Interleukin 10 production in patients undergoing cardiopulmonary bypass: evidence of inhibition of Th-1-type responses

The cardiopulmonary bypass (CPB) procedure has long been associated with a generalized immunosuppression. To understand further the cytokine-mediated regulation of the complex physiological and immunological changes induced by CPB, the authors decided to investigate whether CPB affects the release of interleukin (IL)-10, as well as other cytokines, in correlation to the inhibition of T cell responses. Using phytohaemagglutinin (PHA) as mitogen and peripheral blood mononuclear cells (PBMC) isolated from patients undergoing CPB, we investigated whether this procedure has an effect on the secretion of different patterns of cytokines (Th1- and Th2-type) and PBMC proliferation. In all patients, CPB significantly enhances IL-10 and IL-6 production in resting and PHA-stimulated PBMC. On the other hand, IL-2 production, in response to PHA, was significantly diminished. Reduced IL-2 and enhanced IL-10 production were associated with a significant decrease in PBMC proliferation. Immunosuppression was also associated to lymphopenia, while neutrophil counts were significantly enhanced. These results show that after CPB there is a transient but clear unbalanced immune response demonstrated by a differentiated production of Th1- and Th2-type cytokines. The release of different patterns of cytokines observed after CPB may be helpful in understanding and preventing the development of infectious and immune complications in surgical procedure employing CPB.     

Inhibition of dopamine uptake by D2 antagonists: an in vivo study

D(2)-like antagonists potentiate dopamine release. They also inhibit dopamine uptake by a mechanism yet to be clarified. Here, we monitored dopamine uptake in the striatum of anesthetized mice. The dopamine overflow was evoked by brief electrical stimulation of the medial forebrain bundle (four pulses at 100 Hz) and was monitored with carbon fiber electrodes combined with continuous amperometry. The decay phase of evoked overflows reflects dopamine half-life, which entirely depends on uptake. The D(2)-like antagonists haloperidol and eticlopride enhanced the half-life by 45% and 48%, respectively, a moderate effect as compared to the uptake blocker nomifensine (528%). Both D(2)-like antagonists did not affect dopamine uptake in mice lacking D(2) receptors. Inhibition of tonic dopamine release by gamma-butyrolactone did not mimic the enhancing effect of D(2) antagonists on dopamine half-life. However, prolonged stimulation boosted dopamine uptake and this effect was not observed after haloperidol treatment or in mice lacking D(2) receptors. Therefore, dopamine uptake is accelerated in conditions of excessive D(2) stimulation but not finely tuned in resting conditions. Inhibition of dopamine uptake by D(2) antagonists synergizes with the potentiation of dopamine release to strongly alter the phasic dopamine signaling.     

Dihydroartemisinin-Piperaquine Treatment of Multidrug Resistant Falciparum and Vivax Malaria in Pregnancy

Background

Artemisinin combination therapy (ACT) is recommended for the treatment of multidrug resistant malaria in the second and third trimesters of pregnancy, but the experience with ACTs is limited. We review the exposure of pregnant women to the combination dihydroartemisinin-piperaquine over a 6 year period.

Methods

From April 2004–June 2009, a prospective hospital-based surveillance screened all pregnant women for malaria and documented maternal and neonatal outcomes.

Results

Data were available on 6519 pregnant women admitted to hospital; 332 (5.1%) women presented in the first trimester, 324 (5.0%) in the second, 5843 (89.6%) in the third, and in 20 women the trimester was undocumented. Peripheral parasitaemia was confirmed in 1682 women, of whom 106 (6.3%) had severe malaria. Of the 1217 women admitted with malaria in the second and third trimesters without an impending adverse outcome, those treated with DHP were more likely to be discharged with an ongoing pregnancy compared to those treated with a non-ACT regimen (Odds Ratio OR = 2.48 [1.26–4.86]); p = 0.006. However in the first trimester 63% (5/8) of women treated with oral DHP miscarried compared to 2.6% (1/38) of those receiving oral quinine; p<0.001. Of the 847 women admitted for delivery those reporting a history of malaria during their pregnancy who had been treated with quinine-based regimens rather than DHP had a higher risk of malaria at delivery (adjusted OR = 1.56 (95%CI 0.97–2.5), p = 0.068) and perinatal mortality (adjusted OR = 3.17 [95%CI: 1.17–8.60]; p = 0.023).

Conclusions

In the second and third trimesters of pregnancy, a three day course of DHP simplified antimalarial treatment and had significant benefits over quinine-based regimens in reducing recurrent malaria and poor fetal outcome. These data provide reassuring evidence for the rational design of prospective randomized clinical trials and pharmacokinetic studies.     

Assessing SSU rDNA Barcodes in Foraminifera: A Case Study using Bolivina quadrata

The Small Subunit Ribosomal RNA gene (SSU rDNA) is a widely used tool to reconstruct phylogenetic relationships among foraminiferal species. Recently, the highly variable regions of this gene have been proposed as DNA barcodes to identify foraminiferal species. However, the resolution of these barcodes has not been well established, yet. In this study, we evaluate four SSU rDNA hypervariable regions (37/f, 41/f, 43/e, and 45/e) as DNA barcodes to distinguish among species of the genus Bolivina, with particular emphasis on Bolivina quadrata for which ten new sequences ( KY468817 – KY468826 ) were obtained during this study. Our analyses show that a single SSU rDNA hypervariable sequence is insufficient to resolve all Bolivina species and that some regions (37/f and 41/f) are more useful than others (43/e and 45/e) to distinguish among closely related species. In addition, polymorphism analyses reveal a high degree of variability. In the context of barcoding studies, these results emphasize the need to assess the range of intraspecific variability of DNA barcodes prior to their application to identify foraminiferal species in environmental samples; our results also highlight the possibility that a longer SSU rDNA region might be required to distinguish among species belonging to the same taxonomic group (i.e. genus).     

Development of acute thermotolerance in L929 cells: lack of HSP28 synthesis and phosphorylation.

We investigated the correlation between the development of acute thermotolerance and the phosphorylation, synthesis, and expression of the HSP28 family in murine L929 cells. Following heating at 43 degrees C for 30 min, thermotolerance developed rapidly in exponential-phase cells and reached its maximum 4-9 h after heat shock. Maximal thermal resistance was maintained for 24 h and then gradually decayed. However, heat-induced phosphorylation of HSP28 was not detected. Furthermore, HSP28 synthesis during incubation at 37 degrees C for 12 h following heat shock was not detected by [3H]-leucine labeling followed by two-dimensional polyacrylamide gel electrophoresis. In addition, Northern blots failed to demonstrate expression of the HSP28 gene. Unlike HSP28, the expression of constitutive and inducible HSP70 genes, along with the synthesis of their proteins, was observed during incubation at 37 degrees C after heat shock. These results demonstrate that HSP28 synthesis and its phosphorylation are not required to develop acute thermotolerance in L929 cells.     

Mutations in signal transduction pathways and inherited diseases.

Intracellular signal transduction pathways have central roles in processes such as growth, differentiation, neurotransmission and development. The aberrant expression of components of various signal transduction pathways has profound consequences for cellular functions. Recent findings indicate that many cases of neoplasia and inherited diseases have, at their roots, mutations in key steps of signalling pathways.     

NKD, a developmentally regulated tachykinin receptor in Drosophila.

A number of neuropeptides have been described which are present in the insect nervous system. The physiological role of these neuropeptides has not yet been clarified. We have characterized a Drosophila melanogaster cDNA coding for a protein, NKD, whose sequence resembles that of mammalian G protein-coupled neuropeptide receptors. This protein shows 38% homology with the mammalian tachykinin NK3 receptor within the transmembrane domain region. Stable cell lines expressing this cDNA are responsive to Locusta migratoria tachykinin but not to other peptides of the tachykinin family. The expression of this gene is detected principally in adult fly heads, but also in the adult body and in embryos. Interestingly, NKD mRNA is detected at very early stages of Drosophila embryonic development (3 h) and reaches the highest level of expression at 12-16 h, a time which correlates with the period of major neuronal development. In situ hybridization experiments demonstrate that NKD is expressed in the central nervous system, as well as in subsets of neurons in each segment of the developing ventral ganglia. The cytological localization of this gene is at position 86C on the Drosophila third chromosome.     

Electroporation of extraneous proteins into CHO cells: Increased efficacy by utilizing centrifugal force and microsecond electrical pulses

A novel electroporation system employing an oscillating electric pulse and centrifugal force was used to introduce extraneous proteins into CHO cells. Following the electrical pulse, the compression and subsequent rebound induced by the centrifugal acceleration and deceleration, respectively, enhanced protein uptake, presumably by a hydrodynamic pumping of extracellular solutions through the permeabilized membrane. Protein uptake was quantitated by measuring the amount of radiolabeled, extraneous, CHO proteins introduced into unlabeled CHO cells. The amount of protein introduced into electroporated CHO cells was enhanced up to four-fold by a combination of electric pulse and centrifugal force compared to that introduced by electric pulse only. The optimum gradient of centrifugal force (GCF, temporal change of centrifugal force) was 590 and -470 g/s during acceleration and deceleration, respectively. The optimum electric field was 5 kV/cm with a 30-microsecond pulse length. At this optimum electroporation condition, approximately 5 pg of proteins (up to 200 kDa molecular weight) were introduced per CHO cell. These same settings also permitted electroporation of other membrane impermeable substances including propidium iodide and ethidium bromide. Introduction of extraneous materials into the cytoplasm during electroporation was confirmed by the ability of anti alpha-tubulin to stain the microtubules and propidium iodide and ethidium bromide to stain the nuclei. Cells electroporated with optimum device settings exhibited no significant decrease in clonogenic survival.