Effects of 30 day simulated microgravity and recovery on fluid homeostasis and renal function in the rat

Transition from a normal gravitational environment to that of microgravity eventually results in decreased plasma and blood volumes, increasing with duration of exposure to microgravity. This loss of vascular fluid is presumably due to negative fluid and electrolyte balance and most likely contributes to the orthostatic intolerance associated with the return to gravity. The decrease in plasma volume is presumed to be a reflection of a concurrent decrease in extracellular fluid volume with maintenance of normal plasma-interstitial fluid balance. In addition, the specific alterations in renal function contributing to these changes in fluid and electrolyte homeostasis are potentially responding to neuro-humoral signals that are not consistent with systemic fluid volume status. We have previously demonstrated an early increase in both glomerular filtration rate and extracellular fluid volume and that this decreases towards control values by 7 days of simulated microgravity. However, longer duration studies relating these changes to plasma volume alterations and the response to return to orthostasis have not been fully addressed. Male Wistar rats were chronically cannulated, submitted to 30 days head-down tilt (HDT) and followed for 7 days after return to orthostasis from HDT. Measurements of renal function and extracellular and blood volumes were performed in the awake rat.     

Inhibition of MAO-B by (−)-deprenyl alters dopamine metabolism in the macaque (Macaca facicularis) brain

The present study has examined whether MAO-B has a role in DA metabolism in the primate CNS in situ. Eleven macaques (macaca facicularis) were used in this study to examine the effects of (-)-deprenyl (1 mg/kg, i.v., 2 and 24 hours). (-)-Deprenyl administration completely and selectively blocked MAO-B activity and blocked DA metabolism in the caudate nucleus and frontal cortex. DA metabolism in the substantia nigra was not affected by MAO-B inhibition. Changes in DA metabolism were accompanied by changes in 5-hydroxytryptamine (5HT) turnover: 5-hydroxyindole acetic acid (5HIAA) levels increased in the caudate and decreased in the frontal cortex. Levels of 2-phenylethylamine (PE), a putative modulator of dopaminergic transmission, were increased by MAO-B inhibition in all three brain regions examined. It is concluded that in some regions of the primate brain, in contrast to the rat, MAO-B has an important role in DA metabolism.     

The Presence of Clitoromegaly in the Nonclassical Form of 21-Hydroxylase Deficiency Could Be Partially Modulated by the CAG Polymorphic Tract of the Androgen Receptor Gene

Background

In the nonclassical form (NC), good correlation has been observed between genotypes and 17OH-progesterone (17-OHP) levels. However, this correlation was not identified with regard to the severity of hyperandrogenic manifestations, which could depend on interindividual variability in peripheral androgen sensitivity. Androgen action is modulated by the polymorphic CAG tract (nCAG) of the androgen receptor (AR) gene and by polymorphisms in 5α-reductase type 2 (SRD5A2) enzyme, both of which are involved in the severity of hyperandrogenic disorders.

Objectives

To analyze whether nCAG-AR and SRD5A2 polymorphisms influence the severity of the nonclassical phenotype.

Patients

NC patients (n = 114) diagnosed by stimulated-17OHP ≥10 ng/mL were divided into groups according to the beginning of hyperandrogenic manifestations (pediatric and adolescent/adult) and CYP21A2 genotypes (C/C: homozygosis for mild mutations; A/C: compound heterozygosis for severe/mild mutations).

Methods

CYP21A2 mutations were screened by allelic-specific PCR, MLPA and/or sequencing. HpaII-digested and HpaII-undigested DNA samples underwent GeneScan analysis to study nCAG, and the SRD5A2 polymorphisms were screened by RLFP.

Results

Mean nCAG did not differ among pediatric, adolescent/adult and asymptomatic subjects. In the C/C genotype, we observed a significantly lower frequency of longer CAG alleles in pediatric patients than in adolescent/adults (p = 0.01). In patients carrying the A/C genotype, the frequencies of shorter and longer CAG alleles did not differ between pediatric patients and adolescent/adults (p>0.05). Patients with clitoromegaly had significantly lower weighted CAG biallelic mean than those without it: 19.1±2.7 and 21.6±2.5, respectively (p = 0.007), independent of the CYP21A2 genotype''s severity. The SRD5A2 polymorphisms were not associated with the variability of hyperandrogenic NC phenotypes.

Conclusions

In this series, we observed a modulatory effect of the CAG-AR tract on clinical manifestations of the NC form. Although the NC form is a monogenic disorder, our preliminary data suggested that the interindividual variability of the hyperandrogenic phenotype could arise from polygenic interactions.     

Salmonellae as an Index of Pollution of Surface Waters

Screening enrichments of surface water specimens by means of a polyvalent fluorescent antibody reagent for the salmonellae yielded approximately 60% more positive specimens than was obtained by cultural procedures. It is not known what fraction of the excess of fluorescent antibody-positive over culturally positive specimens represents staining of non-salmonellae or non-arizonae as opposed to the staining of non-cultivatable organisms of these two genera. Cotton gauze and rayon-polypropylene fiber swabs were equally sensitive for collecting salmonellae from the streams examined. Tetrathionate enrichment incubated at 41.5 C appeared to be superior to selenite-cystine for isolation of salmonellae from surface waters. Twenty-eight serotypes of Salmonella and two serotypes of Arizona were identified in the 121 positive specimens. In water rated moderately polluted, 65% of all specimens tested were positive; in minimally polluted waters, 38% were positive; and in unpolluted streams, 44% were positive.     

Male pseudohermaphroditism due to nonsalt-losing 3 beta-hydroxysteroid dehydrogenase deficiency: gender role change and absence of gynecomastia at puberty

Adrenal and gonadal functions were evaluated on two adult cousins with male pseudohermaphroditism due to congenital 3 beta-hydroxysteroid dehydrogenase deficiency (3 beta-HSD) without clinical salt-losing. Both patients had been reared as females since birth. Case 1 presented at age 17 with perineal hypospadias virilization without gynecomastia and a female to male gender role change at puberty. Case 2 had previously undergone bilateral orchidectomy in childhood and presented "primary amenorrhea", absence of virilization and a female gender role at the age of 24. In the basal state, as well as after ACTH and hCG stimulation, 3 beta-hydroxy-5-ene-steroid levels were disproportionately elevated, resulting in abnormal 3 beta-hydroxy-5-ene: 3-oxi-4-ene steroids ratios. Normal basal serum cortisol with inadequate cortisol response to ACTH was observed in both patients. Elevated basal plasma renin activity (PRA) and normal basal serum aldosterone (ALDO) were present in both subjects. After ACTH stimulation serum ALDO rose adequately in Case 1 but subnormally in Case 2. Salt restriction resulted in an increase in serum ALDO and no salt loss in Case 1 whereas in Case 2 the substantial rise in PRA and serum ALDO were unable to prevent slight urinary sodium loss. Case 1 had normal basal serum testosterone with subnormal response to hCG stimulation. Incubation of testicular tissue in vitro with [3H]DHEA resulted in large Androstenediol production but diminished testosterone conversion confirming the 3 beta-HSD deficiency in the testes. We conclude that (1) absence of gynecomastia and a female to male gender role change may be observed in the male pubertal presentation of nonsalt-losing 3 beta-HSD deficiency and (2) the different functional behavior of zona glomerulosa in our patients suggests the presence of variable degrees of 3 beta-HSD deficiency in the zona glomerulosa of the nonsalt-losing form.     

Induction of sexual receptivity in the female broad-headed skink, Eumeces laticeps, by estradiol-17 beta

Sexual receptivity in the female scincid lizard Eumeces laticeps occurs naturally only during the spring breeding season, which is also when maximal follicular development occurs. The presumption that high estrogen levels are coincidentally present and the need for a reliable method of inducing sexual receptivity for behavioral studies prompted tests of the hypothesis that estrogen induces sexual behavior. A series of experiments established that estradiol-17 beta induces sexual behavior. A series of experiments established that estradiol-17 beta induces sexual receptivity within 4 days when injected every other day at 2.0 micrograms in 20 microliters peanut oil in intact or ovariectomized females. In behavioral tests conducted during August, all control females (intact or ovariectomized injected with vehicle only) rejected courtship whereas all females receiving estrogen copulated. Estrogen injections also induced a statistically significant change from rejection to receptivity within individuals. Initial attempts to implant estradiol-17 beta in Silastic tubes killed all females so treated.     

Current Status of Immunofluorescence Techniques for Rapid Detection of Shigellae in Fecal Specimens

Polyvalent Shigella conjugates were prepared for Shigella groups A, B, C, and D. After preliminary testing with pure cultures of both homologous and heterologous organisms, these reagents were used in three evaluation studies. Fecal specimens from patients hospitalized with diarrhea, from children involved in an institutional outbreak of dysentery due to S. sonnei, and from patients with diarrhea in Arizona were screened by fluorescent-antibody (FA) tests and were cultured. Specimens were examined at various periods of time after collection and after incubation in broth and saline. Results showed that shigellae were detected most frequently when specimens were cultured immediately after collection. FA tests revealed more positive results when the specimens were incubated in either saline or broth than when they were examined immediately after collection. The S. sonnei conjugate gave the most reliable results of any of the Shigella FA reagents used in these investigations. It proved to be both sensitive and specific.     

Recovery from potentially lethal damage and recruitment time of noncycling clonogenic cells in 9L confluent monolayers and spheroids

Cells that have been grown as multicell tumor spheroids exhibit radioresistance compared to the same cells grown in monolayers. Comparison of potentially lethal damage (PLD) repair and its kinetics was made between 9L cells grown as spheroids and confluent monolayers. Survival curves of cells plated immediately after irradiation showed the typical radioresistance associated with spheroid culture compared to plateau-phase monolayers. The dose-modification factor for spheroid cell survival is 1.44. Postirradiation incubations in normal phosphate-buffered saline (PBS), conditioned media, or 0.5 M NaCl in PBS reduced the differences in radiosensitivity between the two culture conditions. Postirradiation treatment in PBS or conditioned medium promoted repair of potentially lethal damage, and 0.5 M NaCl prevented the removal of PLD and allowed the fixation of damage resulting in lower survival. Survival of spheroid and monolayer cells after hypertonic NaCl treatment was identical. NaCl treatment reduced Do more than it did the shoulder (Dq) of the survival curve. PLD repair kinetics measured after postirradiation incubation in PBS followed by hypertonic NaCl treatment was the same for spheroids and for plateau-phase monolayers. The kinetics of PLD repair indicates a biphasic phenomenon. There is an initial fast component with a repair half-time of 7.9 min and a slow component with a repair half-time of 56.6 min. Most of the damage (59%) is repaired slowly. Since the repair capacity and kinetics are the same for spheroids and monolayers, the radioresistance of spheroids cannot be explained on this basis. Evidence indicates that the time to return from a Go (noncycling G1 cells) state to a proliferative state (recruitment) for cells from confluent monolayers and from spheroids after dissociation by protease treatment may be the most important determinant of the degree of PLD repair that occurs. Growth curves and flow cytometry cell cycle analysis indicate that spheroid cells have a lag period for reentry into a proliferative state. Since plating efficiency remains high and unchanging during this period, one cannot account for the delay on the basis of the existence of a large fraction of Go cells which are not potentially clonogenic. The cell cycle progression begins in 6-8 h for monolayer cells and in 14-15 h for spheroids. It is hypothesized that the slower reentry of spheroid cells into a cycling phase allows more time for repair than for the rapidly proliferating monolayer cells.     

Somatomedin levels in the diagnosis and therapy of growth hormone deficiency

Serum somatomedin-C (SM-C) and somatomedin (SM) concentrations were measured by, respectively, radioimmuno (SM-C RIA) and radioreceptor assays (SM RRA) in 3 groups of children with short stature. The patient population was different from previously reported series in that it was urban Brazilian, low income, and significantly older. Group A consisted of 6 male and 3 female children, aged 7.7-16.0 years, whose average peak plasma immunoreactive growth hormone (GH) was above 10 ng/ml. Group B contained 8 male and 5 female untreated GH-deficient patients, ranging in age from 9.5 to 21.0 years. In Group C there were 4 male and 1 female GH-deficient subjects treated with I.M. injections of GH (0.1 U/kg) from 1 month to 7 years. The mean +/- SE basal RIA SM-C (ng/ml) concentrations were significantly lower in groups B (34.2 +/- 8.8) and C (43.8 +/- 13.7) than A (214.3 +/- 42.7): A X B, P less than 0.001 and A X C, P less than 0.02. Likewise the mean +/- SE basal RRA SM (ng/ml) concentrations were significantly lower in groups B (78.9 +/- 17.6) and C (90.8 +/- 19.3) than group A (316.3 +/- 43.0): A X B, P less than 0.001 and A X C, P less than 0.002. A significant linear correlation was observed between RIA and RRA in group B (r = 0.84; P less than 0.001) and C (r = 0.96; P less than 0.01), but not for A (r = 0.61; P greater than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Plating efficiency as a function of time postirradiation: evidence for the delayed expression of lethal mutations

The plating efficiency (PE) of a gamma-irradiated (7 Gy) human cell hybrid line (HeLa X skin fibroblast, designated as CGL1) has been measured as a function of time postirradiation and compared to that of unirradiated cells at similar cell densities and under the same growth conditions. The results indicate that following irradiation, the PE of the irradiated cells initially increases but never returns to that of unirradiated cells during the experimental period that we have examined. Furthermore, after a period of 9 to 10 days (equivalent to at least 10 cell doublings) postirradiation and plating, the PE of the irradiated cells begins to decrease and continues to do so over the next 5 days. A decrease does not occur in unirradiated cells until much later (i.e., Day 15) corresponding to at least 5 additional cell doublings. The data are discussed in terms of a delayed expression of lethal mutations. The possible impact of these observations on the estimation of radiation-induced transformation frequencies is also considered.