Paracoccidioides brasilinsis-Induced Migration of Dendritic Cells and Subsequent T-Cell Activation in the Lung-Draining Lymph Nodes

Paracoccidioidomycosis is a mycotic disease caused by a dimorphic fungus, Paracoccidioides brasiliensis (Pb), that starts with inhalation of the fungus; thus, lung cells such as DC are part of the first line of defense against this microorganism. Migration of DC to the lymph nodes is the first step in initiating T cell responses. The mechanisms involved in resistance to Pb infection are poorly understood, but it is likely that DC play a pivotal role in the induction of effector T cells that control Pb infection. In this study, we showed that after Pb Infection, an important modification of lung DC receptor expression occurred. We observed an increased expression of CCR7 and CD103 on lung DC after infection, as well as MHC-II. After Pb infection, bone marrow-derived DC as well lung DC, migrate to lymph nodes. Migration of lung DC could represent an important mechanism of pathogenesis during PCM infection. In resume our data showed that Pb induced DC migration. Furthermore, we demonstrated that bone marrow-derived DC stimulated by Pb migrate to the lymph nodes and activate a T helper (Th) response. To the best of our knowledge, this is the first reported data showing that Pb induces migration of DC and activate a T helper (Th) response.

Leukotriene B(4)-loaded microspheres: a new therapeutic strategy to modulate cell activation

Background: Leukotriene B(4) (LTB(4)) is a potent inflammatory mediator that also stimulates the immune response. In addition, it promotes polymorphonuclear leukocyte phagocytosis, chemotaxis, chemokinesis and modulates cytokines release. Regarding chemical instability of the leukotriene molecule, in the present study we assessed the immunomodulatory activities conferred by LTB(4) released from microspheres (MS). A previous oil-in-water emulsion solvent extraction-evaporation method was chosen to prepare LTB(4)-loaded MS. Results: In the mice cremasteric microcirculation, intraescrotal injection of 0.1 ml of LTB(4)-loaded MS provoked significant increases in leukocyte rolling flux, adhesion and emigration besides significant decreases in the leukocyte rolling velocity. LTB(4)-loaded MS also increase peroxisome proliferator-activated receptor-alpha (PPAR alpha) expression by murine peritoneal macrophages and stimulate them to generate nitrite levels. Monocyte chemoattractant protein-I (MCP-I) and nitric oxide (NO) productions were also increased when human umbilical vein and artery endothelial cells (HUVECs and HUAECs, respectively) were stimulated with LTB(4)-loaded MS. Conclusion: LTB(4)-loaded MS preserve the biological activity of the encapsulated mediator indicating their use as a new strategy to modulate cell activation, especially in the innate immune response.

Structural basis of GC-1 selectivity for thyroid hormone receptor isoforms

Background: Thyroid receptors, TRa and TR beta, are involved in important physiological functions such as metabolism, cholesterol level and heart activities. Whereas metabolism increase and cholesterol level lowering could be achieved by TR beta isoform activation, TRa activation affects heart rates. Therefore, beta-selective thyromimetics have been developed as promising drug-candidates for treatment of obesity and elevated cholesterol level. GC-1 [ 3,5-dimethyl-4-(4'-hydroxy- 3'-isopropylbenzyl)-phenoxy acetic acid] has ability to lower LDL cholesterol with 600-to 1400-fold more potency and approximately two-to threefold more efficacy than atorvastatin (Lipitor(C)) in studies in rats, mice and monkeys. Results: To investigate GC-1 specificity, we solved crystal structures and performed molecular dynamics simulations of both isoforms complexed with GC-1. Crystal structures reveal that, in TRa Arg228 is observed in multiple conformations, an effect triggered by the differences in the interactions between GC-1 and Ser277 or the corresponding asparagine (Asn331) of TR beta. The corresponding Arg282 of TR beta is observed in only one single stable conformation, interacting effectively with the ligand. Molecular dynamics support this model: our simulations show that the multiple conformations can be observed for the Arg228 in TR alpha, in which the ligand interacts either strongly with the ligand or with the Ser277 residue. In contrast, a single stable Arg282 conformation is observed for TR beta, in which it strongly interacts with both GC-1 and the Asn331. Conclusion: Our analysis suggests that the key factors for GC-1 selectivity are the presence of an oxyacetic acid ester oxygen and the absence of the amino group relative to T(3). These results shed light into the beta-selectivity of GC-1 and may assist the development of new compounds with potential as drug candidates to the treatment of hypercholesterolemia and obesity.

Data collapse, scaling functions, and analytical solutions of generalized growth models

We consider a nontrivial one-species population dynamics model with finite and infinite carrying capacities. Time-dependent intrinsic and extrinsic growth rates are considered in these models. Through the model per capita growth rate we obtain a heuristic general procedure to generate scaling functions to collapse data into a simple linear behavior even if an extrinsic growth rate is included. With this data collapse, all the models studied become independent from the parameters and initial condition. Analytical solutions are found when time-dependent coefficients are considered. These solutions allow us to perceive nontrivial transitions between species extinction and survival and to calculate the transition's critical exponents. Considering an extrinsic growth rate as a cancer treatment, we show that the relevant quantity depends not only on the intensity of the treatment, but also on when the cancerous cell growth is maximum.

High levels of T lymphocyte activation in Leishmania-HIV-1 co-infected individuals despite low HIV viral load

Background: Concomitant infections may influence HIV progression by causing chronic activation leading to decline in T-cell function. In the Americas, visceral (AVL) and tegumentary leishmaniasis (ATL) have emerged as important opportunistic infections in HIV-AIDS patients and both of those diseases have been implicated as potentially important co-factors in disease progression. We investigated whether leishmaniasis increases lymphocyte activation in HIV-1 co-infected patients. This might contribute to impaired cellular immune function. Methods: To address this issue we analyzed CD4(+) T absolute counts and the proportion of CD8(+) T cells expressing CD38 in Leishmania/HIV co-infected patients that recovered after anti-leishmanial therapy. Results: We found that, despite clinical remission of leishmaniasis, AVL co-infected patients presented a more severe immunossupression as suggested by CD4(+) T cell counts under 200 cells/mm(3), differing from ATL/HIV-AIDS cases that tends to show higher lymphocytes levels (over 350 cells/mm(3)). Furthermore, five out of nine, AVL/HIV-AIDS presented low CD4(+) T cell counts in spite of low or undetectable viral load. Expression of CD38 on CD8(+) T lymphocytes was significantly higher in AVL or ATL/HIV-AIDS cases compared to HIV/AIDS patients without leishmaniasis or healthy subjects. Conclusions: Leishmania infection can increase the degree of immune system activation in individuals concomitantly infected with HIV. In addition, AVL/HIV-AIDS patients can present low CD4(+) T cell counts and higher proportion of activated T lymphocytes even when HIV viral load is suppressed under HAART. This fact can cause a misinterpretation of these laboratorial markers in co-infected patients.

High Iodine Concentration Attenuates RET/PTC3 Oncogene Activation in Thyroid Follicular Cells

Background: Papillary thyroid carcinoma (PTC) is frequently associated with a RET gene rearrangement that generates a RET/PTC oncogene. RET/PTC is a fusion of the tyrosine kinase domain of RET to the 50 portion of a different gene. This fusion results in a constitutively active MAPK pathway, which plays a key role in PTC development. The RET/PTC3 fusion is primarily associated with radiation-related PTC. Epidemiological studies show a lower incidence of PTC in radiation-exposed regions that are associated with an iodine-rich diet. Since the influence of excess iodine on the development of thyroid cancer is still unclear, the aim of this study is to evaluate the effect of high iodine concentrations on RET/PTC3-activated thyroid cells. Methods: PTC3-5 cells, a rat thyroid cell lineage harboring doxycycline-inducible RET/PTC3, were treated with 10(-3) M NaI. Cell growth was analyzed by cell counting and the MTT assay. The expression and phosphorylation state of MAPK pathway-related (Braf, Erk, pErk, and pRet) and thyroid-specific (natrium-iodide symporter [Nis] and thyroid-stimulating hormone receptor [Tshr]) proteins were analyzed by Western blotting. Thyroid-specific gene expression was further analyzed by quantitative reverse transcription (RT)-polymerase chain reaction. Results: A significant inhibition of proliferation was observed, along with no significant variation in cell death rate, in the iodine-treated cells. Further, iodine treatment attenuated the loss of Nis and Tshr gene and protein expression induced by RET/PTC3 oncogene induction. Finally, iodine treatment reduced Ret and Erk phosphorylation, without altering Braf and Erk expression. Conclusion: Our results indicate an antioncogenic role for excess iodine during thyroid oncogenic activation. These findings contribute to a better understanding of the effect of iodine on thyroid follicular cells, particularly how it may play a protective role during RET/PTC3 oncogene activation.

Activation of endogenous p53 by combined p19Arf gene transfer and nutlin-3 drug treatment modalities in the murine cell lines B16 and C6

Background: Reactivation of p53 by either gene transfer or pharmacologic approaches may compensate for loss of p19Arf or excess mdm2 expression, common events in melanoma and glioma. In our previous work, we constructed the pCLPG retroviral vector where transgene expression is controlled by p53 through a p53-responsive promoter. The use of this vector to introduce p19Arf into tumor cells that harbor p53wt should yield viral expression of p19Arf which, in turn, would activate the endogenous p53 and result in enhanced vector expression and tumor suppression. Since nutlin-3 can activate p53 by blocking its interaction with mdm2, we explored the possibility that the combination of p19Arf gene transfer and nutlin-3 drug treatment may provide an additive benefit in stimulating p53 function. Methods: B16 (mouse melanoma) and C6 (rat glioma) cell lines, which harbor p53wt, were transduced with pCLPGp19 and these were additionally treated with nutlin-3 or the DNA damaging agent, doxorubicin. Viral expression was confirmed by Western, Northern and immunofluorescence assays. p53 function was assessed by reporter gene activity provided by a p53-responsive construct. Alterations in proliferation and viability were measured by colony formation, growth curve, cell cycle and MTT assays. In an animal model, B16 cells were treated with the pCLPGp19 virus and/or drugs before subcutaneous injection in C57BL/6 mice, observation of tumor progression and histopathologic analyses. Results: Here we show that the functional activation of endogenous p53wt in B16 was particularly challenging, but accomplished when combined gene transfer and drug treatments were applied, resulting in increased transactivation by p53, marked cell cycle alteration and reduced viability in culture. In an animal model, B16 cells treated with both p19Arf and nutlin-3 yielded increased necrosis and decreased BrdU marking. In comparison, C6 cells were quite susceptible to either treatment, yet p53 was further activated by the combination of p19Arf and nutlin-3. Conclusions: To the best of our knowledge, this is the first study to apply both p19Arf and nutlin-3 for the stimulation of p53 activity. These results support the notion that a p53 responsive vector may prove to be an interesting gene transfer tool, especially when combined with p53- activating agents, for the treatment of tumors that retain wild-type p53.

Leukotriene B(4) amplifies NF-kappa B activation in mouse macrophages by reducing SOCS1 inhibition of MyD88 expression

Activation of NF-kappa B and 5-lipoxygenase-mediated (5-LO-mediated) biosynthesis of the lipid mediator leukotriene B(4) (LTB(4)) are pivotal components of host defense and inflammatory responses. However, the role of LTB(4) in mediating innate immune responses elicited by specific TLR ligands and cytokines is unknown. Here we have shown that responses dependent on MyD88 (an adaptor protein that mediates signaling through all of the known TLRs, except TLR3, as well as IL-1 beta and IL-18) are reduced in mice lacking either 5-LO or the LTB(4) receptor BTL1, and that macrophages from these mice are impaired in MyD88-dependent activation of NF-kappa B. This macrophage defect was associated with lower basal and inducible expression of MyD88 and reflected impaired activation of STAT1 and overexpression of the STAT1 inhibitor SOCS1. Expression of MyD88 and responsiveness to the TLR4 ligand LPS were decreased by Stat1 siRNA silencing in WT macrophages and restored by Socs1 siRNA in 5-LO-deficient macrophages. These results uncover a pivotal role in macrophages for the GPCR BLT1 in regulating activation of NF-kappa B through Stat1-dependent expression of MyD88.

Comparative Analysis of Activation Phenotype, Proliferation, and IFN-gamma Production by Spleen NK1.1(+) and NK1.1(-) T Cells During Plasmodium chabaudi AS Malaria

The NK1.1 molecule participates in NK, NKT, and T-cell activation, contributing to IFN-gamma production and cytotoxicity. To characterize the early immune response to Plasmodium chabaudi AS, spleen NK1.1(+) and NK1.1(-) T cells were compared in acutely infected C57BL/6 mice. The first parasitemia peak in C57BL/6 mice correlated with increase in CD4(+)NK1.1(+)TCR-alpha beta(+), CD8(+)NK1.1(+)TCR-alpha beta(+), and CD4(+)NK1.1(-)TCR-alpha beta(+) cell numbers per spleen, where a higher increment was observed for NK1.1(+) T cells compared to NK1.1(-) T cells. According to the ability to recognize the CD1d-alpha-GalCer tetramer, CD4(+)NK1.1(+) cells in 7-day infected mice were not predominantly invariant NKT cells. At that time, nearly all NK1.1(+) T cells and around 30% of NK1.1(-) T cells showed an experienced/activated (CD44(HI)CD69(HI)CD122(HI)) cell phenotype, with high expression of Fas and PD-L1 correlating with their low proliferative capacity. Moreover, whereas IFN-gamma production by CD4(+)NK1.1(+) cells peaked at day 4 p.i., the IFN-gamma response of CD4(+)NK1.1(-) cells continued to increase at day 5 of infection. We also observed, at day 7 p.i., 2-fold higher percentages of perforin(+) cells in CD8(+)NK1.1(+) cells compared to CD8(+)NK1.1(-) cells. These results indicate that spleen NK1.1(+) and NK1.1(-) T cells respond to acute P. chabaudi malaria with different kinetics in terms of activation, proliferation, and IFN-gamma production.

Impaired Innate Immunity in Tlr4(-/-) Mice but Preserved CD8(+) T Cell Responses against Trypanosoma cruzi in Tlr4-, Tlr2-, Tlr9- or Myd88-Deficient Mice

The murine model of T. cruzi infection has provided compelling evidence that development of host resistance against intracellular protozoans critically depends on the activation of members of the Toll-like receptor (TLR) family via the MyD88 adaptor molecule. However, the possibility that TLR/MyD88 signaling pathways also control the induction of immunoprotective CD8(+) T cell-mediated effector functions has not been investigated to date. We addressed this question by measuring the frequencies of IFN-gamma secreting CD8(+) T cells specific for H-2K(b)-restricted immunodominant peptides as well as the in vivo Ag-specific cytotoxic response in infected animals that are deficient either in TLR2, TLR4, TLR9 or MyD88 signaling pathways. Strikingly, we found that T. cruzi-infected Tlr2(-/-), Tlr4(-/-), Tlr9(-/-) or Myd88(-/-) mice generated both specific cytotoxic responses and IFN-gamma secreting CD8(+) T cells at levels comparable to WT mice, although the frequency of IFN-gamma(+)CD4(+) cells was diminished in infected Myd88(-/-) mice. We also analyzed the efficiency of TLR4-driven immune responses against T. cruzi using TLR4-deficient mice on the C57BL genetic background (B6 and B10). Our studies demonstrated that TLR4 signaling is required for optimal production of IFN-gamma, TNF-alpha and nitric oxide (NO) in the spleen of infected animals and, as a consequence, Tlr4(-/-) mice display higher parasitemia levels. Collectively, our results indicate that TLR4, as well as previously shown for TLR2, TLR9 and MyD88, contributes to the innate immune response and, consequently, resistance in the acute phase of infection, although each of these pathways is not individually essential for the generation of class I-restricted responses against T. cruzi.

Segregation and activation of myosin IIB creates a rear in migrating cells

We have found that MLC-dependent activation of myosin IIB in migrating cells is required to form an extended rear, which coincides with increased directional migration. Activated myosin IIB localizes prominently at the cell rear and produces large, stable actin. lament bundles and adhesions, which locally inhibit protrusion and de. ne the morphology of the tail. Myosin IIA forms de novo. laments away from the myosin IIB-enriched center and back to form regions that support protrusion. The positioning and dynamics of myosin IIA and IIB depend on the self-assembly regions in their coiled-coil C terminus. COS7 and B16 melanoma cells lack myosin IIA and IIB, respectively; and show isoform-specific front-back polarity in migrating cells. These studies demonstrate the role of MLC activation and myosin isoforms in creating a cell rear, the segregation of isoforms during. lament assembly and their differential effects on adhesion and protrusion, and a key role for the noncontractile region of the isoforms in determining their localization and function.

Charm and longitudinal structure functions within the Kharzeev-Levin-Nardi model

We use the Kharzeev-Levin-Nardi (KLN) model of the low x gluon distributions to fit recent HERA data on F(L) and F(2)(c)(F(2)(b)). Having checked that this model gives a good description of the data, we use it to predict F(L) and F(2)(c) to be measured in a future electron-ion collider. The results are similar to those obtained with the de Florian-Sassot and Eskola-Paukkunen-Salgado nuclear gluon distributions. The conclusion of this exercise is that the KLN model, simple as it is, may still be used as an auxiliary tool to make estimates for both heavy-ion and electron-ion collisions.

Use of correlated potential harmonic basis functions for the description of the (4)He trimer and small clusters

A correlated many-body basis function is used to describe the (4)He trimer and small helium clusters ((4)HeN) with N = 4-9. A realistic helium dimer potential is adopted. The ground state results of the (4)He dimer and trimer are in close agreement with earlier findings. But no evidence is found for the existence of Efimov state in the trimer for the actual (4)He-(4)He interaction. However, decreasing the potential strength we calculate several excited states of the trimer which exhibit Efimov character. We also solve for excited state energies of these clusters which are in good agreement with Monte Carlo hyperspherical description. (C) 2011 American Institute of Physics. [doi:10.1063/1.3583365]

Deformed Gaussian-orthogonal-ensemble description of small-world networks

The study of spectral behavior of networks has gained enthusiasm over the last few years. In particular, random matrix theory (RMT) concepts have proven to be useful. In discussing transition from regular behavior to fully chaotic behavior it has been found that an extrapolation formula of the Brody type can be used. In the present paper we analyze the regular to chaotic behavior of small world (SW) networks using an extension of the Gaussian orthogonal ensemble. This RMT ensemble, coined the deformed Gaussian orthogonal ensemble (DGOE), supplies a natural foundation of the Brody formula. SW networks follow GOE statistics until a certain range of eigenvalue correlations depending upon the strength of random connections. We show that for these regimes of SW networks where spectral correlations do not follow GOE beyond a certain range, DGOE statistics models the correlations very well. The analysis performed in this paper proves the utility of the DGOE in network physics, as much as it has been useful in other physical systems.

Balance functions from Au+Au, d+Au, and p+p collisions at root s(NN)=200 GeV

Balance functions have been measured for charged-particle pairs, identified charged-pion pairs, and identified charged-kaon pairs in Au + Au, d + Au, and p + p collisions at root s(NN) = 200 GeV at the Relativistic Heavy Ion Collider using the STAR detector. These balance functions are presented in terms of relative pseudorapidity, Delta eta, relative rapidity, Delta y, relative azimuthal angle, Delta phi, and invariant relative momentum, q(inv). For charged-particle pairs, the width of the balance function in terms of Delta eta scales smoothly with the number of participating nucleons, while HIJING and UrQMD model calculations show no dependence on centrality or system size. For charged-particle and charged-pion pairs, the balance functions widths in terms of Delta eta and Delta y are narrower in central Au + Au collisions than in peripheral collisions. The width for central collisions is consistent with thermal blast-wave models where the balancing charges are highly correlated in coordinate space at breakup. This strong correlation might be explained by either delayed hadronization or limited diffusion during the reaction. Furthermore, the narrowing trend is consistent with the lower kinetic temperatures inherent to more central collisions. In contrast, the width of the balance function for charged-kaon pairs in terms of Delta y shows little centrality dependence, which may signal a different production mechanism for kaons. The widths of the balance functions for charged pions and kaons in terms of q(inv) narrow in central collisions compared to peripheral collisions, which may be driven by the change in the kinetic temperature.