Is pregnancy in diabetic women associated with folate deficiency?

OBJECTIVE: To determine whether folate metabolism in pregnant diabetic women is significantly different from that in pregnant nondiabetic women, thus predisposing them to having offspring with major congenital anomalies. RESEARCH DESIGN AND METHODS: A total of 31 pregnant diabetic women and 54 pregnant nondiabetic control subjects were studied at their first prenatal visits. Dietary folate intake, serum folate, red blood cell folate, urinary folate, and homocysteine were measured and compared after controlling for folate supplementation. Among diabetic women the relationships among parameters of folate metabolism and glycemic control were also assessed. RESULTS: There were no significant differences between the pregnant diabetic and non-diabetic women for any measures of folate metabolism after accounting for folate supplementation. In addition, among diabetic women, there were no associations among parameters of folate metabolism and glycemic control. CONCLUSIONS: Abnormal folate metabolism does not appear to occur in pregnant diabetic women. It is unlikely that deranged folate metabolism explains the higher incidence of major anomalies in infants of diabetic mothers. These results do not diminish the importance of periconception folate supplementation or preclude other possible scenarios that could restrict folate use by the embryo, leading to congenital anomalies.

Control of the preovulatory gonadotropin surge in the ewe.

The aims of this study were to determine the effect of ovariectomy on the release of LH and FSH during the preovulatory gonadotropin surge and to ascertain, by the use of sodium pentobarbitone (NaPb), if the secretion of these pituitary hormones requires continuous stimulation from the hypothalamus. Sheep were treated with NaPb for 2 h beginning 1) immediately before the gonadotropin surge, 2) during the ascending limb of the gonadotropin surge, and 3) during the descending limb of the gonadotropin surge. Ewes were ovariectomized (ovx) at each of the time periods listed above, and intact ewes included were at times 2 and 3. A group of intact ewes was given 100 microgram gonadotropin-releasing hormone (GnRH) in addition to NaPb at time 2, NaPb given during the ascending limb of the gonadotropin surge caused a transient fall in peripheral LH and FSH; however, the release of gonadotropins was reinitiated and the surge continued when the ewes recovered from anesthesia. Treatment with NaPb after the apex of the gonadotropin surge did not affect circulating levels of LH and FSH. Ewes given NaPb and ovx before the initiation of the gonadotropin surge released significantly less LH and FSH during the surge than the other treatment groups. The total amounts of LH and FSH released in intact and ovx ewes treated with NaPb after the surge was initiated were not different than those levels in the saline-treated controls. Intact ewes treated with 100 micrograms GnRH also released an amount of LH similar to that in the control group. We conclude that gonadotropin release from the pituitary gland requires the continual presence of GnRH during the ascending limb of the preovulatory gonadotropin surge, and that once the surge has been triggered, the ovaries do not appear to be required for further hypothalamic stimulation.

Follicular steroidogenesis and gonadotropin binding to ovine follicles during the estrous cycle.

The interrelationships among [125I]hCG binding in thecal and granulosa cells, antral fluid steroid concentrations, follicular size, and ovarian steroid secretion were examined at three different stages of the estrous cycle. Group 1 ewes were ovariectomized during the luteal phase of the estrous cycle, and the other groups were ovariectomized before (group 2), or after (group 3) the peak of the preovulatory LH surge. Times of luteolysis and the LH surge were assessed by measurement of peripheral concentrations of progesterone and LH. Three patterns of [125I]hCG binding to follicles were noted: 1) binding to both thecal and granulosa cells (activated follicle), 2) binding to the thecal cell layer only, and 3) no observed binding. In general, there was one active follicle per ewe, or one per ovary, and the number of active follicles was not different from the number of corpora lutea in each of the three groups. The active follicles were significantly larger than the other two classes of follicles. Antral fluid estradiol concentrations were significantly greater in the active follicles and were higher in group 2 ewes than in the other two groups. In group 2, antral fluid testosterone concentrations were significantly higher in follicles with LH receptors in the thecal cell layer only. Ovarian secretion of testosterone and estradiol increased during the early follicular phase (group 2), with the major secretion coming from the ovary containing the active follicle. Ovarian progesterone secretion was high in ovaries containing active corpora lutea which prevented the assessment of ovarian follicular secretion of progesterone. The follicle with LH receptors in thecal and granulosa cells was responsible for the increased estradiol secretion observed during the preovulatory period and is presumed to be the ovulatory follicle.

Development and characterization of a monoclonal antibody which distinguishes the beta-subunit of human chorionic gonadotropin (beta hCG) in the presence of the hCG.

A monoclonal antibody specific for the beta-subunit of human chorionic gonadotropin (beta hCG) has been developed which has a cross reactivity of 0.23% with intact hCT. There was no cross reactivity with other glycoprotein pituitary hormones in the range in which they might be expected to be present in serum. The beta-subunits of hFSH, hTSH and hLH did not inhibit binding of [125I]iodo-beta hCG to the antibody at up to 250 ng/tube. The dissociation constant (Kd) with [125I]iodo-beta hCG was 3.3 x 10(-10), and the reaction had reached equilibrium within 1 h.

Plasma substance-P in neuroendocrine tumors and idiopathic flushing: the value of pentagastrin stimulation tests and the effects of somatostatin analog.

We examined the role of the potent vasoactive kinin substance-P (SP) in flushing derived from various causes. SP was measured in plasma after acetone/ether extraction using an antiserum directed at the carboxy-terminal 5-11 amino acid region of undecapeptide SP. The antiserum had less than 1% cross-reaction with the other neurokinins, neurokinin-A and neuropeptide-K, that derive from the beta-preprotachykinin gene and share carboxy-terminal residues. Basal and pentagastrin-stimulated SP levels were measured in 22 healthy controls, 11 patients with histologically proven carcinoid tumors, 8 patients with tumors other than carcinoid, and 7 patients with idiopathic flushing (IF). Basal SP levels were less than 10 pg/mL in normal subjects. All patients with midgut carcinoid tumors had SP levels greater than 25 pg/mL, as did 7 of 8 patients with noncarcinoid tumors and 5 of 7 patients with IF. Using 50 pg/mL as the cutoff point, the sensitivity was 63% for detection of a tumor, and 100% of nontumor patients were excluded. Pentagastrin administration uniformly induced flushing and caused a rise in SP levels greater than 150 pg/mL in 5 of 10 patients with carcinoid tumors, 3 of 8 with noncarcinoid tumors, and 0 of 7 with IF, i.e. a SP rise of more than 100 pg/mL suggests a tumor. Administration of somatostatin (150 micrograms) 0.5 h before the pentagastrin abolished flushing in all carcinoid patients and reduced SP levels, but not into the normal range. Long term treatment with SMS significantly reduced flushing and lowered SP levels, but did not restore these to normal. We conclude that 90% of patients with carcinoid/noncarcinoid tumor have raised COOH-terminal SP levels. A basal level above 50 pg/mL or a pentagastrin-stimulated rise of more than 100 pg/mL distinguishes carcinoid from IF. The dissociation between SP concentrations and flushing suggests that SP may not be the only kinin involved in the flushing associated with carcinoid tumors.

Differentiation of human pituitary adenomas determines the pattern of chromogranin/secretogranin messenger ribonucleic acid expression.

The distribution of chromogranin/secretogranin (Cg/Sg) mRNAs, determined by Northern and in situ hybridization, was analyzed in 14 cultured pituitary adenomas characterized by immunohistochemistry and hormone secretion in a defined medium in vitro. There were 5 functional GH adenomas, 1 silent GH adenoma, 7 null cell adenomas, and 1 oncocytoma. The null cell adenomas, oncocytoma, and silent GH adenomas were also analyzed by electron microscopy. Most null cell adenomas and the oncocytoma secreted FSH and LH into the culture medium. GH adenomas, which are examples of well differentiated tumors based on morphological examination, expressed significantly more SgIII mRNA compared to the null cell adenomas and oncocytoma (70 +/- 6% vs. 22 +/- 5%; P < 0.001). GH adenomas also expressed significantly less CgA mRNA compared to the less well differentiated null cell adenomas and oncocytoma (27 +/- 6% vs. 67 +/- 4%; P < 0.001), which could be considered less well differentiated based on ultrastructural morphological features. After treatment with phorbol 12-myristate 13-acetate (10(-7) M) for 7 days, there was an increase in the mRNA for CgB and SgII mRNAs in GH and null cell tumors, while dexamethasone treatment for 7 days increased CgA mRNA in GH and null cell adenomas. GnRH treatment for 7 days increased CgB mRNA in null cell adenomas. Phorbol 12-myristate 13-acetate also decreased the percentage of immunoreactive GH cells and GHm RNA, determined by in situ and Northern hybridization analyses. These results indicate that pituitary adenomas have a distinct pattern of Cg/Sg mRNA expression, which appears to be related to the degree of morphological differentiation of these neoplasms, and suggest that the effects of secretagogues on various Cg/Sg mRNA levels may be related to the stimulation of hormone secretion.

Gonadotropin-releasing hormone regulates gonadotropin beta-subunit and chromogranin-B messenger ribonucleic acids in cultured chromogranin-A-positive pituitary adenomas.

Chromogranin-A-positive pituitary adenomas include glycoprotein hormone-producing adenomas, null cell adenomas, and a few other pituitary adenomas. We studied the effects of GnRH, CRF, dexamethasone, and phorbol 12-myristate 13-acetate on FSH and LH secretion and on FSH beta and chromogranin-A and -B mRNA expression in 10 chromogranin-A-positive adenomas in vitro to analyze the regulation of FSH and chromogranin-A and -B expression in these neoplasms. Most adenomas responded to GnRH stimulation during 7 days in culture with a 2- to 10-fold increase in FSH and LH secretion and a 2- to 7-fold increase in FSH beta mRNA compared to control values. CRF and phorbol 12-myristate 13-acetate also stimulated FSH and LH secretion 2- to 5-fold in five of seven and three of three cases, respectively, during 7 days in culture. Dexamethasone stimulated both FSH and LH secretion in two of three cases as well as FSH beta mRNA in vitro in the one case examined. GnRH treatment consistently produced a 2-fold increase in chromogranin-B mRNA, but not in chromogranin-A mRNA, after 7 days of culture. These results indicate that many chromogranin-A-positive adenomas respond to GnRH and CRF in vitro by increased hormone secretion and that GnRH stimulation leads to increased amounts of FSH beta and chromogranin-B mRNAs. The differential response of chromogranin-A and -B mRNAs after GnRH stimulation indicates that the chromogranin genes are highly regulated in these tumors.

Serum percent-free PSA does not predict extraprostatic spread of prostate cancer.

Percent-free prostate-specific antigen (proportion of free prostate-specific antigen [PSA] to total PSA) has been shown recently in studies on frozen serum samples to be more useful than total PSA alone in distinguishing prostate cancer from benign conditions of the prostate gland. The primary purpose of our study was to determine whether percent-free PSA could predict extraprostatic spread of prostate cancer. We also sought to evaluate the freeze-thaw stability of free PSA. Percent-free PSA values in fresh serum samples were compared with those in aliquots subjected to one to five freeze-thaw cycles. Percent-free PSA values in frozen serum samples from 130 men undergoing radical prostatectomy for clinically localized prostate cancer were compared across pathologic stages. Free PSA levels remained stable for up to five freeze-thaws. Great overlap was found in percent-free PSA values for men with organ-confined disease and those with extraprostatic spread. These results indicate that multiple freeze-thaw cycles do not significantly affect free PSA levels and percent-free PSA is not useful in identifying ideal candidates for radical prostatectomy.

Intraoperative parathyroid hormone monitoring fails to detect double parathyroid adenomas: a 2-institution experience.

BACKGROUND: We hypothesized that intraoperative parathyroid hormone monitoring (IOPTH) reliably would detect double parathyroid adenomas. METHODS: This was a retrospective study of 20 patients undergoing conventional parathyroidectomy with resection of exactly 2 abnormal glands. Full exploration was performed regardless of IOPTH values, which were measured after anesthetic induction and 5 and 10 minutes following removal of the first abnormal parathyroid gland. Failure to fall below 50% of baseline value by 10 minutes following resection of the first gland indicated the presence of multiglandular disease. RESULTS: All patients were cured. All excised glands were hypercellular on histology. Mean IOPTH values in 9 of the 20 patients with true negative results (noncurative decrease, another gland present) were 66% +/- 7% at 5 minutes and 83% +/- 15% at 10 minutes. The IOPTH values in 11 of the 20 patients with false positive results (curative decrease, another gland present) were 28% +/- 4% at 5 minutes and 18% +/- 2% at 10 minutes. The false positive rate of IOPTH was 55%. CONCLUSIONS: We found that IOPTH failed to reliably detect the presence of double parathyroid adenomas. These data suggest that caution should be exercised when terminating limited parathyroid exploration based on a curative fall in IOPTH values.

Half-life determination of serum free prostate-specific antigen following radical retropubic prostatectomy.

OBJECTIVES: Prostate-specific antigen (PSA) continues to be the the most clinically useful tumor marker for prostate cancer. Recently, several molecular forms of PSA have been detected and characterized. These specific forms, including free PSA and PSA complexed to alpha 1-antichymotrypsin, can be measured and their proportions determined. In doing so, the sensitivity of PSA as a tumor marker can be maintained while the specificity is improved. In order to maximize the clinical utility of free PSA, the half-life and elimination kinetics of free PSA from the serum were determined. METHODS: Twenty-five patients, ages 43-74 years (mean 60 years) with biopsy proven, organ-confined adenocarcinoma of the prostate who underwent anatomic radical retropubic prostatectomy, were identified. For each patient, venous blood samples were obtained preoperatively, and at 60-minute intervals beginning 1 hour after the prostate was removed. The specimens were handled and stored in a consistent fashion. Using the AxSYM immunoassay analyzer (Abbott Diagnostics, Abbott Park, IL), the serum free PSA values were determined and plotted as a function of time for each patient. From the 25 individual elimination curves that were generated, the half-life of serum free PSA was determined. RESULTS: The mean half-life of serum free PSA was 110 minutes +/- 18.6 minutes (SD). Analysis of the individual and cumulative elimination curves indicates that the elimination of free PSA from the serum following radical prostatectomy follows a biphasic pattern. CONCLUSIONS: Unlike PSA, which has a half life of 2-3 days, the half-life of serum free PSA is 110 minutes (1.83 hours). This short half-life may have significant implications for the use of percentage of free PSA as a clinically useful tool in distinguishing patients with early, curable prostate cancer from men with benign prostatic hyperplasia (BPH) only.

Serum thyroglobulin levels after thyroxine withdrawal in patients with low risk papillary thyroid carcinoma.

We hypothesized that elevated levels of serum thyroglobulin (Tg) are frequently found as the only index of residual neoplasm in patients with low-risk papillary thyroid carcinoma. The records of patients operated on for papillary thyroid carcinoma over a 2-year period were reviewed, and the patients were allocated to risk groups by a validated staging method that does not include Tg levels. Of the 35 patients who manifested a low-risk carcinoma, 9 (26%) exhibited elevated Tg concentrations (11-53 ng/mL) during thyroxine withdrawal after therapies, while clinical, scintigraphic, and radiographic studies at least 1 year later showed no evidence of tumor. Prior scintigraphic imaging of therapeutic doses of 131I in 8 of 9 patients demonstrated no distant metastases, further confirming the low-risk status of this group. The staging method predicts that only 0.9% of patients with low-risk papillary carcinoma will have a cause specific death in 20 years. Elevated Tg concentrations have not been shown to forecast independently the survival of patients with low-risk papillary carcinoma. Thus, although frequently encountered, elevated Tg concentrations are unlikely to predict shortened survival in patients with papillary carcinoma for whom low risk has been determined from other data.

A chemical approach to solving bridging phenomena in steroid radioimmunoassays.

Steroid radioimmunoassays (RIA) employ antibodies raised against a carrier protein-steroid conjugate. Individual antibodies may recognize the steroid, the protein or the chemical bridge used to join them together. Use of the same bridge in the tracer results in higher affinity binding of the tracer than the native ligand which in turn results in a loss of sensitivity and precision. We have greatly reduced bridge-binding in a RIA for androstenedione. Conjugates and radioiodinated labels were prepared with either an ester or either chemical bridge. By using an antibody and the corresponding label with the heterologous bridge very sensitive assays were obtained.

A specific radioimmunoassay for androstenedione with reduced bridge-binding.

Antibody used in a steroid radioimmunoassay raised against a steroid hapten-carrier protein conjugate may recognize both the hapten and the chemical bridge to the protein. Use of the same bridge in the radio-isotopic label may lead to higher affinity binding to the label than to the native steroid. Inhibition curves under these conditions are shallow and generally not acceptable for radioimmunoassay procedures. We have developed a radioimmunoassay for androstenedione that employs different bridges at the 11 beta position of the steroid for the protein conjugate and label. The resulting assay has greatly reduced bridge-binding, has an acceptable slope for the standard curve and is very specific as evidenced by low crossreactivies to other steroids.

A specific, non-chromatographic radioimmunoassay for human plasma cortisol.

A radioimmunoassay technique has been developed for the measurement of cortisol in a single methylene chloride extract of human plasma without chromatography. The antiserum, obtained by immunizing rabbits with cortisol-3-carboxymethyl-oxime conjugated to bovine serum albumin, had a high affinity (KA = 1.8 X 10(9) 1/mole) and capacity (2.3 X 10(-6) moles/L undiluted serum) for cortisol. The minimum detectable amount determined at the lower 95% confidence limit of the buffer control tubes was 8.3 +/- 4.7 pg/tube and a log dose - logit response standard curve was linear between 20 pg and 20 ng/tube. The antiserum was highly specific for cortisol with only corticosterone, cortisone, 11-deoxycortisol and 21-deoxycortisol showing significant cross-reaction (12.4, 6.6, 3.8 and 3.7%, respectively). The cross-reaction for the other tested naturally occurring and synthetic steroids did not exceed 1%. Regression analysis of cortisol concentration estimates obtained on 20 samples before and after Sephadex LH-20 column chromatography gave a coefficient of correlation (r) of 0.995 and a regression coefficient (b) of 1.04. Recovery of cortisol added to plasma samples was quantitative. The intra-assay error was 8.5% and the inter-assay error averaged 5.7%. The method is simple requiring a single solvent extraction of plasma, therefore permitting large numbers of samples to be handled efficiently by a single technician.

Male-female differences in effects of parental absence on glucocorticoid stress response.

This study examines the family environments and hormone profiles of 316 individuals aged 2 months-58 years residing in a rural village on the east coast of Dominica, a former British colony in the West Indies. Fieldwork was conducted over an eight-year period (1988-1995). Research methods and techniques include radioimmunoassay of cortisol and testosterone from saliva samples (N=22,340), residence histories, behavioral observations of family interactions, extensive ethnographic interview and participant observation, psychological questionnaires, and medical examinations.Analyses of data indicate complex, sex-specific effects of family environment on endocrine function. Male endocrine profiles exhibit greater sensitivity to presence of father than do female endocrine profiles. Father-absent males tend to have (a) low cortisol levels during infancy, (b) high or abnormal cortisol profiles during childhood and adolescence, and (c) high cortisol and low testosterone levels during adulthood compared with those of males raised with a resident father. These results indicate that early family environment has significant effects on endocrine response throughout male life histories.

The delivery of testosterone and dihydrotestosterone by ALCAP ceramic implants in rats.

The objectives of this investigation were to evaluate the release of testosterone (T) and dihydrotestosterone (DHT) from nonimpregnated and polylactic acid (PLA) impregnated ALCAP ceramic reservoirs implanted in rats, and to study the effects of delivered androgens on the reproductive system of male rats. A total of 120 Sprague-Dawley albino male rats were distributed equally into three groups Two ALCAP capsules, one nonimpregnated and the other impregnated with PLA, were implanted into each rat in groups I and II. Capsules implanted into group I rats were loaded with a mixture of 20 mg T and 20 mg DHT. Group II rats were implanted with two empty capsules (sham group), and group III animals served as unimplanted controls. Eight rats from each group were euthanized at the end of 3, 6, 9, and 12 months following the implantation of the ceramics. No significant change in the weights of vital organs of rats was observed among any of the three different groups. Vas deferens and epididymal fluid were devoid of normal spermatozoa within 3 months of implanting the steroid-containing ceramics. Testicular weights decreased significantly in the rats implanted with ALCAP containing steroids and the seminiferous tubules became oligospermic after 1 month and azoospermic after 3 months. The data collected in this study suggest that (1) ALCAP ceramic capsules are capable of delivering T and DHT in combined form, and (2) T and DHT delivered by ALCAP capsules can be used effectively to regulate spermatogenesis in rats.

Changes in pelvic conformation and peripheral estrone concentrations in pre- and post-partum beef cows.

An experiment was conducted to investigate the temporal relationship of peripheral estrone (E1) concentration to changes in the size of the pelvic opening preceding and immediately following parturition. Twenty-six multiparous beef cows were observed from approximately 50 d prepartum to 7 d postpartum. Blood samples were collected at 7 d intervals preceding calving and at 1, 3 and 7 d following for E1 quantitation. Estimates of pelvic opening area were made at the time of blood sampling. Peripheral E1 concentrations were elevated beginning at approximately 25 d prepartum. Dams bearing male fetuses had greater (P less than 0.01) concentrations of E1 than did dams with female fetuses. Calf birth weight was correlated (r = 0.44, P less than 0.01) with E1 levels from 10 d prepartum through parturition. Postpartum pelvic area was greater for cows giving birth to male calves, with no significant differences for calf birth weights by sex. Correlations were observed between E1 concentration, and pelvic area measured from 50 d prepartum to 7 d postpartum (r = 0.26, P less than 0.01), 10 d prepartum to calving (r = 0.42, P less than 0.01), and from calving to 7 d postpartum (r = 0.33, P less than 0.01). Percentage increase in E1 concentration from 50 d prepartum to calving was significantly correlated (r = 0.75, P less than 0.01) to percentage pelvic area increase over the same period. A correlation also exists between maternal E1 concentrations and fetal sex and pelvic area. In summary, the increased estrogen concentrations in cows with male calves may facilitate pelvic spread, resulting in a larger pelvic opening.

Ovarian function in the cycling cow: relationship between gonadotropin binding to theca and granulosa and steroidogenesis in individual follicles.

Thirty-six Angus and Angus crossbred cows were used in a 3 x 2 x 2 factorial experiment designed to determine the effects of stage of the estrous cycle (day 6, 12 or 18), a long term with a single ovary (2 years) and side of previous ovulation or remaining ovary on the development of large ovarian follicles (LF; diameter greater than or equal to 8 mm). Follicular fluid estradiol-17 beta (FFE) concentration, localization of gonadotropin binding sites and granulosa cell condition were determined for each LF by radioimmunoassay, autoradiography and histological examination, respectively. No side differences were noted. However, the sample was biased by the use of equal numbers of left and right side ovulators. Compared to intact controls, one-ovary cows showed 100% compensation of LF development. Eighty-five percent of the LF studied bound follicle stimulating hormone (FSH) in the granulosa and human chorionic gonadotropin (hCG) in the theca. Thirty-eight percent also bound hCG in the granulosa. Both binding patterns were found on all days studied. Atretic follicles had lower (P less than .05) FFE concentrations than healthy follicles (r = -.59). The number of LF per cow increased (P less than .05) from day 6 (1.3) to days 12 (1.8) and 18 (2.1). Conversely, their FFE content per cow decreased (P less than .05) from 109 ng/cow on day 6 to 20 ng on; day 12 and 34 ng on day 18. The data suggested the emergency of two LF classes on day 18, one preovulatory, with exceptionally high FFE levels, the other atretic or becoming atretic, with low FFE levels.

Effects of high altitude on plasma concentrations of testosterone and pituitary gonadotropins in man.

Ten healthy male volunteers were rapidly taken to the top of Pike's Peak, altitude 14,100 ft (4,300 m) where they remained at the research station for 6 d. There were no significant changes, compared to their values at sea-level, in the plasma concentrations of testosterone, FSH, and LH during this period. There was a highly significant direct correlation between individual changes in testosterone and the gonadotropins on Day 1. Our data provide no evidence for an acute effect of high-altitude per se on the secretion of testosterone and the pituitary gonadotropins.