(1/892) Skeletal muscle type ryanodine receptor is involved in calcium signaling in human B lymphocytes.
The regulation of intracellular free Ca2+ concentration ([Ca2+]i) in B cells remains poorly understood and is presently explained almost solely by inositol 1,4,5-triphosphate (IP3)-mediated Ca2+ release, followed by activation of a store-operated channel mechanism. In fact, there are reports indicating that IP3 production does not always correlate with the magnitude of Ca2+ release. We demonstrate here that human B cells express a ryanodine receptor (RYR) that functions as a Ca2+ release channel during the B cell antigen receptor (BCR)-stimulated Ca2+ signaling process. Immunoblotting studies showed that both human primary CD19(+) B and DAKIKI cells express a 565-kDa immunoreactive protein that is indistinguishable in molecular size and immunoreactivity from the RYR. Selective reverse transcription-polymerase chain reaction, restriction fragment length polymorphism, and sequencing of cloned cDNA indicated that the major isoform of the RYR expressed in primary CD19(+) B and DAKIKI cells is identical to the skeletal muscle type (RYR1). Saturation analysis of [3H]ryanodine binding yielded Bmax = 150 fmol/mg of protein and Kd = 110 nM in DAKIKI cells. In fluo-3-loaded CD19(+) B and DAKIKI cells, 4-chloro-m-cresol, a potent activator of Ca2+ release mediated by the ryanodine-sensitive Ca2+ release channel, induced Ca2+ release in a dose-dependent and ryanodine-sensitive fashion. Furthermore, BCR-mediated Ca2+ release in CD19(+) B cells was significantly altered by 4-chloro-m-cresol and ryanodine. These results indicate that RYR1 functions as a Ca2+ release channel during BCR-stimulated Ca2+ signaling and suggest that complex Ca2+ signals that control the cellular activities of B cells may be generated by cooperation of the IP3 receptor and RYR1. (+info)
(2/892) Increased expression of regeneration and tolerance factor in individuals with human immunodeficiency virus infection.
Regeneration and tolerance factor (RTF) plays a pivotal role in successful pregnancy outcome and has potent immunomodulating properties. During pregnancy, it is abundantly expressed in the placenta and on peripheral B lymphocytes. Several lines of evidence suggest that both successful pregnancy outcome and progression from human immunodeficiency virus (HIV) infection to AIDS are associated with a Th2-type response. As a result, we hypothesized that the cellular expression of RTF may also be increased during infection with HIV. Using flow cytometric analysis, we showed a significantly (P < 0.01) increased expression of RTF on CD3(+) cells obtained from individuals with HIV over that for individuals without HIV. On average, 32.1% of the CD3(+) cells from individuals with HIV expressed high levels of RTF. In contrast, an average of only 6.7% of the CD3(+) cells from individuals without HIV expressed high levels of RTF. Similar results were obtained when CD19(+) cells from individuals with (mean, 44.1%) and without (mean, 25.8%) HIV were evaluated. Linear regression analysis suggested that high levels of RTF expression by CD3(+) cells correlated better with viral load (r value, 0.46) than with absolute CD4 count (r value, 0.09). While additional experiments are necessary to delineate the precise immunologic role of RTF, our current data suggest that RTF expression during HIV infection may be a useful marker of immune activation. (+info)
(3/892) Predictive value of CD19 measurements for bacterial infections in children infected with human immunodeficiency virus.
We investigated the predictive value of CD19 cell percentages (CD19%) for times to bacterial infections, using data from six pediatric AIDS Clinical Trials Group protocols and adjusting for other potentially prognostic variables, such as CD4%, CD8%, immunoglobulin (IgA) level, lymphocyte count, prior infections, prior zidovudine treatment, and age. In addition, we explored the combined effects of CD19% and IgG level in predicting time to infection. We found that a low CD19% is associated with a nonsignificant 1.2-fold increase in hazard of bacterial infection (95% confidence interval: 0.97, 1.49). In contrast, a high IgG level is associated with a nonsignificant 0.87-fold decrease in hazard of infection (95% confidence interval: 0.68, 1.12). CD4% was more prognostic of time to bacterial infection than CD19% or IgG level. Low CD19% and high IgG levels together lead to a significant (P < 0. 01) 0.50-fold decrease in hazard (95% confidence interval: 0.35, 0. 73) relative to low CD19% and low IgG levels. Similarly, in a model involving assay result changes (from baseline to 6 months) as well as baseline values, the effect of CD19% by itself is reversed from its effect in conjunction with IgG. In this model, CD19% that are increasing and high are associated with decreases in hazard of infection (P < 0.01), while increasing CD19% and increasing IgG levels are associated with significant (at the P = 0.01 level) fourfold increases in hazard of infection relative to stable CD19% and decreasing, stable, or increasing IgG levels. Our data suggest that CD19%, in conjunction with IgG level, provides a useful prognostic tool for bacterial infections. It is highly likely that T-helper function impacts on B-cell function; thus, inclusion of CD4% in such analyses may greatly enhance the assessment of risk for bacterial infection. (+info)
(4/892) Long-term fetal microchimerism in peripheral blood mononuclear cell subsets in healthy women and women with scleroderma.
Fetal CD34(+) CD38(+) cells have recently been found to persist in maternal peripheral blood for many years after pregnancy. CD34(+) CD38(+) cells are progenitor cells that can differentiate into mature immune-competent cells. We asked whether long-term fetal microchimerism occurs in T lymphocyte, B lymphocyte, monocyte, and natural-killer cell populations of previously pregnant women. We targeted women with sons and used polymerase chain reaction for a Y-chromosome-specific sequence to test DNA extracted from peripheral blood mononuclear cells (PBMC) and from CD3, CD19, CD14, and CD56/16 sorted subsets. We also asked whether persistent microchimerism might contribute to subsequent autoimmune disease in the mother and included women with the autoimmune disease scleroderma. Scleroderma has a peak incidence in women after childbearing years and has clinical similarities to chronic graft-versus-host disease that occurs after allogeneic hematopoietic stem-cell transplantation, known to involve chimerism. Sixty-eight parous women were studied for male DNA in PBMC and 20 for PBMC subsets. Microchimerism was found in PBMC from 33% (16 of 48) of healthy women and 60% (12 of 20) women with scleroderma, P =.046. Microchimerism was found in some women in CD3, CD19, CD14, and CD56/16 subsets including up to 38 years after pregnancy. Microchimerism in PBMC subsets was not appreciably more frequent in scleroderma patients than in healthy controls. Overall, microchimerism was found in CD3, CD19, and CD14 subsets in approximately one third of women and in CD56/16 in one half of women. HLA typing of mothers and sons indicated that HLA compatibility was not a requirement for persistent microchimerism in PBMC subsets. Fetal microchimerism in the face of HLA disparity implies that specific maternal immunoregulatory pathways exist that permit persistence but prevent effector function of these cells in normal women. Although microchimerism in PBMC was more frequent in women with scleroderma than healthy controls additional studies will be necessary to determine whether microchimerism plays a role in the pathogenesis of this or other autoimmune diseases. (+info)
(5/892) Cutting edge: recruitment of the CD19/CD21 coreceptor to B cell antigen receptor is required for antigen-mediated expression of Bcl-2 by resting and cycling hen egg lysozyme transgenic B cells.
Recruitment of the CD19/CD21 coreceptor is thought to lower the threshold for effective signaling through the B cell Ag receptor. We provide evidence supporting a second role for coreceptor recruitment, and that is to enhance the survival/proliferative potential of the responding B cells. We show that B cell Ag receptor signaling in the absence of coreceptor recruitment induces cellular accumulation of the anti-apoptotic protein Bcl-xL, whereas CD19-mediated signals are required for Bcl-2 accumulation. The expression of both anti-apoptotic proteins correlates with the enhanced responsiveness of both resting and cycling B cells to growth-promoting signals delivered through CD40. These results provide further evidence for the necessity of coreceptor recruitment during Ag-dependent B cell activation and indicate that Ags derived from inflammatory sites function as better thymus-dependent Ags than their counterparts not coated with complement fragments. (+info)
(6/892) Phosphorylation of CD19 Y484 and Y515, and linked activation of phosphatidylinositol 3-kinase, are required for B cell antigen receptor-mediated activation of Bruton's tyrosine kinase.
Bruton's tyrosine kinase (Btk) plays a critical role in B cell Ag receptor (BCR) signaling, as indicated by the X-linked immunodeficiency and X-linked agammaglobulinemia phenotypes of mice and men that express mutant forms of the kinase. Although Btk activity can be regulated by Src-family and Syk tyrosine kinases, and perhaps by phosphatidylinositol 3,4,5-trisphosphate, BCR-coupled signaling pathways leading to Btk activation are poorly understood. In view of previous findings that CD19 is involved in BCR-mediated phosphatidylinositol 3-kinase (PI3-K) activation, we assessed its role in Btk activation. Using a CD19 reconstituted myeloma model and CD19 gene-ablated animals we found that BCR-mediated Btk activation and phosphorylation are dependent on the expression of CD19, while BCR-mediated activation of Lyn and Syk is not. Wortmannin preincubation inhibited the BCR-mediated activation and phosphorylation of Btk. Btk activation was not rescued in the myeloma by expression of a CD19 mutant in which tyrosine residues previously shown to mediate CD19 interaction with PI3-K, Y484 and Y515, were changed to phenylalanine. Taken together, the data presented indicate that BCR aggregation-driven CD19 phosphorylation functions to promote Btk activation via recruitment and activation of PI3-K. Resultant phosphatidylinositol 3,4,5-trisphosphate probably functions to localize Btk for subsequent phosphorylation and activation by Src and Syk family kinases. (+info)
(7/892) Expression of Epstein-Barr virus BamHI-A rightward transcripts in latently infected B cells from peripheral blood.
In addition to the Epstein-Barr virus (EBV) EBNA and LMP latency genes, there is a family of alternatively spliced BamHI-A rightward transcripts (BARTs). These latency transcripts are highly expressed in the EBV-associated malignancies nasopharyngeal carcinoma and Burkitt's lymphoma, and are expressed at lower levels in latently EBV-infected B-cell lines. The contribution of the BARTs to EBV biology or pathogenesis is unknown. Resting B cells have recently been recognized as a reservoir for EBV persistence in the peripheral blood. In these cells, EBV gene expression is tightly restricted and the only viral gene known to be consistently expressed is LMP2A. We used cell sorting and reverse-transcriptase polymerase chain reaction (RT-PCR) to examine whether BARTs are expressed in the restricted form of in vivo latency. Our results demonstrated that RNAs with splicing diagnostic for transcripts containing the BART RPMS1 and BARFO open-reading frames (ORFs) were expressed in CD19(+) but not in CD23(+) B cells isolated from peripheral blood of healthy individuals. The product of the proximal RPMS1 ORF has not previously been characterized. The RPMS1 ORF was shown to encode a 15-kD protein that localized to the nucleus of transfected cells. Expression of the BARTs in peripheral blood B cells suggests that the proteins encoded by these transcripts are likely to be important for maintenance of in vivo latency. (+info)
(8/892) Distribution of lymphocytes and adhesion molecules in human cervix and vagina.
Knowledge of the histological distribution of leucocytes and adhesion molecules in the human genital tract is scarce although local immunity in this region is important. Using immunohistochemical methods, we here describe the organization of CD3+, CD8+ and CD4+ T cells, CD19+ B cells, CD38+ plasma cells, major histocompatibility complex (MHC) class II+ antigen-presenting cells and CD14+ monocytes, as well as the expression of endothelial addressins in normal human ecto-cervical and vaginal mucosa. T cells were clustered in a distinct band beneath the epithelium and were also dispersed in the epithelium and the lamina propria, whereas CD38+ plasma cells were present only in the lamina propria. MHC class II+ cells were numerous in the lamina propria and in the epithelium, where they morphologically resembled dendritic cells. Lymphoid aggregates containing CD19+ and CD20+ B cells as well as CD3+, CD4+ and CD8+ cells were also found in the cervix. The mucosal addressin cell adhesion molecule-1 (MAdCAM-1) was not expressed on the vascular endothelium in the cervical or vaginal mucosa. In contrast, intercellular adhesion molecule-1 (ICAM-1), vascular adhesion protein-1 (VAP-1) and P-selectin were expressed in all tissue samples, and vascular cell adhesion molecule-1 (VCAM-1) and E-selectin were found in four of seven samples. We conclude that the distribution of leucocytes and adhesion molecules is very similar in the ecto-cervical and the vaginal mucosa and that the regulation of lymphocyte homing to the genital tract is different from that seen in the intestine. Our results also clearly suggest that the leucocytes are not randomly scattered in the tissue but organized in a distinct pattern. (+info)