Stimulation of haematopoiesis in primates by continuous infusion of recombinant human GM-CSF

Certain proteins are known to play an important part in the proliferation, differentiation and functional activation of haematopoietic progenitor cells in vitro. These proteins include erythropoietin and various colony-stimulating factors (CSFs), one of which is granulocyte-macrophage colony-stimulating factor (GM-CSF). Recently, both murine and human GM-CSF have been purified to homogeneity and complementary DNAs encoding them have been cloned. Although the in vitro activity of recombinant human GM-CSF has been investigated intensively, little is known about the functional activity of this protein in vivo. There is strong evidence that colony-stimulating activities produced by various human and murine tumour tissues and cell lines can stimulate granulopoiesis in mice, as can human urinary extracts. A partially purified preparation of human urinary colony-stimulating factor, however, proved only marginally effective in stimulating granulopoiesis in humans. All these studies suffer from the lack of a homogeneous preparation of colony-stimulating factor. It has recently been shown that recombinant murine multi-CSF or interleukin-3 can stimulate haematopoiesis in mice in vivo. Large-scale production of recombinant human GM-CSF now permits us to examine its effects in vivo using a primate model. We find that the continuous infusion of GM-CSF in healthy monkeys rapidly elicits a dramatic leukocytosis and a substantial reticulocytosis. A similar effect has been observed in one pancytopenic, immunodeficient rhesus macaque. These results suggest that GM-CSF could prove useful in several clinical situations.     

The effects of triaminopyrimidine on the short circuit current and transmembrane potentials of the isolated rat visceral yolk sac in vitro

Summary Intracellular potentials in the cells from 17.5-day old rat visceral yolk sacs were measured by a glass microelectrode. When penetrated from the maternal side, the cells have potentials of about 50.2±1.9 mV (inside negative) which were reduced by increasing the external K⁺ concentration and increased by removing Na⁺ ions from the bathing fluid. Triaminopyrimidine (TAP) which inhibited Na⁺ transport caused a dose-dependent depolarization of the cell membrane. The depolarization was dependent on the presence of extracellular Ca²⁺ ions. It is proposed that TAP may inhibit Na⁺ transport by increasing the intracellular concentration of calcium ions.This work was supported by the University of Hong Kong (grant number 335. 034.5105).Acknowledgment. Triaminopyrimidine was synthesized by Dr. Barbara Roth of the Wellcome Research Laboratories.     

Induction of vanadium accumulation and nuclear sequestration causing cell suicide in human Chang liver cells

Very little is known about the modulation of vanadium accumulation in cells, although this ultratrace element has long been seen as an essential nutrient in lower life forms, but not necessarily in humans where factors modulating cellular uptake of vanadium seem unclear. Using nuclear microscopy, which is capable of the direct evaluation of free and bound (total) elemental concentrations of single cells we show here that an NH₄Cl acidification prepulse causes distinctive accumulation of vanadium (free and bound) in human Chang liver cells, concentrating particularly in the nucleus. Vanadium loaded with acidification but leaked away with realkalinization, suggests proton-dependent loading. Vanadyl(4), the oxidative state of intracellular vanadium ions, is known to be a potent source of hydroxyl free radicals (OH^.). The high oxidative state of nuclei after induction of vanadyl(4) loading was shown by the redox indicator methylene blue, suggesting direct oxidative damage to nuclear DNA. Flow cytometric evaluation of cell cycle phase-specific DNA composition showed degradation of both 2N and 4N DNA phases in G₁, S and G₂/M cell cycle profiles to a solitary 1N DNA peak, in a dose-dependent manner, effective from micromolar vanadyl(4) levels. This trend was reproduced with microccocal nuclease digestion in a time response, supporting the notion of DNA fragmentation effects. Several other approaches confirmed fragmentation occurring in virtually all cells after 4 mM V(4) loading. Ultrastructural profiles showed various stages of autophagic autodigestion and well defined plasma membrane outlines, consistent with programmed cell death but not with necrotic cell death. Direct intranuclear oxidative damage seemed associated with the induction of mass suicide in these human Chang liver cells following vanadium loading and nuclear sequestration.     

粉煤灰残炭吸附-电化学复合法净化印染废水的研究

作者提出一种粉煤灰残炭吸附-电化学复合法净化印染废水的新方法,研究了各因素对净化效果的影响。通过模拟和实际印染废水的试验,得到复合法的工艺条件,取得了良好净化效果,处理后的水质达到排放标准。     

Crystal structure of the anthrax lethal factor.

Lethal factor (LF) is a protein (relative molecular mass 90,000) that is critical in the pathogenesis of anthrax. It is a highly specific protease that cleaves members of the mitogen-activated protein kinase kinase (MAPKK) family near to their amino termini, leading to the inhibition of one or more signalling pathways. Here we describe the crystal structure of LF and its complex with the N terminus of MAPKK-2. LF comprises four domains: domain I binds the membrane-translocating component of anthrax toxin, the protective antigen (PA); domains II, III and IV together create a long deep groove that holds the 16-residue N-terminal tail of MAPKK-2 before cleavage. Domain II resembles the ADP-ribosylating toxin from Bacillus cereus, but the active site has been mutated and recruited to augment substrate recognition. Domain III is inserted into domain II, and seems to have arisen from a repeated duplication of a structural element of domain II. Domain IV is distantly related to the zinc metalloprotease family, and contains the catalytic centre; it also resembles domain I. The structure thus reveals a protein that has evolved through a process of gene duplication, mutation and fusion, into an enzyme with high and unusual specificity.     

Effect of mutation in ribosomal protein on translation elongation

Translation accuracy, translation elongation rate and translation processivity are three important parameters for each elongation cycle. To study the role of ribosomal protein in translation elongation, these parameters were measured simultaneously for the first time in threeEscherichia coli T83 mutantsrpsL, rplX andrpmA. The results suggested that the mutants of ribosomal protein S12(rpsL) and ribosomal protein L24(rplX) decreased the processivity of ribosome while the mutant of ribosomal protein L27(rpmA) had little effect on the processivity;rplX mutant andrpmA mutant had higher accuracy; the elongation rate ofrplX mutant was faster than that of the wild type, whereas the elongation rate ofrpmA mutant was the same as that of the wild type.     

Synthesis and properties of crosslinked recombinant pro-resilin

Resilin is a member of a family of elastic proteins that includes elastin, as well as gluten, gliadin, abductin and spider silks. Resilin is found in specialized regions of the cuticle of most insects, providing low stiffness, high strain and efficient energy storage; it is best known for its roles in insect flight and the remarkable jumping ability of fleas and spittle bugs. Previously, the Drosophila melanogaster CG15920 gene was tentatively identified as one encoding a resilin-like protein (pro-resilin). Here we report the cloning and expression of the first exon of the Drosophila CG15920 gene as a soluble protein in Escherichia coli. We show that this recombinant protein can be cast into a rubber-like biomaterial by rapid photochemical crosslinking. This observation validates the role of the putative elastic repeat motif in resilin function. The resilience (recovery after deformation) of crosslinked recombinant resilin was found to exceed that of unfilled synthetic polybutadiene, a high resilience rubber. We believe that our work will greatly facilitate structural investigations into the functional properties of resilin and shed light on more general aspects of the structure of elastomeric proteins. In addition, the ability to rapidly cast samples of this biomaterial may enable its use in situ for both industrial and biomedical applications.     

New developments in enzymatic peptide synthesis.

This review article describes new enzymatic methods developed for the efficient and irreversible synthesis of peptides based on native and modified proteases, and for the synthesis of polypeptides containing D- and/or unnatural amino acids. Potential opportunities for future developments in the field based on new enzymes, tailor-made catalytic antibodies, and on the technique of in vitro mutagenesis are also described.     

Further studies on the effect of red and far red light on rat retinal development

Summary The influence on the development of the outer segments of the rat retina of far red and red light in different sequences was investigated. The far red treatment appeared to be dominating, and for animals treated with far red light first, further treatment of red light could not bring the outer segment growth back to normal. The treatments also initiated different dopamine uptakes in the retinas.