The regulatory network that controls the differentiation of T lymphocytes

There is a vast amount of molecular information regarding the differentiation of T lymphocytes, in particular regarding in vitro experimental treatments that modify their differentiation process. This publicly available information was used to infer the regulatory network that controls the differentiation of T lymphocytes into CD4⁺ and CD8⁺ cells. Hereby we present a network that consists of 50 nodes and 97 regulatory interactions, representing the main signaling circuits established among molecules and molecular complexes regulating the differentiation of T cells. The network was converted into a continuous dynamical system in the form of a set of coupled ordinary differential equations, and its dynamical behavior was studied. With the aid of numerical methods, nine fixed point attractors were found for the dynamical system. These attractors correspond to the activation patterns observed experimentally for the following cell types: CD4⁻CD8⁻, CD4⁺CD8⁺, CD4⁺ naive, Th1, Th2, Th17, Treg, CD8⁺ naive, and CTL. Furthermore, the model is able to describe the differentiation process from the precursor CD4⁻CD8⁻ to any of the effector types due to a specific series of extracellular signals.     

Oral or nasal antigen induces regulatory T cells that suppress arthritis and proliferation of arthritogenic T cells in joint draining lymph nodes

The propagation of mucosal tolerance as a therapeutic approach in autoimmune diseases remains a difficult goal to achieve, and therefore further mechanistic studies are necessary to develop potential clinical protocols to induce mucosal regulatory T cells (Tr cells). In this study we addressed whether oral or nasal proteoglycan induced functional Tr cells in the cartilage proteoglycan-induced chronic arthritis model. Both nasal and oral application of human proteoglycan before induction of disease suppressed arthritis severity and incidence. Tolerized mice showed enhanced numbers of IL-10 producing CD4(+) cells in the paw-draining lymph nodes. Furthermore, CD4(+) spleen cells displayed enhanced expression of molecules associated with Tr cells, such as IL-10, Foxp3, and TGF-beta. Transfer of CD4(+) spleen cells from mucosally tolerized donors into proteoglycan-immunized mice abolished arthritis and reduced humoral responses, indicative of Tr cells with the capacity to inhibit already induced immune responses. Tr cells were activated upon transfer, because enhanced proliferation was observed in the joint draining lymph nodes compared with activated T cells from nontolerized donors. Upon cotransfer with naive proteoglycan-specific T cells, mucosally induced Tr cells inhibited proliferation of these arthritogenic T cells in vivo. Herein we show that both oral and nasal Ag application induced Tr cells, which had a direct inhibitory effect on already established pathogenic B and T cell responses.     

Identification of oncofetal antigen/immature laminin receptor protein epitopes that activate BALB/c mouse OFA/iLRP-specific effector and regulatory T cell clones

During tumor development in mice and humans, oncofetal Ag/immature laminin receptor (OFA/iLRP)-specific Th1, CTL, and IL-10-secreting T (Ts) cells are induced. The presence of too many Ts or too few effector T cells appears to predict a poor prognosis. We established clones of OFA/iLRP-specific splenic Th1, CTL, and Ts cells from the OFA/iLRP+ MCA1315 fibrosarcoma-bearing BALB/c mice or from BALB/c mice vaccinated with 1 or 10 microg of rOFA/iLRP. The MCA1315 tumor cell-reactive T cell clones were characterized as to surface Ag phenotype, cytokine secretion profile, and specificity for OFA/iLRP presented by syngeneic splenic APC. OFA/iLRP-specific Th1 and Ts clones were established from all mice. OFA/iLRP-specific CTL could be established from all mice except for mice immunized with 10 microg of rOFA/iLRP. Analysis of the proliferation profile of the OFA/iLRP-specific clones to overlapping OFA/iLRP 12-mer peptides that spanned the OFA/iLRP protein sequence defined the epitopes to which the T cell clones responded. There was a similar spatial distribution of the epitopes to which the two types of CD8 T cell clones responded. The nonapeptide epitopes of the Ts clones were located between aa 36 and 147 of OFA/iLRP, while the epitopes of the CTL clones were located between aa 52 and 163. Even though the CTL and Ts epitopes shared part of the protein, all of the CD8 CTL epitopes were distinct and separable from those of CD8 Ts cells.     

Properties of bovine nephritogenic antigen that induces anti-GBM nephritis in rats and its similarity to the Goodpasture antigen.

The nephritogenic antigen that induces antiglomerular basement membrane antibody-induced glomerulonephritis (anti-GBM nephritis) in rats was isolated from collagenase-solubilized bovine renal basement membranes. Purification was achieved using antibody-coupled affinity columns which were originally used for the purification of trypsin-solubilized nephritogenic antigen (Sado et al. 1984a). The nephritogenic antigen was a heteropolymer composed of P2 (Mr 28 kDa) and P3 (Mr 30 kDa) polypeptides as monomers and their dimers in sodium-dodecyl-sulfate (SDS) polyacrylamide gel electrophoresis. The P3 polypeptide was considered to be the nephritogenic epitope, since a fraction composed of the P2 polypeptide alone was not nephritogenic. The properties of the nephritogenic epitope were the same as those of the Goodpasture epitope (M2*), which is a noncollagenous domain of the alpha 3 chain of type IV collagen (Butkowski et al. 1985; Saus et al. 1988), indicating that the nephritogenic antigen is the same as the Goodpasture antigen.     

Leptin induces a novel form of NMDA receptor-dependent long-term depression

It is becoming apparent that the hormone leptin plays an important role in modulating hippocampal function. Indeed, leptin enhances NMDA receptor activation and promotes hippocampal long-term potentiation (LTP). Furthermore, obese rodents with dysfunctional leptin receptors display impairments in hippocampal synaptic plasticity. Here we demonstrate that under conditions of enhanced excitability (evoked in Mg2+-free medium or following blockade of GABA(A) receptors), leptin induces a novel form of long-term depression (LTD) in area CA1 of the hippocampus. Leptin-induced LTD was markedly attenuated in the presence of D-(-)-2-Amino-5-Phosphonopentanoic acid (D-AP5), suggesting that it is dependent on the synaptic activation of NMDA receptors. In addition, low-frequency stimulus-evoked LTD occluded the effects of leptin. In contrast, metabotropic glutamate receptors (mGluRs) did not contribute to leptin-induced LTD as mGluR antagonists failed to either prevent or reverse this process. The signalling mechanisms underlying leptin-induced LTD were independent of the Ras-Raf-mitogen-activated protein kinase signalling pathway, but were markedly enhanced following inhibition of either phosphoinositide 3-kinase or protein phosphatases 1 and 2A. These data indicate that under conditions of enhanced excitability, leptin induces a novel form of homosynaptic LTD, which further underscores the proposed key role for this hormone in modulating NMDA receptor-dependent hippocampal synaptic plasticity.     

Thy-1 is a differentiation antigen that characterizes immature murine erythroid and myeloid hematopoietic progenitors

To determine the role of Thy-1 antigen in murine hematopoietic differentiation, bone marrow was treated with anti-Thy-1.2 antibody and complement or complement alone. Growth of immature hematopoietic progenitors, erythroid burst-forming units (BFU-E), and granulocyte/macrophage colony-forming units (CFU-GM) was greatly reduced following antibody and complement treatment and was not restored by mitogen-stimulated spleen cell supernatants. In contrast, more mature erythroid and myeloid progenitors, the erythroid colony-forming unit (CFU-E) and the macrophage progenitor stimulated by L-cell-conditioned media (LCM), were spared by anti-Thy-1.2 antibody and complement treatment. Here, to separate the effects of anti-Thy-1.2 antibody treatment on accessory cells from those on progenitors, splenic T cells and thymocytes were added to treated marrow at ratios of up to 200%. Growth of BFU-E and CFU-GM was not restored. To more precisely replace required accessory cells, male complement-treated marrow was cocultured with female anti-Thy-1.2 antibody and complement-treated marrow. Even marrow cells failed to restore female BFU-E and CFU-GM growth. Fluorescent-activated cell sorting (FACS) and immune sheep red cell rosetting with anti-Thy-1.2-labeled marrow were then performed to determine if immature hematopoietic progenitors bear Thy-1.2. These techniques revealed enrichment of BFU-E and CFU-GM in the Thy-1.2-positive fraction, demonstrating the presence of Thy-1.2 on early murine hematopoietic progenitors. CFU-E and CFU-M were present in the Thy-1.2-negative fraction following FACS separation. These data demonstrate that Thy-1.2 is a differentiation antigen, present on at least some murine BFU-E and CFU-GM and lost as they mature to CFU-E and CFU-M.     

Defining the region of troponin-I that binds to troponin-C

The kinetics and energetics of the binding of three troponin-I peptides, corresponding to regions 96-131 (TnI96-131), 96-139 (TnI96-139), and 96-148 (TnI96-148), to skeletal chicken troponin-C were investigated using multinuclear, multidimensional NMR spectroscopy. The kinetic off-rate and dissociation constants for TnI96-131 (400 s-1, 32 microM), TnI96-139 (65 s-1, <1 microM), and TnI96-148 (45 s-1, <1 microM) binding to TnC were determined from simulation and analysis of the behavior of 1H,15N-heteronuclear single quantum correlation NMR spectra taken during titrations of TnC with these peptides. Two-dimensional 15N-edited TOCSY and NOESY spectroscopy were used to identify 11 C-terminal residues from the 15N-labeled TnI96-148 that were unperturbed by TnC binding. TnI96-139 labeled with 13C at four positions (Leu102, Leu111, Met 121, and Met134) was complexed with TnC and revealed single bound species for Leu102 and Leu111 but multiple bound species for Met121 and Met134. These results indicate that residues 97-136 (and 96 or 137) of TnI are involved in binding to the two domains of troponin-C under calcium saturating conditions, and that the interaction with the regulatory domain is complex. Implications of these results in the context of various models of muscle regulation are discussed.     

A computational framework for gene regulatory network inference that combines multiple methods and datasets

Background  

Reverse engineering in systems biology entails inference of gene regulatory networks from observational data. This data typically include gene expression measurements of wild type and mutant cells in response to a given stimulus. It has been shown that when more than one type of experiment is used in the network inference process the accuracy is higher. Therefore the development of generally applicable and effective methodologies that embed multiple sources of information in a single computational framework is a worthwhile objective.