Cytokine production in the immune response to murine gammaherpesvirus 68.

Cytokine profiles were determined following intranasal infection of C57BL/6J mice with murine gammaherpesvirus 68 (MHV-68). Spleen, mediastinal, and cervical lymph node cells from infected mice produced high levels of interleukin 6 (IL-6) and gamma interferon (IFN-gamma) and lower levels of IL-2 and IL-10 following in vitro restimulation. Little or no IL-4 or IL-5 was detected. Cytokine production was generally maximal at 10 days after infection, correlating with viral clearance from the lung, although significant levels were seen as early as 3 days after administration of the virus. In vitro infection of naive splenocytes induced B-cell- dependent secretion of IL-6 and IL-10, whereas IFN-gamma and IL-2 were produced only by cells that had been primed in vivo. Depletion of B lymphocytes from primed splenocyte populations did not, however, abrogate IL-6 and IL-10 production. Highly purified immune T cells made IL-6, IL-10. and IFN-gamma following in vitro restimulation with MHV-68. Thus, IL-6 and IL-10 are components of both the acquired and the innate host response. These cytokines have potential roles in the establishment and maintenance of persistent infection.     

Induction of cytokines in mice with parainfluenza pneumonia.

The possible involvement of cytokines in the acute viral pneumonia induced by the murine parainfluenza type 1 virus, Sendai virus, was studied. Cytokine profiles for both the respiratory tract and the draining mediastinal lymph node (MLN) of virus-infected C57BL/6J mice were quantified by using the single-cell cytokine (ELISPOT) assay with freshly isolated cell populations and enzyme-linked immunosorbent assay for lung lavage fluids and culture supernatants. Maximal levels of interleukin 2 (IL-2), gamma interferon (IFN-gamma), tumor necrosis factor, IL-6, and IL-10 were detected at the inflammatory site 7 to 10 days after infection, about the time that virus is cleared from the lung. The frequencies of cells producing IL-2, IL-4, IL-6, IL-10, IFN-gamma, and tumor necrosis factor were much higher for the bronchoalveolar lavage (BAL) cell population than for the MLN cell population. Cytokine production after in vitro restimulation of MLN cells was dominated by IL-2 and IFN-gamma, with low levels of IL-10 and IL-6 also being present. Most of the cytokine was produced by the CD4+ cells, although the CD8+ subset was also involved. No IL-4 was found in the BAL fluid or in culture supernatants from restimulated BAL or MLN cells, although a high frequency of IL-4-producing cells was demonstrated in the BAL population by ELISPOT analysis.     

Differential antigen burden modulates the gamma interferon but not the immunoglobulin response in mice that vary in susceptibility to Sendai virus pneumonia.

Sendai virus, a paramyxovirus which causes murine pneumonia, grew to approximately 10-fold higher titers and was cleared less rapidly from the lungs of 129/J (129) than H-2b-compatible C57BL/6J (B6) mice. The more susceptible 129 mice also made higher titers of gamma interferon (IFN-gamma) and immunoglobulin G2a (IgG2a) virus-specific antibody. Analysis with acutely irradiated (950 rads) mice and immunologically reconstituted bone marrow (BM) radiation chimeras indicated that the enhanced virus growth was a function of the radiation-resistant respiratory epithelium. Prolonged exposure to more virus in turn influenced the magnitude of IFN-gamma production, most of which was made by CD4+ T lymphocytes. Somewhat surprisingly, however, the 129 pattern of a higher virus-specific serum Ig response skewed towards IgG2a mapped to the reconstituting BM. Thus, the characteristics of the humoral response are at least partly dissociated from both the antigen load, resulting from viral replication, and the level of IFN-gamma production. Further analysis of double chimeras (B6+129 BM-->B6 recipients) confirmed that the divergent humoral immune response to Sendai virus in B6 and 129 mice is largely determined by the inherent characteristics of the lymphoid cells.     

Separation of growth-stimulating activity of BSA fraction V from the bulk of albumin using Heparin Sepharose Chromatography

Bovine serum albumin (BSA) is a potential source of biological contamination in cell culture medium. The aim of this work was to attempt to replace BSA in low serum and serum-free medium (SFM). BSA fraction V was subjected to a variety of processes in order to determine if the growth promoting activity observed for NRK cells could be extracted from the BSA molecule. These included solvent extractions, diafiltration, reverse phase HPLC and affinity chromatography using heparin sepharose. Solvent extraction and diafiltration failed to remove the activity from the BSA. Affinity chromatography using heparin sepharose indicated that all of the activity observed with BSA was retained in the 0.5 M NaCl fraction and was associated with less than 3% of the original protein. The major protein band in the 0.5 M NaCl fraction had the same apparent molecular weight as albumin (as seen by SDS-PAGE and analytical reverse phase HPLC). Unlike the untreated BSA, the 0.5 M NaCl fraction was partially susceptible to proteolytic digestion and to variations in pH.Abbreviations HS heparin sepharose - DHS donor horse serum - SFM serum free-medium     

Microhabitat use by 0+ and older fishes in a small English chalk stream

The microhabitat use of two size/age classes of fish (0 +, ≥1+) in the River Lee, based on measurements of 14 environmental variables, was studied using point abundance sampling by electrofishing over the summer and autumn of 1995. Microhabitat use by all cyprinid species (barbel Barbus barbus, minnow Phoxinus phoxinus , chub Leuciscus cephalus and gudgeon Gobio gobio ) differed between 0+ and older (≥1+) with lentic, shallow, littoral environments being important for 0+ fishes, whereas deeper, faster areas in mid-channel were important for ≥ 1 + fishes. There was more overlap in microhabitat use by 0+ juvenile cyprinids in the River Lee than in larger systems such as the River Danube (Slovakia/Hungary) and River Great Ouse (U.K.).     

Modulation of murine AIDS-related pathology by concurrent antibody treatment and coinfection with Leishmania major.

Infection of C57BL/6 mice with a mixture of murine leukemia viruses (MuLVs) designated LP-BM5 MuLV leads to a disease characterized by progressive immunodeficiency and lymphoproliferation, known as murine AIDS (MAIDS). The development of MAIDS is associated with increased B-cell lymphoblast proliferation, but there is reason to believe that T-cell function and, particularly, T-cell-derived cytokines may also play a role. We have previously shown that concurrent infection with Leishmania major (which induces a strongly polarized Th1 response in C57BL/6 mice) and LP-BM5 MuLV modulates the disease induced by both infections. Here we show by treatment of mice with anticytokine antibodies that this modulation is largely exerted through the balance of Th1 and Th2 cytokines. Infected mice treated with antibodies to interleukin-4 and interleukin-10 exhibited a delayed development of MAIDS-related pathology and maintained T-cell responsiveness longer than mice treated with control antibody. Gamma interferon induced by coinfection with L. major synergized with anti-IL-4 treatment to inhibit the development of MAIDS pathology. Conversely, treatment with anti-gamma interferon led to a significant increase in splenomegaly and lymphadenopathy and slightly exacerbated loss of T-cell function. These data suggest that the production of Th2-associated cytokines may promote MAIDS pathology, while Th1-associated cytokines may help control the disease.     

The cytotoxic T-lymphocyte response to Sendai virus is unimpaired in the absence of gamma interferon.

Sendai virus is eliminated from the respiratory tract of gamma interferon (IFN-gamma) -/- BALB/c mice with normal kinetics. The level of virus-specific cytotoxic T-lymphocyte (CTL) activity in the cell population recovered by bronchoalveolar lavage is unimpaired, the prevalence of interleukin-4 (IL-4)-producing cells is increased, and the titers of virus-specific immunoglobulins IgG1 and IgG2b are higher in the IFN-gamma -/- mice. The emergence of this T-helper 2 response profile in both lymphoid tissue and the pneumonic lung has no obvious deleterious consequences. Virus clearance is slightly delayed following depletion of the CD4+ subset, with the effect being similar in magnitude for IFN-gamma -/- and +/+ mice. However, the generation of CTL precursors (CTLp) is diminished in the IFN-gamma -/- (but not +/+) mice in the absence of concurrent CD4+ T help. Apparently the clonal expansion of the CTLp population can be promoted either by a cytokine (perhaps IL-2) produced by the IFN-gamma -/- CD4+ T cells or by IFN-gamma made by other cell types in the +/+ mice.     

GENETIC AND COLOR INTERACTIONS AT A CONTACT ZONE OF ACANTHOCHROMIS POLYACANTHUS: A MARINE FISH LACKING PELAGIC LARVAE

Abstract. — Acanthochromis polyacanthus is an unusual tropical marine damselfish that uniquely lacks pelagic larvae and has lost the capacity for broad-scale dispersal among coral reefs. Different color morphs exist in different regions of the Great Barrier Reef, and morphs from northern and southern regions are genetically distinct. In the Hydrographers Passage area, which is a large break through the reef matrix in the central Great Barrier Reef that may have acted as a bottleneck on the migration of these animals during sea level rise, three morphs recognized from other regions were found on neighboring reefs. The transition between them is abrupt with three loci (AAT-2*, GPI-1*, and PGM*) showing allelic frequency patterns close to fixation between opposite alleles within a few kilometers. On two reefs (Hyde, Bebe), a pair of morphs was found to coexist and exhibited a habitat partitioning pattern with each morph restricted to one side on the reef and steep transitions in between. Outside these transition zones, phenotypes and genotypes matched those on surrounding reefs without coexistence and were little changed from reefs several hundred kilometers away. An electrophoretic survey across one transition zone on Hyde Reef showed steep genetic gradients along one kilometer of reef slope. Significant linkage disequilibria in samples collected in Hyde Reef as a result of dispersal of parental combinations of alleles into the center or because parental combinations of alleles confer greater fitness, allowed us to estimate the dispersal rate (189 m/generation) and the selection pressure on the marker loci (0.411). Finally, we investigated models that could lead to such a steep transition in genotypic and phenotypic combinations. Both contact zones on each side of Hyde Reef were associated with geomorphological discontinuities in the reef structure. We suggest that assortative mating may be a proximal mechanism for maintaining isolated each color morph, which could be reinforced by selective predation against hybrids outside the zone of their formation (i.e., the frequency-dependent selection model of Mallet and Barton (1989). Acanthochromis is a midwater planktivore and, when in coexistence, the two morphs forage in different habitats amid multispecific flocks of other damselfishes of matching colors.     

The nfkb1 promoter is controlled by proteins of the Ets family.

The gene encoding NFKB1 is autoregulated, responding to NF-kappa B/Rel activation through NF-kappa B binding sites in its promoter, which also contains putative sites for Ets proteins. One of the Ets sites, which we refer to as EBS4, is located next to an NF-kappa B/Rel binding site, kB3, which is absolutely required for activity of the promoter in Jurkat T cells in response to activation by phorbol 12-myristate 13-acetate (PMA), PMA/ionomycin, or the Tax protein from human T cell leukemia virus type I. We show that EBS4 is, required for the full response of the nfkb1 promoter to PMA or PMA/ionomycin in Jurkat cells. EBS4 is bound by Ets-1, Elf-1, and other species. Overexpression of Ets-1 augments the response to PMA/ionomycin and this is reduced by mutation of EBS4. Elf-1 has less effect in conjunction with PMA/ionomycin, but by itself activates the promoter 12-fold. This activation is only partly affected by mutation of EBS4, and a mutant promoter that binds Ets-1, but not Elf-1, at the EBS4 site responds to PMA/ionomycin as efficiently as the wild-type. Ets proteins may be responsible for fine-tuning the activity of the nfkb1 gene in a cell-type-specific manner.