An enzyme with properties similar to those of beta-N-acetylhexosaminidase S is expressed in the promyelocytic cell line HL-60.

Extracts of the human promyelocytic cell line HL-60 contain a form of beta-N-acetylhexosaminidase that is not retained on columns of benzeneboronate-agarose ('phenylboronate-agarose') and has a pI value lower than that of beta-N-acetylhexosaminidase A. It is clearly distinct from beta-N-acetylhexosaminidase A in its behaviour on DEAE-cellulose columns, and it requires a higher concentration of salt for its elution. This 'extra' form has a higher ratio of activity towards 4-methylumbelliferyl beta-N-acetylglucosaminide 6-sulphate and 4-methylumbelliferyl beta-N-acetylglucosaminide than has beta-N-acetylhexosaminidase A and is less stable when heated at 50 degrees C. It has a pH optimum of 4.5 and is therefore not beta-N-acetylglucosaminidase C. Anti-(human beta-N-acetylhexosaminidase alpha-subunit) serum precipitated both beta-N-acetylhexosaminidase A and the 'extra' form, whereas an anti-(beta-subunit) serum precipitated beta-N-acetylhexosaminidase A but not the 'extra' form. Western blotting and immunodetection of polypeptides derived from the 'extra' form revealed a band corresponding in size to mature alpha-subunits. On the basis of this and of its behaviour on isoelectric focusing, chromatofocusing and its kinetic properties, we conclude that the 'extra' form is composed of alpha-subunits and resembles beta-N-acetylhexosaminidase S, the residual form in Sandhoff's disease.     

Distribution of Thiamine, Thiamine Phosphates, and Thiamine Metabolizing Enzymes in Neuronal and Glial Cell Enriched Fractions of Rat Brain

The distribution of thiamine, thiamine phosphoesters, and the thiamine pyrophosphate synthetizing [thiamine-pyrophosphokinase (TPKase)] as well as hydrolyzing [thiamine pyrophosphatase (TPPase) and thiamine monophosphatase (TMPase)] enzymes was determined in neuronal and glial enriched fractions prepared from rat brain. Nucleoside diphosphatases [inosine diphosphatase (IDPase) and uridine diphosphatase (UDPase)] and nucleoside monophosphatases [uridine monophosphatase (UMPase) and inosine monophosphatase (IMPase)] were also determined. Thiamine and thiamine mono- and pyrophosphate were present in neuronal enriched fractions at concentrations 2.8, 3.6, and 4.6 times higher than in glial fractions. TMPase was found only in glial enriched fractions, whereas the levels of TPKase, UMPase, IMPase, IDPase, UDPase, and TPPase were 2.0-, 2.2-, 1.3-, 2.8-, 3.7-, and 20.8-fold higher in neuronal than in glial fractions.     

Changes in erythropoiesis due to radiation or chemotherapy as studied by flow cytometric determination of peripheral blood reticulocytes

Flow cytometric determination of time dependent changes of numbers of reticulocytes in peripheral blood were investigated as a parameter for changes in erythropoiesis induced by radiation- or chemotherapy. Rats irradiated or treated with drugs (such as e.g. cyclophosphamide 100 mg/kg, vincristin 0.2 mg/kg, or mitomycin C 1.0 mg/kg) showed clear changes in erythropoietic activity. Reticulocyte numbers decreased rapidly until day 3-4 after treatment; this period was followed by a gradual increase and normal control values were seen at day 8-11. Radiation effects of doses as low 0.5 Gy could be detected in such a way. Similar studies were performed with patients with ovarian tumors treated with cis-platinum, a drug that may cause non-immune haemolysis. During prolonged treatment some patients showed increasing numbers of reticulocytes, measured at the first day of each hospitalization period, whereas leucocyte and platelet counts stayed more or less constant. Increasing numbers of reticulocytes generally indicates stimulation of erythropoietic activity of the bone marrow (due to increased blood loss); in this study increasing numbers often preceeded a decrease in hemoglobin values later on. Flow cytometric analysis of reticulocytes is therefore a potentially useful tool to detect changes in erythropoiesis, and considered more sensitive for the early recognition of patients that develop anemia, than hemoglobin measurements only.     

Tubular structures associated with the endothelial endoplasmic reticulum in glomerular capillaries of Rhesus monkey and nephritic man

Summary Round bodies, tubular profiles and crystalline images have been studied by electron microscopy in the endothelium of seven normal young Rhesus monkeys and in the renal glomerular endothelium of two nephritic human patients. The crystalline images are most frequently formed by aggregation of round bodies, 200–240 Å in diameter. In Rhesus monkeys a variety of crystalline images are seen. On the contrary, in nephritic patients round bodies and tubular profiles are rare and less organized. In the glomerular endothelium of two normal men they were not found.The results obtained suggest that the round bodies, the tubular profiles and the crystalline images result from sectioning of a system of undulating tubules associated with the smooth endoplasmic reticulum. In nephritic patients the formation of such a tubular system could represent a change taking place within the affected cells as a pathologic response to the disease.This work was supported by U.S. Public Health Service (N.H.I.), Grant AI-04527-03-04, and by the Consiglio Nazionale delle Ricerche (C.N.R.), Contributo 115/0815/0-1365. The Authors are greatly indebted to Miss Hermina Spiele for skilful technical assistance.     

A re-evaluation of the mesangial cells of the renal glomerulus

Summary The topographical localization of the mesangial cells in renal glomeruli of rats, and their relationships with the structures at the hilus of the glomerulus were studied in this investigation. It was observed that the mesangial cells occupy a parietal position in the wall of the glomerular capillaries, and that they are anatomically continuous with the smooth muscle cells of the tunica media of the glomerular arterioles.This study was supported by a United States Public Health Service Grant AM 08628 (Institute of Arthritis and Metabolic Diseases).     

Induced Autolysis of Aspergillus oryzae (A. niger group): IV. Carbohydrates

The examination of substances formed during induced autolysis by Aspergillus niger was continued in this work, which dealt in particular with carbohydrates. The autolysate contained a large amount of d-glucose (14 to 20% dry wt) and traces of glycolic aldehyde, dihydroxyacetone, ribose, xylose, and fructose. It also contained glycopeptides (about 10% dry wt), which were split from the cell wall during autolysis and which differed from one another in their level of polymerization and their composition. They were constituted by glucose and mannose, glucose and galactose, or mannose, glucose, and galactose (mannose being the most abundant in this case), and amino acids (chiefly alanine, serine, glutamic acid, and aspartic acid). During autolysis, only a part of the cell wall was dissolved, since it retained its shape. Upon further chemical hydrolysis, it produced mostly glucose and glucosamine, and smaller amounts of mannose, galactose, and amino acids. Presumably, glucomannoproteins and glucogalactoproteins were present in the intact cell as a macromolecular complex, constituting, together with chitin, the major part of the cell wall of Aspergillus.     

Stomatal and cuticular transpiration of beans (Phaseolus vulgaris M.) attacked by rust (Uromyces appendiculatus [Pers.] Link)

The changes of stomatal and cuticular transpiration of bean plants were investigated by graphical transpiration curves method (Slavík 1958). Bean leaves were infected by fungusUromyces appendiculatus (Pers.) Link. After the infection the intensity of stomatal transpiration had a decreasing tendency. Beginning with the sixth day after infection, the proportion of stomatal and cuticular transpirations becomes more expressive, i.e. the leaves transpire more by cuticles than by stomata. The higher share of cuticular transpiration brings extensive water relations to the diseased plants.     

Electrotransformation of Streptococcus agalactiae with plasmid DNA

Abstract A protocol for efficient electrotransformation of Streptococcus agalactiae (group B streptococcus) Lancefield's strain O90R (NTCT 9993) (an unencapsulated derivative of type Ia strain O90) was developed. The Escherichia coli - Streptococcus shuttle vector pDP28 (7.8 kb) carrying the ermB gene for resistance to erythromycin was used as donor DNA. Frozen 'electrocompetent' cells were prepared by repeated washes in 10% glycerol. A 50-μl aliquot containing about 5×10⁹ colony forming units of bacteria was subjected to the electric pulse. Optimal conditions for electrotransformation were determined using different media, harvesting cells at different points of the growth curve, and using different field strengths. The dose-response curve for transformation of S. agalactiae with pDP28 showed one-hit kinetics as donor DNA varied between 0.01 and 3 μg. The efficiency of electrotransformation for this range of amounts of donor DNA was 1.2×10⁴ cfu μg⁻¹. The transformation frequencies obtained with this electroporation protocol are high enough to allow both subcloning and shotgun cloning of streptococcal DNA in S. agalactiae .     

Seasonal succession of phytoplankton in a lake of the Paraná river floodplain,Argentina

Phytoplankton biomass, morphological and taxonomic composition, species diversity and productivity were analyzed in a shallow lake of the Middle Paraná River floodplain (El Tigre, 31 ° 41 S and 60° 42 W), between November 1986 and July 1988. Lake inundation (filling and through-flow phases) constituted an intense long-term perturbation in the physical and chemical environment. As the lake filled with river water, K-selected species (netplanktonic filamentous bluegreens, > 37 µm, with low surface area/volume (SA/V) ratios) that had existed prior to filling (late spring 1986) were replaced in summer-fall by r-selected species (nannoplanktonic chlorophytes and cryptophytes, < 37 µm, mainly stout forms with high SA/V ratios). During the through-flow phase, lentic phytoplankton was replaced by lotic flagellate populations due to the direct flushing by river water. During the period of falling water (drainage and isolation phases), nanoplanktonic algae with similar characteristics to those of the filling phase dominated in late winter-spring. Later in the isolation phase, these were succeeded by K-selected species (netplanktonic algae, mainly motile spherical dinoflagellates and filamentous bluegreens with low SA/V ratios). Simultaneously, primary production per unit biomass decreased and total biomass and specific diversity increased. Seasonal changes of phytoplankton in floodplain lakes can be interpreted as the interaction between true successional development (as observed in the drainage and isolation phases) and intermediate disturbance. Using Reynolds' terminology, short-term disturbance (slight inflow of nutrient-rich river water) caused reversion to an earlier stage in the former succession, and long-term disturbance (lake inundation) truncated the successional progression and a new (or shifted) succession was initiated.     

Presence of telomeric and subtelomeric sequences at the fusion points of ring chromosomes indicates that the ring syndrome is caused by ring instability

In situ hybridization of a telomeric (TTA-GGG) _n sequence to metaphases from three cases of ring chromosome, involving respectively chromosomes 4, 16, and 20, showed the presence of the cognate sequences in all three rings. To investigate whether these ring chromosomes originated by telomere-telomere fusion, we determined, by in situ hybridization, whether telomere-associated sequences and/or specific distal sequences were still present in the ring chromosomes. The finding that these sequences were preserved in all the ring chromosomes strongly indicates that they originated by telomere-telomere fusion. All three subjects carrying the ring chromosomes are affected by the so-called ring syndrome, with failure to thrive, minor dysmorphic signs and no major anomalies. The r(4) patient has the ring in mosaic form with a normal cell line and has normal intelligence. The r(16) and the r(20) patients have moderate mental retardation and suffer from seizures. We conclude that the ring syndrome, even in its more severe manifestation, is caused by ring chromosome instability.