Altered morphogenesis of the immature hamster uterus following neonatal exposure to diethylstilbestrol

Abstract. In previous studies, we found that a single neonatal exposure to diethylstilbestrol (DES) resulted in severe hyperplasia and a high incidence of endometrial adenocarcinoma in the uterus of adult hamsters. These observations prompted us to investigate the consequences of DES exposure on earlier stages of uterine morphogenesis. After neonates were treated within 6 h of birth (day 1) with 100 μg of DES or oil vehicle, uterine tissue morphometry plus cell labelling indices following in vivo pulse labeling with [³H]thymidine were determined on days 3–21 of life. The sequential findings were: (1) a precocious (day 3) burst of cellular proliferation throughout the uterus, (2) an early period (days 3–9) of hypertrophy and increased cell density in the luminal epithelium, (3) an extreme acceleration of uterine growth resulting in a persistent increase in total uterine mass (threefold enhancement on days 5–21), (4) precocious development of endometrial glands (day 9) that were sites of intense but transient proliferative activity, (5) a middle period (days 9–15) when the percentage of stromal cells engaged in proliferative activity was reduced, (6) a second wave (days 15–21) of enhanced proliferative activity in the luminal epithelium, and (7) later development (day 21) of reduced cell density in the uterine stroma, apparently due to increased intercellular collagen accumulation. These results support our working hypothesis that the acute uterotropic response to neonatal DES treatment initiates a change in the developing hamster uterus, and later estrogenic stimulation promotes neoplastic progression in the DES-altered adult organ, perhaps due to disruption of stromal-epithelial interactions.     

A modified conductance medium for the detection of Salmonella spp

Selenite-cystine/trimethylamine oxide/dulcitol medium has been used in conjunction with conductance instruments to detect the presence of Salmonella spp. in foods and faeces. However, a small but significant number of salmonella strains were missed by this method. The majority of these strains were detected when dulcitol was substituted by mannitol and tested on two separate Malthus conductance instruments. Some strains of Citrobacter freundii and Escherichia coli continued to give false positive results. Attempts are made to explain why the substitution of mannitol for dulcitol gives an improved medium.     

Determination of the enantiomeric composition of (R*,S*)-1-ethylpropyl 2-methyl-3-hydroxypentanoate, the male-produced aggregation pheromone of Sitophilus granarius

The enantiomeric composition of sitophilate, the granary weevil [Sitophilus granarius (L.)] male-produced aggregation pheromone [(R^*,S^*)-1-ethylpropyl 2-methyl-3-hydroxypentanoate)], was determined by three methods: (1) bioassaying the synthetic (2S,3R) and (2R,3S) enantiomers of the active (R^*,S^*) diastereomer; (2) ¹H NMR spectroscopy of Mosher ester derivatives of the natural pheromone and synthetic (2S,3R)-and (2R,3S)-sitophilate; and (3) capillary GLC comparisons of the retention times of derivatized natural pheromone and the two synthetic enantiomers. The combined methods confirmed the (2S,3R) enantiomer as the active form of sitophilate. Male granary weevils were shown to produce >96% (2S,3R)-sitophilate. No significant attraction of S. granarius by the (2R,3S) enantiomer was observed. Rice and maize weevils [S. oryzae (L.) and S. zeamais Motschulsky] were not attracted by (2S,3R)-sitophilate. S. granarius L. est un déprédateur important des grains stockés. Le (R^*,S^*)-1-éthylpropyl 2-méthyl-3-hydroxypentanoate a été identifié en 1987 comme le principal composé du sitophilate, la phéromone mâle d'agrégation de S. granarius. La composition énantiométrique du sitophilate a été déterminée par 3 méthodes:

Advantages of firefly luciferase as a reporter gene: application to the interleukin-2 gene promoter

The chloramphenicol acetyltransferase (CAT) gene is widely used in recombinant constructs employed to study promoter and enhancer control of gene expression. However, CAT-based assays require a laborious, multi-step procedure for quantitation of promoter activity. We have applied the recently described firefly luciferase (LUC) reporter gene to the study of the interleukin-2 (IL2) promoter and have further defined the properties of this reporter gene system. We find that IL2-LUC constructs have multiple advantages over IL2-CAT constructs. The LUC assay is highly sensitive and requires 1/10 the cells used in the CAT system. A final quantitative measure of promoter activity can be obtained within 25 h following transfection with IL2-LUC, compared to 108-160 h with IL2-CAT. Light emission significantly (fourfold) above background is detectable 3 h after induction in a direct assay of extracts from transfected cells. We have described the variability of the assay, the minimum number of transfected cells required to detect light, the stability of luciferase in cell extracts, the effect of Triton X-100 on the assay, and a rapid cell lysis procedure. The luciferase system is a simple, rapid, and sensitive method for the study of promoter activity in transfected cells, particularly for weakly expressed genes such as IL2 which give low activity in the CAT assay.     

Quantitative assessment of various fixatives on nuclear and cytoplasmic areas in oral cytology

Summary Both nuclear and cytoplasmic areas are parameters known to be of significance in the diagnosis of malignancy. However, few studies have assessed the effect of fixation on exfoliative cytology and none has looked at such influences upon oral smears. Hence the method of fixation may influence directly diagnostic cytology. The effect of three methods of fixation upon the nuclear and cytoplasmic areas of cells removed from the buccal mucosa was quantitatively assessed. The three methods employed, prior to Papanicolaou staining, were: direct immersion in diethylether and ethanol (11 v/v), spray fixation (Vale Smear Fix) and air drying. Three smears from each of 21 patients were used, each slide being allocated randomly a, method of fixation. After 24h all smears were processed for Papanicolaou's stain.The nuclear and cytoplasmic areas were calculated using semi-automated image analysis. No significant differences were found in the two areas whichever method of fixation was used.     

Nasal trigeminal responses to toluene presented by an automated delivery system

An apparatus was developed which permits the automated deliveryof volatile chemical stimuli for use in neurophysiology experiments.A computer-controlled olfactometer, incorporating electronicmass flow controllers (EMFCs) and Teflon-lined solenoid valves,generated and delivered clean or odorized air. Neural and respiratorysignals from the animal were amplified and stored, along withtrial information (e.g. odorant concentration) on a chart recorderand video cassette recorder, both of which were controlled bythe computer. This apparatus was used to measure responses totoluene from the rat ethmoid nerve, a part of the ophthalmicdivision of the trigeminal nerve. Multi-unit responses to thiscompound were first observed at 2000 p.p.m. The magnitude ofthe response increased linearly with logarithmic increases inconcentration up to vapour saturation. Changes in respirationin response to toluene also were observed, although neural responsesoften were seen in the absence of respiratory changes.     

The replicative origin of the E. coli chromosome binds to cell membranes only when hemimethylated

DNA from the E. coli replicative origin binds with high affinity to outer membrane preparations. Specific binding regions are contained within a 463 bp stretch of origin DNA between positions -46 and +417 on the oriC map. This region of DNA contains an unusually high number of GATC sites, the recognition sequence for the E. coli DNA adenine methylase. We show here that oriC DNA binds to membrane only when it is hemimethylated. The E. coli chromosomal origin is hemimethylated for 8-10 min after initiation of replication, and origin DNA binds to membranes only during this time period. Based on these results, we propose a speculative model for chromosome segregation in E. coli.     

Polyamines Differentially Inhibit Cyclic AMP-Dependent Protein Kinase-Mediated Phosphorylation in the Brain of the Tobacco Hornworm, Manduca sexta

The effects of the naturally occurring polyamines spermine and spermidine on phosphorylation promoted by cyclic AMP (cAMP)-dependent protein kinase (PK) (cAMP-PK; EC 2.7.1.37) were studied using the brain of the tobacco hornworm, Manduca sexta. Four particulate-associated peptides (280, 34, 21, and 19 kilodaltons) in day 1 pupal brains are endogenous substrates for a particulate type II cAMP-PK. These phosphoproteins are present in brain synaptosomal, as well as microsomal, particulate fractions but are not present in the cytosol. They are distributed throughout the CNS and PNS and are present in several nonneuronal tissues as well. Phosphorylation of these proteins via cAMP-PK was inhibited markedly by micromolar concentrations of spermine and spermidine. Other particulate-associated peptides phosphorylated via a Ca2+/calmodulin-PK or Ca2+ and cAMP-independent PKs were unaffected by polyamines, whereas the phosphorylation of a 260-kilodalton peptide was markedly enhanced. Spermine did not exert its inhibitory effect indirectly by enhancement of cAMP or ATP hydrolysis or via proteolysis, but its action appears to involve a substrate-directed inhibition of cAMP-PK-promoted phosphorylation as well as enhanced dephosphorylation. Although addition of spermine resulted in marked ribosome aggregation in synaptosomal and microsomal particulate fractions, this phenomenon was not involved in the inhibition of cAMP-PK-promoted phosphorylation.     

Ultrastructure of the vegetative organisation and initial stages of silica plate deposition in the soil testate amoebaCorythion dubium

Summary The siliceous body plates ofCorythion dubium are bound by a band of organic cement which is thickest at the lateral margins. The anterior vacuolar cytoplasm is separated by a pigment zone, which forms a dark band in the mid-body region, from the compact posterior region containing a typical vesicular nucleus surrounded by a region of dense endoplasmic reticulum. A pellicular basket of microtubules surrounds the posterior cytoplasm. The large Golgi complex lies between the nucleus and the fundus. Numerous coated and uncoated vesicles from the Golgi cisternae are seen in the peripheral cytoplasm alongside developing plates. These small siliceous plates are enclosed in silicon deposition vesicles lying in surface ruffles of the plasmalemma, often in association with a pair of microtubules. Observations are made on the formation of these vesicles and the early stages of silica deposition. A comparison is drawn between silica deposition inC. dubium and choanoflagellates where there is a similar association between silicon deposition vesicles and microtubules.     

Karyotype of Saguinus labiatus labiatus (red-bellied marmosets)

The karyotype of Saguinus labiatus labiatus was determined by the Giemsa-banding technique on leukocytes cultured from 10 marmosets. The diploid chromosome number (2n = 46) was the same and the chromosome complement similar to other marmosets of genus Saguinus. Small karyotypic differences were found between S. l. labiatus and white-lipped marmosets (Saguinus fuscicollis) in the size of the X chromosome and in the banding pattern of one pair of metacentric chromosomes. A karyotypic variant was detected in 1 S. l. labiatus, characterized by a diploid chromosome number of 45 with balanced autosomal translocation involving two pairs of acrocentric chromosomes (T 16/19).