B Cell Receptor Signaling-Based Index as a Biomarker for the Loss of Peripheral Immune Tolerance in Autoreactive B Cells in Rheumatoid Arthritis

This study examines the loss of peripherally induced B cell immune tolerance in Rheumatoid arthritis (RA) and establishes a novel signaling-based measure of activation in a subset of autoreactive B cells - the Induced tolerance status index (ITSI). Naturally occurring naïve autoreactive B cells can escape the “classical” tolerogenic mechanisms of clonal deletion and receptor editing, but remain peripherally tolerized through B cell receptor (BCR) signaling inhibition (postdevelopmental “receptor tuning” or anergy). ITSI is a statistical index that numerically determines the level of homology between activation patterns of BCR signaling intermediaries in B cells that are either tolerized or activated by auto antigen exposure, and thus quantifies the level of peripheral immune tolerance. The index is based on the logistic regression analysis of phosphorylation levels in a panel of BCR signaling proteins. Our results demonstrate a new approach to identifying autoreactive B cells based on their BCR signaling features.     

Endogenous and Uric Acid-Induced Activation of NLRP3 Inflammasome in Pregnant Women with Preeclampsia

Preeclampsia (PE) is a specific syndrome of pregnancy, characterized by hypertension and proteinuria. This pathology is associated with hyperuricemia and elevated serum levels of inflammatory cytokines. Uric acid crystals may activate an intracellular complex called inflammasome, which is important for processing and release of inflammatory cytokines. This study investigated the state of monocyte activation, both endogenous and stimulated with monosodium urate (MSU), by gene expression of NLRP1 and NLRP3 receptors as well as their association with inflammatory cytokines expression. Monocytes were obtained from peripheral blood of 23 preeclamptic pregnant women, 23 normotensive pregnant women (NT) and 23 healthy non-pregnant women (NP). Inflammasome activation was evaluated by the gene expression of NLRP1, NLRP3, caspase-1, IL-1β, IL-18 and TNF-α by RT-qPCR in unstimulated monocytes (endogenous expression), or after cell stimulation with MSU (stimulated expression). The concentration of cytokines was assessed by ELISA. In preeclamptic pregnant women, gene expression of NLRP1, NLRP3, caspase-1, IL-1β and TNF-α by monocytes stimulated or not with MSU was significantly higher than in NT and NP groups. Stimulation of monocytes from preeclamptic and non-pregnant women with MSU induced increased gene expression of NLRP3, caspase-1 and TNF-α in relation to the endogenous expression in these groups, while this was not observed in the NT group. The cytokine determination showed that monocytes from women with PE produced higher endogenous levels of IL-1β, IL-18 and TNF-α compared to the other groups, while the stimulus with MSU led to higher production of these cytokines in preeclamptic group than in the NT group. In conclusion, the results showed increased basal gene expression of NLRP1 and NLRP3 receptors in monocytes from PE group. These cells stimulation with MSU demonstrates that uric acid plays a role in NLRP3 inflammasome activation, suggesting the participation of this inflammatory complex in the pathogenesis of preeclampsia.     

Hypertension in response to CD4(+) T cells from reduced uterine perfusion pregnant rats is associated with activation of the endothelin-1 system

We have shown that adoptive transfer of CD4(+) T cells from placental ischemia (reduction in uteroplacental perfusion, RUPP) rats causes hypertension and elevated inflammatory cytokines during pregnancy. In this study we tested the hypothesis that adoptive transfer of RUPP CD4(+) T cells was associated with endothelin-1 activation as a mechanism to increase blood pressure during pregnancy. CD4(+) T cells from RUPP or normal pregnant (NP) rats were adoptively transferred into NP rats on gestational day 13. Mean arterial pressure (MAP) was analyzed on gestational day 19, and tissues were collected for endothelin-1 analysis. MAP increased in placental ischemic RUPP rats versus NP rats (124.1 ± 3 vs. 96.2 ± 3 mmHg; P = 0.0001) and increased in NP recipients of RUPP CD4(+) T cells (117.8 ± 2 mmHg; P = 0.001 compared with NP). Adoptive transfer of RUPP CD4(+) T cells increased placental preproendothelin-1 mRNA 2.1-fold compared with NP CD4(+) T cell rats and 1.7-fold compared with NP. Endothelin-1 secretion from endothelial cells exposed to NP rat serum was 52.2 ± 1.9 pg·mg(-1)·ml(-1), 77.5 ± 4.3 pg·mg(-1)·ml(-1) with RUPP rat serum (P = 0.0003); 47.2 ± .16 pg·mg(-1)·ml(-1) with NP+NP CD4(+) T cell serum, and 62.2 ± 2.1 pg·mg(-1)·ml(-1) with NP+RUPP CD4(+) T cell serum (P = 0.002). To test the role of endothelin-1 in RUPP CD4(+) T cell-induced hypertension, pregnant rats were treated with an endothelin A (ET(A)) receptor antagonist (ABT-627, 5 mg/kg) via drinking water. MAP was 92 ± 2 mmHg in NP+ET(A) blockade and 108 ± 3 mmHg in RUPP+ET(A) blockade; 95 ± 5 mmHg in NP+NP CD4(+) T cells+ET(A) blockade and 102 ± 2 mmHg in NP+RUPP CD4(+) T cells+ET(A) blockade. These data indicate the importance of endothelin-1 activation to cause hypertension via chronic exposure to activated CD4(+) T cells in response to placental ischemia.     

Activating autoantibodies to the angiotensin II type I receptor play an important role in mediating hypertension in response to adoptive transfer of CD4+ T lymphocytes from placental ischemic rats

Hypertension in rats with chronic placental ischemia (reduced uterine perfusion pressure, RUPP) is associated with elevated inflammatory cytokines, agonistic autoantibodies to the angiotensin II type I receptor (AT1-AA) and CD4(+) T cells; all of which are elevated in preclamptic women. Additionally, we have shown that adoptive transfer of RUPP CD4(+) T cells increases blood pressure, inflammatory cytokines, and sFlt-1. The objective of this study was to determine the long-term effects of RUPP CD4(+) T cells on AT1-AA, renal and systemic hemodynamics in pregnant rats. To answer this question CD4(+) T splenocytes were magnetically isolated on day 19 of gestation from control RUPP and normal pregnant (NP) rats and injected into a new group of NP rats at day 13 of gestation. On day 19 of gestation mean arterial pressure (MAP) and renal function (glomerular filtration rates, GFR) were analyzed and serum collected for AT1-AA analysis. To determine a role for AT1-AA to mediate RUPP CD4(+) T cell-induced blood pressure increases, MAP was analyzed in a second group of rats treated with AT1 receptor blockade losartan (10 mg·kg(-1)·day(-1)) and in a third group of rats treated with rituximab, a B cell-depleting agent (250 mg/kg) we have shown previously to decrease AT1-AA production in RUPP rats. MAP increased from 101 ± 2 mmHg NP to 126 ± 2 mmHg in RUPP rats (P < 0.001) and to 123 ± 1 mmHg in NP rats injected with RUPP CD4(+) T cells (NP+RUPP CD4(+)T cells) (P < 0.001). Furthermore, GFR decreased from 2.2 ml/min (n = 7) in NP rats to 1.0 ml/min (n = 5) NP+RUPP CD4(+)T cell. Circulating AT1-AA increased from 0.22 ± 0.1 units in NP rats to 13 ± 0.7 (P < 0.001) units in NP+RUPP CD4(+)T cell-treated rats but decreased to 8.34 ± 1 beats/min in NP+RUPP CD4(+) T cells chronically treated with rituximab. Hypertension in NP+RUPP CD4(+)T cell group was attenuated by losartan (102 ± 4 mmHg) and with B cell depletion (101 ± 5 mmHg). Therefore, we conclude that one mechanism of hypertension in response to CD4(+) T lymphocytes activated during placental ischemia is via AT1 receptor activation, potentially via AT1-AA during pregnancy.     

Cutting edge: CD40 engagement eliminates the need for Bruton's tyrosine kinase in B cell receptor signaling for NF-kappa B

The Tec kinase Bruton's tyrosine kinase (Btk) represents a key intermediary for B cell receptor (BCR) signaling. Btk mutation produces B cell deficiency in mice with X-linked immunodeficiency (xid), and surface Ig-mediated responses of mature B cells are seriously deranged. The central role that Btk plays in directing downstream events produced by BCR engagement is demonstrated by the complete failure of NF-kappa B induction and cellular proliferation following anti-Ig treatment of B cells obtained from xid mice. In this study, we report that the block in BCR signaling produced by Btk mutation is reversed by CD40 engagement. Prior treatment with CD40 ligand normalized subsequent responses of xid B cells to BCR cross-linking, so that typical outcomes of BCR signaling such as NF-kappa B activation and cell cycle progression occurred in a Btk-independent fashion. These results demonstrate that a specific genetic lesion interrupting BCR-mediated intracellular signaling is circumvented through stimulation of CD40.     

Tolerance induction of IgG+ memory B cells by T cell-independent type II antigens

Memory B cells generated during a T cell-dependent immune response rapidly respond to a secondary immunization by producing abundant IgG Abs that bind cognate Ag with high affinity. It is currently unclear whether this heightened recall response by memory B cells is due to augmented IgG-BCR signaling, which has only been demonstrated in the context of naive transgenic B cells. To address this question, we examined whether memory B cells can respond in vivo to Ags that stimulate only through BCR, namely T cell-independent type II (TI-II) Ags. In this study, we show that the TI-II Ag (4-hydroxy-3-nitrophenyl) acetyl (NP)-Ficoll cannot elicit the recall response in mice first immunized with the T cell-dependent Ag NP-chicken γ-globulin. Moreover, the NP-Ficoll challenge in vivo as well as in vitro significantly inhibits a subsequent recall response to NP-chicken γ-globulin in a B cell-intrinsic manner. This NP-Ficoll-mediated tolerance is caused by the preferential elimination of IgG(+) memory B cells binding to NP with high affinity. These data indicate that BCR cross-linking with a TI-II Ag does not activate IgG(+) memory B cells, but rather tolerizes them, identifying a terminal checkpoint of memory B cell differentiation that may prevent autoimmunity.     

Engagement of B cell receptor regulates the invariant chain-dependent MHC class II presentation pathway

The intracellular sites in which Ags delivered by the B cell receptor (BCR) are degraded and loaded onto class II molecules remain poorly defined. To address this issue, we generated wild-type and invariant chain (Ii)-deficient H-2k mice bearing BCR specific for hen egg lysozyme. Our results show that, 1) unlike Ags taken up from the fluid phase, Ii is required for presentation of hen egg lysozyme internalized through the BCR in a manner independent of the peptide analyzed; 2) BCR ligation induces intracellular accumulation of MHC class II molecules only in Ii-positive B cells; and 3) these class II molecules reach intracellular compartments where BCR targets exogenous Ag. No differences in expression of adhesion and costimulatory molecules or in the presentation of soluble peptides were detectable between Ii-positive and -negative B cells. Therefore, the BCR delivers its ligand to compartments containing MHC class II-Ii complexes and bypasses the Ii-independent presentation pathway. The linked roles of Ag internalization and B cell activation of the BCR leads to potent Ii-dependent presentation in splenic B cells.     

Autoreactive B cells discriminate CpG-rich and CpG-poor DNA and this response is modulated by IFN-alpha

Autoreactive B cells are activated by DNA, chromatin, or chromatin-containing immune complexes (ICs) through a mechanism dependent on dual engagement of the BCR and TLR9. We examined the contribution of endogenous DNA sequence elements to this process. DNA sequence can determine both recognition by the BCR and by TLR9. DNA fragments containing CpG islands, a natural source of unmethylated CpG dinucleotides, promote the activation of DNA-reactive B cells derived from BCR transgenic mice as well as DNA-reactive B cells present in the normal repertoire. ICs containing these CpG island fragments are potent ligands for AM14 IgG2a-reactive B cells. In contrast, ICs containing total mammalian DNA, or DNA fragments lacking immunostimulatory motifs, fail to induce B cell proliferation, indicating that BCR crosslinking alone is insufficient to activate low-affinity autoreactive B cells. Importantly, priming B cells with IFN-alpha lowers the BCR activation threshold and relaxes the selectivity for CpG-containing DNA. Taken together, our findings underscore the importance of endogenous CpG-containing DNAs in the TLR9-dependent activation of autoreactive B cells and further identify an important mechanism through which IFN-alpha can contribute to the pathogenesis of systemic lupus erythematosus.     

Modulation of B cell activation threshold mediated by BCR/CD40 costimulation by targeting Cbl-b for ubiquitination

It has been shown that CD40 is required for optimal B cell activation. Casitas-B-lineage lymphoma-b (Cbl-b), a RING finger E3 ubiquitin ligase, inhibits B cell activation. In this report, we demonstrate that CD40 stimulation markedly enhances IgM-induced B cell proliferation in wild-type (WT) mice, whereas this cell proliferation was reduced in CD40-deficient (Cd40^?/?) mice. Interestingly, CD40 ligation strongly augments IgM-induced Cbl-b ubiquitination and degradation in primary mouse B cells, which closely correlates with their proliferation capacity. Cbl-b deficiency uncouples BCR-induced B cell proliferation from CD40 costimulation. Our results indicate that Cbl-b negatively regulates costimulation of BCR and CD40, possibly by setting the threshold for B cell activation via controlling Cbl-b expression.     

Rosetting in Plasmodium vivax: A Cytoadhesion Phenotype Associated with Anaemia

Background

Plasmodium vivax can potentially lead to life-threatening episodes but the mechanisms underlying severe disease remain poorly defined. Cytoadhesion of infected erythrocytes may contribute to P. vivax sequestration and organ injury although its physiological impact is still unknown. Here, we aimed to describe clinically-relevant cytoadhesive phenotypes of P. vivax isolates.

Methodology/Principal findings

Rosetting and adhesion to CSA, CD36, ICAM1, placental and brain cryosections were determined in P. vivax peripheral isolates from 12 pregnant women, 24 non-pregnant women and 23 men from Manaus (Brazil). P. falciparum co-infection was excluded by PCR and P. vivax isolates were genotyped by assessing the size polymorphism of microsatellites ms2, ms20 and msp1F3 through capillary electrophoresis of PCR products. P. vivax monoinfection was confirmed by PCR in 59 isolates, with 50 (85%) of them being single-clone infections. One P. vivax haplotype was more frequently found among pregnant women (33%) than in non-pregnant women (0%) and men (4%; p = 0.010). Rosetting was observed in 64% of the isolates, adhesion to CSA in 15%, to ICAM1 in 12% and to placental cryosections in 9%, being similar among pregnant and non-pregnant groups. Intensity of rosetting was higher among anaemic individuals compared to non-anaemic (p = 0.010) and decreased with increasing haematocrit (p = 0.033) and haemoglobin levels (p = 0.015).

Conclusions/Significance

P. vivax peripheral isolates from pregnant women do not exhibit a prominent adhesion to CSA, although other parasite phenotypes still unknown may increase the propagation of certain P. vivax clones observed among pregnant hosts. Rosetting is a frequent cytoadhesive phenotype in P. vivax infections that may contribute to the development of anaemia.     

Non-invasive faecal steroid monitoring of ovarian and adrenal activity in farmed blue fox (Alopex lagopus) females during late pregnancy, parturition and lactation onset

In the semi-domesticated blue fox, handling stress may influence reproductive performance and increase perinatal pup loss. Ovarian and adrenal steroids were analysed in faecal samples collected from mid-gestation through the first week of lactation in 40 female blue foxes to characterize hormone patterns during this important reproductive period. Daily faecal samples were collected from 40 foxes during 30 pregnancies, one late abortion and nine bred-matched non-pregnancies. Mean concentrations of faecal progestagens over the 10 days before birth were significantly higher in pregnant compared to non-pregnant females (51+/-1.50 microg/g versus 36+/-3.72 microg/g, respectively; P < 0.01). From 10 to 3 days before whelping, total faecal oestrogen concentrations also were higher (P < 0.01) in pregnant (1082+/-41.69 ng/g) than non-pregnant (628+/-72.43 ng/g) foxes, before declining to non-pregnant values (402+/-24.88 ng/g) after parturition. Overall mean faecal corticoid concentrations from 3 to 20 days before whelping differed between pregnant and non-pregnant foxes (128+/-3.11 ng/g versus 103+/-5.86 ng/g, respectively; P < 0.01). Furthermore, in pregnant foxes, corticoid excretion increased further from 2 days before to 3 days after whelping (216+/-13.71 ng/g; P < 0.01). Thereafter, corticoid concentrations were similar between pregnant and non-pregnant females (P > 0.05). In sum, the faecal steroid hormone patterns for oestrogens and progestagens were similar to those previously obtained by analyses of fox serum hormones, with both steroids being higher in pregnant than non-pregnant foxes at the end of gestation. The elevation in corticoid concentrations in pregnant females suggests that adrenal activation is involved in the initiation of parturition in the blue fox. Thus, faecal steroid analyses can be used to monitor ovarian activity during pregnancy and pseudopregnancy in farmed blue fox females.     

Swimming of pregnant rats at different water temperatures

We studied the chronic effect of exercise during water immersion, associated with thermal stress (water temperature at 22, 35 and 40 degrees C) at an intensity of 80% of maximal work load supported in pregnant rats (P) and non-pregnant female rats (NP). P and NP were subdivided into three subgroups according to water temperature during exercise (P22 and NP22; P35 and NP35; P40 and NP40). The animals were submitted to daily swimming sessions of 10-15 min, for 19 days of pregnancy (P) or experimental conditions (NP). Plasma concentration of triglycerides, cholesterol, glucose, total protein, albumin and corticosterone were determined 24 h after the last exercise session. Weight gain and rectal temperature pre- and post-swimming session were also determined. The offspring were examined just after caesarian section on the 20th day of pregnancy to check weight, length and litter size. Pregnant rats showed an increase of triglycerides, reduction of glycemia, total protein and albumin and cholesterol (at 35 degrees C) when compared to non-pregnant animals. Such effects probably lead to an adequate delivery of substrate to the fetus and prepare the mother for lactation. Daily thermal stress did not modify metabolic responses to exercise in pregnant rats. Results also show a deleterious effect on offspring when the mother is exposed daily to extreme temperatures during swimming. These results suggest that water temperature (cold and hot) in swimming have to be considered to avoid damage in fetal development.     

Development of Physiologically Based Pharmacokinetic/Pharmacodynamic Model for Indomethacin Disposition in Pregnancy

Findings of a recent clinical study showed indomethacin has lower plasma levels and higher steady-state apparent clearance in pregnant subjects when compared to those in non-pregnant subjects reported in separate studies. Thus, in the current work we developed a pregnancy physiological based pharmacokinetic/pharmacodynamic (PBPK/PD) model for indomethacin to explain the differences in indomethacin pharmacokinetics between pregnancy and non-pregnancy. A whole-body PBPK model with key pregnancy-related physiological changes was developed to characterize indomethacin PK in pregnant women and compare these parameters to those in non-pregnant subjects. Data related to maternal physiological and biological changes were obtained from literature and incorporated into the structural PBPK model that describes non-pregnant PK data. Changes in indomethacin area under the curve (AUC), maximum concentration (C_max) and average steady-state concentration (C_ave) in pregnant women were predicted. Model-simulated PK profiles were in agreement with observed data. The predicted mean ratio (non-pregnant:second trimester (T₂)) of indomethacin C_ave was 1.6 compared to the observed value of 1.59. In addition, the predicted steady-state apparent clearance (CL/F_ss) ratio was almost similar to the observed value (0.46 vs. 0.42). Sensitivity analysis suggested changes in CYP2C9 activity, and to a lesser extent UGT2B7, as the primary factor contributing to differences in indomethacin disposition between pregnancy and non-pregnancy. The developed PBPK model which integrates prior physiological knowledge, in vitro and in vivo data, allowed the successful prediction of indomethacin disposition during T₂. Our PBPK/PD model suggested a higher indomethacin dosing requirement during pregnancy.     

复发性自然流产患者外周血中Th17及Treg细胞及其效应因子的变化

目的:探讨复发性自然流产患者外周血中Th17、Treg细胞以及相关细胞因子的变化。方法:选择复发性自然流产患者25例,正常妊娠妇女25例以及正常非妊娠育龄妇女25例,流式细胞术测定外周血中Th17和Treg细胞数量,ELISA法测定血清IL-10、TGF-β及IL-17的浓度。结果:复发性自然流产患者外周血中Th17细胞百分率显著高于正常妊娠以及正常非孕妇女(P0.05),Treg细胞百分率显著低于正常非孕妇女(P0.05),但与正常妊娠妇女相比无显著性差异(P0.05);复发性自然流产患者外周血IL-10及TGF-β水平显著低于正常妊娠妇女以及正常非孕妇女(P0.05),而IL-17水平显著高于正常妊娠妇女以及正常非孕妇女(P0.05)。结论:外周血Th17细胞数和IL-17水平的升高以及抑制性细胞因子IL-10和TGF-β水平的下降可能是复发性自然流产发生的重要原因。     

Augmented B Lymphocyte Response to Antigen in the Absence of Antigen-Induced B Lymphocyte Signaling in an IgG-Transgenic Mouse Line

IgG-containing B cell antigen receptor (IgG-BCR), the BCR mostly expressed on memory B cells, contains a distinct signaling function from IgM-BCR or IgD-BCR expressed on naïve B cells. Because naïve B cells transgenic for IgG exhibit augmented response to antigens similar to memory B cells, the distinct signaling function of IgG-BCR appears to play a role in augmented antibody responses of memory B cells. However, how IgG-BCR signaling augments B cell responses is not yet well understood. Here we demonstrate that B cells from IgG-transgenic mice are anergic with defect in generation of BCR signaling upon BCR ligation. However, these IgG-transgenic B cells generate markedly augmented antibody response to a T cell-dependent antigen, probably due to hyper-responsiveness to a T cell-derived signal through CD40. Both BCR signaling defect and augmented response to CD40 ligation are partially restored in xid IgG-transgenic mice in which BCR signaling is down-modulated due to a loss-of-function mutation in the tyrosine kinase Btk crucial for BCR signaling. Thus, IgG-BCR induces augmented B cell responses in the absence of antigen-induced BCR signaling probably through high ligand-independent BCR signaling that may “idle” B cells to make them ready to respond to T cell help. This finding strongly suggests a crucial role of ligand-independent signaling in receptor function.     

Asymptomatic Plasmodium falciparum Malaria in Pregnant Women in the Chittagong Hill Districts of Bangladesh

Background

Pregnancy is a known risk factor for malaria which is associated with increased maternal and infant mortality and morbidity in areas of moderate-high malaria transmission intensity where Plasmodium falciparum predominates. The nature and impact of malaria, however, is not well understood in pregnant women residing in areas of low, unstable malaria transmission where P. falciparum and P. vivax co-exist.

Methods

A large longitudinal active surveillance study of malaria was conducted in the Chittagong Hill Districts of Bangladesh. Over 32 months in 2010–2013, the period prevalence of asymptomatic P. falciparum infections was assessed by rapid diagnostic test and blood smear and compared among men, non-pregnant women and pregnant women. A subset of samples was tested for infection by PCR. Hemoglobin was assessed. Independent risk factors for malaria infection were determined using a multivariate logistic regression model.

Results

Total of 34 asymptomatic P. falciparum infections were detected by RDT/smear from 3,110 tests. The period prevalence of asymptomatic P. falciparum infection in pregnant women was 2.3%, compared to 0.5% in non-pregnant women and 0.9% in men. All RDT/smear positive samples that were tested by PCR were PCR-positive, and PCR detected additional 35 infections that were RDT/smear negative. In a multivariate logistic regression analysis, pregnant women had 5.4-fold higher odds of infection as compared to non-pregnant women. Malaria-positive pregnant women, though asymptomatic, had statistically lower hemoglobin than those without malaria or pregnancy. Asymptomatic malaria was found to be evenly distributed across space and time, in contrast to symptomatic infections which tend to cluster.

Conclusion

Pregnancy is a risk factor for asymptomatic P. falciparum infection in the Chittagong Hill Districts of Bangladesh, and pregnancy and malaria interact to heighten the effect of each on hemoglobin. The even distribution of asymptomatic malaria, without temporal and spatial clustering, may have critical implications for malaria elimination strategies.     

The dynamics of protein kinase B regulation during B cell antigen receptor engagement.

This study has used biochemistry and real time confocal imaging of green fluorescent protein (GFP)-tagged molecules in live cells to explore the dynamics of protein kinase B (PKB) regulation during B lymphocyte activation. The data show that triggering of the B cell antigen receptor (BCR) induces a transient membrane localization of PKB but a sustained activation of the enzyme; active PKB is found in the cytosol and nuclei of activated B cells. Hence, PKB has three potential sites of action in B lymphocytes; transiently after BCR triggering PKB can phosphorylate plasma membrane localized targets, whereas during the sustained B cell response to antigen, PKB acts in the nucleus and the cytosol. Membrane translocation of PKB and subsequent PKB activation are dependent on BCR activation of phosphatidylinositol 3-kinase (PI3K). Moreover, PI3K signals are both necessary and sufficient for sustained activation of PKB in B lymphocytes. However, under conditions of continuous PI3K activation or BCR triggering there is only transient recruitment of PKB to the plasma membrane, indicating that there must be a molecular mechanism to dissociate PKB from sites of PI3K activity in B cells. The inhibitory Fc receptor, the FcgammaRIIB, mediates vital homeostatic control of B cell function by recruiting an inositol 5 phosphatase SHIP into the BCR complex. Herein we show that coligation of the BCR with the inhibitory FcgammaRIIB prevents membrane targeting of PKB. The FcgammaRIIB can thus antagonize BCR signals for PKB localization and prevent BCR stimulation of PKB activity which demonstrates the mechanism for the inhibitory action of the FcgammaRIIB on the BCR/PKB response.     

Association of cytokines in hepatitis E with pregnancy outcome

AimsThe aim of this study was to evaluate tumour necrosis factor-alpha (TNF-α), interleukin (IL)-6, interferon gamma (IFN-γ) and transforming growth factor-beta1 (TGF-β1) in hepatitis E infection during pregnancy and its relation with pregnancy outcome.MethodsA total of 272 pregnant and 219 non-pregnant women with hepatitis and 262 age and gestational age matched healthy pregnant women and 208 age matched, healthy non-pregnant women were evaluated on the basis of history, clinical examination, liver function profile. Serological tests of hepatitis A, B, C and E and cytokines using commercially available (ELISA) kits. The patients with hepatitis E were further evaluated for viral load by Real Time PCR. All these were followed till delivery for pregnancy outcome.ResultsHEV viral load in acute viral hepatitis (AVH) and fulminant hepatic failure (FHF) of pregnant women were comparatively higher than non-pregnant women. Significantly higher levels of TNF-α, IL-6, IFN-γ and TGF-β1 were present in HEV infected pregnant women compared to non-pregnant women and controls. TNF-α, IL-6 and IFN-γ had significant positive correlation with viral load, serum bilirubin and prothrombin time in pregnant women. Higher levels of all four cytokines were found in pregnant women with HEV infection having adverse pregnancy outcome compared to that of pregnant women with non-HEV infection and controls.ConclusionIn conclusion, severity of HEV infection and associated adverse pregnancy outcome might be mediated by cytokine in pregnancy.     

SRC-like adaptor protein regulates B cell development and function

The avidity of BCRs and TCRs influences signal strength during processes of lymphocyte development. Avidity is determined by both the intrinsic affinity for Ag and surface levels of the Ag receptor. The Src-like adaptor protein (SLAP) is a regulator of TCR levels on thymocytes, and its deficiency alters thymocyte development. We hypothesized that SLAP, which is expressed in B cells, also is important in regulating BCR levels, signal strength, and B cell development. To test this hypothesis, we analyzed the B cell compartment in SLAP-deficient mice. We found increased splenic B cell numbers and decreased surface IgM levels on mature, splenic B cells deficient in SLAP. Immature bone marrow and splenic B cells from BCR-transgenic, SLAP-deficient mice were found to express higher surface levels of IgM. In contrast, mature splenic B cells from BCR-transgenic mice expressed decreased levels of surface BCR associated with decreased calcium flux and activation-induced markers, compared with controls. These data suggest that SLAP regulates BCR levels and signal strength during lymphocyte development.