In vivo kinetics of eosinophils and mast cells in experimental murine Schistosomiasis

During the schistosomiasis infection there is a [quot ]dance of the cells[quot ], varying from site to site and related to the time of infection. 1 - Eosinophil levels exhibit a bimodal pattern, with the first peak related to the egg deposition and maturation and increased Kupfferian hyperplasia; the second peak precedes the death of some adult worms; 2 - The peritoneal eosinophilic levels are inversely proportional to the blood eosinophilic levels; 3 - Eosinopoiesis in the bone marrow begins at day 40, reaching the highest levels at day 50 and coincides with hepatic eosinophilic and neutrophilic metaplasia; 4 - Peritoneal mast cell levels present a bimodal pattern similar to the blood eosinophils, and inverse to the peritoneal eosinophils. They also show a cyclic behaviour within the hepatic and intestinal granulomas. Integral analysis of the events related to the eosinophils in the blood, bone marrow, peritoneal cavity and hepatic and intestinal granulomas allows the detection of two important eosinophilic phases: the first is due to mobilization and redistribution of the marginal pool and the second originates from eosinophilic production in the bone marrow and liver. The productive phase is characterized by an increase in the number of eosinophils and monocyte/macrophages, and a decrease in neutrophils and stabilization of megakariocytes and erithroid lineages.

Kinetics of antigen specific and non-specific polyclonal B-cell responses during lethal Plasmodium yoelii malaria

In order to study the kinetics and composition of the polyclonal B-cell activation associated to malaria infection, antigen-specific and non-specific B-cell responses were evaluated in the spleens of mice infected with Plasmodium yoelii 17 XL or injected with lysed erythrocytes or plasma from P. yoelii infected mice or with P. falciparum culture supernatants. Spleen/body weigth ratio, numbers of nucleated spleen cells and Immunoglobulin-containing and Immunoglobulin-secreting cells increased progressively during the course of infection,in parallel to the parasitemia. A different pattern of kinetics was observed when anti-sheep red blood cell and anti-trinitrophenylated-sheep red blood cell plaque forming cells response were studied: maximum values were observed at early stages of infection, whereas the number of total Immunoglobulin-containing and Immunoglobulin-secreting cells were not yet altered. Conversely, at the end of infection, when these latter values reached their maximum, the anti-sheep red blood cell and anti-trinitrophenylated-sheep red blood cell specific responses were normal or even infranormal. In mice injected with Plasmodium-derived material, a higher increase in antigen-specific PFC was observed, as compared to the increase of Immunoglobulin-containing and Immunoglobulin-secreting cell numbers. This suggested a "preferential" (antigen-plus mitogen-induced) stimulation of antigen-specific cells rather than a generalized non-specific (mitogen-induced) triggering of B-lymphocytes. On the basis of these and previous results, it is suggested that polyclonal B-cell activation that takes place during the course of infection appears as a result of successive waves of antigen-specific B-cell activation.

Reverse transcriptase-dependent and -independent phases of infection with mouse mammary tumor virus: implications for superantigen function.

Mouse mammary tumor virus (MMTV) encodes a superantigen (SAg) that promotes stable infection and virus transmission. Upon subcutaneous MMTV injection, infected B cells present SAg to SAg-reactive T cells leading to a strong local immune response in the draining lymph node (LN) that peaks after 6 d. We have used the reverse transcriptase inhibitor 3'-azido-3'-deoxythymidine (AZT) to dissect in more detail the mechanism of SAg-dependent enhancement of MMTV infection in this system. Our data show that no detectable B or T cell response to SAg occurs in AZT pretreated mice. However, if AZT treatment is delayed 1-2 d after MMTV injection, a normal SAg-dependent local immune response is observed on day 6. Quantitation of viral DNA in draining LN of these infected mice indicates that a 4,000-fold increase in the absolute numbers of infected cells occurs between days 2 and 6 despite the presence of AZT. Furthermore MMTV DNA was found preferentially in surface IgG+ B cells of infected mice and was not detectable in SAg-reactive T cells. Collectively our data suggest that MMTV infection occurs preferentially in B cells without SAg involvement and is completed 1-2 d after virus challenge. Subsequent amplification of MMTV infection between days 2 and 6 requires SAg expression and occurs in the absence of any further requirement for reverse transcription. We therefore conclude that clonal expansion of infected B cells via cognate interaction with SAg-reactive T cells is the predominant mechanism for increasing the level of MMTV infection. Since infected B cells display a memory (surface IgG+) phenotype, both clonal expansion and possibly longevity of the virus carrier cells may contribute to stable MMTV infection.

Retrovirus-induced target cell activation in the early phases of infection: the mouse mammary tumor virus model.

Mouse mammary tumor virus (MMTV) infects B lymphocytes and expresses a superantigen on the cell surface after integration of its reverse-transcribed genome. Superantigen-dependent B- and T-cell activation becomes detectable 2 to 3 days after infection. We show here that before this event, B cells undergo a polyclonal activation which does not involve massive proliferation. This first phase of B-cell activation is T cell independent. Moreover, during the first phase of activation, when only a small fraction of B cells is infected by MMTV(SW), viral DNA is detected only in activated B cells. Such a B-cell activation is also seen after injection of murine leukemia virus but not after injection of vaccinia virus, despite the very similar kinetics and intensity of the immune response. Since retroviruses require activated target cells to induce efficient infection, these data suggest that the early polyclonal retrovirus-induced target cell activation might play an important role in the establishment of retroviral infections.

Monitoring procalcitonin in febrile neutropenia: what is its utility for initial diagnosis of infection and reassessment in persistent Fever?

Background: Management of febrile neutropenic episodes (FE) is challenged by lacking microbiological and clinical documentation of infection. We aimed at evaluating the utility of monitoring blood procalcitonin (PCT) in FE for initial diagnosis of infection and reassessment in persistent fever.Methods: PCT kinetics was prospectively monitored in 194 consecutive FE (1771 blood samples): 65 microbiologically documented infections (MDI, 33.5%; 49 due to non-coagulase-negative staphylococci, non-CNS), 68 clinically documented infections (CDI, 35%; 39 deep-seated), and 61 fever of unexplained origin (FUO, 31.5%).Results: At fever onset median PCT was 190 pg/mL (range 30-26'800), without significant difference among MDI, CDI and FUO. PCT peak occurred on day 2 after onset of fever: non-CNS-MDI/deep-seated-CDI (656, 80-86350) vs. FUO (205, 33-771; p<0.001). PCT >500 pg/mL distinguished non-CNS-MDI/deep-seated-CDI from FUO with 56% sensitivity and 90% specificity. PCT was >500 pg/ml in only 10% of FUO (688, 570-771). A PCT peak >500 pg/mL (1196, 524-11950) occurred beyond 3 days of persistent fever in 17/21 (81%) invasive fungal diseases (IFD). This late PCT peak identified IFD with 81% sensitivity and 57% specificity and preceded diagnosis according to EORTC-MSG criteria in 41% of cases. In IFD responding to therapy, median days to PCT <500 pg/mL and defervescence were 5 (1-23) vs. 10 (3-22; p = 0.026), respectively.Conclusion: While procalcitonin is not useful for diagnosis of infection at onset of neutropenic fever, it may help to distinguish a minority of potentially severe infections among FUOs on day 2 after onset of fever. In persistent fever monitoring procalcitonin contributes to early diagnosis and follow-up of invasive mycoses

Rapid IL-4 production by Leishmania homolog of mammalian RACK1-reactive CD4(+) T cells in resistant mice treated once with anti-IL-12 or -IFN-gamma antibodies at the onset of infection with Leishmania major instructs Th2 cell development, resulting in nonhealing lesions.

Rapid production of IL-4 by Leishmania homolog of mammalian RACK1 (LACK)-reactive CD4(+) T cells expressing the V beta 4-V alpha 8 TCR chains has been shown to drive aberrant Th2 cell development and susceptibility to Leishmania major in BALB/c mice. In contrast, mice from resistant strains fail to express this early IL-4 response. However, administration of either anti-IL-12 or -IFN-gamma at the initiation of infection allows the expression of this early IL-4 response in resistant mice. In this work we show that Leishmania homolog of mammalian RACK1-reactive CD4(+) T cells also expressing the V beta 4-V alpha 8 TCR chains are the source of the early IL-4 response to L. major in resistant mice given anti-IL-12 or -IFN-gamma Abs only at the onset of infection. Strikingly, these cells were found to be required for the reversal of the natural resistance of C57BL/6 mice following a single administration of anti-IL-12 or -IFN-gamma Abs. Together these results suggest that a deficiency in mechanisms capable of down-regulating the early IL-4 response to L. major contributes to the exquisite susceptibility of BALB/c mice to L. major.

Different inflammatory stimuli in the footpad of mice influence the kinetics of resident peritoneal cells

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Kinetics of ocular and systemic antigen-specific T-cell responses elicited during murine cytomegalovirus retinitis

Cytomegalovirus (CMV) reactivation in the retina of immunocompromized patients is a cause of significant morbidity as it can lead to blindness. The adaptive immune response is critical in controlling murine CMV (MCMV) infection in MCMV-susceptible mouse strains. CD8(+) T cells limit systemic viral replication in the acute phase of infection and are essential to contain latent virus. In this study, we provide the first evaluation of the kinetics of anti-viral T-cell responses after subretinal infection with MCMV. The acute response was characterized by a rapid expansion phase, with infiltration of CD8(+) T cells into the infected retina, followed by a contraction phase. MCMV-specific T cells displayed biphasic kinetics with a first peak at day 12 and contraction by day 18 followed by sustained recruitment of these cells into the retina at later time points post-infection. MCMV-specific CD8(+) T cells were also observed in the draining cervical lymph nodes and the spleen. Presentation of viral epitopes and activation of CD8(+) T cells was widespread and could be detected in the spleen and the draining lymph nodes, but not in the retina or iris. Moreover, after intraocular infection, antigen-specific cytotoxic activity was detectable and exhibited kinetics equivalent to those observed after intraperitoneal infection with the same viral dose. These data provide novel insights of how and where immune responses are initiated when viral antigen is present in the subretinal space.

Long-term kinetics of T cell production in HIV-infected subjects treated with highly active antiretroviral therapy

The long-term kinetics of T cell production following highly active antiretroviral therapy (HAART) were investigated in blood and lymph node in a group of HIV-infected subjects at early stage of established infection and prospectively studied for 72 wk. Before HAART, CD4 and CD8 T cell turnover was increased. However, the total number of proliferating CD4+ T lymphocytes, i.e., CD4+Ki67+ T lymphocytes, was not significantly different in HIV-infected (n = 73) and HIV-negative (n = 15) subjects, whereas proliferating CD8+Ki67+ T lymphocytes were significantly higher in HIV-infected subjects. After HAART, the total body number of proliferating CD4+Ki67+ T lymphocytes increased over time and was associated with an increase of both naive and memory CD4+ T cells. The maximal increase (2-fold) was observed at week 36, whereas at week 72 the number of proliferating CD4+ T cells dropped to baseline levels, i.e., before HAART. The kinetics of the fraction of proliferating CD4 and CD8 T cells were significantly correlated with the changes in the total body number of these T cell subsets. These results demonstrate a direct relationship between ex vivo measures of T cell production and quantitative changes in total body T lymphocyte populations. This study provides advances in the delineation of the kinetics of T cell production in HIV infection in the presence and/or in the absence of HAART.

The Kinetics of Myeloid Derived Suppressor Cell Frequency and Function in Chronic Retroviral Infections

Thesis (Ph.D.)--University of Washington, 2016-06

Thermal behavior and decomposition kinetics of rifampicin polymorphs under isothermal and non-isothermal conditions

The thermal behavior of two polymorphic forms of rifampicin was studied by DSC and TG/DTG. The thermoanalytical results clearly showed the differences between the two crystalline forms. Polymorph I was the most thermally stable form, the DSC curve showed no fusion for this species and the thermal decomposition process occurred around 245 ºC. The DSC curve of polymorph II showed two consecutive events, an endothermic event (Tpeak = 193.9 ºC) and one exothermic event (Tpeak = 209.4 ºC), due to a melting process followed by recrystallization, which was attributed to the conversion of form II to form I. Isothermal and non-isothermal thermogravimetric methods were used to determine the kinetic parameters of the thermal decomposition process. For non-isothermal experiments, the activation energy (Ea) was derived from the plot of Log β vs 1/T, yielding values for polymorph form I and II of 154 and 123 kJ mol-1, respectively. In the isothermal experiments, the Ea was obtained from the plot of lnt vs 1/T at a constant conversion level. The mean values found for form I and form II were 137 and 144 kJ mol-1, respectively.

Study of infection by Rickettsiae of the spotted fever group in humans and ticks in an urban park located in the City of Londrina, State of Paraná, Brazil

INTRODUCTION: Spotted fevers are emerging zoonoses caused by Rickettsia species in the spotted fever group (SFG). Rickettsia rickettsii is the main etiologic agent of Brazilian spotted fever (BSF) and it is transmitted by Amblyomma spp. ticks. METHODS: The study aimed to investigate SFG rickettsiae in the Arthur Thomas Municipal Park in Londrina, PR, by collecting free-living ticks and ticks from capybaras and blood samples from personnel working in these areas. Samples from A. dubitatum and A. cajennense were submitted for PCR in pools to analyze the Rickettsia spp. gltA (citrate synthase gene). RESULTS: All the pools analyzed were negative. Human sera were tested by indirect immunofluorescence assay with R. rickettsii and R. parkeri as antigens. Among the 34 sera analyzed, seven (20.6%) were reactive for R. rickettsii: four of these had endpoint titers equal to 64, 2 titers were 128 and 1 titer was 256. None of the samples were reactive for R. parkeri. An epidemiological questionnaire was applied to the park staff, but no statistically significant associations were identified. CONCLUSIONS: The serological studies suggest the presence of Rickettsiae related to SFG that could be infecting the human population studied; however, analysis of the ticks collected was unable to determine which species may be involved in transmission to humans.

Lipid Metabolism in Subclinical Hypothyroidism: Plasma Kinetics of Triglyceride-Rich Lipoproteins and Lipid Transfers to High-Density Lipoprotein Before and After Levothyroxine Treatment

Background: Subclinical hypothyroidism (SCH) has been associated with atherosclerosis, but the abnormalities in plasma lipids that can contribute to atherogenesis are not prominent. The aim of this study was to test the hypothesis that patients with normocholesterolemic, normotriglyceridemic SCH display abnormalities in plasma lipid metabolism not detected in routine laboratory tests including abnormalities in the intravascular metabolism of triglyceride-rich lipoproteins, lipid transfers to high-density lipoprotein (HDL), and paraoxonase 1 activity. The impact of levothyroxine (LT4) treatment and euthyroidism in these parameters was also tested. Methods: The study included 12 SCH women and 10 matched controls. Plasma kinetics of an artificial triglyceride-rich emulsion labeled with radioactive triglycerides and cholesteryl esters as well as in vitro transfer of four lipids from an artificial donor nanoemulsion to HDL were determined at baseline in both groups and after 4 months of euthyroidism in the SCH group. Results: Fractional clearance rates of triglycerides (SCH 0.035 +/- 0.016 min(-1), controls 0.029 +/- 0.013 min(-1), p=0.336) and cholesteryl esters (SCH 0.009 +/- 0.007 min(-1), controls 0.009 +/- 0.009 min(-1), p=0.906) were equal in SCH and controls and were unchanged by LT4 treatment and euthyroidism in patients with SCH, suggesting that lipolysis and remnant removal of triglyceride-rich lipoproteins were normal. Transfer of triglycerides to HDL (SCH 3.6 +/- 0.48%, controls 4.7 +/- 0.63%, p=0.001) and phospholipids (SCH 16.2 +/- 3.58%, controls 21.2 +/- 3.32%, p=0.004) was reduced when compared with controls. After LT4 treatment, transfers increased and achieved normal values. Transfer of free and esterified cholesterol to HDL, HDL particle size, and paraoxonase 1 activity were similar to controls and were unchanged by treatment. Conclusions: Although intravascular metabolism of triglyceride-rich lipoproteins was normal, patients with SCH showed abnormalities in HDL metabolism that were reversed by LT4 treatment and achievement of euthyroidism.

Kinetics of the xylitol crystallization in hydro-alcoholic solution

Nyvlt method Was used to determine the kinetic parameters of commercial xylitol in ethanol:water (50:50 %w/w) Solution by batch cooling crystallization. The kinetic exponents (n, g and in) and the system kinetic constant (B(N)) were determined. Model experiments were carried Out in order to verify the combined effects of saturation temperatures (40, 50 and 60 degrees C) and cooling rates (0.10, 0.25 and 0.50 degrees C/min) on these parameters. The fitting between experimental and Calculated crystal sizes has 11.30% mean deviation. (C) 2007 Elsevier B.V. All rights reserved.

Characterization and kinetics of sulfide-oxidizing autotrophic denitrification in batch reactors containing suspended and immobilized cells

Sulfide-oxidizing autotrophic denitrification is an advantageous alternative over heterotrophic denitrification, and may have potential for nitrogen removal of low-strength wastewaters, such as anaerobically pre-treated domestic sewage. This study evaluated the fundamentals and kinetics of this process in batch reactors containing suspended and immobilized cells. Batch tests were performed for different NO(x)(-)/S(2-) ratios and using nitrate and nitrite as electron acceptors. Autotrophic denitrification was observed for both electron acceptors, and NO(x)(-)/S(2-) ratios defined whether sulfide oxidation was complete or not. Kinetic parameter values obtained for nitrate were higher than for nitrite as electron acceptor. Zero-order models were better adjusted to profiles obtained for suspended cell reactors, whereas first-order models were more adequate for immobilized cell reactors. However, in the latter, mass transfer physical phenomena had a significant effect on kinetics based on biochemical reactions. Results showed that sulfide-oxidizing autotrophic denitrification can be successfully established for low-strength wastewaters and have potential for nitrogen removal from anaerobically pre-treated domestic sewage.