Biomedical subjects

K Pfeffer

Publications and source records attributed to K Pfeffer.

At least 19 recordsLinked to original sources

PubMed2005

Major histocompatibility complex (MHC2+) perivascular macrophages in the axotomized facial motor nucleus are regulated by receptors for interferon-gamma (IFNgamma) and tumor necrosis factor (TNF).

The major histocompatibility complex (MHC) glycoproteins, MHC1 and MHC2, play a key role in the presentation of antigen and the development of the immune response. In the current study we examined the regulation of the MHC2 in the mouse brain after facial axotomy. The normal facial motor nucleus showed very few slender and elongated MHC2+ cells. Transection of the facial nerve led to a gradual but strong upregulation in the number of MHC2+ cells, beginning at day 2 and reaching a maximum 14 days after axotomy, correlated with the induction of mRNA for tumor necrosis factor (TNF) alpha, interleukin (IL) 1beta and interferon-gamma (IFNgamma) and a peak in neuronal cell death. In almost all cases, MHC2 immunoreactivity was restricted to perivascular macrophages that colocalized with vascular basement membrane laminin and macrophage IBA1-immunoreactivity, with no immunoreactivity on phagocytic microglia, astrocytes or invading T-cells. Heterologous transplantation and systemic injection of endotoxin or IFNgamma did not affect this perivascular MHC2 immunoreactivity, and transgenic deletion of the IL1 receptor type I, or TNF receptor type 1, also had no effect. However, the deletion of IFNgamma receptor subunit 1 caused a significant increase, and that of TNF receptor type 2 a strong reduction in the number of MHC2+ macrophages, pointing to a counter-regulatory role of IFNgamma and TNFalpha in the immune surveillance of the injured nervous system.

Animals↗

PubMed2004

Microglial major histocompatibility complex glycoprotein-1 in the axotomized facial motor nucleus: regulation and role of tumor necrosis factor receptors 1 and 2.

Presentation of antigen is key to the development of the immune response, mediated by association of antigen with major histocompatibility complex glycoproteins abbreviated as MHC1 and MHC2. In the current study, we examined the regulation of MHC1 in the brain after facial axotomy. The normal facial motor nucleus showed no immunoreactivity for MHC1 (MHC1-IR). Transection of the facial nerve led to a strong and selective up-regulation of MHC1-IR on the microglia in the affected nucleus, beginning at day 2 and reaching a maximum 14 days after axotomy, coinciding with a peak influx of the T lymphocytes that express CD8, the lymphocyte coreceptor for MHC1. Specificity of the MHC1 staining was confirmed in beta2-microglobulin-deficient mice, which lack normal cell surface MHC1-IR. MHC1-IR was particularly strong on phagocytic microglia, induced by delayed neuronal cell death, and correlated with the induction of mRNA for tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, and interferon-gamma and the influx of T lymphocytes. Mice with severe combined immunodeficiency (scid), lacking T and B cells, showed an increase in the number of MHC1-positive nodules but no significant effect on overall MHC1-IR. Transgenic deletion of the IL1 receptor type I, or the interferon-gamma receptor type 1 subunit, did not affect the microglial MHC1-IR. However, a combined deletion of TNF receptors 1 and 2 (TNFR1&2-KO) led to a decrease in microglial MHC1-IR and to a striking absence of the phagocytic microglial nodules. Deletion of TNFR2 (p75) did not have an effect; deletion of TNFR1 (p55) reduced the diffuse microglial staining for MHC1-IR but did not abolish the MHC1(+) microglial nodules. In summary, neural injury leads to the induction of MHC1-IR on the activated, phagocytic microglia. This induction of MHC1 precedes the interaction with the immune system, at least in the facial motor nucleus model. Finally, the impaired induction of these molecules, up to now, only in the TNFR-deficient mice underscores the central role of TNF in the immune activation of the injured nervous system.

Animals↗

PubMed2004

B7.2 on activated and phagocytic microglia in the facial axotomy model: regulation by interleukin-1 receptor type 1, tumor necrosis factor receptors 1 and 2 and endotoxin.

Co-stimulatory factors are involved in different forms of brain pathology and play an important role in the activation of T-cells. In the current study, we explored the regulation of B7.2, a prominent member of the B7 family of costimulatory factors, in the facial motor nucleus (FMN) following facial axotomy and systemic application of lipopolysaccharide (LPS, endotoxin) using light and electron immunohistochemistry and cytokine-receptor-deficient mice. Facial axotomy led to a gradual increase of B7.2 immunoreactivity (IR) on microglial cell surface; similar effects were also observed following application of LPS, but both effects were not additive, suggesting overlapping or saturated signaling pathways. Some B7.2-IR was already present on activated microglia surrounding injured neurons at days 1-4 after injury, but became particularly intense during neuronal cell death, peaking at day 14. Previous studies revealed that these late microglial changes are accompanied by a strong increase in the expression of proinflammatory cytokines such as interleukin-1 beta (IL1beta) tumor necrosis factor-alpha (TNFalpha) and interferon gamma (IFNgamma) [J. Neurosci. 18 (1998a) 5804]. Here, deletion of the receptors for these cytokines-IL1R1, TNFR1 or TNFR2, but not IFNgammaR1-caused a strong and significant reduction in B7.2-IR in reactive microglial cells, compared with their wild type (WT) controls on the same genetic strain background, with a 31% decrease in IL1R1-/- , 39% in TNFR1-/- and 49% in TNFR2-/- mice. These data underscore the significance of IL1beta, TNFalpha and LPS, and their receptors, as potent inflammatory signals that regulate the cellular response in the injured brain as well as the interaction with the rapidly recruited immune system. The broad susceptibility of B7.2 regulation to a wide range of different inflammatory signals also points to its role as a sensor of molecular pathology, and a factor that plays an important accessory role in allowing and shaping the microglia/T-cell interaction in the injured central nervous system.

Animals↗

PubMed2004

Children's perceptions of dangerous substances.

The aim of this research was to examine age differences in children's perceptions of dangerous substances. Children's responses to photographs of child models encountering alcohol, a syringe, medicine, glue, and household bleach were coded for recognition of substance, awareness of potential danger, and understanding of danger or harm. Responses from 59 children (28 girls and 31 boys, who were all children of the required age in an urban primary school) were compared across three age groups (6-7 years, 8-9 years, and 10-11 years). 15 to 18 of 20 children in each of three age groups recognised all the dangerous substances. Significant differences across age groups were found for awareness of the potential harm from alcohol, glue, and medicine, but not for the syringe or bleach. Children reported less serious consequences from ingesting alcohol than other substances, the consequences of sniffing glue tended to be confused with the sticking properties of glue, children's understanding of transmission of germs, disease, and AIDS through syringes was varied, and there was a tendency among the older children to underestimate the potential harm of self-administration of medicines. Findings were discussed in relation to children's concepts of danger and safety education.

Attitude↗

PubMed2003

Choosing a safe place to cross the road: the relationship between attention and identification of safe and dangerous road-crossing sites.

BACKGROUND: Safe pedestrian behaviour relies on cognitive skills, including the ability to focus attention on the traffic environment and ignore irrelevant stimuli. An important pedestrian skill that young children find difficult is the ability to find a safe place to cross the road. The aim of this study was to examine the relationship between attention and children's ability to identify safe and dangerous road-crossing sites. METHODS: Participants were 95 children (aged 6.5 years, 8.6 years and 10.4 years) and 33 adults. Ability to identify safe and dangerous road-crossing sites was assessed using computer presentations of five safe and five dangerous sites. Attention was assessed using the Stroop test for resistance to interference. Correlations were calculated between Stroop test measures and pedestrian task measures (accuracy and speed of identifying safe and dangerous road-crossing sites) for each age group separately. RESULTS: The ability to identify safe and dangerous road-crossing sites and the ability to resist interference increased with age. Significant correlations were observed between identification of safe and dangerous road-crossing sites and performance on the Stroop test for children but not for adults. DISCUSSION: The results indicated that attention is required for identifying road-crossing sites quickly and accurately, especially for younger children. Road safety training programmes for children may need to take into account the development of children's attention.

Accident Prevention↗

PubMed2001

Erythropoiesis in the absence of janus-kinase 2: BCR-ABL induces red cell formation in JAK2(-/-) hematopoietic progenitors.

The receptor-associated protein tyrosine kinase janus-kinase 2 (JAK2) is essential for normal red cell development and for erythropoietin receptor (EpoR) signaling. JAK2(-/-) embryos are severely deficient in erythropoiesis and die at an early stage of development from fetal anemia. The binding of erythropoietin (Epo) to the EpoR triggers the activation of JAK2, the phosphorylation of the EpoR, and the initiation of the EpoR signaling cascade. In addition to Epo binding to its receptor, signaling pathways downstream of the EpoR can also be stimulated by the BCR-ABL oncoprotein. This study explored whether JAK2 is required for BCR-ABL-mediated stimulation of erythropoiesis. Here, it is shown that JAK2 is constitutively tyrosine phosphorylated in cultured and primary erythroid cells expressing BCR-ABL. However, BCR-ABL effectively supports normal erythroid proliferation, differentiation, and maturation in JAK2-deficient fetal liver cells. Using mutants of BCR-ABL, this study shows that certain signaling pathways activated by BCR-ABL segments distinct from its tyrosine kinase domain are essential for rescue of erythropoiesis in JAK2(-/-) progenitors. The consequences of these multiple signaling pathways for normal erythroid development are discussed.

Amino Acid Substitution↗

PubMed2001

Signal via lymphotoxin-beta R on bone marrow stromal cells is required for an early checkpoint of NK cell development.

NK cells play an important role in the immune system but the cellular and molecular requirements for their early development are poorly understood. Lymphotoxin-alpha (LTalpha)(-/-) and LTbetaR(-/-) mice show a severe systemic reduction of NK cells, which provides an excellent model to study NK cell development. In this study, we show that the bone marrow (BM) or fetal liver cells from LTalpha(-/-) or LTbetaR(-/-) mice efficiently develop into mature NK cells in the presence of stromal cells from wild-type mice but not from LTalpha(-/-) or LTbetaR(-/-) mice. Direct activation of LTbetaR-expressing BM stromal cells is shown to promote to early NK cell development in vitro. Furthermore, the blockade of the interaction between LT and LTbetaR in adult wild-type mice by administration of LTbetaR-Ig impairs the development of NK cells in vivo. Together, these results indicate that the signal via LTbetaR on BM stromal cells by membrane LT is an important pathway for early NK cell development.

Adoptive Transfer↗

PubMed2001

PUMA-G, an IFN-gamma-inducible gene in macrophages is a novel member of the seven transmembrane spanning receptor superfamily.

IFN-gamma is a key immunoregulatory cytokine that plays a predominant role in innate immunity. By employing PCR-Select to search for genes differentially expressed in IFN-gamma/TNF-alpha stimulated macrophages, we identified a novel IFN-gamma-induced transcript designated PUMA-G (protein up-regulated in macrophages by IFN-gamma). PUMA-G codes for a protein with seven transmembrane helices, a feature commonly shared with the G protein-coupled receptor superfamily (GPCR). The PUMA-G protein is most similar to the human orphan GPCR HM74 (73 % identity) and GPR31 (30 % identity). PUMA-G mRNA was readily induced in macrophages after stimulation with IFN-gamma, LPS, polyIC and CpG oligonucleotides. In vivo PUMA-G was up-regulated in mice suffering from microbial sepsis or from Listeria monocytogenes infection. Characterization of the genomic locus revealed an intronless PUMA-G open reading frame. Genomic Southern blot analysis indicates that PUMA-G is a single-copy gene. PUMA-G maps to mouse chromosome 5F. Confocal microscopy of transiently transfected 264.7 RAW macrophages and 293T cells with a PUMA-G-EGFP fusion construct showed predominant fluorescence at the cell surface, suggesting a localization at the cell membrane. Taken together, our data indicate that PUMA-G is a new inducible representative of GPCR, with potential importance in macrophage functions.

Amino Acid Sequence↗

PubMed2001

The battle of two genomes: genetics of bacterial host/pathogen interactions in mice.

Genetic factors strongly determine the outcome of infectious diseases caused by various pathogens. The molecular mechanisms of resistance and susceptibility in humans, however, remains largely unknown. Complex interactions of multiple genes that control the host response to a pathogen further complicate the picture. Animal models have a tremendous potential to dissect the complex genetic system of host-pathogen interaction into single components. This is particularly true for the mouse, which will continue to develop into an invaluable tool in the identification and cloning of host resistance genes. Three main approaches have been taken to establish mouse models for human infectious diseases: 1) Production of mouse mutants by gene targeting; 2) positional cloning of host-resistance genes in mutant mice; and 3) mapping and characterization of quantitative trait loci (QTL) controlling the complex aspects of host-pathogen interactions. The contribution of all three methods to the understanding of infectious diseases in humans will be reviewed in this work, with a special emphasis on the studies of resistance/susceptibility mechanism in bacterial infections.

Animals↗

PubMed2001

Inhibition of natural killer cells results in acceptance of cardiac allografts in CD28-/- mice.

Successful transplantation of allogeneic organs is an important objective in modern medicine. However, sophisticated immune defense mechanisms, primarily evolved to combat infections, often work against medical transplantation. To investigate the roles of natural and adaptive immune responses in transplant rejection, we functionally inactivated key effector systems of the innate (NK cells) and the adaptive immune system (CD28-mediated costimulation of T cells) in mice. Neither of these interventions alone led to acceptance of allogeneic vascularized cardiac grafts. In contrast, inhibition of NK-receptor-bearing cells combined with CD28-costimulation blockade established long-term graft acceptance. These results indicate a concerted interplay between innate and adaptive immune surveillance for graft rejection. Thus we suggest that inactivation of NK-receptor-bearing cells could be a new strategy for successful survival of solid-organ transplants.

Animals↗

PubMed2001

Lsc is required for marginal zone B cells, regulation of lymphocyte motility and immune responses.

Lsc (the murine homolog of human p115 Rho GEF) is a member of the Dbl-homology family of GTP exchange factors and is a specific activator of Rho. Lsc is activated by the G alpha(13) subunit of heterotrimeric G proteins and contains a regulator of G protein signaling domain that downmodulates G alpha(12) and G alpha(13). Lsc is expressed primarily in the hematopoietic system and links the activation of G alpha(12) and G alpha(13)-coupled receptors to actin polymerization in B and T cells. Lsc is essential for marginal zone B (MZB) cell homeostasis and for the generation of immune responses. Although Lsc-deficient lymphocytes show reduced basal motility, MZB cells show enhanced migration after serum activation. Thus, Lsc is a critical regulator of MZB cells and immune functions.

Animals↗

PubMed2001

Intact lysosome transport and phagosome function despite kinectin deficiency.

The mechanism of cargo coupling to kinesin motor proteins is a fundamental issue in organelle transport along microtubules. Kinectin has been postulated to function as a membrane anchor protein that attaches various organelles to the prototype motor protein kinesin. To verify the biological relevance of kinectin in vivo, the murine kinectin gene was disrupted by homologous recombination. Unexpectedly, kinectin-deficient mice were viable and fertile, and no gross abnormalities were observed up to 1 year of age. The assembly of the endoplasmic reticulum was essentially unaffected in kinectin-deficient cells. Mitochondria appeared to be correctly distributed throughout the cytoplasm along the microtubules. Furthermore, the stationary distribution and the bidirectional movement of lysosomes did not depend on kinectin. Kinectin-deficient phagocytes internalized and cleared bacteria, indicating that phagosome trafficking and maturation are functional without kinectin. Thus, these data unequivocally indicate that kinectin is not essential for trafficking of lysosomes, phagosomes, and mitochondria in vivo.

Alleles↗

PubMed2000

Lethal granuloma disintegration in mycobacteria-infected TNFRp55-/- mice is dependent on T cells and IL-12.

Genetically susceptible, TNFRp55 gene-deficient (TNFRp55-/-) mice succumb to infection with Mycobacterium avium. Before their death, M. avium-infected TNFRp55-/- mice develop granulomatous lesions that, in contrast to granulomas in wild-type syngeneic mice, undergo acute disintegration. To determine the factors involved in these events, we depleted T cell subsets or neutralized the inflammatory cytokines IFN-gamma, IL-12, or TNF in TNFRp55-/- mice infected i.v. with M. avium. Infected TNFRp55-/- mice treated with a control mAb became moribund between days 26 and 34 postinfection, showing widespread inflammatory cell apoptosis within disintegrating granulomas. In contrast, TNFRp55-/- mice depleted of either CD4+ or CD8+ cells after granuloma initiation stayed healthy until at least day 38 postinfection and showed no signs of granuloma destruction. Neutralization of IL-12, but not of IFN-gamma or TNF, also protected M. avium-infected TNFRp55-/- mice from granuloma decomposition and from premature death. Treatment with dexamethasone or with a specific inhibitor of inducible NO synthase did not prevent granuloma dissolution or death of TNFRp55-/- mice. In conclusion, granuloma disintegration in TNFRp55-/- mice is a lethal event that is dependent on IL-12 and that is mediated by an excess of T cells.

Animals↗

PubMed2000

Inflammatory and immunological parameters in children with haemolytic uremic syndrome (HUS) and gastroenteritis-pathophysiological and diagnostic clues.

The objective of this study was to identify parameters indicating a risk for developing typical haemolytic uremic syndrome (D+HUS) during the prodromal phase of diarrhea caused by enterohaemorrhagic Escherichia coli (EHEC). Forty-eight children were studied prospectively with regard to inflammatory serum factors on admission to hospital. Ten patients developed D+HUS (group I), 15 suffered from viral-gastroenteritis (group IIa) and 23 from other types of bacterial gastroenteritis (group IIb). Mean levels of IL-8 tended to be elevated in group I compared to groups IIa and IIb. Neopterin and IL-10 levels particularly were significantly decreased in HUS in comparison to both gastroenteritis groups. Low IL-10 levels indicate a substantial disregulation of the immune response in HUS, as IL-10 downregulates the pro-inflammatory response and suppresses pro-coagulant activity in experimental endotoxemia. Our results suggest low neopterin, high IL-8 and especially low IL-10 levels are indicators of a high risk for developing HUS.

Bacterial Infections↗

PubMed2000

Identification of immunological relevant phenotypes in ENU mutagenized mice.

The immunology screen focuses on the identification of novel gene products involved in the mammalian immune response and on the establishment of mouse models for immunological disorders. For this purpose, high throughput and semi-automated techniques were developed and optimized for low cost per sample and reproducibility. All assays are designed to be nonconsumptive and are based on peripheral blood or direct PCR amplification.

Animals↗

PubMed2000

Up-regulation of BOB.1/OBF.1 expression in normal germinal center B cells and germinal center-derived lymphomas.

The BOB.1/OBF.1/OCAB.1 protein is a lymphocyte-specific transcriptional coactivator. It interacts with the Oct1 and Oct2 transcription factors and contributes to the transcriptional activity of octamer motifs. The analysis of established B cell lines had suggested that BOB.1/OBF.1 is constitutively expressed at all stages of B cell development. Here we show that expression of BOB. 1/OBF.1 is regulated within the B cell lineage. Specifically, germinal center B cells show highly increased BOB.1/OBF.1 levels. We can induce the up-regulation by stimulating primary splenic B cells, eg, by triggering CD40 signaling in the presence of interleukin-4. Expression of BOB.1/OBF.1 is detectable but reduced in spleens from mice unable to undergo the germinal center reaction due to mutations in the TNF receptor p55 or lymphotoxin beta (LTbeta) receptor genes. Furthermore, we demonstrate that BOB.1/OBF.1 expression is highly regulated in human B cell lymphomas. Whereas lymphomas representing pre- and postfollicular B cell developmental stages are negative for BOB.1/OBF.1, high-level expression of BOB.1/OBF.1 is characteristic of germinal center-derived tumors. In these tumors BOB.1/OBF.1 is typically coexpressed with high levels of Bcl6. These results imply that overexpression of BOB.1/OBF.1, like overexpression of Bcl6, might play a role in the pathogenesis of germinal center-derived B cell lymphomas. Furthermore, overexpression of BOB.1/OBF.1 represents a characteristic feature of these tumors that is useful in their identification.

Animals↗

PubMed2000

Interferon-gamma and tumor necrosis factor-alpha determine resistance to Paracoccidioides brasiliensis infection in mice.

To investigate the role of interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha) in the resistance to Paracoccidioides brasiliensis (Pb) infection, mice with homologous disruption of the IFN-gamma (GKO) or TNF-alpha receptor p55 (p55KO) were infected with the parasite. GKO and p55KO, but not wild-type (WT) mice, were unable to control the growth of yeast cells and the mice succumbed to infection by days 16 and 90 after infection, respectively. Typical inflammatory granulomas were found only in WT mice. In contrast, knockout mice presented an inflammatory infiltrate composed of a few neutrophils, mononuclear, epithelioid, and multinuclear giant cells forming incipient granulomas in GKO mice and without granuloma formation in p55KO mice. Besides, both groups of knockout mice exhibited elevated numbers of yeast forms in agreement with colony-forming unit counts in organs. Compared with WT, splenocytes from infected GKO mice cultured with the Pb F1 fraction produced lower TNF-alpha levels, whereas leukocytes from infected p55KO mice produced similar amounts of TNF-alpha but higher levels of IFN-gamma. Moreover, splenocytes from infected WT mice produced higher levels of nitric oxide (NO) resulting in a lower T-cell proliferative response to Con A than uninfected WT, or infected p55KO and GKO mice. On the contrary, the addition of IFN-gamma to splenocytes from infected GKO mice resulted in higher NO production and lower T cell proliferation. Taken together, these findings suggests that endogenous TNF-alpha, acting through the p55 receptor, and IFN-gamma mediate resistance to Pb infection and induce NO production that determines marked T cell unresponsiveness.

Animals↗

PubMed2000

Partial impairment of cytokine responses in Tyk2-deficient mice.

To assess the role of the Janus kinase (Jak) family member Tyk2, we have generated Tyk2-/- mice. In contrast to other Jaks, where inactivation leads to a complete loss of the respective cytokine receptor signal, Tyk2-/- mice display reduced responses to IFNalpha/beta and IL-12 and a selective deficiency in Stat3 activation in these pathways. Unexpectedly, IFNgamma signaling is also impaired in Tyk2-/- mice. Tyk2-/- macrophages fail to produce nitric oxide upon lipopolysaccharide induction. Tyk2-/- mice are unable to clear vaccinia virus and show a reduced T cell response after LCMV challenge. These data imply a selective contribution of Tyk2 to the signals triggered by various biological stimuli and cytokine receptors.

Animals↗