Characterization of specific HLA-DQ alpha allospecificities by genomic, biochemical, and serologic analysis

Bgl II restriction endonuclease digestion of genomic DNA from lymphoblastoid cell lines homozygous for HLA DR and DQ serological specificities, followed by hybridization with a DQ alpha cDNA probe, identified a genomic polymorphism characterized by two reciprocal patterns, one associated with DR 3, 5 and 8 and the other with DR 1, 2, 4, 7, and 9. The former pattern corresponded precisely to the reactivity of monoclonal antibody SFR20-DQ alpha 5, shown by Western blotting to react with isolated alpha-chains, but not with beta-chains. Additional variants of the DQ alpha genes were identified by using a locus-specific oligonucleotide probe for the DQ alpha gene, indicating differences among the DQ alpha 5-negative set of alleles. This analysis defines a set of DQ alpha allelic markers that are distinct from the well-established DQ serologic specificities DQw1, 2, 3 or "blank." Although most DQ alpha 5+ cells carry the DRw52 specificity associated with the DR beta 2 gene, analysis of DQ alpha polymorphisms on DR5, DQw1; DR8, DQw1; and DRw13, DQw1 cells verified that this DQ alpha family of alleles was not invariably linked to the DR beta 2 locus.     

Subcellular organization of Streptomyces aureofaciens and overproduction of chlortetracycline

Biosynthesis of chlortetracycline is localized differently under low- and high-production conditions (standard low-production strain and its high-production variant). The experimental evidence was based on the assay of anhydrotetracycline oxygenase in subcellular fractions, ultracytochemical localization and electron-probe X-ray microanalysis of the product in the mycelium. Overproduction of chlortetracycline is closely associated with compartmentation of biosynthetic enzymes and with an efficient export of the antibiotic out of the cell.     

Timing of laparoscopic aspiration of preovulatory oocytes in heifers

Twenty-one heifers were synchronized with PGF(2) alpha and 22 heifers were stimulated with FSH-P in decreasing doses and synchronized with PGF(2) alpha. The beginning of LH rise was observed to be 47.6+/-14.0 h and 35.9+/-1.3 h (P < 0.05) and the peak of LH rise was observed at 53.8 +/- 13.4 h and 40.14+/-1.4 h (P < 0.05) after the luteolyticum administration in the synchronized and superovulated group respectively. The beginning of LH rise was observed 6.9+/-6.7 h and 4.8+/-2.6 h (P < 0.05) and the LH peak was observed 11.8+/-7.7 h and 8.3+/-3.2 h (P < 0.05) after the frist symptoms of oestrus in the synchronized and superovulated group respectively. Ovulation was not observed in stimulated heifers in the period of 23-25 h after the preovulatory LH rise. Compact cumulus oophorus was seen at 30.5%, expanded at 67.0 and partial at 2.5% during this interval of 23-25h. Within this same interval 26.9%, 51.3% and 21.8% oocytes without perivitelline space, with perivitelline space and with extruded first polar body were aspirated respectively. It may be concluded from the reported results that to recover fully mature oocytes for in vitro fertilization, it will be necessary to monitor the preovulatory period of the donor cow in great detail.     

Autologous B lymphoblastoid cell lines and long-term cultured T cells as stimulators in the mixed lymphocyte reaction: analysis of the role of HLA class II antigens as stimulatory molecules

Human activated T cells, long-term cultured in the presence of interleukin 2 (IL 2), were compared with autologous Epstein Barr virus-transformed B lymphoblastoid cell lines for expression of human leukocyte (HLA)-HLA-DR and -DQ antigens and for ability to induce proliferative responses in autologous and allogeneic lymphocytes. Immunofluorescence analysis performed with a panel of monoclonal antibodies (mAb) specific for HLA-DR or -DQ antigens did not reveal any significant difference in the expression of HLA-DR antigens but revealed reduced expression of HLA-DQ antigens on two out of four T cell lines tested. No obvious difference could be detected in the two-dimensional gel electrophoretic profile of HLA-DR and -DQ beta-chains synthesized by the autologous pairs of B and T cell lines. In contrast with previous reports, the IL 2-dependent cell lines consistently induced alloproliferative responses in standard 6-day mixed lymphocyte cultures; however, these responses were severalfold lower than those elicited by the autologous B lymphoid lines. Both anti-HLA-DR and anti-HLA-DQ mAb blocked the proliferative responses induced by the B cell lines but did not affect those generated by the T cell lines, suggesting that the latter cells induce T lymphocyte activation via a mechanism independent of HLA-DR or -DQ antigen expression on their surface. Addition of IL 2 to the mixed cultures with B cell lines as stimulators did not affect the outcome of the proliferative responses but partially or completely reversed the blocking activity of the mAb. In contrast, IL 2 significantly enhanced the alloproliferation induced by the T lymphoblastoid cell lines, and the anti-HLA class II mAb partially antagonized this effect. Taken together, these data suggest that unlike the HLA-DR and -DQ gene products on B cells, those on IL 2-dependent long-term cultured T cells do not play a direct or primary stimulatory role in the mixed lymphocyte reaction; the reduced levels of alloproliferation induced by the T cell lines are, at least in part, due to a defective production of endogenous IL 2 by the responder lymphocytes rather than to a defective expression of IL 2 receptors by the alloproliferative T cell subset; and the anti-HLA class II mAb in these cultures act only at the responder cell level, since they can efficiently block the enhancement of T cell proliferation triggered by exogenous IL 2, but not the proliferative responses induced by T cell lines in standard conditions.     

An improved method to isolate lichen algae by gel filtration

Photobiont cells of the lichen Evernia prunastri have completely been separated from their fungal partner by filtration through a bed of Sepharose 2B. Both mannitol and ribitol have been quantified by gas-liquid chromatography in the different steps of the isolation procedure. Absence of mannitol, which is exclusively produced by the mycobiont, has been used as the best probe to monitor isolation.     

Invertase immobilization via its carbohydrate moiety

After periodate oxidation of its glycosidic component, invertase was covalently bound onto three types of modified solid supports: glycidyl methacrylate, styrene-divinylbenzene copolymers, and bead cellulose. Direct reaction of the invertase aldehyde groups that were formed with amino groups of the support and use of the modified Ugi reaction have been employed as immobilization procedures. Apart from binding methods, the important effects of the buffer, support, conditions of periodate oxidation, and the length of the spacer on the activity of the enzyme conjugate have been investigated. Superior conjugate activity was obtained, via modified Ugi reaction, by the immobilization of a suitably oxidized invertase to a styrene-divinylbenzene copolymer having free amino groups.     

New C-nucleoside analogs by dehydration of 1-benzyl-4,5,6,7-tetrahydro-6,6-dimethyl-2-(d-galacto-pentitol-1-yl)-indol-4-one

The reaction between 2-(benzylamino)-2-deoxy-d-glycero-l-gluco-heptose and 5,5-dimethyl-1,3-cyclohexanedione yields 1-benzyl-4,5,6,7-tetrahydro-6,6-dimethyl-2-(d-galacto-pentitol-1-yl)-indol-4-one (2). Acid-catalyzed, intramolecular dehydration of 2 under kinetically controlled conditions gives 1-benzyl-4,5,6,7-tetrahydro-2-α-d-lyxofuranosyl-6,6-dimethylindol-4-one; the anomeric configuration of this compound is only suggested. When the dehydration reaction is conducted under thermodynamically controlled conditions, it produces a 1:1 mixture of the α- and β-d-lyxopyranosyl compounds. The structures of the new compounds were elucidated by chemical and physical methods.     

The biology of Butomus umbellatus in shallow waters with fluctuating water level

Butomus umbellatus L. is a plant species typical of littoral communities of river and stream shores. It can form continuous stands in shallow reservoirs with fluctuating water level. Their expansion is promoted by: (a) intensive vegetative reproduction of plants, (b) crowded sprouting from rhizome fragments on emerged pond bottom, (c) shallow water layer in the year following summer drainage. Expansion of B. umbellatus depends on ploidy level: two cytotypes were found in the Czech and Slovak Republics, differing in their reproductive ability. Seed production of triploids is strongly limited (they are self-incompatible within clones), while diploids can be fully fertile. Nevertheless, even in diploids, the efficiency of seed reproduction under natural conditions is low. Triploids spread by intensive vegetative reproduction, which is decisive for clonal growth of populations and their regeneration after scraping of bottom surface. During seasonal development, maximum of aboveground biomass is produced in early summer, while underground biomass increases till autumn. Growth of the plants is limited by cutting before maximum underground biomass is attained, or by duck grazing.