Recombinant soluble tumor necrosis factor receptor proteins protect mice from lipopolysaccharide-induced lethality.

The in vivo efficacy of human recombinant soluble tumor necrosis factor (TNF) receptor protein to prevent and to treat lipopolysaccharide (LPS)-induced lethal toxicity in D-galactosamine-treated mice was investigated. Chimeric proteins of the receptor extracellular domains fused to the hinge region of human IgG3 were expressed in myeloma cells (rsTNFR-h gamma 3). The fusion proteins had a disulfide-bonded dimeric structure. Upon intravenous injection, their serum concentration decreased relatively slowly after an initial phase of rapid elimination. D-galactosamine-sensitized mice were fully protected from the toxic effects of LPS, if the animal were pretreated with rsTNFR-h gamma 3 at 20 micrograms/animal. Partial protection was seen at significantly lower doses and when rsTNFR-h gamma 3 was given up to 3 h after LPS.     

Involvement of tumour necrosis factor and other cytokines in immune-mediated vascular pathology

Vascular endothelial cells are actively involved in coagulation and inflammation processes and appear to represent an important element in cell-mediated immune responses. In this paper, the possible role of endothelial cells as a target for immunopathological reactions was analyzed. Experimental neurovascular lesions were studied in a model of cerebral malaria, with particular attention to the role of cytokine interactions in vivo.     

CD4-independent protective cytotoxic T cells induced in early life by a non-replicative delivery system based on virus-like particles

The relative immaturity of the neonatal immune system limits CD4(+) Th1 and cytotoxic T lymphocyte (CTL) responses, and represents a significant challenge for the development of vaccines against intracellular pathogens. In this report, we demonstrate the ability of a non-replicative delivery system based on parvovirus-like particles (VLP) to induce CTL responses in the neonatal period. A single immunization of 1-week-old BALB/c mice with recombinant VLP carrying a CD8(+) T cell determinant from lymphocytic choriomeningitis virus (VLP-LCMV) induced antigen-specific CD8(+) cytotoxic T cells that were similar to those elicited by adult immunization, as assessed by cytotoxic activity, interferon (IFN)-gamma secretion, cytotoxic precursor cell frequencies, in vitro avidity for antigen and protective activity against viral challenge. These CTL responses are elicited within 2 weeks of a single immunization, in the absence of adjuvant and independently of the presence and help of CD4(+) T cells, highlighting the potential of VLP as candidate vaccine vectors in early life.     

A role for lymphotoxin beta receptor in host defense against Mycobacterium bovis BCG infection

To investigate the role of membrane lymphotoxin (LT)alpha1 / beta2 and its LTbeta receptor (LTbetaR) in the protective immune response to Mycobacterium bovis bacillus Calmette-Guérin (BCG) infection, we have used a soluble fusion molecule (LTbetaR-IgG1). LTbetaR-Ig treatment interferes with granuloma formation mainly in the spleen by inhibiting macrophage activation and nitric oxide synthase activity. In addition, a large accumulation of eosinophils was observed in the spleen of LTbetaR-Ig-treated infected mice. Decreased blood levels of IFN-gamma and increased IL-4 were also observed, suggesting that the LTbetaR pathway is important in BCG infection to favor a Th1 type of immune response. The treatment of transgenic mice expressing high blood levels of a soluble TNFR1-IgG3 fusion protein with LTbetaR-Ig resulted in a still higher sensitivity to BCG infection, and extensive necrosis in the spleen. In conclusion, these results suggest that the LTbetaR and the TNFR pathways are not redundant in the course of BCG infection and protective granuloma formation: the LTbetaR pathway appears to be important in spleen granuloma formation, whereas the TNFR pathway has a predominant role in other tissues.     

Presence of a very small population of Thy-1+, L3T4+ cells producing large amounts of IL-3 in young athymic nude mice.

A mixture of phorbol myristate acetate (PMA) and ionomycin was found to stimulate spleen and lymph node cells (LNC) from 6 to 8 week-old-athymic BALB/c nude mice, as well as from control +/+ mice, to secrete interleukin-3 (IL-3) in vitro. The specificity of the IL-3 bioassay was attested to by the use of rabbit anti-IL-3 antibodies, and by the detection of an accumulation of IL-3 mRNA. Cytotoxic treatment with relevant antibodies showed that the cells responsible for the IL-3 production in athymic nude mice was Thy-1+, L3T4+, Ly2-, while both L3T4+ and Ly 2+ cells produced IL-3 in control +/+ mice. Although the levels of IL-3 secreted by nude LNC varied among experiments, nude LNC were able to produce IL-3 at a level comparable to or higher than +/+ LNC. In addition, nude LNC consistently secreted two to three times more granulocyte-macrophage colony-stimulating factor (GM-CSF) than +/+ LNC, and the majority of GM-CSF secretion was dependent on the presence of L3T4+ cells. In contrast, IL-2 production by nude LNC was markedly limited. Since the flow microfluorometry analysis failed to demonstrate the presence of L3T4+ cells (less than 1%) in nude LNC, compared with 40-50% L3T4+ cells in +/+ LNC, our results suggest that athymic nude mice have a small population of Thy-1+, L3T4+ cells characterized by its ability to secrete IL-3 and GM-CSF at a very high rate.     

Aggravation of experimental cutaneous leishmaniasis in mice by administration of interleukin 3

Previous studies from our laboratory have shown that some in vitro maintained Leishmania major-specific L3T4+ T cells were capable of exacerbating cutaneous leishmaniasis after adoptive transfer to normal syngeneic mice. Results presented in this report show that these cells released substantial amounts of interleukin 3 (IL 3) and granulocyte-macrophage colony-stimulating factors after specific stimulation in vitro. In order to assess the involvement of such lymphokines in the exacerbation of cutaneous leishmaniasis by these L3T4+ T cells, the effect of the administration of important doses of IL 3 on the course of infection with L. major was investigated. The treatment of genetically susceptible BALB/c mice with IL 3 resulted in an enhancement of the size of lesions and favored the multiplication of parasites at anatomical sites distant from the primary lesion. Although IL 3 did not modify the development of lesions in genetically resistant CBA mice, this lymphokine promoted the growth of Leishmania in lymph node draining the lesion. Finally, the addition of IL 3 to macrophages parasitized in vitro enhanced the survival of intracellular Leishmania major.     

Association of RANTES G-403A gene polymorphism with increased risk of coronary arteriosclerosis

Aims Polymorphisms in the RANTES (G-403A), monocyte chemoattractantprotein-1 (MCP-1; A-2518G), stromal cell-derived factor-1ß(SDF-1ß; G801A), and C–C chemokine receptor-5(CCR5; 32) genes have been associated with functional effects.These chemokines have been implicated in leucocyte recruitmentto arterial lesions. In a case-control study, we explored relationsbetween these polymorphisms and coronary artery disease (CAD),with respect to angiographic abnormalities and acute coronarysyndromes (ACS). Methods and Results The LUdwigshafen Risk and Cardiovascularhealth (LURIC) cohort was genotyped by RFLP-PCR. Based on coronaryangiography, individuals were sub-divided into CAD cases and controls . RANTES-403 genotype frequencies were significantly different in cases and controls, as were A allele carrier frequencies (36.01% vs. 30.19%, OR=1.30 [95%-CI=1.06–1.60], ). By multivariate analysis, RANTES A-403 retained significantassociation with CAD . RANTES A-403 was associated with increased ACS prevalence (OR=1.36 [95%-CI=1.08–1.71],). MCP-1 G-2518, SDF-1ß A801, and CCR5 32 were not associated with CAD. Conclusions RANTES A-403 was associated with CAD independentlyfrom conventional risk factors and CRP or fibrinogen as inflammatorybiomarkers. The association was enhanced in smokers and ACS,conditions where platelet activation and inflammation predominate.RANTES A-403 may increase genetic susceptibility to CAD.     

What is the meaning of colorectal transit time measurement?

This study was done to understand the different available methods used to calculate colorectal transit times. A single abdominal radiograph is taken following six successive daily ingestions of the same number of identical radiopaque markers. This method correlates well (P < 0.001) with that using a single ingestion of markers with daily x-ray films until total expulsion. In techniques used to measure colorectal transit time with multiple ingestion of markers, the number of days of ingestion depends on the kinetics of marker defecation. This was found to differ markedly in various groups of control subjects and constipated patients (P < 0.001) and can be used to obtain reliable data, even in subjects with severe constipation. When they ingest 20 markers, constipated patients are found to retain eight or more markers three days after ingestion, and taking a plain film of the abdomen on that day is sufficient to make a diagnosis of constipation. Transit time studies are reproducible from month to month in patients with an irritable bowel syndrome. Control subjects who claim that their bowel habits are not modified by stress have shorter transit times, similar in both sexes, than those who say they are (P <0.001). This may explain why a large percentage of constipated patients have been found by most authors to have normal colorectal transit times. The choice of control subjects is thus a key element in studies of functional bowel motor disorders. Stool frequency and consistency, in health, correlate only to rectosigmoid transit time.     

Evidence for hydrophobic region within heavy chains of mouse B lymphocyte membrane-bound IgM

The gel filtration behavior, in the presence of detergents, of membrane-bound IgM from normal mouse spleen B lymphocytes was compared to that of secretory IgM from mouse plasma cells. The proteins were labeled either by surface radioiodination or biosynthetically with radioactive amino acids. Cell lysates were fractionated on calibrated Sepharose 6B columns in the presence of the detergents Nonidet P-40 or deoxycholate. Eluted fractions were immunoprecipitated and the reduced or unreduced precipitates were analyzed by sodium dodecyl sulfate gel electrophoresis followed by radioautography. Surface (125)I-labeled 8S IgM exhibited a gel filtration pattern in Nonidet P-40 corresponding to much higher apparent molecular weight than that of secretory 8S IgM, a difference that almost disappeared when gel filtration was performed in the presence of deoxycholate, which forms much smaller micelles than does Nonidet P-40. Biosynthetically labeled lymphocytes contain two types of IgM molecules differing in their gel filtration behavior and fate: one identical to secretory 8S IgM of plasma cells and secreted in the medium during chase periods, and the other identical to surface (125)I-labeled IgM and remaining cell-associated. Because the surface-bound 8S IgM was not found to be associated with other labeled molecules, it is likely that the detergent-binding behavior of surface IgM is due to a hydrophobic segment carried by these Ig molecules. That lymphocytes synthesize two types of mu chains was also shown by the use of tunicamycin, an inhibitor of glycosylation. In its presence, two unglycosylated mu chains were observed: one identical in size to that made by tunicamycin-treated plasma cells, and the second slightly larger. Gel filtration in Nonidet P-40 of the cell lysates of tunicamycin-treated lymphocytes showed that the nonsecretory 8S IgM contains this second type of mu chains, whereas the IgM molecules of the secretory type contain plasma cell-like mu chains. It is suggested that membrane IgM mu chains contain a hydrophobic segment which is responsible for its association to the membrane.