered alone to OVX animals whereas no tumor progressed with EM-800. The present data strongly suggest that at least part of the initial lack of response and resistance to tamoxifen during tamoxifen treatment in women is due to the estrogenic activity of this compound, whereas the new antiestrogen EM-800 exerts pure antagonistic action.

Effect of combined treatment with the pure antiestrogen EM-800 and radiotherapy on the growth of human ZR-75-1 breast cancer xenografts in nude mice.

Human breast tumors are usually composed of heterogeneous cell populations that exhibit different sensitivities to therapeutic agents. We therefore investigated the effect of treatment with various regimens of the novel pure antiestrogen EM-800, alone or in combination with external beam radiation therapy, on the growth of human ZR-75-1 xenografts in athymic mice. The animals received a maximal dose of EM-800 (300 microg, p.o.) and/or radiotherapy at the dose of 10 Gy. 2.5 Gy fractions were administered over a 9-day period in four sessions of 13.7 min each (250-kilovolt Siemens with 2-mm aluminum filtration at 90 cm from the source origin). EM-800 was administered p.o. once daily, whereas radiotherapy was repeated every 35 days. Tumor size was expressed as a percentage of the initial tumor size, which was assigned a value of 100%. Average tumor size increased by 514% in ovariectomized mice supplemented with estrone alone for 259 days compared with the pretreatment value. Treatment with radiotherapy or EM-800 alone resulted in 11 and 73% decreases in mean tumor size, respectively, whereas combined treatment given simultaneously at the beginning caused a dramatic 98% decrease in tumor size. The start of radiotherapy on day 35 in EM-800-treated mice, or conversely, the start of EM-800 in irradiated mice at the 35-day time interval, resulted in somewhat lower, 88% and 95%, decreases in tumor size, respectively. In animals receiving EM-800 alone, 40% of tumors disappeared, thus indicating a cytotoxic effect caused by the estrogen blockade achieved with the pure antiestrogen. Eighty-six % of the original tumors disappeared under continuous combined treatment. Most importantly, no tumor reappeared under estrogenic stimulation after stopping treatment, thus indicating cure of 86% of the tumors in the group of animals who received the combination therapy. The present data indicate that combined treatment with EM-800 and radiotherapy yields a faster response, a greater decrease in tumor size, and a higher percentage of complete responses or tumor disappearance (cure) than either treatment used alone. The present data also suggest that maximal benefits are achieved when the pure antiestrogen is administered continuously, starting at the same time as radiation therapy and continued without interruption as adjuvant therapy. The present data also clearly show that efficient blockade of estrogens with a potent and pure antiestrogen is not only cytostatic but is cytotoxic and can lead to the disappearance of an important proportion of tumors or cure.

Effects of the antiestrogen EM-800 (SCH 57050) and cyclophosphamide alone and in combination on growth of human ZR-75-1 breast cancer xenografts in nude mice.

Human breast cancer proliferates as heterogeneous cell populations that exhibit different sensitivities to therapeutic agents. A logical approach to control these different cancer cell populations is the use of combined treatment with agents that block cell proliferation or induce apoptosis via different mechanisms. We therefore investigated the effect of treatment with the novel pure antiestrogen EM-800, alone or in combination with chemotherapy, on the growth of ZR-75-1 human breast tumors in nude mice, a well-recognized model of human breast cancer. Mice bearing estrone-releasing silastic implants as estrogenic stimulus received EM-800 or cyclophosphamide alone or in combination for 227 days. Cyclophosphamide (256 mg/kg/2 weeks) was administered by i.p. injection in 64 mg/kg fractions over 4 consecutive days with repetition of the cycle every 14 days. EM-800 was administered p.o. once daily at the maximally effective dose of 300 microg/mouse. After 227 days of treatment, average tumor size in mice receiving estrone alone was 192% higher than pretreatment. The average tumor size of mice treated with chemotherapy was reduced by 47%, whereas on the other hand, EM-800 caused a 81% decrease of the value of the same parameter. The combined treatment (EM-800 + cyclophosphamide), on the other hand, resulted in a 95% decrease in tumor size compared with control estrogen alone. In fact, EM-800 alone decreased tumor size to 55% of the value at the start of treatment, whereas the addition of cyclophosphamide to the antiestrogen further decreased tumor size to as low as 15% of the pretreatment value. The combination of EM-800 and cyclophosphamide resulted in 95% of complete or partial responses compared with 61 and 27% with EM-800 and cyclophosphamide alone, respectively. In fact, in the combination therapy group, only one tumor remained stable, while 17 regressed >50% and four disappeared. It is noteworthy that no tumor progressed with EM-800 alone or in combination with cyclophosphamide. The present data show, for the first time, that the addition of cyclophosphamide to a pure antiestrogen used at a maximal dose causes a more potent inhibition of human breast tumor growth, thus suggesting that combined treatment using a maximal dose of a pure antiestrogen and a chemotherapeutic agent(s), two classes of compounds having different mechanisms of action, could further improve breast cancer therapy above the results achieved with a potent and pure antiestrogen alone in estrogen-sensitive breast cancer.

Novel compounds inhibit estrogen formation and action.

Estrogens are well known to play a predominant role in human breast cancer. The current endocrine therapy of breast cancer consists in administering an antiestrogen which blocks the action of estrogens at the receptor level. However, the currently available antiestrogens possess mixed estrogenic and antiestrogenic activity, thus limiting their potential therapeutic efficacy. The present data show that a series of new estrogen derivatives demonstrate not only pure antiestrogenic activity in the sensitive in vivo mouse uterus assay, but simultaneously exert potent inhibitory effects on 17 beta-hydroxysteroid dehydrogenase activity, the enzyme responsible for the formation of 17 beta-estradiol from estrone, the last step in estrogen formation. Such compounds having a dual site of inhibitory action, namely on estrogen formation and on the estrogen receptor, could well lead to an improved endocrine therapy of breast and other estrogen-sensitive cancers as well as other nonmalignant estrogen-sensitive diseases.

Blockade of the stimulatory effect of estrogens, OH-tamoxifen, OH-toremifene, droloxifene, and raloxifene on alkaline phosphatase activity by the antiestrogen EM-800 in human endometrial adenocarcinoma Ishikawa cells.

Although temporary benefits of tamoxifen therapy are observed in up to 40% of women with breast cancer, this compound, which is known to possess mixed estrogenic and antiestrogenic activities, has been associated with increased risk of endometrial carcinoma. This study compares the effects of the novel nonsteroidal pure antiestrogen EM-800 and related compounds with those of a series of antiestrogens on the estrogen-sensitive alkaline phosphatase (AP) activity in human endometrial adenocarcinoma Ishikawa cells. Exposure to increasing concentrations of up to 1000 nM EM-800 or its active metabolite EM-652 alone failed to affect basal AP activity. In contrast, incubation with 10 nM (Z)-4-OH-tamoxifen, (Z)-4-OH-toremifene, droloxifene, or raloxifene increased the value of this estrogen-sensitive parameter by 3.3-, 3.5-, 2.2-, and 1.6-fold, respectively, a stimulatory effect that was completely reversed by simultaneous exposure to 30 nM EM-800. Moreover, the stimulation of AP activity induced by 1 nM 17beta-estradiol was completely reversed by EM-800, EM-652, or ICI-182780, at the IC50 value of 1.98 +/- 0.23, 1.01 +/- 0.16, and 5.64 +/- 0.59 nM, respectively, whereas the partial blockade exerted by (Z)-4-OH-tamoxifen, (Z)-4-OH-toremifene, or raloxifene was observed at IC50 values of 13.5 +/- 3.80, 41.0 +/- 7.2, and 3.74 +/- 0.43 nM, respectively. Thus, as assessed by their activity in the human Ishikawa endometrial carcinoma cells, EM-800 and EM-652 are the most potent known antiestrogens in Ishikawa cells, and, most importantly, they are devoid of the estrogenic activity observed in these human endometrial cancer cells with (Z)-4-OH-tamoxifen, (Z)-4-OH-toremifene, droloxifene, and raloxifene.

Resting and exercise hyperemic pulsatile arterial blood flow in insulin-dependent diabetic subjects.

Pulsatile arterial blood flow was studied in 20 normal (N), 20 short-term (STIDDM; mean: 5.17 yr), and 20 long-term insulin-dependent diabetic patients (LTIDDM; mean: 14.76 yr) between the ages of 18 and 30 yr with no clinically detectable peripheral vascular disease. Measurements were taken from waveforms obtained noninvasively using an electromagnetic flowmeter at rest and immediately after a 3-min isometric exercise challenge of the right leg. At rest, both groups of diabetics exhibited minute flow values similar to those in the normal group. This was achieved, however, by increased vasodilation in peripheral tissues as indicated by a difference in waveform configuration. Diabetic subjects showed a significantly smaller peak flow, a less steep ascending and descending slope, and a higher minute heart rate than normal controls. After 3 min of isometric exercise, the diabetic groups exhibited significantly less minute flow, flow/pulse, and a more vasodilated flow pattern similar to that recorded at rest. In addition, the LTIDDM group showed significantly less arterial elasticity than N or STIDDM groups as indicated by a shorter propagation time. These findings imply that apparent functional changes in pulsatile arterial blood flow occur early in the time course of diabetes and are independent of duration.

Is dehydroepiandrosterone a hormone?

Dehydroepiandrosterone (DHEA) is not a hormone but it is a very important prohormone secreted in large amounts by the adrenals in humans and other primates, but not in lower species. It is secreted in larger quantities than cortisol and is present in the blood at concentrations only second to cholesterol. All the enzymes required to transform DHEA into androgens and/or estrogens are expressed in a cell-specific manner in a large series of peripheral target tissues, thus permitting all androgen-sensitive and estrogen-sensitive tissues to make locally and control the intracellular levels of sex steroids according to local needs. This new field of endocrinology has been called intracrinology. In women, after menopause, all estrogens and almost all androgens are made locally in peripheral tissues from DHEA which indirectly exerts effects, among others, on bone formation, adiposity, muscle, insulin and glucose metabolism, skin, libido and well-being. In men, where the secretion of androgens by the testicles continues for life, the contribution of DHEA to androgens has been best evaluated in the prostate where about 50% of androgens are made locally from DHEA. Such knowledge has led to the development of combined androgen blockade (CAB), a treatment which adds a pure anti-androgen to medical (GnRH agonist) or surgical castration in order to block the access of the androgens made locally to the androgen receptor. In fact, CAB has been the first treatment demonstrated to prolong life in advanced prostate cancer while recent data indicate that it can permit long-term control and probably cure in at least 90% of cases of localized prostate cancer. The new field of intracrinology or local formation of sex steroids from DHEA in target tissues has permitted major advances in the treatment of the two most frequent cancers, namely breast and prostate cancer, while its potential use as a physiological HRT could well provide a physiological balance of androgens and estrogens, thus offering exciting possibilities for women's health at menopause.

Role of 17 beta-hydroxysteroid dehydrogenases in sex steroid formation in peripheral intracrine tissues.

In postmenopausal women, almost 100% of active sex steroids are synthesized in peripheral target tissues from inactive steroid precursors and, in adult men, approximately 50% of androgens are made locally in target tissues. This new field of endocrinology has been called intracrinology. The last and key step in the formation of all estrogens and androgens is catalyzed by a series of substrate-specific, cell-specific and unidirectional 17 beta-hydroxysteroid dehydrogenases (17 beta-HSDs). To date, seven human 17 beta-HSDs have been cloned, sequenced and characterized. The 17 beta-HSDs provide each cell with the means of precisely controlling the intracellular concentration of each sex steroid according to local needs.

Modulation by sex steroids and [D-Trp6, Des-Gly-NH2(10)]luteinizing hormone (LH)-releasing hormone ethylamide of alpha-subunit and LH beta messenger ribonucleic acid levels in the rat anterior pituitary gland.

LHRH and sex steroids play a major and direct regulatory role in the secretion of LH by the anterior pituitary gland. The aim of the present study was to investigate the interactions between sex steroids, more especially the potentiating effect of progesterone (P) in the presence or absence of a low dose of 17 beta-estradiol (E2) and/or dihydrotestosterone (D) on mRNA levels encoding the alpha- and beta-subunits of LH in both female and male rats. We also studied the effect of 2-week treatment with the LHRH agonist [D-Trp6, des-Gly-NH2(10)]LHRH ethylamide on the same parameters. After treatment with the LHRH agonist (5 micrograms daily), the accumulation of mRNA encoding the alpha-subunit was stimulated by approximately 3-fold while the LH beta mRNA concentration remained unchanged. Ovariectomy performed 14 days earlier, increased pituitary alpha and LH beta mRNA levels by 3.7- and 8.8-fold, respectively, while orchiectomy performed 14 days earlier increased alpha and LH beta mRNA levels by 6- and 6.5-fold, respectively. The present data demonstrate that although P alone exerts no effect on alpha and LH beta mRNA levels in castrated animals, treatment with P markedly potentiates the inhibitory effect of E2 on both mRNA levels in female as well as male rats. In addition, P potentiates the inhibitory effect of D on LH beta mRNA levels in castrated female rats. Furthermore, the present study illustrates the importance of the cumulative inhibitory effects of relatively low doses of E2 and D on mRNAs encoding both LH subunits. Moreover, the present observation of a differential modulation of alpha-subunit and LH beta mRNA levels after chronic treatment with an LHRH agonist offers an explanation for the high plasma levels of free alpha-subunit found in patients treated with LHRH agonists.

Regulation of pro-gonadotropin-releasing hormone gene expression by sex steroids in the brain of male and female rats.

The fine modulation of gonadotropin gene expression and secretion is well recognized to be regulated by sex steroids through their direct action both at the anterior pituitary level and on the pulsatile pattern of GnRH secretion at the hypothalamic level. Since the influence of sex steroids on hypothalamic GnRH mRNA levels remains to be elucidated, quantitative in situ hybridization was used to study the effect of sex steroids on cellular levels of pro-GnRH mRNA in adult rats of both sexes. The effects of 14-day gonadectomy as well as administration of 17 beta-estradiol (E2, 0.25 micrograms) or dihydrotestosterone (DHT, 100 micrograms) twice a day during 14 days to gonadectomized animals were evaluated. In addition, the effect of progesterone (P, 2 mg, twice daily) alone or in the presence of E2 was also studied in ovariectomized animals. Hybridization was performed using a 35S-labeled cDNA probe encoding rat pro-GnRH and the corresponding mRNA levels were assessed by counting the number of silver grains overlying labeled neurons. In male rats, castration induced a highly significant 65% increase (compared to intact rats) in the mean number of grains per neuron. Administration of E2 or DHT to castrated animals completely prevented the post castration rise in pro-GnRH mRNA levels. In female animals, the effect of ovariectomy was less striking than in the male, a 25% increase (P less than 0.001) being observed. Treatment with E2 or DHT also completely prevented the increase in pro-GnRH mRNA levels induced by ovariectomy. Moreover, treatment with P in ovariectomized animals markedly potentiated the inhibitory effect of E2 on pro-GnRH mRNA levels.(ABSTRACT TRUNCATED AT 250 WORDS)