Effects of seed-irradiation on meiosis in M1 plants of Lactuca and related plants

Meiosis was studied in control and seed-irradiated materials of Lactuca sativa, cultivars Tom, Cos and Webbs, L. serriola from the wild, and Cichorium intybus. In meiosis of the M₁ plants from irradiated seed, observations of univalent and multivalent formation, bridge formation at Anaphase I and II, micronucleus formation and persistence, and pollen fertility, were carried out, and compared with controls. Quantitative estimations of bridge formation were made in L. sativa Tom and Cos and in L. serriola, at anaphase I only. It was found that the frequency of anaphase I bridges and reduction of pollen fertility in the M₁ and also M₂ generation, were different for each variety or species; these could be sequenced radiation-resistant to sensitive in the order Tom, Cos and L. serriola.     

Low degree of ubiquitination of histone 2A in the dipteran Chironomus tentans

A polyclonal antibody to ubiquitin has been prepared and shown to react with both ubiquitin and ubiquitinated histone 2A (uH2A). Applying this antibody in Western blotting experiments, we have observed that the salivary glands of Chironomus tentans contain an unusually low amount of uH2A (1% of histone 2A), while the amount of free ubiquitin is as abundant as in other animal cells, e.g. HeLa cells. The same low content of uH2A was also found in diploid epidermal cells of Chironomus origin suggesting that the low amount is not a characteristic of the polytene state of chromatin in salivary gland cells but rather a property of C. tentans as a species. The significance of the low degree of ubiquitination is discussed in relation to the information available on the organization of Chironomus chromatin into unusually large chromomeric entities.     

Determination of dialkyl phosphates in human hair for the biomonitoring of exposure to organophosphate pesticides

A new, simple, fast and sensitive method that enables the measurement of four dialkyl phosphates (DAPs) in human head hair is presented in the current study. The dialkyl phosphates, dimethyl phosphate (DMP), diethyl phosphate (DEP), diethyl thiophosphate (DETP) and diethyl dithiophosphate (DEDTP) are non-selective metabolites of the organophosphate pesticides (OPs). The extraction of DAPs from hair matrix was achieved by one step methanolic extraction. Head hair samples from general population and population occupationally exposed to OPs were analysed using gas chromatography–mass spectrometry (GC–MS) after derivatization with pentafluorobenzylbromide. The recovery of the target compounds was estimated at 84.3% for DMP, 116.1% for DEP, 109.0% for DETP and 91.5% for DEDTP. The limit of quantitation (LOQ) and detection (LOD) was 20 and 6 pg/mg for DMP, 10 and 5 pg/mg for DEP and DETP and 5 and 3 pg/mg for DEDTP, respectively. With-run and between-run precision as well as accuracy was estimated. The percentage of positive hair samples for DMP, DEP, DETP and DEDTP for the group of general population was 63.0%, 96.3%, 66.7%, and 70.4% respectively. The samples from the group with occupational exposure were positive for all dialkyl phosphates analysed. The median concentrations for DMP were 165.0 and 181.7 pg/mg, for DEP were 51.2 and 812.9 pg/mg, for DETP were 54.0 and 660.1 pg/mg, and for DEDTP were 40.0 and 60.6 pg/mg for the general population group and the group with occupational exposure respectively. Significant differences in the levels of the total dialkyl phosphates amongst exposed and not exposed groups were observed (p < 0.001). More specifically, the total ethyl phosphate (DEPs) and DAPs median concentrations were 119.5 and 301.5 pg/mg for the general population group and 1498.8 and 1694.4 pg/mg for the group with occupational exposure.     

Intragroup serotyping of meningococci

The method for obtaining antisera to meningococci of different serotypes are described and the scheme for the preparation of serotyping is presented, as well as the method for the preparation of the determinate fraction of serotype 2. Antisera to typing antigens 1, 2, 2-7, 2-10, 4, 5, 6, 8 (1) have been obtained, their specificity tested in parallel experiments with American and French typing sera. When typing meningococci, the use of antisera to purified protein antigen 2 is recommended.     

A Yersinia pseudotuberculosis protein which cross-reacts with HLA-B27

The most-debated question in the investigation of the spondyloarthropathies has been whether there is molecular mimicry between host HLA-B27 antigens and the arthritis-causing pathogens. We have generated a monoclonal anti-HLA-B27 antibody in our laboratory and have used a radioimmunoassay to screen a panel of bacterial species. Two strains of Yersinia pseudotuberculosis were found to be highly reactive. The cross-reactive Yersinia component was identified by Western blot to be a 19,000 component. A preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis chromatography apparatus was constructed to isolate milligram quantities of this component. To verify that the component carried the HLA-B27-specific epitope, rabbits were hyperimmunized with the purified materials. Affinity-purified antibodies from one of the immunized rabbits indeed carried anti-HLA-B27 activity. Last, antibodies generated against synthetic peptides derived from the HLA-B27.1 amino acid sequence were tested against the Yersinia component. Positive reactivity was found with antibodies generated against a peptide spanning residues 69-83 of the HLA-B27.1 protein. Since this resides in the segment responsible for the allotypic specificity of the antigen, these experiments establish the presence of molecular mimicry to a high degree of confidence.     

Relationship between cell density and prolidase activity in human skin fibroblasts: effects of ascorbate and fructose

To evaluate the influence of cell density on the activity of fibroblast prolidase (EC 3.4.13.9), we determined this activity in sparse and dense cultures. We also investigated, the effects of different concentrations of β-d(?) fructose and l(+) ascorbate, which both increased cell density at confluency. For a fructose concentration of 25 mM, we observed that in the absence of glucose, intracellular total proteins increased 1.5-fold and prolidase specific activity, 1.8-fold. For ascorbate, a broad optimum concentration was found (range 0.01 – 0.50 mM). Addition to cultures of 0.1 mM ascorbate increased total proteins 1.4-fold, and doubled prolidase activity. This investigation was prompted by our previous results [J. Metab. Dis. 1983, 6, 27–31], confirmed here, and suggesting that increased prolidase activity at confluency was due to a rise in cell density.     

Possible involvement of DNA breaks in epigenetic regulation of cell differentiation

The review summarizes the authors’ and literature data on accumulation of DNA breaks in differentiating cells. Large 50-kb free DNA fragments were observed by several research teams in non-apoptotic insect, mammal, and plant cells. More intense DNA breakage was observed during maturation of spermatides, embryo development, and differentiation of myotubes, epidermal cells, lymphocytes, and neutrophils. In general, accumulation of DNA breaks in differentiating cells cannot be attributed to a decrease in the DNA repair efficiency. Poly(ADP)ribose synthesis often follows the DNA breakage in differentiating cells. We hypothesize that DNA fragmentation is an epigenetic tool for regulating the differentiation process. Scarce data on localization of the differentiation-associated DNA breaks indicate their preferable accumulation in specific DNA sequences including the nuclear matrix attachment sites. The same sites are degraded at early stages of apoptosis. Recent data on non-apoptotic function of caspases provide more evidence for possible existence of a DNA breakage mechanism in differentiating cells, resembling the initial stage of apoptosis. Excision of methylated cytosine and recombination are other possible explanations of the phenomenon. Elucidation of mechanisms of differentiation-induced DNA breaks appears to be a prospective research direction.     

Spongelike alginate nanoparticles as a new potential system for the delivery of antisense oligonucleotides.

The aim of this study was to design a new antisense oligonucleotide (ON) carrier system based on alginate nanoparticles and to investigate its ability to protect ON from degradation in the presence of serum. Pharmacokinetics and tissue distribution of ON-loaded nanoparticles have been determined after intravenous administration. An original and dynamic process for ON loading into polymeric nanoparticles has been applied. It is based on the diffusion of ON or ON/polylysine complex into the nanoparticle or the alginate gel, respectively. Indeed, the single coincubation of ON with nanoparticles led, within a few days, to an extremely efficient association. The diffusion kinetic of ON was shown to be dependent on several parameters, incubation temperature, ON concentration, presence or absence of polylysine, polylysine molecular weight, and nanoparticle preparation procedure. This new alginate-based system was found to be able to protect [33P]-radiolabeled ON from degradation in bovine serum medium and to modify their biodistribution, as an important accumulation of radioactivity was observed in the lungs, in the liver, and in the spleen after intravenous administration into mice. ON may be associated efficiently with calcium alginate in a colloidal state. Such nanosponges are promising carriers for specific delivery of ON to lungs, liver, and spleen.