The benefit of culture-independent methods to detect bacteria and fungi in re-infected root filled teeth: a pilot study.

AIM: To identify dominant microorganisms in root filled teeth with apical periodontitis by Pan-PCRs in comparison with a culture-dependent approach, focusing on fungal species profiling. METHODOLOGY: The root filling material (gutta-percha) removed from 42 teeth with periapical radiolucencies undergoing root canal retreatments was analysed by molecular genetics techniques. Real-Time Pan-PCRs were conducted for the diagnosis of predominant bacteria (targeting 16S rDNA) and fungi (targeting ITS1-2 region). Identification of microorganisms was performed by Sanger sequencing of the PCR products and BLAST analysis. Additionally, subgingival plaque samples were collected and cultured to review the composition of the microbial flora. The McNemar test and the repeated measures anova were used for statistical analyses (significance level was set at P < 0.05). RESULTS: Overall, 42/42 plaque samples had bacterial growth, whereas 32/42 gutta-percha samples had bacterial growth with a dominance of Streptococcus spp. (12/42) and Enterococcus faecalis (9/42). The mean number of bacterial taxa per gutta-percha sample was 1.6 cultivatable taxa, significantly lower than in the plaque sample that had six taxa/sample (P < 0.001). Fungus-specific cultures were negative for gutta-percha samples, and only one plaque sample had growth of a fungus. In total, 36/42 plaque samples were positive in bacterial Pan-PCRs. In bacterial Pan-PCRs of 31/42 gutta-percha samples, dominant microorganisms were identified including Streptococcus spp. (5/42) and E. faecalis (4/42). Moreover, in 7/42 gutta-percha samples, DNA of bacteria which are difficult-to-cultivate in microbiology routine culture (Acinetobacter,Pyramidobacter,Bacteroidetes,Synergistes,Atopobium and Pseudoramibacter) was found. DNA of Candida spp. was detected in 5/42 root canals by fungal Pan-PCR (1/5) and genus-specific Candida-PCR (5/5). CONCLUSIONS: Pan-PCR assays remain appropriate as a broad-range approach for the detection of a dominant pathogen in gutta-percha samples which have less diverse microbial composition. The molecular genetic Pan-PCR approach has the advantage of detecting microorganisms that are as-yet-uncultivable or difficult-to-cultivate and should be therefore complement conventional microbiological diagnostics.

A novel mba-based Real time PCR approach for genotyping of Ureaplasma parvum validated in a cohort of Mongolian mothers and offspring.

The role of Ureaplasma parvum in abnormal outcomes of human pregnancy has been discussed controversially in the past. Of the 14 known ureaplasma serovars, the Ureaplasma parvum serovars 1, 3, 6 and 14, have been found to derive from smaller genomes. Serovars 3 and 6 have been described more often to cause complications in pregnancy. To elucidate the serovar distribution in U. parvum positive specimens of 200 Mongolian mothers and their offspring, a new set of mba-targeting PCRs was developed enabling a fast and reliable serovar differentiation by melting peak analysis in a Real time PCR approach or by conventional agarose gel electrophoresis. 92% maternal and 55% neonatal samples were retrospectively genotyped and a dominance of serovars 3 and 6 was detected while serovar 14 was almost absent. Transmission from mothers to newborns was detected in 83% of U. parvum positive neonates exhibiting serovar patterns identical to their mothers. No statistically significant correlation between a distinct serovar and pregnancy outcome could be detected. However, neonatal colonization with serovar 1 declined with progressing pregnancy suggesting that a higher ureaplasma load shortened pregnancy and thereby had a potential negative effect on offspring health. Our novel mba-based Real time PCR approach, which can also be used in conventional PCR and gel electrophoretic analysis, provides the proof of principle that the four U. parvum serovars 1, 3, 6 and 14 can be differentially detected and quantified. A larger scale study outside the scope of this work should be conducted to clarify the impact of serovar 1 on pregnancy outcome.

IRF4 and BATF are critical for CD8⁺ T-cell function following infection with LCMV.

CD8(+) T-cell functions are critical for preventing chronic viral infections by eliminating infected cells. For healthy immune responses, beneficial destruction of infected cells must be balanced against immunopathology resulting from collateral damage to tissues. These processes are regulated by factors controlling CD8(+) T-cell function, which are still incompletely understood. Here, we show that the interferon regulatory factor 4 (IRF4) and its cooperating binding partner B-cell-activating transcription factor (BATF) are necessary for sustained CD8(+) T-cell effector function. Although Irf4(-/-) CD8(+) T cells were initially capable of proliferation, IRF4 deficiency resulted in limited CD8(+) T-cell responses after infection with the lymphocytic choriomeningitis virus. Consequently, Irf4(-/-) mice established chronic infections, but were protected from fatal immunopathology. Absence of BATF also resulted in reduced CD8(+) T-cell function, limited immunopathology, and promotion of viral persistence. These data identify the transcription factors IRF4 and BATF as major regulators of antiviral cytotoxic T-cell immunity.

The effects of peer influence on adolescent pedestrian road-crossing decisions.

OBJECTIVE: Adolescence is a high-risk period for pedestrian injury. It is also a time of heightened susceptibility to peer influence. The aim of this research was to examine the effects of peer influence on the pedestrian road-crossing decisions of adolescents. METHODS: Using 10 videos of road-crossing sites, 80 16- to 18-year-olds were asked to make pedestrian road-crossing decisions. Participants were assigned to one of 4 experimental conditions: negative peer (influencing unsafe decisions), positive peer (influencing cautious decisions), silent peer (who observed but did not comment), and no peer (the participant completed the task alone). Peers from the adolescent's own friendship group were recruited to influence either an unsafe or a cautious decision. RESULTS: Statistically significant differences were found between peer conditions. Participants least often identified safe road-crossing sites when accompanied by a negative peer and more frequently identified dangerous road-crossing sites when accompanied by a positive peer. Both cautious and unsafe comments from a peer influenced adolescent pedestrians' decisions. CONCLUSIONS: These findings showed that road-crossing decisions of adolescents were influenced by both unsafe and cautious comments from their peers. The discussion highlighted the role that peers can play in both increasing and reducing adolescent risk-taking.

Phosphorylation of Atg5 by the Gadd45ß-MEKK4-p38 pathway inhibits autophagy.

Autophagy is a lysosomal degradation pathway important for cellular homeostasis, mammalian development, cancer and immunity. Many molecular components of autophagy have been identified, but little is known about regulatory mechanisms controlling their effector functions. Here, we show that, in contrast to other p38 MAP kinase activators, the growth arrest and DNA damage 45 beta (Gadd45ß)-MAPK/ERK kinase kinase 4 (MEKK4) pathway specifically directs p38 to autophagosomes. This process results in an accumulation of autophagosomes through p38-mediated inhibition of lysosome fusion. Conversely, autophagic flux is increased in p38-deficient fibroblasts and Gadd45ß-deficient cells. We further identified the underlying mechanism and demonstrate that phosphorylation of the autophagy regulator autophagy-related (Atg)5 at threonine 75 through p38 is responsible for inhibition of starvation-induced autophagy. Thus, we show for the first time that Atg5 activity is controlled by phosphorylation and, moreover, that the spatial regulation of p38 by Gadd45ß/MEKK4 negatively regulates the autophagic process.

Antimicrobial effects of murine mesenchymal stromal cells directed against Toxoplasma gondii and Neospora caninum: role of immunity-related GTPases (IRGs) and guanylate-binding proteins (GBPs).

Mesenchymal stromal cells (MSCs) have a multilineage differentiation potential and provide immunosuppressive and antimicrobial functions. Murine as well as human MSCs restrict the proliferation of T cells. However, species-specific differences in the underlying molecular mechanisms have been described. Here, we analyzed the antiparasitic effector mechanisms active in murine MSCs. Murine MSCs, in contrast to human MSCs, could not restrict the growth of a highly virulent strain of Toxoplasma gondii (BK) after stimulation with IFN-γ. However, the growth of a type II strain of T. gondii (ME49) was strongly inhibited by IFN-γ-activated murine MSCs. Immunity-related GTPases (IRGs) as well as guanylate-binding proteins (GBPs) contributed to this antiparasitic effect. Further analysis showed that IFN-γ-activated mMSCs also inhibit the growth of Neospora caninum, a parasite belonging to the apicomplexan group as well. Detailed studies with murine IFN-γ-activated MSC indicated an involvement in IRGs like Irga6, Irgb6 and Irgd in the inhibition of N. caninum. Additional data showed that, furthermore, GBPs like mGBP1 and mGBP2 could have played a role in the anti-N. caninum effect of murine MSCs. These data underline that MSCs, in addition to their regenerative and immunosuppressive activity, function as antiparasitic effector cells as well. However, IRGs are not present in the human genome, indicating a species-specific difference in anti-T. gondii and anti-N. caninum effect between human and murine MSCs.

Adult pedestrian behavior when accompanying children on the route to school.

OBJECTIVE: Pedestrian injuries are a major cause of morbidity and mortality to children, especially boys. Adults serve as pedestrian role models and provide learning opportunities for children when walking to school. The research objectives were to investigate adult pedestrian behavior when accompanying boys and girls. METHODS: Behavioral observation of 140 adult pedestrians accompanying 4- to 9-year-old children was done in British residential locations. Observations took place at light-controlled crossings, speed-restricted school safety zones, and mid-block unmarked crossing sites. Behaviors observed included stopping at the curb, waiting at the curb, looking left and right before and during road crossing, holding hands, talking, and walking straight across. RESULTS: In general, adults modeled safe road crossing behaviors. Adult safe behavior scores were higher when accompanying girls than when accompanying boys. No statistically significant differences were found by child age group. The fewest safe pedestrian behaviors were observed at light-controlled crossings. CONCLUSIONS: Adult pedestrians behave differently when with boys and girls and at different types of road crossing site. Interventions aimed at reducing pedestrian injuries to children may need to take these different everyday experiences into consideration.

Risk and injury portrayal in boys' and girls' favourite television programmes.

OBJECTIVES: To analyse the injury-related content of children's television programmes preferred by boys and by girls, and to determine whether there are more televised models of unsafe behaviour in programmes preferred by boys. METHODS: Parents of 4-11-year-old children identified their children's favourite television programmes. Content analysis of 120 episodes of children's favourite programmes was used to quantify safe and risky behaviours, actual injuries and potential injuries. The gender of the characters portraying the behaviours was also analysed. RESULTS: More risky behaviour was portrayed in the boys' favourite programmes (mean per episode = 6.40) than in the girls' favourite programmes (mean = 2.57). There were almost twice as many potential injuries (n = 310) as actual injuries (n = 157). Potential injuries were portrayed more often by male characters (mean = 1.92) than female characters (mean = 0.98), mostly in the boys' favourite programmes. Actual injuries occurred more often to male characters (mean = 1.04) than to female characters (mean = 0.27) overall. CONCLUSIONS: Television programmes preferred by this sample of boys portrayed male role models engaging in risky behaviours and injuries more often than the programmes preferred by the sample of girls.

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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

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.