Josephine B. Miller

Menopausal status, moderate-intensity walking, and symptoms in midlife women.

The purpose of this randomized clinical trial study was to determine the effectiveness of a 24-week, home-based, moderate-intensity, walking intervention in improving symptoms (vasomotor, uro-genital/sexual, sleep, psychological, cognitive, physical) experienced by midlife women. One hundred and seventy-three Caucasian and African American women aged 45 to 65 who were not on hormone therapy, had no major signs or symptoms of cardiovascular disease, and were sedentary in their leisure activity were randomly assigned to the moderate-intensity walking group or the nonexercise control group. The exercise prescription was walking at a frequency of 4 times a week for a duration of 20 to 30 minutes. The symptom impact inventory included the frequency, intensity, and bothersomeness of 33 symptoms collected at baseline and 24 weeks. Adherence was measured with a heart rate monitor and exercise log. Average adherence to frequency of walking was 71.6% of the expected walks. After 24 weeks, there were no differences between the walking and control group on change in symptoms. However, multiple regression revealed that frequency of adherence to walking along with change in physical symptoms and menopausal status were significant predictors of change in sleep symptoms. While walking did not improve most symptoms experienced by midlife women, frequency of walking may improve sleep.

A Baboon (Papio anubis) simulated-pregnant model: cell specific expression of insulin-like growth factor binding protein-1 (IGFBP-1), type I IGF receptor (IGF-1 R) and retinol binding protein (RBP) in the uterus

In order to test the hypothesis that the baboon conceptus/placenta regulates the synthesis of specific proteins in the endometrium, we developed a simulated-pregnant baboon model. Baboons (n=2–6/group) were treated with increasing amounts of human chorionic gonadotrophin (hCG) for 10 or 12 days beginning on day 6 or 7 PO. Uterine tissues were obtained at day 18 PO following 12 days of hCG treatment. Animals in the day 25 and 32 PO group were treated for 10 days with hCG. Following the hCG treatment, estradiol (E) and progesterone (P) implants were inserted subcutaneously. Control groups consisted of E and P treatment only (day 25 PO), or ovariectomy on day 6 or 7 PO followed by hCG plus E and P treatment (days 18 and 25 PO). Serum samples were obtained daily or once every 2 days and analysed for E and P by radioimmunoassay. hCG activity in serum was determined by a Leydig cell bioassay. Portions of the endometrial tissue were either subjected to organ explant culture, analysed by immunocytochemistry or extracted for RNA. Peripheral serum levels of hCG, E and P in the experimental groups fell within the 95% confidence interval limits of hormone concentrations achieved during pregnancy. The morphology of the endometrium in the hCG treated baboons and pregnant baboons was similar i.e., distended convoluted glands, many spiral artery beds, a loose vacuolized stroma, and increased collagen staining. However, in the absence of hCG (E+P treatment only) the glands tended to be straight rather than corkscrew-shaped, and decreased stromal vacuolization and collagen staining was evident.35S-methionine labeled proteins in explant culture conditioned media (TCM) were analysed by two-dimensional SDS-PAGE and fluorography. A comparable pattern of protein synthesis was apparent in all treatment groups except for a low molecular weight (27 000–30 000 daltons) group of polypetides which only was evident in TCM from the hCG treated baboons. A similar group of proteins are also secreted by the baboon endometrium during pregnancy. The immunocytochemical localization of estrogen (ER) and progesterone receptors (PR) was comparable to that observed in pregnant baboons. IGFBP-1 localization was confined to the glandular epithelium in the hCG treated groups (intact and ovariectomized) and was virtually undetectable in the E and P treated group. The intensity of IGFBP-1 staining was variable within each of the hCG treatment groups on days 18, 25 and 32 PO. This variability was also apparent by Western blot analysis, immunoassay of proteins in TCM and on Northern blots of total RNA from the same animals. In contrast, IGF-I R immunostaining was evident in both glandular and surface epithelium of all treatment groups. Expression of RBP was confined to the basal glands. The characteristic upregulation of RBP synthesis in the functionalis observed during early pregnancy was not apparent in any of the treatment groups. In summary, these studies indicate that exogenous hCG in conjunction with E and P, can induce the general morphological and biosynthetic changes the baboon endometrium undergoes during early pregnancy. In addition, this hormonal treatment is also capable of maintaining the epithelial expression of IGFBP-1, IGF-1 and RBP. However, other factors from the conceptus appear to be necessary to induce the cell specific changes in the expression of these three proteins that are observed during pregnancy.

CALCIUM-INDEPENDENT PHOSPHOLIPID/DIOLEIN-DEPENDENT PHOSPHORYLATION OF A SOLUBLE OVARIAN MR 80 000 SUBSTRATE PROTEIN : BIOCHEMICAL CHARACTERISTICS

Soluble ovarian extracts were incubated with protein kinase effectors in the presence of [gamma 32P]ATP and proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Autoradiograms revealed phosphorylation of an ovarian Mr = 80,000 substrate in the presence of EGTA ([ethylenebis(oxyethylenenitrilo)]tetraacetic acid), phosphatidylserine and 1,2-diolein. In contrast to a classical response pattern to C-kinase effectors, the ovarian Mr = 80,000 phosphorylation was inhibited by 2 x 10(-7) M or greater free Ca2+. The ovarian Mr = 80,000 substrate was distinguished from the myristoylated acidic Mr = 80,000 C-kinase substrate of brain tissue on the basis of heat stability and phosphorylative response to effectors. Phosphorylation of the exogenous substrate myelin basic protein by DEAE-resolved ovarian kinase showed the variant effector dependence, maximal in the presence of EGTA, phosphatidylserine and 1,2-diolein. Finally, the effect of Ca2+ on ovarian Mr = 80,000 [32P]phosphate content could not be accounted for by post-phosphorylation activities, or by DEAE-resolvable or hydroxylapatite-resolvable inhibitory activities.

Characterization and physiological variation in prostaglandin, prostacyclin and thromboxane synthesis by corpora lutea, non-luteal and uterine tissues during pseudopregnancy in the rabbit

To determine if luteal as well as uterine prostaglandin production is associated with luteal regression, the conversion of (14C) arachidonic acid into prostaglandin (PG) F2 alpha, PGE2, PGD2, prostacyclin (measured as its stable metabolite 6-keto-PGF1 alpha) and thromboxane (measured as its stable metabolite TXB2) was characterized and measured in microsomes from the uterus, corpora lutea and nonluteal tissue of the rabbit on days 8, 12 and 15 of pseudopregnancy. PGF2 alpha production was increased on day 15 compared to days 8 or 12 of pseudopregnancy in corpora lutea and in the uterus. In uterine microsomes, 6-keto-PGF1 alpha production was also elevated on day 15. No significant changes in the production of PGF2 alpha, TXB2 or PGD2 were observed with uterine or luteal microsomes. Arachidonic acid metabolism by microsomes from nonluteal tissue was not significantly different on any day examined. These results suggest that intraluteal, as well as uterine PGF2 alpha production may contribute physiologically to the final phase of luteal regression.

Variant Diacylglycerol—Dependent Protein Phosphotransferase Activity in Ovarian Tissues

Protein phosphorylation has been implicated as a key response to gonadotropic stimulation of ovarian cells. The second messenger systems, which would act by control of protein phosphorylation, include the adenylate cyclase/cAMP/A-kinase system (1) and the phosphoinositide hydrolysis system (2). Hydrolysis of phospatidylinositol 4,5-bisphosphate leads to the generation of 1,2-diacylglycerol, activation of diacylglycerol-dependent protein kinases (the C-kinases) (2, 3), and the generation of inositol 1,4,5- triphosphate, promoting release of Ca++(2) and activation of Ca++/ calmodulin-dependent kinases (1). Ovarian tissues have been shown to contain protein kinases in each of the above classes. The functional significance of each class of protein kinases is not yet fully defined, but the existence of hormone-regulated parameters for each class of kinase in the ovary suggests that the kinases participate in the mediation of or modulation of hormone-regulated cellular responses. Ovarian A-kinases are regulated acutely (4) and chronically (5) by gonadotropic stimulation. Estrogen is also required for stimulated levels of mRNA for A-kinase regulatory subunit (RIIβ) in the rat granulosa cell (6). Ca++/calmodulin kinase III, which phosphorylates the protein synthesis elongation factor EF2, is regulated by estrogen in the rat corpus luteum of pregnancy (7). Levels of lipid-derived second messengers, 1,2-diacylglycerol (DAG) and inositol 1,4,5-triphosphate, are regulated by gonadotropin, prostaglandin, and GnRH exposure in ovarian cells (8–11).