Truncated thioredoxin (Trx80) exerts unique mitogenic cytokine effects via a mechanism independent of thiol oxido-reductase activity

Recently we discovered that a naturally occurring C-terminally truncated thioredoxin (Trx80) is a potent mitogenic cytokine stimulating IL-12 production from CD40(+) monocytes. To further characterise Trx80 we have engineered cysteine to serine mutants of Trx80 corresponding to the active site cysteines of Trx (Trx80SGPS) and to the structural cysteine at position 72 (Trx80C72S). Trx80SGPS and Trx80C72S retained the cell stimulatory activity of Trx80 and increased peripheral blood mononuclear cell (PBMC) proliferation three- to five-fold in vitro (P<0.01, n=18). Both Trx80SGPS and Trx80C72S significantly stimulated IL-12 and IFN-gamma secretion from PBMCs in the same manner as Trx80 (P<0.01, n=9 and 10). The previously described Trx80 dimer is caused by non-covalent interactions, and not by any intermolecular disulphide bonds.     

Eclipse-synchrony relationship in Escherichia coli strains with mutations affecting sequestration, initiation of replication and superhelicity of the bacterial chromosome

Initiation of replication from oriC on the Escherichia coli chromosomes occurs once and only once per generation at the same cell mass per origin. During rapid growth there are overlapping replication cycles, and initiation occurs synchronously at two or more copies of oriC. Since the bacterial growth can vary over a wide range (from three divisions per hour to 2.5 hours or more per division) the frequency of initiation should change in coordination with bacterial growth. Prevention of reinitiation from a newly replicated origin by temporary sequestration of the hemi-methylated GATC-sites in the origin region provides the molecular/genetic basis for the maintenance of the eclipse period between two successive rounds of replication. Sequestration is also believed to be responsible for initiation synchrony, since inactivation of either the seqA or the dam gene abolishes synchrony while drastically reducing the eclipse. In this work, we attempted to examine the functional relationship(s) between the eclipse period and the synchrony of initiation in E.coli strains by direct measurements of these parameters by density-shift centrifugation and flow-cytometric analyses, respectively. The eclipse period, measured as a fraction of DNA-duplication times, varied continuously from 0.6 for the wild-type E.coli K12 to 0.1 for strains with mutations in seqA, dam, dnaA, topA and gyr genes (all of which have been shown to cause asynchrony) and their various combinations. The asynchrony index, a quantitative indicator for the loss of synchrony of initiation, changed from low (synchronous) to high (asynchronous) values in a step-function-like relationship with the eclipse. An eclipse period of approximately 0.5 generation time appeared to be the critical value for the switch from synchronous to asynchronous initiation.     

Activation of ERK1/2 after neonatal rat cerebral hypoxia-ischaemia

Activation of extracellular signal-related kinases (ERK1/2), also known as p42/44 mitogen-activated protein kinase (MAPK), is considered important for neuronal survival, cell proliferation and apoptosis. In the present study, activation (phosphorylation) of ERK1/2 (P-ERK) was investigated in brains of 7-day-oldrats after hypoxia-ischaemia (HI). In damaged areas, P-ERK-positive neurons appeared immediately after HI and the staining remained for at least 8 h. At later time points, 24 and 72 h post-HI, P-ERK-positive neurons were found in the core of the infarct and in the border zone to undamaged tissue. These cells also showed signs of DNA damage and calpain-induced fodrin breakdown, indicative of injury. At 72 h post-HI, P-ERK was also observed in microglia in the border zone to the damaged area and in astrocytes and oligodendrocytes in white matter of both hemispheres. P-ERK was strongly expressed in the subventricular zone in both hemispheres after HI at most time points, although the staining in the ipsilateral (damaged hemisphere) was stronger than in the contralateral (non-damaged hemisphere). In summary, ERK1/2 activation occurred early in neurons after HI in the neonatal brain, and mainly in cells displaying signs of damage.     

Glucagon-like peptide-1 (7-36) amide prevents the accumulation of pyruvate and lactate in the ischemic and non-ischemic porcine myocardium

Glucagon-like peptide-1 (7-36) amide (GLP-1) has been studied as a treatment option in diabetic patients. We investigated the effect of recombinant GLP-1 infusion on hemodynamic parameters, myocardial metabolism, and infarct size during normoxic conditions as well as during ischemia and reperfusion using an open-chest porcine heart model. In the presence of rGLP-1, interstitial levels of pyruvate and lactate decreased during ischemia and reperfusion both in ischemic and non-ischemic tissue. Moreover, rGLP-1 infusion resulted in increased plasma insulin levels and decreased blood glucose levels. Neither hemodynamic variables nor the consequent infarct size were influenced by rGLP-1 infusion. We conclude that rGLP-1 altered myocardial glucose utilization during ischemia and reperfusion. It did not exert any untoward hemodynamic effects.     

Area specificity and topography of thalamocortical projections are controlled by ephrin/Eph genes

The mechanisms generating precise connections between specific thalamic nuclei and cortical areas remain poorly understood. Using axon tracing analysis of ephrin/Eph mutant mice, we provide in vivo evidence that Eph receptors in the thalamus and ephrins in the cortex control intra-areal topographic mapping of thalamocortical (TC) axons. In addition, we show that the same ephrin/Eph genes unexpectedly control the inter-areal specificity of TC projections through the early topographic sorting of TC axons in an intermediate target, the ventral telencephalon. Our results constitute the first identification of guidance cues involved in inter-areal specificity of TC projections and demonstrate that the same set of mapping labels is used differentially for the generation of topographic specificity of TC projections between and within individual cortical areas.     

A missense mutation in ftsZ differentially affects vegetative and developmentally controlled cell division in Streptomyces coelicolor A3(2)

Streptomyces coelicolor A3(2) undergoes at least two kinds of cell division: vegetative septation leading to cross-walls in the substrate mycelium; and developmentally regulated sporulation septation in aerial hyphae. By isolation and characterization of a non-sporulating ftsZ mutant, we demonstrate a difference between the two types of septation. The ftsZ17(Spo) allele gave rise to a classical white phenotype. The mutant grew as well as the parent on plates, and formed apparently normal hyphal cross-walls, although with a small reduction in frequency. In contrast, sporulation septation was almost completely abolished, resulting in a phenotype reminiscent of whiH and ftsZdelta2p mutants. The ftsZ17(Spo) allele was partially dominant and had no detectable effect on the cellular FtsZ content. As judged from both immunofluorescence microscopy of FtsZ and translational fusion of ftsZ to egfp, the mutation prevented correct temporal and spatial assembly of Z rings in sporulating hyphae. Homology modelling of S. coelicolor FtsZ indicated that the mutation, an A249T change in the C-terminal domain, would be expected to alter the protein on the lateral face of FtsZ protofilaments. The results suggest that cytokinesis may be developmentally controlled at the level of Z-ring assembly during sporulation of S. coelicolor A3(2).     

Growth polarity and cell division in Streptomyces

Streptomycetes are mycelial bacteria that resemble filamentous fungi in their apical growth, branching, and morphogenetic development. One inroad into the largely unknown mechanisms underlying this prokaryotic growth polarity is provided by Streptomyces DivIVA, a protein localized at hyphal tips and involved in tip extension. Another aspect is a proposed migration of nucleoids. During sporulation, the modes of growth and cell division are reorganised. This involves dynamic assembly of FtsZ into a multitude of cytokinetic rings. Controlled by developmental regulators and intriguingly coordinated with chromosome segregation, this leads to spores with a single chromosome each. Genome sequences have shed new light on these aspects and reinforced the role of Streptomyces in bacterial cell biology.     

Mast cell activation and tissue cell proliferation

Summary The effect of mast cell activation and degranulation on the proliferation in the intact mesentery was studied in Sprague-Dawley rats. Mast cell activation was achieved by a single intraperitoneal injection of Compound 48/80.The proliferation was studied using three independent methods for estimation of cell production and DNA synthesis: 1. the mitotic index, 2. the relative number of cells having a DNA content in the S and G2 regions, by Feulgen photometric measurement in individual cells, and 3. the specific DNA activity, employing a method which combines a liquid scintillation technique after an intravenous injection of ³H-thymidine and Feulgen photometric determination of the DNA content per membrane preparation.It was found that the proliferation of the normal mesenchymal cells adjacent to the activated and degranulated mast cells in the mesentery was significantly increased within 24 and 32 h, the maximum increase being more than 20-fold compared to untreated controls. The results suggest that the common type of mast cell may have a pathophysiological function related to stimulation of local cell proliferation.Supported by grants from the Swedish Medical Research Council (Project 12X-2235) and from the Medical Faculty, University of LinköpingWe thank Brita Söderlund, Margareta Odenö and Iréne Svensson for skilful technical assistance, and Erik Leander, Ph. D., for help with statistical methodsPart of this work was presented at the 7th Meeting of the European Study Group for Cell Proliferation, 5–9 May 1975, in Amsterdam, The Netherlands