Decrease in insulin-containing secretory granules and mitochondrial gene expression in mouse pancreatic islets maintained in culture following streptozotocin exposure

We have previously described a preferential reduction in the secretory response to nutrient secretagogues in pancreatic mouse islets maintained in culture after in vitro exposure to streptozotocin (SZ). This reduction was associated with an impaired substrate metabolism at the mitochondrial level. To further clarify this issue, mouse pancreatic islets were exposed in vitro to 2.2 mM SZ for 30 min. At 4 h after SZ treatment ultrastructural changes were apparent in the endoplasmic reticulum and Golgi areas of the B-cells. However, 2 and 6 days following SZ exposure the B-cells appeared well preserved, except for a marked decrease in the number of insulin-containing secretory granules. A morphometric analysis of the B-cells 6 days after SZ exposure showed a normal B-cell size and a normal volume fraction of B-cell mitochondria. However, there was a decrease in total islet size and a 13% decrease in the volume fraction of B-cells in the islets. These mouse islets exhibited a decreased content of the mitochondrial DNA-encoded cytochrome b mRNA, as evaluated by dot-blot analysis. As a whole, the data obtained indicate that SZ treatment does not induce a decrease in the number of mitochondria or long-lasting ultrastructural damage to this organelle. However, there is a clear decrease in the cytochrome b mRNA, suggesting that SZ can induce damage to the mitochondrial DNA.     

DNA replication and H5 histone exchange during reactivation of chick erythrocyte nuclei in heterokaryons

Fusion of terminally differentiated chick erythrocytes (CE) with replicating quail myoblasts or established L6J1 rat myoblasts results in reactivation of DNA synthesis in the dormant CE nuclei and in suppression of DNA synthesis in the myoblast nuclei. The nuclei of primary quail myoblasts are more effectively inhibited than the nuclei of established rat myoblasts. Inhibition of DNA replication occurs not only by preventing G1 nuclei from entering S-phase but also by blocking nuclei in S-phase and by delaying nuclei in G2 from undergoing mitosis and starting a new DNA replication cycle. No inhibition of DNA synthesis could be observed when mouse erythrocytes, i.e., erythrocytes lacking nuclei, were fused with rat myoblasts to generate mouse-globin-containing L6J1 cybrids. — Reactivation of CE nuclei is associated with a loss of the tissuespecific H5 histone variant. Complete elimination of H5 histone, however, does not seem to be a necessary prerequisite for the initiation or completion of DNA replication in CE nuclei since H5 antigens are found on reactivated G1, S, and G2 nuclei.     

Covalent immobilization of the multienzyme enniatin synthetase

Summary The multienzyme enniatin synthetase was covalently immobilized to N-hydroxysuccinimide activated agarose. The stability of the immobilized enzyme at 25°C was enhanced compared to the soluble enzyme. Immobilization experiments also indicated that the enniatins are synthesized by a single molecule and thus do not require interactions of several enzyme molecules.     

Production and characterization of KDO-specific monoclonal antibodies recognizing lipopolysaccharides from heptoseless mutants of Salmonella

Abstract Hybrid cell lines producing monoclonal antibodies with specificity for the lipopolysaccharide (LPS) from the deep rough mutant Salmonella minnesota R595 have been established. Spleen cells from BALB/c mice immunized with live R595 bacteria were fused with Sp 2/0 myeloma cells and three hybridomas producing antibodies specific for heptoseless LPS from Salmonella were selected. All three monoclonal antibodies were shown to bind only to heptoseless, but 3-deoxy- d -manno-octulosonic acid (KDO) containing LPS when tested in enzyme-linked immunosorbent assay (ELISA) against a set of structurally defined LPS and lipid A from Salmonella, Shigella and Escherichia coli . Synthetic KDO was an efficient inhibitor of the antibody-R595 LPS interaction defining that KDO is in an immunodeterminant position interacting with the monoclonal antibodies.     

Dolichol and Dolichyl Phosphate Levels in Brain Tissue from English Setters with Ceroid Lipofuscinosis

Human ceroid lipofuscinosis (CL) is an inherited disease marked by cerebromacular degeneration and early death. We have utilized the canine model to investigate the possible role of dolichol and dolichyl phosphate in the developmental pathology of CL. We found that while brain levels of dolichol increase with age in both affected and unaffected dogs, the amount in the diseased animal was similar to that in controls. Brain levels of dolichyl phosphate ranged from 20 to 35 micrograms/g in control dogs at all ages examined, but increased substantially during development in the affected dogs, a value of 113 +/- 24 micrograms/g (mean +/- SD) being obtained in the end-stage animals. In addition to the results obtained in the canine model, dolichyl phosphate levels in human brain tissues from a 5-year-old with late infantile CL and from a 19-year-old with juvenile CL were found to be 153 and 382 micrograms/g, respectively, compared with a control that assayed 26 micrograms/g. The preliminary findings with human tissues provide further evidence for an association of elevated brain dolichyl phosphate levels with CL. Whether the increase is primary or secondary remains to be determined.     

Studies of Electrophoresis on Cellulose Acetate Membrane of Serum Proteins from Normal Horses,Sheep and Pigs

A method for the rapid electrophoresis on a cellulose acetate membrane of serum proteins from horses, sheep and pigs is discussed. The various main globulin fractions in the serum of these animals were experimentally identified. Normal values for the percentage composition of serum from normal horses, sheep and pigs were calculated. In the horse there was great individual variation in the shape of the β-fraction, assumed to be due to different transferrin types. The mean value for β-globulin of 19.5 % in the horse was higher than for the other two species. The albumin percentage was highest in the sheep and lowest in the pig, 48.5 % and 43.2 % respectively. The sheep had the highest γ-globulin percentage, 22.8 %, while the horse had the lowest with 19.0 %. Finally the values were compared with corresponding figures reported by other authors and the results discussed.