The influence of age and sex on phagocyte chemiluminescence

The process of ageing is associated with increased susceptibility to infection. Phagocytes form the primary defence mechanism against infecting microorganisms, but the influence of ageing on phagocyte function remains controversial. In this study we have applied a microtitre plate phagocyte chemiluminescence (CL) assay suitable for clinical use to compare phagocyte oxidative metabolism in younger healthy subjects (age 20–60 years) and healthy older (60–70 years) subjects. Polymorphonuclear leukocytes (PMNL) and monocytes were stimulated using phorbol myristate acetate (PMA), serum opsonized zymosan (SOZ), and non-opsonized zymosan (ZYM) in the presence of both lucigenin and luminol. Monocytes showed a higher luminolenhanced CL response to PMA in males compared with females in the younger age group. No PMNL differences were observed between the sexes. Although no difference were found in relation to age when cells were stimulated with PMA and SOZ, significantly lower background (unstimulated) CL was obtained from PMNL with luminol. PMNL luminol-enhanced CL responses were also lower in response to ZYM. The findings suggest a reduced response of PMNL from older subjects to minimal stimulation. This could be related to abnormalities in the triggering of the respiratory burst or myeloperoxidase release due to ageing. The influence of age and sex should be taken into account in clinical studies of phagocyte CL.     

Dynamic resource allocation between pre- and postcopulatory episodes of sexual selection determines competitive fertilization success

In polyandrous mating systems, male reproductive success depends on both mate-acquisition traits (precopulatory) and sperm competitive abilities (postcopulatory). Empirical data on the interaction between these traits are inconsistent; revealing positive, negative or no relationships. It is generally expected that the investment in pre- and postcopulatory traits is mediated by environmental conditions. To test how dietary resource availability affects sexual ornamentation, sperm quality and their interrelationship in three-spined sticklebacks (Gasterosteus aculeatus), full-sibling groups were raised under three conditions differing in food quantity and/or quality (i.e. carotenoid content): (i) high-quantity/high-quality, (ii) high-quantity/low-quality or (iii) low-quantity/low-quality. After 1 year of feeding, food-restricted males developed a more intense breeding coloration and faster sperm compared with their well-fed brothers, indicating that they allocated relatively more in pre- and postcopulatory traits. Moreover, they outcompeted their well-fed, carotenoid-supplemented brothers in sperm competition trials with equal numbers of competing sperm, suggesting that food-restricted males maximize their present reproductive success. This may result in reduced future reproductive opportunities as food-restricted males suffered from a higher mortality, had an overall reduced body size, and sperm number available for fertilization. In accordance with theory, a trade-off between the investment in pre- and postcopulatory traits was observed in food-restricted males, whereas well-fed males were able to allocate to both traits resulting in a significantly positive relationship.     

5S ribosomal gene clusters in wheat: pulsed field gel electrophoresis reveals a high degree of polymorphism

Summary The long-range structure of 5S rRNA gene clusters has been investigated in wheat (Triticum aestivum L.) by means of pulsed field gel electrophoresis. Using aneuploid stocks, 5S rRNA gene clusters were assigned to sites on chromosomes 1B, 1D, 513 and 5D. Cluster sizes were evaluated and the copy number of 5S DNA repeats was estimated at 4700-5200 copies for the short repeating unit (410 bp) and about 3100 copies for the long repeat (500 bp) per haploid genome. A comparison of wheat cultivars revealed extremely high levels of polymorphism in the 5S rRNA gene clusters. With one restriction enzyme digest all varieties tested gave unique banding patterns and, on a per fragment basis, 21-fold more polymorphism was detected among cultivars for 5S DNA compared to standard restriction fragment length polymorphisms (RFLPs) detected with single copy clones. Experiments with aneuploid stocks suggest that the 5S rRNA gene clusters at several chromosomal sites contribute to this polymorphism. A number of previous reports have shown that wheat cultivars are not easily distinguished by isozymes or RFLPs. The high level of variation detected in 5S rRNA gene clusters therefore offers the possibility of a sensitive fingerprinting method for wheat. 5S DNA and other macro-satellite sequences may also serve as hypervariable Mendelian markers for genetic and breeding experiments in wheat.     

Mycolactone toxin induces an inflammatory response by targeting the IL-1β pathway: Mechanistic insight into Buruli ulcer pathophysiology

Mycolactone, a lipid-like toxin, is the major virulence factor of Mycobacterium ulcerans, the etiological agent of Buruli ulcer. Its involvement in lesion development has been widely described in early stages of the disease, through its cytotoxic and immunosuppressive activities, but less is known about later stages. Here, we revisit the role of mycolactone in disease outcome and provide the first demonstration of the pro-inflammatory potential of this toxin. We found that the mycolactone-containing mycobacterial extracellular vesicles produced by M. ulcerans induced the production of IL-1β, a potent pro-inflammatory cytokine, in a TLR2-dependent manner, targeting NLRP3/1 inflammasomes. We show our data to be relevant in a physiological context. The in vivo injection of these mycolactone-containing vesicles induced a strong local inflammatory response and tissue damage, which were prevented by corticosteroids. Finally, several soluble pro-inflammatory factors, including IL-1β, were detected in infected tissues from mice and Buruli ulcer patients. Our results revisit Buruli ulcer pathophysiology by providing new insight, thus paving the way for the development of new therapeutic strategies taking the pro-inflammatory potential of mycolactone into account.     

Tumor necrosis factor induces acute phase proteins in rats

Inoculation of WAG rats with recombinant mouse tumor necrosis factor results in a rapid and marked increase in several acute phase proteins in the serum (haptoglobin, alpha 1 acid glycoprotein, alpha 2 macroglobulin) and in the plasma (fibrinogen). We conclude that TNF may play an important role in the inflammatory response in vivo and possibly in the pathogenesis of inflammatory disorders.     

The gene coding for the human T-lymphocyte CD2 antigen is located on chromosome 1p

Summary A cDNA clone encoding the human T lymphocyte sheep erythrocyte receptor [the CD2 (T11) antigen] was used as a probe to define the chromosomal location of the gene. The signal, revealed by hybridisation to Southern blots of genomic DNA from somatic cell hybrids, showed a high degree of concordance for human chromosome 1. In particular, the hybrid F4Sc13C19 which contained the short arm only of human chromosome 1 was positive. The location of the CD2 gene to 1p13 was confirmed by in situ hybridisation.     

Antibody to myelin constituents: A possible factor in induction of cell-mediated demyelination

Results from this laboratory have demonstrated that¹⁴C-labeled myelin opsonized with antibodies raised to purified CNS myelin in rabbit is phagocytized by cultured macrophages in larger amounts than untreated myelin or myelin opsonized with preimmune serum. The cultured macrophages produced high amounts of radioactive cholesterol ester and triglyceride from the antibody-treated myelin while much less was formed from preimmune serum-treated or untreated myelin. Antiserum to galactocerebroside also greatly enhanced the formation of radioactive cholesterol ester, while that to myelin basic protein as well as to other myelin constituents had little or no effect. Serum from Lewis rats with acute EAE 13–14 days after immunization with whole CNS myelin also stimulated radioactive cholesterol ester formation compared to serum from Freund's adjuvant-injected controls (FAC). Serum from EAE rats as a result of myelin basic protein injection was as active as that from rats with whole myelin injection. No galactocerebroside antibody could be demonstrated in the EAE sera, although a strong immunostaining to myelin basic protein and proteolipid protein was demonstrated. IgG prepared from EAE serum also showed stimulatory effects compared to IgG from FAC serum, but much of the activity was lost, and the possibility that other factors may be involved is discussed. These experiments provide evidence that myelin phagocytosis and digestion by macrophages is enhanced by the presence of antibody to myelin. In EAE this antibody may leak into CNS with the breakdown of the blood-brain barrier. A humoral involvement in demyelination in EAE is implicated, and these findings may be extended eventually to the demyelinative mechanism in multiple sclerosis where IgG is found in large amounts in the CNS.Special Issue Dedicated to Dr. Abel Lajtha.     

Carbonic anhydrase activity in acetate grown Methanosarcina barkeri

Cell extracts (27000xg supernatant) of acetate grown Methanosarcina barkeri were found to have carbonic anhydrase activity (0.41 U/mg protein), which was lost upon heating or incubation with proteinase K. The activity was inhibited by Diamox (apparent K _i=0.5 mM), by azide (apparent K _i=1 mM), and by cyanide (apparent K _i=0.02 mM). These and other properties indicate that the archaebacterium contains the enzyme carbonic anhydrase (EC 4.2.1.1). Evidence is presented that the protein is probably located in the cytoplasm. Methanol or H₂/CO₂ grown cells of M. barkeri showed no or only very little carbonic anhydrase activity. After transfer of these cells to acetate medium the activity was induced suggesting a function of this enzyme in acetate fermentation to CO₂ and CH₄. Interestingly, Desulfobacter postgatei and Desulfotomaculum acetoxidans, which oxidize acetate to 2 CO₂ with sulfate as electron acceptor, were also found to exhibit carbonic anhydrase activity (0.2 U/mg protein).     

Sporomusa malonica sp. nov., a homoacetogenic bacterium growing by decarboxylation of malonate or succinate

A new strictly anaerobic bacterium was isolated from an enrichment culture with glutarate as sole substrate and freshwater sediment as inoculum, however, glutarate was not metabolized by the pure culture. The isolate was a mesophilic, spore-forming, Gram-negative, motile curved rod. It fermented various organic acids, alcohols, fructose, acetoin, and H₂/CO₂ to acetate, usually as the only product. Other acids were fermented to acetate and propionate or acetate and butyrate. Succinate and malonate were decarboxylated to propionate or acetate, respectively, and served as sole sources of carbon and energy for growth. No inorganic electron acceptors except CO₂ were reduced. Yeast extract (0.05% w/v) was required for growth. Small amounts of cytochrome b were detected in membrane fractions. The guanine-plus-cytosine content of the DNA was 44.1±2 mol%. The isolate is described as a new species of the genus Sporomusa, S. malonica.