Commensal Microbiota Are Required for Systemic Inflammation Triggered by Necrotic Dendritic Cells

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FADD-deficient T cells exhibit a disaccord in regulation of the cell cycle machinery

FADD is an adapter protein that was originally isolated as a transducer of apoptotic signals for death domain-containing receptors. However, FADD-deficient mice are embryonic lethal and FADD(-/-) T cells developed from FADD(-/-) embryonic stem cells in the RAG-1(-/-) hosts lack the full potential to proliferate when stimulated through their T-cell receptor complex, suggesting that FADD protein might play a dualistic role in mediating not only cell death signaling but other non-apoptotic cellular pathways as well. Here we show that a substantial number of freshly isolated FADD(-/-) peripheral T cells are cycling but are defective in their co-stimulatory response when stimulated. Analysis of several cell cycle proteins shows normal down-regulation of p27 inhibitor but increased levels of p21, decreased levels of cyclin D2, and constitutive activation of several cyclin-dependent kinases in activated T cells. These data suggest that FADD is involved in the regulation of cell cycle machinery in T lymphocytes.     

The orphan steroid receptor Nur77 family member Nor-1 is essential for early mouse embryogenesis

Nur77 and its family members, Nor-1 and Nurr1, are orphan steroid receptors implicated in a wide variety of biological processes, including apoptosis and dopamine neuron agenesis. Expression of these family members can be detected at low levels in many tissues but they are expressed at very high levels when cells are stimulated by outside signals, including serum, nerve growth factor, and receptor engagement. Introduction of a dominant negative Nur77 protein that blocks the activities of all family members led to inhibition of apoptosis in T cells. Nur77-deficient mice, however, exhibit no phenotype, and a line of Nor-1 mutant mice was reported to exhibit a mild ear development phenotype but no other gross abnormalities. Here, we report the generation of Nor-1-deficient mice with a block in early embryonic development. Nor-1 is expressed early during embryogenesis, and its loss leads to embryonic lethality around embryonic day 8.5 of gestation. The mutant embryos fail to complete gastrulation and display distinct morphological abnormalities, including a decrease in overall size, developmental delay and an accumulation of mesoderm in the primitive streak during gastrulation. Abnormal expression of a number of early developmental markers and defects in growth or distribution of emerging mesoderm cells were also detected. These data suggest that Nor-1 plays a crucial role in gastrulation.     

Deletion of FADD in Macrophages and Granulocytes Results in RIP3- and MyD88-Dependent Systemic Inflammation

Myeloid cells, which include monocytes, macrophages, and granulocytes, are important innate immune cells, but the mechanism and downstream effect of their cell death on the immune system is not completely clear. Necroptosis is an alternate form of cell death that can be triggered when death receptor-mediated apoptosis is blocked, for example, in stimulated Fas-associated Death Domain (FADD) deficient cells. We report here that mice deficient for FADD in myeloid cells (mFADD^-/-) exhibit systemic inflammation with elevated inflammatory cytokines and increased levels of myeloid and B cell populations while their dendritic and T cell numbers are normal. These phenotypes were abolished when RIP3 deficiency was introduced, suggesting that systemic inflammation is caused by RIP3-dependent necroptotic and/or inflammatory activity. We further found that loss of MyD88 can rescue the systemic inflammation observed in these mice. These phenotypes are surprisingly similar to that of dendritic cell (DC)-specific FADD deficient mice with the exception that DC numbers are normal in mFADD^-/- mice. Together these data support the notion that innate immune cells are constantly being stimulated through the MyD88-dependent pathway and aberrations in their cell death machinery can result in systemic effects on the immune system.     

Mass transport in a microchannel bioreactor with a porous wall

A two-dimensional flow model has been developed to simulate mass transport in a microchannel bioreactor with a porous wall. A two-domain approach, based on the finite volume method, was implemented. For the fluid part, the governing equation used was the Navier-Stokes equation; for the porous medium region, the generalized Darcy-Brinkman-Forchheimer extended model was used. For the porous-fluid interface, a stress jump condition was enforced with a continuity of normal stress, and the mass interfacial conditions were continuities of mass and mass flux. Two parameters were defined to characterize the mass transports in the fluid and porous regions. The porous Damkohler number is the ratio of consumption to diffusion of the substrates in the porous medium. The fluid Damkohler number is the ratio of the substrate consumption in the porous medium to the substrate convection in the fluid region. The concentration results were found to be well correlated by the use of a reaction-convection distance parameter, which incorporated the effects of axial distance, substrate consumption, and convection. The reactor efficiency reduced with reaction-convection distance parameter because of reduced reaction (or flux), and smaller local effectiveness factor due to the lower concentration in Michaelis-Menten type reactions. The reactor was more effective, and hence, more efficient with the smaller porous Damkohler number. The generalized results could find applications for the design of bioreactors with a porous wall.     

Cloning of GT box-binding proteins: a novel Sp1 multigene family regulating T-cell receptor gene expression.

Analysis of a T-cell antigen receptor (TCR) alpha promoter from a variable gene segment (V) revealed a critical GT box element which is also found in upstream regions of several V alpha genes, TCR enhancer, and regulatory elements of other genes. This element is necessary for TCR gene expression and binds several proteins. These GT box-binding proteins were identified as members of a novel Sp1 multigene family. Two of them, which we term Sp2 and Sp3, were cloned. Sp2 and Sp3 contain zinc fingers and transactivation domains similar to those of Sp1. Like Sp1, Sp2 and Sp3 are expressed ubiquitously, and their in vitro-translated products bind to the GT box in TCR V alpha promoters. Sp3, in particular, also binds to the Sp1 consensus sequence GC box and has binding activity similar to that of Sp1. As the GT box has also previously been shown to play a role in gene regulation of other genes, these newly isolated Sp2 and Sp3 proteins might regulate expression not only of the TCR gene but of other genes as well.     

A mouse Fas-associated protein with homology to the human Mort1/FADD protein is essential for Fas-induced apoptosis.

The Fas cell surface receptor belongs to the tumor necrosis factor receptor family and can initiate apoptosis in a variety of cell types. Using the Fas cytoplasmic domain as bait in a yeast two-hybrid screening, we isolated a mouse cDNA encoding a 205-amino-acid protein. Its predicted protein sequence shows 68% identity and 80% similarity with the sequence of recently described human Mort/FADD. This protein, most likely the mouse homolog of human FADD, associates with Fas in vivo only upon the induction of cell death. A fraction of this protein is highly phosphorylated at serine/threonine residues, with both phosphorylated and unphosphorylated forms being capable of binding to FAS. Stable expression of a truncated form of the Mort/FADD protein protects cells from Fas-mediated apoptosis by interfering with the wild-type protein-Fas interaction. Thus, mouse Mort/FADD is an essential downstream component that mediates Fas-induced apoptosis.