Effect of lysosomotropic amines on the secretory pathway and on the recycling of the asialoglycoprotein receptor in human hepatoma cells

We studied the intracellular transport of secretory and membrane proteins in the human hepatoma cell line HepG-2 infected with vesicular stomatitis virus. Cells were pulse-labeled in the presence of [35S]methionine and chased in the presence of the lysosomotropic agent primaquine. At a concentration of 0.3 mM primaquine effectively inhibited the secretion of albumin and, to a lesser extent, that of orosomucoid and transferrin. The drug also prevented the budding of virus particles at the cell surface. The intracellular transport to the Golgi complex of the membrane protein VSV-G was not affected by primaquine as it acquires resistance to endo-beta-N-acetylglucosaminidase H at the same rate as in control cells. Addition of primaquine at various times after the initiation of the chase period indicates that the effect of primaquine occurs just before secretion. In confirmation of the biochemical data, immunocytochemical localization of albumin in cells treated with NH4Cl demonstrated that albumin accumulated in vesicles at the trans side of the Golgi complex. The effect of primaquine on secretion was also compared with its effect on receptor recycling. The dose-response characteristics of the effect of primaquine on receptor recycling are identical to those of the effects on protein secretion and virus budding. These results indicate that both processes involve the same transport mechanism, and/or that they occur via at least one identical intracellular compartment.     

The influence of dolichol, dolichol esters, and dolichyl phosphate on phospholipid polymorphism and fluidity in model membranes

The effect of dolichols, polyprenols, dolichol esterified with fatty acids, and dolichyl phosphate on the structure and fluidity of model membranes was studied using 31P NMR, small-angle x-ray scattering, differential scanning calorimetry, and freeze-fracture electron microscopy. These studies suggest that dolichol and dolichol derivatives destabilize unsaturated phosphatidylethanolamine containing bilayer structures and promote hexagonal II phase formation; high concentrations of dolichol induce lipid structures characterized by "isotropic" 31P NMR and particulate fracture faces; dolichol, contrary to cholesterol, has no effect on the thermotropic behavior of membranes consisting of phosphatidylcholine, while dolichyl-P incorporation abolishes the transition from the gel to liquid crystalline phase in 1,2-dimyristoyl-sn-glycero-3-phosphocholine; both dolichol and dolichyl-P increase the fatty acid fluidity in phosphatidylethanolamine mixtures; the effect of dolichol on bilayer structure and fluidity is more pronounced with increasing number of isoprene residues; dolichol esters are only soluble to a limited extent in the bilayer and segregates into domains at low concentrations; the results are consistent with a localization of dolichyl-P in which the phosphate group is oriented to the water interphase. The induction of hexagonal II phase by dolichyl-P may elicit the transmembrane movement of glycosylated lipid intermediate.     

Interferon affects intracellular calmodulin levels

Interferon lowers calmodulin levels in two cell lines sensitive to its antiproliferative effect. Further, in synchronized cells, interferon strongly inhibits the increase in calmodulin observed when control cells enter the S phase, and concomitantly inhibits DNA synthesis. Calmodulin has been implicated in the control of cell proliferation and an increase in this protein seems to be necessary for the progression of cells into the S phase of the cell cycle. Therefore, the effect of interferon on calmodulin content might constitute part of the molecular mechanism by which interferon inhibits DNA synthesis.     

MHC-like molecules in some nonmammalian vertebrates can be detected by some cross-reactive xenoantisera

Rabbit antisera raised to human and chicken MHC molecules were used to immunoprecipitate cross-reactive molecules from biosynthetically and cell surface-labeled spleen and/or blood cells of representative vertebrate species. Five major points emerged: 1) There were many nonspecific cross-reactions using these techniques, so various criteria were developed to distinguish these from true MHC-like molecules. 2) Only very small subpopulations of immunogen-specific antibodies cross-reacted with MHC-like molecules in other nonmammalian species. These subpopulations were different for each species and even within a species, sometimes being so limited as to behave like alloantisera. This led to a very scattered pattern of true cross-reactions that sometimes failed to reflect the properties of the bulk antibody population. 3) Antisera containing antibodies to class II beta- and class I alpha-chains cross-reacted better and more widely than those to B-G, class II alpha and, in general, beta 2-microglobulin. 4) Some cross-reactive antibodies were clearly directed to epitopes on the surface of the mature heterodimers, but many seemed to recognize nonlinear cryptic determinants, presumably in the contact regions between the chains. These latter antibodies recognized biosynthetic intermediates and also a variety of unusual cell surface MHC-like molecules present in reptile and amphibian, but absent in the mammal and chicken cells tested. These included E homodimers whose relationship to chicken B-G molecules is unknown. 5) MHC-like molecules were identified in a bird, three reptiles, and two amphibians, but no molecules with the expected properties were found with these reagents in any of the fish tested.     

Identification of a crotoxin-binding protein in membranes from guinea pig brain by photoaffinity labeling

Crotoxin is a neurotoxic phospholipase A₂ capable of blocking synaptic transmission by inhibiting the release of neurotransmitters. The photoaffinity labeling technique was used to identify the neural membrane molecules involved in the binding of crotoxin. A photoactivatable, radioactive derivative of crotoxin was synthesized by reacting crotoxin withN-hydroxysuccinimidyl-4-azidobenzoate and with Na[¹²⁵I]. Photoirradiation of synaptosomes from guinea pig brains in the presence of the crotoxin derivative resulted in the formation of a major radioactive conjugate of 100,000 daltons as revealed by autoradiography of a sodium dodecyl sulfate-polyacrylamide gel electrophoretic pattern. Pretreatment of the synaptosomes with trypsin,Staphylococcus aureus protease, or papain prevented the formation of this conjugate. The conjugate was not detected when plasma membranes from several nonneural tissues replaced the brain synaptosomes. Unmodified crotoxin inhibited the formation of this adduct with an IC₅₀ of about 10^–8 M. Mojave toxin, caudoxin, notexin,Naja naja PLA, and taipoxin also inhibited adduct formation with different potencies, while -bungarotoxin and pancreatic PLA were ineffective. We concluded that an 85,000-dalton protein is the major component responsible for the binding of crotoxin to synaptosomal membranes.On leave from Department of Biochemistry and Biophysics, University of Hawaii School of Medicine, Honolulu, Hawaii.     

Effect of prolonged high salt intake on atrial natriuretic factor's kinetics in rats

Plasma atrial natriuretic factor (ANF) paradoxically decreases after 5 weeks (but not after 3 weeks) of 8% NaCl intake in normotensive rats. As this phenomenon remains unaccounted for by changes in ANF production, we studied the disappearance of [125I]ANF(99-126) from the circulation as an alternative explanation of plasma ANF decline. Following 5 weeks (but not 3 weeks) of an 8% NaCl diet, plasma concentrations of [125I]ANF were significantly decreased and metabolic clearance rate and volume of distribution were increased compared to control rats fed a 0.8% NaCl diet. By studying [125I]ANF tissue uptake we noted significantly greater peptide uptake after 5 weeks (but not after 3 weeks) of high salt consumption in several tissues. We hypothesize that prolonged (at least 5 weeks) 8% NaCl ingestion increases the density and/or affinity of ANF binding sites. These changes may be responsible for the previously observed decline in plasma ANF concentrations after a prolonged high salt intake.     

旱地作物生态工程的设计

旱地作物生态工程是运用生态学和系统科学的原理和方法而设计和组建的旱地作物生产工艺体系。它把作物及其环境作为一个系统,统筹兼顾,相互协调,全面安排,综合利用。其最终目标是建立高效的,相对平衡的旱地作物生态系统。工程的最大特点在于它的整体性和综合性。它是作物先进生产技术的科学组装     

Evaluation of hydroxyl radical-scavenging property of garlic

While antibiotics are broadly used in dental and medical therapy, little attention has been directed towards the potential toxic side effects of antibiotics on tissue regeneration. Here we examined the effect of a quinolone antibiotic, pefloxacin (Rhone Poulenc) on rat parotid gland responses to chronic isoproterenol treatment. Groups of rats received injections of isoproterenol to induce glandular growth, saline (controls), pefloxacin, or isoproterenol and pefloxacin in combination. Parotid gland weight decreased significantly after pefloxacin treatment for 7 days as well as inhibiting glandular enlargement provoked by isoproterenol. The same trend was observed for the rates of DNA synthesis, with the incorporation of [³H]-thymidine in isoproterenol/pefloxacin-treated rats reduced to 49% of isoproterenol treatment alone levels. Saline-treated animals were 42% of the rate of [³H]-thymidine incorporation into DNA observed in isoproterenol treated rats. While isoproterenol treatment increased steady-state mRNA levels for fos, jun, myc, src, c-erbB-2, ras and topo II, inclusion of pefloxacin with the isoproterenol regimen blocked these increases. Pefloxacin treatment by itself did not alter proto-oncogene mRNA levels in the parotid gland. Glandular amylase activity was decreased in the pefloxacin treated group, while the combination of isoproterenol with pefloxacin did not decrease glandular amylase levels to the extent of that observed with -agonist treatment alone. In acute experiments, pefloxacin significantly decreased the volume of saliva secreted by the parotid gland. These results suggest that quinolone-based antibiotics disturb the secretory function of the parotid gland and can inhibit cell proliferation and regeneration. (Mol Cell Biochem 165: 55–63, 1996)     

胰岛素诱导低血糖大鼠心房肌细胞特殊颗粒的形态计量和心房利钠肽免疫组织化学研究

通过形态计量学和免疫组织化学方法发现胰岛素诱导低血糖大鼠心房肌细胞核周区特殊颗粒（ＡＳＧ）的体密度、面数密度和数密度及平均直径均高于对照组（Ｐ＜０．０５）,但高尔基复合体各参数与对照组比较没有差别（Ｐ＞０．０５）。实验组的心房利钠肽（ＡＮＰ）的免疫反应强度比对照组强（Ｐ＜０．００１）。提示胰岛素诱导低血糖对心房利钠肽的释放具有抑制作用,表明ＡＮＰ作为生理和病理调节递质与代谢刺激相拮抗。