Molecular and flow cytometric characterization of the CD4 and CD8 T-cell repertoire in patients with myelodysplastic syndrome

We studied 18 patients with myelodysplastic syndrome (MDS), measuring clonality and T-cell receptor Vbeta (TCRBV) expression of CD4 and CD8 T cells by polymerase chain reaction and by flow cytometric analysis of TCRBV families. The CD4 and CD8 T-cell repertoire in most MDS patients is characterized by an abnormal TCRBV-restricted expansion of T cells in CD4 and CD8 cells, and increased expression of the CD8 effector marker CD57 of multiple TCRBV in CD8 cells. Clonality analysis of CD4 and CD8 cells showed that seven of 10 patients analysed had a major clone in the CD8 cells but not in CD4 cells. Furthermore, in one patient we found that both the CD57- and CD57+ fraction contained the clone (which was absent from the TCRBV-negative fraction). These data suggest that, in MDS, multiple T-cell expansions can be found in both helper and cytotoxic T cells, and that, in the CD8 cells, T cells functionally differentiate in vivo from memory to effector T cells. Together, these data support the hypothesis of the involvement of T cells in the pathogenesis of MDS.     

Distinct domain responses of R-state human hemoglobins A,C, and S to anions

Anionic regulation of hemoglobin (Hb) is of increasing interest for the design of Hb-based oxygen carriers. Even "external" amino-acid substitutions can alter the nature and extent of anionic control. This was shown by evaluation of the anion sensitivities of liganded, R-state, forms of HbA, HbC (beta6 Glu --> Lys) and HbS (beta6 Glu --> Val). The beta6 mutants differ in the anion-sensitivity of their central cavities, alpha1beta2 interfaces, and heme and beta93 Cys environments. The mutant Hbs also exhibit increased anion-dependent oxidation and surface denaturation. Moreover, differential chloride effects on oxygen binding by Hbs C, S compared to HbA occur after R-state stabilization by fluoresceination of beta93 Cys. It is concluded that the "external" substitutions in the mutant Hbs have structural consequences that are propagated to varying extents to other domains as a result of anion binding, and that these anion-dependent changes may underlie mechanisms leading to the observed increase in oxidation propensity and surface denaturation.     

Depression in perinatally HIV-infected pregnant women compared to non-perinatally HIV-infected and HIV-uninfected pregnant women

“Depression (as noted in chart by a physician)” was compared between HIV infected pregnant women and controls. Perinatally HIV-infected (PHIV), non-perinatally HIV-infected (NPHIV), and HIV-uninfected (HIV-U) pregnant women were all compared using a logistic regression model. Overall, HIV-infected women had higher rates of depression than HIV-U, with PHIV women demonstrating a clinically and statistically significant increased risk compared to HIV-U women [adjusted OR: 15.9, 95% CI?=?1.8–143.8]. Future studies in larger populations are warranted to confirm these findings and further elucidate mental health outcomes of PHIV and NPHIV pregnant women.     

Provider-initiated HIV testing for paediatric inpatients and their caretakers is feasible and acceptable

Objectives  Early diagnosis of HIV-infected children remains a major challenge in Africa. Children who are hospitalised represent an opportunity for HIV diagnosis and appropriate treatment. We introduced HIV Counselling and Testing (HCT) for hospitalised children and their caretakers in Mulago teaching hospital in Uganda to assess its feasibility.
Methods  We analysed routine program data for children and caretakers who were tested between February 2005 and February 2008 to assess the proportion of children and caretakers who were HIV-infected. We also assessed the level of immune suppression (CD4 percentage) in a subset of HIV infected children tested between January 2007 and December 2007.
Results  Caretakers agreed to HIV testing for 8990 (92.8%) of the 9687 children who were offered HIV testing. Among the caretakers, 89.8% agreed to be tested. At the time of hospitalization, 41.3% of the caretakers had previously tested for HIV. Although 313 parents (mothers and fathers) reported that they had previously tested HIV positive, only 113 (36.3%) of these had tested their children prior to hospitalization. Overall HIV prevalence among caretakers was 16.7%. HIV prevalence among children was 12.4%, highest on the nutrition ward (30.8%). Of those children who underwent CD4 counts, 56.4% had a CD4 percentage of <20%.
Conclusion  HCT for hospitalized children and their caretakers identified a significant number of HIV infected children and caretakers. More than half of the children had advanced HIV disease. More intensive efforts are needed to ensure earlier diagnosis and linkage to care for HIV infected children.     

A phase I trial of perillyl alcohol administered four times daily for 14 days out of 28 days

Purpose Perillyl alcohol (POH) has been shown to have both chemopreventative and chemotherapeutic activities in preclinical studies. The underlying mechanism(s) of action of POH have yet to be delineated but may involve effects on the transforming growth factor (TGF) and/or the Ras signaling pathways. A phase I study of POH for 14 days out of every 28 days in subjects with advanced malignancies was performed to evaluate dose escalation, toxicity, pharmacokinetics, and effects on TGF and Ras.Methods POH was administered orally (500 mg capsules containing 250 mg POH) to 20 patients four times a day on a continuous basis for 14 days followed by a 14-day rest period, for up to three courses. The starting dose was 1200 mg/m² per dose. A minimum of three patients were treated and evaluated at each escalating POH dose. Pharmacokinetic analysis was performed on days 1 and 14 of course 1 and day 1 of selected later courses. Plasma TGF levels were measured on days 1 and 14. Peripheral blood lymphocyte (PBLs) Ras levels were assayed on days 1 and 2 of the first course.Results The 20 patients, of whom 15 were evaluable, received doses between 1200 and 2000 mg/m² per dose for a total of 43 courses. The most common observed toxicities were nausea, gastrointestinal distress, and fatigue. Other toxicities included diarrhea or constipation, hypokalemia, and one incidence of acute pancreatitis. Due to these toxicities, four of the patients declined further treatment either during or after the second course. While POH was not detected in plasma, perillic acid (PA) and dihydroperillic acid (DHPA) were detected in plasma, and the peak levels at 2000 mg/m² per dose were approximately 600 M (PA) and 50 M (DHPA). There was some evidence for linearity in the peak plasma levels and area under the concentration–time curve of the metabolites from the starting dose to the highest dose. Metabolite pharmacokinetics were not significantly affected by ingestion in the fed or fasting state, or repeated exposure to POH. No evidence for an effect of POH on plasma TGF or PBL Ras protein was observed. No objective responses were observed.Conclusions In adults with advanced malignancies, an interrupted administration schedule of POH did not reveal significant advantages over continuous dosing schedules.     

The many faces of preparatory control in task switching: Reviewing a decade of fMRI research

A large body of behavioural research has used the cued task‐switching paradigm to characterize the nature of trial‐by‐trial preparatory adjustments that enable fluent task implementation when demands on cognitive flexibility are high. This work reviews the growing number of fMRI studies on the same topic, mostly focusing on the central hypothesis that preparatory adjustments should be indicated by enhanced prefrontal and parietal BOLD activation in task switch when compared with task repeat trials under conditions that enable advance task preparation. The evaluation of this straight‐forward hypothesis reveals surprisingly heterogeneous results regarding both the precise localization and the very existence of switch‐related preparatory activation. Explanations for these inconsistencies are considered on two levels. First, we discuss methodological issues regarding (i) the possible impact of different fMRI‐specific experimental design modifications and (ii) statistical uncertainty in the context of massively multivariate imaging data. Second, we discuss explanations related to the multidimensional nature of task preparation itself. Specifically, the precise localization and the size of switch‐related preparatory activation might depend on the differential interplay of hierarchical control via abstract task goals and attentional versus action‐directed preparatory processes. We argue that different preparatory modes can be adopted relying either on advance goal activation alone or on the advance resolution of competition within action sets or attentional sets. Importantly, while either mode can result in a reduction of behavioral switch cost, only the latter two are supposed to be associated with enhanced switch versus repeat BOLD activation in prepared trial conditions. Hum Brain Mapp, 2013. © 2011 Wiley Periodicals, Inc.     

Metabolism and bioactivation of 3-methylindole by human liver microsomes

Metabolism and bioactivation of 3-methylindole (3MI) were investigated in human liver microsomes. The metabolism of two deuterium-labeled analogues of 3MI permitted a relatively unambiguous identification of multiple metabolites and glutathione (GSH) adducts of reactive intermediates. A total of eight oxidized metabolites were detected, five of which were assigned as previously identified 3-methyloxindole, 3-hydroxy-3-methylindolenine, 3-hydroxy-3-methyloxindole, 5-hydroxy-3-methylindole, and 6-hydroxy-3-methylindole. Among the three new metabolites, one was either 4- or 7-OH-3-methylindole, and the other two were derived from additional oxidation on the phenyl ring of 3-methyloxindole. When GSH was added to the microsomal incubations, seven conjugates that had molecular ions corresponding to the incorporation of GSH and an atom of oxygen at m/z 453 (group I) were produced, and two additional conjugates had molecular ions at m/z 437 that corresponded to the incorporation of GSH with no additional oxygen (group II). Two conjugates in group I (m/z 453) were apparently derived by GSH addition to the 5,6-epoxide metabolite of 3-methyloxindole. These two GSH adducts were tentatively identified as 5-(glutathione-S-yl)-3-methyloxindole and 6-(glutathione-S-yl)-3-methyloxindole. The most abundant conjugate in group I was identified as 3-(glutathione-S-yl)-3-methyloxindole, which substantiated the presence of the putative 2,3-epoxy-3-methylindole intermediate. The remaining four adducts in group I were likely formed by conjugation of GSH at different positions of the phenyl ring, possibly via oxidation of 5-hydroxy-3-methylindole and 6-hydroxy-3-methylindole to two very interesting new electrophilic benzoquinone imine intermediates. For the group II conjugates (m/z 437), two isomers were identified as 2-(glutathione-S-yl)-3-methylindole and 3-(glutathione-S-yl-methyl)-indole. The former adduct was primarily derived from the 2,3-epoxide intermediate by thiol conjugation followed by dehydration. The latter adduct was consistent with our previously published work on the dehydrogenation of 3MI. In those studies, we showed that the reactive intermediate, 3-methylenenindolenine, was formed by hydrogen abstraction at the methyl group and was trapped with GSH. The putative dehydrogenation bioactivation mechanism is also substantiated by the finding that CYP2E1 selectively generated 2-(glutathione-S-yl)-3-methylindole but did not produce 3-(glutathione-S-yl-methyl)-indole. In summary, the results not only confirmed the formation of 2,3-epoxide-3-methylindole in human liver microsomes but also suggested that the phenolic metabolites of 3-methylindole were dehydrogenated to previously uncharacterized reactive intermediates.