B7-1 costimulatory molecule is critical for the development of experimental autoimmune myasthenia gravis

Following immunization with acetylcholine receptor (AChR), MHC class II-restricted, AChR-specific CD4 cell activation is critical for the development of experimental autoimmune myasthenia gravis (EAMG) in C57BL/6 mice. To study the contributions of B7-1 and B7-2 costimulatory molecules in EAMG, B7-1, B7-2, and B7-1/B7-2 gene knockout (KO) mice were immunized with Torpedo AChR in CFA. Compared with wild-type C57BL6 mice, B7-1 and B7-1/2 KO mice were resistant to EAMG development. B7-1 KO mice had reduced anti-AChR Ab compared with C57BL/6 mice. However, neither B7-1 nor B7-2 gene disruption impaired AChR-induced or dominant alpha(146-162) peptide-induced in vitro lymphoproliferative responses. Blocking of the B7-1 or B7-2 molecule by specific mAbs in vivo led to a reduction in the AChR-specific lymphocyte response, and the reduction was more pronounced in mice treated with anti-B7-2 Ab. The findings implicate B7-1 molecules as having a critical role in the induction of EAMG, and the resistance of B7-1 KO mice is associated with suppressed humoral, rather than suppressed AChR-specific, T cell responses. The data also point to B7-2 molecules as being the dominant costimulatory molecules required for AChR-induced lymphocyte proliferation.     

Evidence for glycosylphosphatidylinositol (GPI)-anchored eosinophil-derived neurotoxin (EDN) on human granulocytes

MIST (mast cell immunoreceptor signal transducer; also termed Clnk) is an adaptor protein structurally related to SLP-76-family hematopoietic cell-specific adaptor proteins. We demonstrate here that two major MIST-associated phosphoproteins expressed in mast cell lines are SLAP-130 and SKAP55, adaptors known to interact with the Src-homology (SH) 2 domain of Src-family protein tyrosine kinases (PTKs). MIST directly associated with SLAP-130 via its SH2 domain, and collaboration of SLAP-130 with SKAP55 was required for the recruitment of MIST to Lyn. Furthermore, MIST was preferentially recruited to Fyn rather than Lyn, which is regulated by higher affinity binding of SLAP-130 and SKAP55 with the Fyn-SH2 domain than the Lyn-SH2 domain. Our results suggest that the MIST–SLAP-130–SKAP55 adaptor complex functions downstream of high-affinity IgE receptor-associated Src-PTKs in mast cells.     

Marker-assisted introgression of favorable alleles at quantitative trait loci between maize elite lines

This article reports the marker-assisted introgression of favorable alleles at three quantitative trait loci (QTL) for earliness and grain yield among maize elite lines. The QTL were originally detected in 1992 by means of ANOVA in a population of 96 recombinant inbred lines (RILs). Introgression started from a selected RIL, which was crossed three times to one of the original parents and then self-fertilized, leading to BC(3)S(1) progenies. Markers were used to assist both foreground and background selection at each generation. At the end of the program, the effect of introgression was assessed phenotypically in agronomic trials, and QTL detection was performed by composite interval mapping among BC(3)S(1) progenies. The marker-assisted introgression proved successful at the genotypic level, as analyzed by precision graphical genotypes, although no emphasis was put on the reduction of linkage drag around QTL. Also, QTL positions were generally sustained in the introgression background. For earliness, the magnitude and sign of the QTL effects were in good agreement with those expected from initial RIL analyses. Conversely, for yield, important discrepancies were observed in the magnitude and sign of the QTL effects observed after introgression, when compared to those expected from initial RIL analyses. These discrepancies are probably due to important genotype-by-environment interactions.     

SOX10 mutations in chronic intestinal pseudo-obstruction suggest a complex physiopathological mechanism

The type IV Waardenburg syndrome (WS4), also referred to as Shah-Waardenburg syndrome or Waardenburg-Hirschsprung disease, is characterised by the association of Waardenburg features (WS, depigmentation and deafness) and the absence of enteric ganglia in the distal part of the intestine (Hirschsprung disease). Mutations in the EDN3, EDNRB, and SOX10 genes have been reported in this syndrome. Recently, a new SOX10 mutation was observed in a girl with a neural crest disorder without evidence of depigmentation, but with severe constipation due to a chronic intestinal pseudo-obstruction and persistence of enteric ganglia. To refine the nosology of WS, we studied patients with typical WS4 (including Hirschsprung disease) or with WS and intestinal pseudo-obstruction. We found three SOX10 mutations, one EDNRB and one EDN3 mutations in patients presenting with the classical form of WS4, and two SOX10 mutations in patients displaying chronic intestinal pseudo-obstruction and WS features. These results show that chronic intestinal pseudo-obstruction may be a manifestation associated with WS, and indicate that aganglionosis is not the only mechanism underlying the intestinal dysfunction of patients with SOX10 mutations.     

A Novel Mouse Model for Stable Engraftment of a Human Immune System and Human Hepatocytes

Hepatic infections by hepatitis B virus (HBV), hepatitis C virus (HCV) and Plasmodium parasites leading to acute or chronic diseases constitute a global health challenge. The species tropism of these hepatotropic pathogens is restricted to chimpanzees and humans, thus model systems to study their pathological mechanisms are severely limited. Although these pathogens infect hepatocytes, disease pathology is intimately related to the degree and quality of the immune response. As a first step to decipher the immune response to infected hepatocytes, we developed an animal model harboring both a human immune system (HIS) and human hepatocytes (HUHEP) in BALB/c Rag2^-/- IL-2Rγc^-/- NOD.sirpa uPA^tg/tg mice. The extent and kinetics of human hepatocyte engraftment were similar between HUHEP and HIS-HUHEP mice. Transplanted human hepatocytes were polarized and mature in vivo, resulting in 20–50% liver chimerism in these models. Human myeloid and lymphoid cell lineages developed at similar frequencies in HIS and HIS-HUHEP mice, and splenic and hepatic compartments were humanized with mature B cells, NK cells and naïve T cells, as well as monocytes and dendritic cells. Taken together, these results demonstrate that HIS-HUHEP mice can be stably (> 5 months) and robustly engrafted with a humanized immune system and chimeric human liver. This novel HIS-HUHEP model provides a platform to investigate human immune responses against hepatotropic pathogens and to test novel drug strategies or vaccine candidates.     

Hepatic Proliferation and Angiogenesis Markers Are Increased after Portal Deprivation in Rats: A Study of Molecular,Histological and Radiological Changes

MethodsWe conducted an experimental study comparing portacaval shunt (PCS), total portal vein ligation (PVL), and sham (S) operated rats. Each group were either sacrificed at 6 weeks (early) or 6 months (late). Arterial liver perfusion was studied in vivo using CT, and histopathological changes were noted. Liver mRNA levels were quantified by RT-QPCR for markers of inflammation (Il10, Tnfa), proliferation (Il6st, Mki67, Hgf, Hnf4a), angiogenesis: (Vegfa, Vegfr 1, 2 and 3; Pgf), oxidative stress (Nos2, and 3, Hif1a), and fibrosis (Tgfb). PCS and PVL were compared to the S group.ResultsPeriportal fibrosis and arterial proliferation was observed in late PCS and PVL groups. CT imaging demonstrated increased arterial liver perfusion in the PCS group. RT-QPCR showed increased inflammatory markers in PCS and PVL early groups. Tnfa and Il10 were increased in PCS and PVL late groups respectively. All proliferative markers increased in the PCS, and Hnf4a in the PVL early groups. Mki67 and Hnf4a were increased in the PCS late group. Nos3 was increased in the early and late PCS groups, and Hif1a was decreased in the PVL groups. Markers of angiogenesis were all increased in the early PCS group, and Vegfr3 and Pgf in the late PCS group. Only Vegfr3 was increased in the PVL groups. Tgf was increased in the PCS groups.ConclusionsPortal deprivation in rats induces a sustained increase in intrahepatic markers of inflammation, angiogenesis, proliferation, and fibrosis.     

The Repellent DEET Potentiates Carbamate Effects via Insect Muscarinic Receptor Interactions: An Alternative Strategy to Control Insect Vector-Borne Diseases

Insect vector-borne diseases remain one of the principal causes of human mortality. In addition to conventional measures of insect control, repellents continue to be the mainstay for personal protection. Because of the increasing pyrethroid-resistant mosquito populations, alternative strategies to reconstitute pyrethroid repellency and knock-down effects have been proposed by mixing the repellent DEET (N,N-Diethyl-3-methylbenzamide) with non-pyrethroid insecticide to better control resistant insect vector-borne diseases. By using electrophysiological, biochemichal, in vivo toxicological techniques together with calcium imaging, binding studies and in silico docking, we have shown that DEET, at low concentrations, interacts with high affinity with insect M1/M3 mAChR allosteric site potentiating agonist effects on mAChRs coupled to phospholipase C second messenger pathway. This increases the anticholinesterase activity of the carbamate propoxur through calcium-dependent regulation of acetylcholinesterase. At high concentrations, DEET interacts with low affinity on distinct M1/M3 mAChR site, counteracting the potentiation. Similar dose-dependent dual effects of DEET have also been observed at synaptic mAChR level. Additionally, binding and in silico docking studies performed on human M1 and M3 mAChR subtypes indicate that DEET only displays a low affinity antagonist profile on these M1/M3 mAChRs. These results reveal a selective high affinity positive allosteric site for DEET in insect mAChRs. Finally, bioassays conducted on Aedes aegypti confirm the synergistic interaction between DEET and propoxur observed in vitro, resulting in a higher mortality of mosquitoes. Our findings reveal an unusual allosterically potentiating action of the repellent DEET, which involves a selective site in insect. These results open exciting research areas in public health particularly in the control of the pyrethroid-resistant insect-vector borne diseases. Mixing low doses of DEET and a non-pyrethroid insecticide will lead to improvement in the efficiency treatments thus reducing both the concentration of active ingredients and side effects for non-target organisms. The discovery of this insect specific site may pave the way for the development of new strategies essential in the management of chemical use against resistant mosquitoes.     

Evolution and Design Governing Signal Precision and Amplification in a Bacterial Chemosensory Pathway

Understanding the principles underlying the plasticity of signal transduction networks is fundamental to decipher the functioning of living cells. In Myxococcus xanthus, a particular chemosensory system (Frz) coordinates the activity of two separate motility systems (the A- and S-motility systems), promoting multicellular development. This unusual structure asks how signal is transduced in a branched signal transduction pathway. Using combined evolution-guided and single cell approaches, we successfully uncoupled the regulations and showed that the A-motility regulation system branched-off an existing signaling system that initially only controlled S-motility. Pathway branching emerged in part following a gene duplication event and changes in the circuit structure increasing the signaling efficiency. In the evolved pathway, the Frz histidine kinase generates a steep biphasic response to increasing external stimulations, which is essential for signal partitioning to the motility systems. We further show that this behavior results from the action of two accessory response regulator proteins that act independently to filter and amplify signals from the upstream kinase. Thus, signal amplification loops may underlie the emergence of new connectivity in signal transduction pathways.