Fine structure of early human embryos frozen with 1,2 propanediol

Previous studies by a French group (Fertil Steril 44:645–651, 1985) have shown that two-to eight-cell human embryos can survive slow freeze-thawing with propanediol in a biological freezer. These embryos were assessed for morphological appearance by phase-contrast microscopy. We assessed the structure of 25 frozen-thawed one- to 12-cell embryos, obtained from our in vitro fertilization (IVF) and GIFT programmes, by phase-contrast and electron microscopy, using the same method of cryopreservation. One-fourth of the embryos examined had all cells intact, and more than one-half the embryos had over 50% of their cells well preserved. Some of these embryos had unequal blastomeres and cytoplasmic fragments. Ultrastructural assessment revealed good preservation of fine structure in the intact blastomeres of all embryos and maintenance of cell-to-cell contacts. Most cytoplasmic organelles, cell membranes, and nuclei were well preserved compared to nonfrozen controls. The cells that were cryoinjured showed varying degrees of disorganization of the cell membrane, cytosol, and cellular membranes, including swelling and disruption of the nuclear envelope. Disruption of the zona was somewhat rare. Small cytoplasmic fragments were less prone to cryoinjury than blastomeres. The use of propanediol for embryo cryopreservation seems to be feasible; frozen embryos with more than 50% cells intact have produced 10 pregnancies after embryo transfer (Fertil Steril 46:268–272, 1986). Replacement of 17 frozen embryos in seven patients has resulted in a twin pregnancy in Singapore. However, the effects of freezing on the mitotic spindles of embryonic cells need to be investigated further.     

Laboratory studies on salmonella-contaminated cheese involved in a major outbreak of gastroenteritis

A major outbreak of gastroenteritis was traced to Cheddar cheese contaminated with Salmonella typhimurium. There were no significant differences in pH values of the contaminated (mean pH 5.31) and non-contaminated (mean pH 5.39) cheese. The isolation rates of Salm. typhimurium were about the same when cheese samples were homogenized in lactose broth, lactose broth containing 1% Tween 80, or in aqueous 2% sodium citrate. Salmonella typhimurium was isolated regardless of preenrichment in lactose broth, but required selective enrichment in selenite cystine or tetrathionate brilliant green broth. There were no marked differences in the isolation rates obtained with different selective enrichment media, or after incubation at 36 degrees and 43 degrees C for 24 or 48 h. Contaminated samples of cheese failed to yield Salm. typhimurium consistently despite large and multiple samplings; samples from the interior of cheese blocks yielded positive results more frequently than the samples from the exterior. The number of Salm. typhimurium in factory sealed blocks as well as in samples obtained from the homes of known cases of salmonellosis was found to range from less than 3/100 g to 9/100 g of cheese. The infective dose of Salm. typhimurium in contaminated cheese was probably no greater than 10(4) organisms, and a rapid decline in numbers of Salm. typhimurium must have occurred subsequent to the outbreak.     

Laboratory studies on salmonella-contaminated cheese involved in a major outbreak of gastroenteritis

Dried biomass of Streptomyces cyaneus NCIB 9616 harvested at 1, 3, 7, 10 and 14 d from a liquid culture was degraded by alkaline methanolysis and the fatty acid methyl esters released examined quantitatively by gas chromatography. Isoprenoid quinones were extracted with petroleum ether and methanolic saline, purified by thin-layer chromatography and quantified by high performance liquid chromatography. Data for each component were plotted against time. The results showed that at the beginning of logarithmic growth there was a small quantitative shift in the fatty acid composition but, the fatty acid profiles remained relatively constant during both the logarithmic and stationary phases of growth. In contrast, the isoprenoid quinone composition depended markedly on the stage of growth. These results indicate that valid comparison of isoprenoid quinone profiles requires that sampling take place at defined stages of the growth cycle.     

A vigorous mutant sugarcane (Saccharum sp.) clone Co 527

Summary A vigorous fast growing mutant which ends vegetative growth sixty days earlier than the parent variety Co 527 was isolated from gamma irradiated vegetative buds. The mutant initially segregated but stabilized in the vM₄ generation. Its growth rate was almost fifty per cent higher than Co 527 beginning in the early stages of growth. It produced a significantly higher early shoot population which enabled it to yield a higher number of millable canes at maturity. Economic characters like sucrose content and juice purity remained unaffected. This mutant had a chromosomal basis in that the number was 2 to 3 lower than in the parent variety.Research carried out under the FAO/IAEA Co-ordinated Research Programme on the Improvement of Vegetatively Propagated Plants and Tree Crops through Induced mutations (Research Contract No. 1680)     

Effect of imipramine hydrochloride on the activity of gamma-aminobutyric acid transaminase in regions of the rat brain

The effect of intraperitoneal injection of imipramine hydrochloride on the activity of gamma-aminobutyric acid transaminase was determined in three regions of the rat brain.The cerebral hemispheres did not show a significant change in the activity of gamma-aminobutyric acid transaminase. Cerebellum and brain stem, both, however, showed a very significant decrease in the activity of the enzyme at 15 and 30 minutes after drug administration. At 90 minutes after drug administration, the activity of gamma-aminobutyric acid transaminase had returned to nearly control values.     

A high-throughput turbidometric assay for screening inhibitors of Leishmania major protein disulfide isomerase

The use of a high-throughput technique to perform a pilot screen for Leishmania major protein disulfide isomerase (LmPDI) inhibitors identification is reported. In eukaryotic cells, protein disulfide isomerase (PDI) plays a crucial role in protein folding by catalyzing the rearrangement of disulfide bonds in substrate proteins following their synthesis. LmPDI displays similar domain structure organization and functional properties to other PDI family members and is involved in Leishmania virulence. The authors used a method based on the enzyme-catalyzed reduction of insulin in the presence of dithiothreitol. The screen of a small library of 1920 compounds was performed in a 384-well format and led to the identification of 27 compounds with inhibitory activity against LmPDI. The authors further tested the cytotoxicity of these compounds using Jurkat cells as well as their effect on Leishmania donovani amastigotes using high-content analysis. Results show hexachlorophene and a mixture of theaflavin monogallates inhibit Leishmania multiplication in infected macrophages derived from THP-1 cells, although the inhibitory effect on LmPDI enzymatic activity does not necessarily correlate with the antileishmanial activity.     

Demographic and clinical variables affecting mid‐ to late‐life trajectories of plasma ceramide and dihydroceramide species

It has been increasingly recognized at the basic science level that perturbations in ceramide metabolism are associated with the development and progression of many age‐related diseases. However, the translation of this work to the clinic has lagged behind. Understanding the factors longitudinally associated with plasma ceramides and dihydroceramides (DHCer) at the population level and how these lipid levels change with age, and by sex, is important for the clinical development of future therapeutics and biomarkers focused on ceramide metabolism. We, therefore, examined factors cross‐sectionally and longitudinally associated with plasma concentrations of ceramides and DHCer among Baltimore Longitudinal Study of Aging participants (n = 992; 3960 total samples), aged 55 years and older, with plasma at a mean of 4.1 visits (range 2–6). Quantitative analyses were performed on a high‐performance liquid chromatography‐coupled electrospray ionization tandem mass spectrometer. Linear mixed models were used to assess the relationships between plasma ceramide and DHCer species and demographics, diseases, medications, and lifestyle factors. Women had higher plasma concentrations of most ceramide and DHCer species and showed steeper trajectories of age‐related increases compared to men. Ceramides and DHCer were more associated with waist–hip ratio than body mass index. Plasma cholesterol and triglycerides, prediabetes, and diabetes were associated with ceramides and DHCer, but the relationship showed specificity to the acyl chain length and saturation. These results demonstrate the importance of examining the individual species of ceramides and DHCer, and of establishing whether intra‐individual age‐ and sex‐specific changes occur in synchrony to disease onset and progression.     

Assessing the trypanocidal potential of natural and semi-synthetic diketopiperazines from two deep water marine-derived fungi

Human African trypanosomiasis (HAT, commonly known as African sleeping sickness) is categorized as a neglected disease, as it afflicts >50,000 people annually in sub-saharan Africa, and there are few formal programs in the world focused on drug discovery approaches for this disease. In this study, we examined the crude extracts of two fungal strains (Aspergillus fumigatus and Nectria inventa) isolated from deep water sediment which provided >99% growth inhibition at 1 μg/mL of Trypanosoma brucei, the causative parasite of HAT. A collection of fifteen natural products was supplemented with six semi-synthetic derivatives and one commercially available compound. Twelve of the compounds, each containing a diketopiperazine core, showed excellent activity against T. brucei (IC₅₀ = 0.002–40 μM), with selectivity over mammalian cells as great as 20-fold. The trypanocidal diketopiperazines were also tested against two cysteine protease targets Rhodesain and TbCatB, where five compounds showed inhibition activity at concentrations less than 20 μM. A preliminary activity pattern is described and analyzed.