John K. H. Lu

Biologic effects of transdermal estradiol.

Abstract We conducted a dose–response study in 23 postmenopausal women to compare the physiologic effects of transdermal estradiol and oral conjugated equine estrogens. The doses studied were 25, 50, 100, and 200 μg of transdermal estradiol per 24 hours, and 0.625 and 1.25 μg of oral conjugated estrogens. Transdermal estradiol increased circulating concentrations of estradiol and estrone. Oral conjugated estrogens also raised the levels of estrogen, particularly estrone. Both preparations lowered gonadotropin levels, decreased the percentages of vaginal parabasal cells, increased the percentage of superficial cells, and lowered urinary calcium excretion. The effects of 0.625 and 1.25 mg of oral estrogens were similar to those of 50 and 100 μg of transdermal estradiol per 24 hours, respectively. Oral estrogens significantly increased circulating levels of renin substrate, sex-hormone–binding globulin, thyroxine-binding globulin, and cortisol-binding globulin; transdermal estradiol had no effect. The higher...

The type I BMP receptor BmprIB is essential for female reproductive function

Maintenance of female reproductive competence depends on the actions of several hormones and signaling factors. Recent reports suggest roles for bone morphogenetic proteins (BMPs) in early stages of folliculogenesis. A role for the type I BMP receptor BmprIB as a regulator of ovulation rates in sheep has been described recently, but little is known about the roles of BMP signaling pathways in other aspects of reproductive function. We report here that BMPRIB is essential for multiple aspects of female fertility. Mice deficient in BmprIB exhibit irregular estrous cycles and an impaired pseudopregnancy response. BmprIB mutants produce oocytes that can be fertilized in vitro, but defects in cumulus expansion prevent fertilization in vivo. This defect is associated with decreased levels of aromatase production in granulosa cells. Unexpectedly, levels of mRNA for cyclooxygenase 2, an enzyme required for cumulus expansion, are increased. BmprIB mutants also exhibit a failure in endometrial gland formation. The expression of BmprIB in uterine linings suggests that these defects are a direct consequence of loss of BMP signaling in this tissue. In summary, these studies demonstrate the importance of BMP signaling pathways for estrus cyclicity, estradiol biosynthesis, and cumulus cell expansion in vivo and reveal sites of action for BMP signaling pathways in reproductive tissues.

The Luteinizing Hormone Surge Is Preceded by an Estrogen-Induced Increase of Hypothalamic Progesterone in Ovariectomized and Adrenalectomized Rats

As circulating estrogen levels rise on the afternoon of proestrus, they stimulate the hypothalamo-pituitary axis. This estrogen positive feedback is pivotal to stimulate the luteinizing hormone (LH) surge required for ovulation and luteinization of ovarian follicles. In addition to estrogen, pre-LH surge progesterone is critical for an LH surge as was demonstrated by blocking progesterone synthesis. In ovariectomized (OVX) rats treated with trilostane, a blocker of the enzyme 3β-hydroxysteroid dehydrogenase (3β-HSD) that catalyzes the conversion of pregnenolone to progesterone, estrogen did not induce an LH surge. Further, estrogen induced an LH surge in OVX and adrenalectomized (ADX) rats, indicating that the source of progesterone was neither the ovary nor adrenal gland. This estrogen-only LH surge was inhibited by pretreatment with trilostane, indicating that although the adrenal gland and ovary were not necessary for positive feedback, progesterone synthesis was critical for estrogen-induced positive feedback in an OVX/ADX rat. This suggested that the LH surge is dependent on the pre-LH surge synthesis of progesterone. Estrogen-induced progesterone receptors in the hypothalamus are vital for the LH surge, so a potential location for progesterone synthesis is the hypothalamus. OVX/ADX female rats were treated with 17β-estradiol (50 µg) and progesterone levels were assayed by RIA. Progesterone levels were elevated in hypothalamic tissue following estrogen treatment. No increases in tissue progesterone levels were found in parietal cortex, cerebellum, medulla, pituitary or plasma. Additionally, male rats that do not have an estrogen positive feedback-induced LH surge were examined. Castrated/ADX male rats had no increase in hypothalamic progesterone levels after estrogen treatment. Together, these data strongly suggest that estrogen enhances neuroprogesterone synthesis in the hypothalamus that is involved in the positive feedback regulating the LH surge.

AMG 479, a fully human anti–insulin-like growth factor receptor type I monoclonal antibody, inhibits the growth and survival of pancreatic carcinoma cells

Pancreatic carcinoma is a leading cause of cancer deaths, and recent clinical trials of a number of oncology therapeutics have not substantially improved clinical outcomes. We have evaluated the therapeutic potential of AMG 479, a fully human monoclonal antibody against insulin-like growth factor (IGF) type I receptor (IGF-IR), in two IGF-IR–expressing pancreatic carcinoma cell lines, BxPC-3 and MiaPaCa2, which also differentially express insulin receptor (INSR). AMG 479 bound to IGF-IR (KD 0.33 nmol/L) and blocked IGF-I and IGF-II binding (IC50 < 0.6 nmol/L) without cross-reacting to INSR. AMG 479 completely inhibited ligand-induced (IGF-I, IGF-II, and insulin) activation of IGF-IR homodimers and IGF-IR/INSR hybrids (but not INSR homodimers) leading to reduced cellular viability in serum-deprived cultures. AMG 479 inhibited >80% of basal IGF-IR activity in BxPC-3 and MiaPaCa2 xenografts and prevented IGF-IR and IGF-IR/INSR hybrid activation following challenge with supraphysiologic concentrations of IGF-I. As a single agent, AMG 479 inhibited (∼80%) the growth of pancreatic carcinoma xenografts, and long-term treatment was associated with reduced IGF-IR signaling activity and expression. Efficacy seemed to be the result of two distinct biological effects: proapoptotic in BxPC-3 and antimitogenic in MiaPaCa2. The combination of AMG 479 with gemcitabine resulted in additive inhibitory activity both in vitro and in vivo. These results indicate that AMG 479 is a clinical candidate, both as a single agent and in combination with gemcitabine, for the treatment of patients with pancreatic carcinoma.[Mol Cancer Ther 2009;8(5):1095–105]

Suppression of the ovary using a gonadotropin releasing-hormone agonist prior to stimulation for oocyte retrieval *

Five women with prior suboptimal ovarian stimulation for in vitro fertilization and embryo transfer (IVF-ET) were pretreated with a long-acting gonadotropin-releasing hormone (GnRH) agonist beginning in the midluteal phase of the preceding menstrual cycle. The four women with normal luteal function had castrate estrogen levels following regression of the corpus luteum, whereas one woman with an abnormal luteal phase and perimenopausal levels of gonadotropins had an agonistic response. In the three women with adequate stimulation, 20 oocytes were obtained and one women became pregnant. Initiation of GnRH agonist therapy during the luteal phase of a normal menstrual cycle may be an efficient way of obtaining ovarian suppression without an agonistic response.

Gonadal function in aging rats and its relation to pituitary and mammary pathology

In the female rat, aging is characterized by a high incidence of prolactin (Prl)-secreting pituitary adenomas and mammary tumors. In contrast to this, old males show only a moderate to low incidence of pituitary and mammary pathology. Since gonadal steroids and Prl are thought to be key factors in the genesis of the above neoplastic pathologies, it was of interest to compare the serum levels of progesterone (P), estradiol (E2), testosterone (T) and Prl with the incidence of pituitary and mammary tumors in aging male and female rats. Young (3-4-month; YF), old (25-month; OF) and senescent (33-35-month; SF) female and young (3-4-month; YM) and old (24-26-month; OM) male Sprague-Dawley rats were killed by decapitation and their pituitaries weighed. Serum sex steroids and Prl were measured by RIA. The average life span of females but not males was markedly extended by systematic removal of mammary tumors. Females showed a rising incidence of mammary tumors after 14 months of age. In males, this pathology which began to appear at 16 months, had a much lower incidence than in females at all ages. Serum levels of E2 were (means +/- S.E.M.) 22.0 +/- 1.6; 18.9 +/- 0.8; 32.9 +/- 2.5; 37.3 +/- 2.0 and 32.2 +/- 3.0 pg/ml for YM, OM, YF, OF and SF, respectively. Serum P was 1.4 +/- 0.3; 1.6 +/- 0.2; 10.4 +/- 2.2; 9.7 +/- 3.3 and 6.8 +/- 0.8 ng/ml for YM, OM, YF, OF and SF, respectively. Serum T was 1578.9 +/- 188.7; 807.6 +/- 103.0; 197.5 +/- 11.8; 223.7 +/- 25.5 and 176.9 +/- 20.7 pg/ml for YM, OM, YF, OF and SF, respectively. Finally, serum Prl was 14.9 +/- 1.7; 21.9 +/- 4.0; 15.9 +/- 1.4; 52.4 +/- 9.4 and 170.8 +/- 31.1 ng/ml for YM, OM, YF, OF and SF, respectively. A strong correlation was found between serum Prl and anterior pituitary weight in OM, OF and SF, but not between serum Prl and sex steroid levels or sex steroid ratios. We conclude that, although the sex-related differences in mammary and pituitary tumor incidence during aging in rats can be partially accounted for by the different serum profiles of Prl and gonadal steroids in each sex, sex-associated differences in target tissue susceptibility should also be considered as an important determinant of the level of tumor incidence.

Influences of Age and Ovarian Follicular Reserve on Estrous Cycle Patterns, Ovulation, and Hormone Secretion in the Long-Evans Rat

Abstract This study examined the influences of aging and reduced ovarian follicular reserve on estrous cyclicity, estradiol (E2) production, and gonadotropin secretion. Young virgin and middle-aged (MA) retired breeder female rats were unilaterally ovariectomized (ULO) or sham operated (control). Unilateral ovariectomy of young rats reduced the ovarian follicular reserve by one-half, to a level similar to that found in MA controls. Unilateral ovariectomy of MA females reduced the follicular pool further, to one half of MA controls. The incidence of regular cyclicity was significantly lower in MA ULO females than in young controls, with intermediate cycle frequency in young ULO and MA controls. Among cyclic rats, the magnitude of the proestrous LH surge was highest in young controls, intermediate in young ULO rats and MA controls, and lowest in MA ULO females. Similarly, ovulation rates were highest in young controls, intermediate in young ULO rats and MA controls, and lowest in MA ULO females. While young ULO rats exhibited augmented secondary FSH surges on estrous morning, middle-aged ULO females displayed secondary FSH levels comparable to young controls. The effects of age and reduced follicle number on estrous cyclicity and gonadotropin secretion were not due to altered E2 secretion, as preovulatory E2 levels were similar among all groups. Thus, experimental reduction in the follicular reserve exerts acute effects on the preovulatory LH surge, ovulation rate, and estrous cyclicity in both young and MA rats. However, decreased follicle number increases FSH levels only in young rats, indicating aging-related alterations in the feedback regulation of FSH.

Changes in Ovarian Function and Gonadotropin and Prolactin Secretion in Aging Female Rats

In the female rat, reproductive function is characterized by several interesting features, including (1) estrous cycles and the concomitant changes in neuroendocrine function are repeated at frequent intervals (4–5 days), and (2) reproductive life represents less than one-third of the entire lifespan. Under optimal laboratory conditions, the female rat may have a lifespan of about three years. Puberty occurs around 35–40 days after birth, and regular estrous cycles are usually seen in animals older than 2 months of age. Between the ages of 2–12 months, most adult females display regular estrous cycles of 4- or 5-day duration, and this cyclicity is interrupted by a persistent-diestrous phase when the rat is in gestation, postpartum lactation, or in a pseudopregnant condition.

Impotence in scleroderma

Hormonal, neurologic, and vascular factors affecting potency were evaluated in 10 men with scleroderma and in 10 age-matched men with rheumatoid arthritis. Impotence was reported by 6 of the patients with scleroderma and none with rheumatoid arthritis. Studies of serum testosterone, free testosterone index, follicle-stimulating hormone, luteinizing hormone, prolactin, estradiol, thyroxine, and thyrotropin did not show a hormonal basis for impotence in any patient. Neurologic causes were not found on physical examination. Penile blood pressures were markedly abnormal in 4 impotent patients, intermediate in 2 impotent and 3 potent patients, and normal in 11 potent patients. A history of claudication and diminished ankle blood pressures indicated large vessel disease in 2 impotent patients; the remaining 4 impotent men had normal ankle pressures, suggesting that their poor penile blood pressures and impotence were due to small vessel disease, perhaps the small artery lesions of scleroderma.

Induction of hypothalamic opioid activity with transdermal estradiol* administration in postmenopausal women†‡

The effects of estradiol (E 2 ) administered by a transdermal system on the induction of hypothalamic opioid activity was examined in 18 postmenopausal women. Women were given 25, 50, 100, or 200 μg /d dosages of E 2 for 28 days each. Naloxone at a dosage of 2 mg/h was infused intravenously for 4 hours, and serum was obtained every 15 minutes for 6 hours. With increasing doses, rises of serum E 2 and estrone were elicited across the range seen in premenopausal women. A stepwise reduction of luteinizing hormone (LH) levels was observed with increasing dosages of E 2 . The naloxone infusions resulted in significant increases of LH when E 2 was being given but not during the pretreatment studies. These data indicate E 2 , in physiological concentrations, induces hypothalamic opioid activity.