DNA content of Ovis musimon spermatozoa.

To our knowledge, the value of the haploid DNA content (C-value) of Ovis musimon (mouflon) has not been previously published. Therefore, the aim of the present work was to determine the C-value and the nuclear area of O. musimon sperm cells and compare both parameters with those of Ovis aries. Feulgen reaction, which is specific and stoichiometric for DNA, was carried out on semen smears. The C-value and sperm nuclear area were determined using microspectrophotometry and Gallus domesticus erythrocytes as standard species. The C-value of O. musimon was 3.02 ± 0.04 pg, and the sperm nuclear area was 23.92 ± 0.89 µm(2). The C-value and the sperm nuclear area of O. aries were 3.07 ± 0.03 pg and 22.98 ± 0.86 µm(2) respectively. The O. musimon C-value was not significantly different (P > 0.05) from that of O. aries, indicating that both species may have a very close phylogenetic relation.

Equine sperm nuclei with different ploidy levels: relationship between the nuclear DNA content and the nuclear area.

The aims of this study were to estimate the ability of the Feulgen reaction to identify equine sperm nuclei with different ploidy levels, to determine the frequency of haploid, diploid and polyploid sperm nuclei in the semen of fertile equines and to evaluate the relationship between the nuclear DNA content and the nuclear area. Determination of the ploidy level of Feulgen-stained spermatozoa using a scanning microspectrophotometer was very similar to the subjective estimations made with a light microscope. This indicates that the Feulgen reaction is a simple, inexpensive and reliable technique to recognise the ploidy level of equine spermatozoa. The incidence of diploid and polyploid spermatozoa, determined with a light microscope in 11 fertile equines, was 0.17 ± 0.08% and 0.027 ± 0.027% respectively. DNA content values obtained by microspectrophotometry in the only equine that presented polyploid spermatozoa allowed us to discriminate between haploid, diploid and polyploid subpopulations. Measurement of the nuclear area discriminated only two subpopulations: one including the haploid and diploid subpopulations and the other including the polyploid one. The similarity between the area of the haploid and diploid sperm nuclei suggests that the increase in DNA content is anisotropic, with a privileged direction of growth perpendicular to the nuclear flattening plane.

Discovery of CGS 27023A, a non-peptidic, potent, and orally active stromelysin inhibitor that blocks cartilage degradation in rabbits.

Structure-activity relationships of a lead hydroxamic acid inhibitor of recombinant human stromelysin were systematically defined by taking advantage of a concise synthesis that allowed diverse functionality to be explored at each position in a template. An ex vivo rat model and an in vivo rabbit model of stromelysin-induced cartilage degradation were used to further optimize these analogs for oral activity and duration of action. The culmination of these modifications resulted in CGS 27023A, a potent, orally active stromelysin inhibitor that blocks the erosion of cartilage matrix.

Acute myocardial infarction shortly after a normal exercise stress test. Case reports.

The authors describe 3 cases of AMI occurring shortly after a negative bicycle ergometer stress test. These cases represent an unfortunate but extremely rare complication of a relatively safe diagnostic procedure. The authors also focus on the pathogenesis of the ischemic event, which may be attributed either to intraplaque hemorrhage or to platelet aggregation, both exercise-induced. The prevalence of AMI in this paper (0.06%) is similar to the data described in literature.

Oxcarbazepine adjunctive therapy in infants and young children with partial seizures.

OBJECTIVE: To evaluate the efficacy, safety, and pharmacokinetics of oxcarbazepine as adjunctive therapy in infants and young children (1 month to < 4 years). METHODS: Children 1 month to < 4 years of age with inadequately controlled partial seizures taking up to two concomitant antiepileptic drugs (AEDs) were enrolled in this rater-blind, randomized, parallel-group study. Patients received either high-dose (60 mg/kg/day) or low-dose (10 mg/kg/day) oxcarbazepine as oral suspension. The primary efficacy variable was the absolute change in electrographic partial seizures with a behavioral correlate (type 1 seizure) frequency per 24 hours during the last 72 hours of continuous video-EEG monitoring in the treatment phase compared with baseline seizure frequency. RESULTS: Of 191 patients screened, 128 were randomized: 64 to both oxcarbazepine dose groups. The median absolute change in type 1 seizure frequency per 24 hours was more effective for the high-dose group (-2.00) compared with the low-dose group (-1.37; p = 0.043). The median percentage reduction in type 1 seizure frequency per 24 hours was also greater in the high-dose group (83.33%) than in the low-dose group (46.18%; p = 0.047). The most frequent adverse events (> or = 10%) were somnolence and pyrexia, and most were mild in severity. CONCLUSIONS: In this study, high-dose oxcarbazepine was significantly more effective than low-dose oxcarbazepine in controlling partial seizures in infants and very young children.

A comparison of chondrocyte proteoglycan metabolism in monolayer and agarose cultures.

Bovine chondrocyte cultures were established in agarose and in monolayers to compare the effects of cytokines and drugs on matrix metabolism. The production of sulfated glycosaminoglycans (S-GAG) from the medium and cell surface compartments were measured by a dimethylmethylene blue assay. In the agarose cultures most of the proteoglycan remained in the agar, but was continuously released into the medium for more than 50 days. In the monolayers, the cell surface compartment became saturated with S-GAG in 5-6 days. Then a time-dependent decrease of accumulation occurred in the medium after 8-10 days. The anabolic effects of insulin-like growth factor (IGF) and a protein kinase C activator (PMA) were measured in these cultures. IGF and PMA increased S-GAG accumulation in the medium from monolayers but not from agarose cultures. In the agarose cultures, S-GAG was released into the medium after these cultures were changed to serum-free test conditions. This release overshadowed any increase in S-GAG synthesis. The catabolic effect of IL-1 was more evident in the monolayers than in the agarose cultures. Agarose cultures maintain the chondrocyte phenotype longer than monolayers but for initial drug studies monolayer cultures appear to be more appropriate.

Thymic stromal cells in culture. I. Establishment and characterization of a line which is cytotoxic for normal thymocytes and produces hematopoietic growth factor(s).

Lines of thymic stromal cells have been established. One of these, designated TS-9, has been cloned and studied extensively. This line expresses both acid and alkaline phosphatases. Despite repeated cloning, TS-9 cells remain morphologically heterogeneous. The origin of these cells is not clear. They express low levels of immunologically identifiable cytokeratins, produce laminin, a basement membrane protein, but express antigens typically found on bone marrow stromal cells. The TS-9 cells are MHC Class I+ but Class II-. They express the Thy-1, Pgp-1, and Mac-2 antigens but not other lineage markers of T cells or macrophages. Coculturing TSC with normal thymocytes or with the CTLL-1 cell line leads to a profound inhibition of lectin-induced and/or IL-2 induced T cell proliferation. This requires direct cell-cell contact and ultimately results in the death of the bound lymphocytes. It cannot be reproduced by culturing the thymocytes with TSC culture supernatants. These supernatants do contain hematopoietic growth factor(s) which augment the growth of some T lineage cells and support the growth of monocytic colonies in semi-solid culture medium. Both normal thymocytes and a variety of T cell tumors bind to TSC but only the normal cells are killed as a consequence of this interaction. Neither the binding nor the killing appear to be MHC restricted. We suggest that this killing may provide a model for the effector mechanism of the negative selection imposed by the thymus on developing T cells.

Distribution of 18+28S ribosomal genes in mammalian genomes.

In situ hybridization with 3H 18S and 28S ribosomal RNA from Xenopus laevis has been used to study the distribution of DNA sequences coding for these RNAs (the nucleolus organizing regions) in the genomes of six mammals. Several patterns of distribution have been found: 1) A single major site (rat kangaroo, Seba's fruit bat), 2) Two major sites (Indian muntjac), 3) Multiple sites in centromeric heterochromatin (field vole), 4) Multiple sites in heterochromatic short arms (Peromyscus eremicus), 5) Multiple sites in telomeric regions (Chinese hamster). - The chromosomal sites which bind 3H 18S and 28S ribosomal RNA correspond closely to the sites of secondary constrictions where these are known. However, the correlation is not absolute. Some secondary constrictions do not appear to bind 3H ribosomal RNA. Some regions which bind ribosomal RNA do not appear as secondary constrictions in metaphase chromosomes. - Although the nucleolus organizing regions of most mammalian karyotypes are found on the autosomes, the X chromosomes in Carollia perspicillata and C. castanea carry large clusters of sequences complementary to ribosomal RNA. In situ hybridization shows that the Y chromosome in C. castanea also has a large nucleolus organizing region.

Release of cell surface proteoglycan from chondrocytes by interleukin-1.

A cell culture model was developed to assess factors that can protect cartilage from recombinant human interleukin-1 alpha (IL-1)-induced breakdown. IL-1 (0.01-10 ng/ml) caused a dose-dependent release of cell surface proteoglycan into the medium of bovine and rabbit articular chondrocytes. Sulfated glycosaminoglycan (S-GAG) was detected in the medium and on the cell surface by a dimethlymethylene blue assay. The redistribution of the cell surface S-GAG to the medium compartment required protein synthesis because it was inhibited by cycloheximide and was time-dependent (> 5 h). Although the release was most likely due to de novo synthesis of proteases, standard protease inhibitors failed to prevent the release even when used in combination. TGF-beta and IGF-I increased the amount of S-GAG in both the medium and cell surface compartments, but did not protect from IL-1-induced release. This method has an advantage over a cartilage model of IL-1-induced matrix release because the drug and cytokine exposure time is reduced and the variability is less.

Chromatin cytophotometric analysis of abnormal bovine spermatozoa.

Sperm head morphology is basically conditioned by the nuclear structure. The aim of the present work was to study the relation between nuclear morphological features, DNA content and chromatin distribution in morphologically normal vs. abnormal bovine spermatozoa. To this end, individual Feulgen-reacted spermatozoa were cytophotometrically studied. Chromatin compactation was evaluated by means of nuclear area, as well as mean and maximal absorbance of each nucleus. Morphological abnormality analysed included large, small, pear, narrow and round shapes, together with presumably 'diploid' sperms. Both large and small spermatozoa have a DNA content that does not differ significantly from normal values, but their area and mean and maximal absorbance are significantly different. Size variation seems basically due to altered chromatin compactation. The pear shapes have a narrower neck and a significant increase in maximal absorbance alone, which is invariably recorded in the neck zone whose increase would indicate a change in distribution and/or compactation. The narrow and round shapes fail to present significant variations in studied parameters. The possible 'diploids' differ significantly from normal cells in all studied variables, with a little area increase.