Killer B Cells: Regulation of the Growth and Function of Fas Ligand-Expressing B Lymphocytes.

The immune system is a highly-specialized cellular network that must respond to a wide variety of microbial species while remaining tolerant of an organism’s own cells and tissues. Loss of self-tolerance can lead to immune-mediated tissue destruction and disease, and therefore regulatory mechanisms have evolved that inhibit immune responses to self-antigens. The work presented in this dissertation sought to better understand the biology of B cells that express the death-inducing molecule Fas ligand (FasL), a potentially important regulatory B cell population. Conceptually, FasL+ “killer” B cells are unique amongst regulatory B cells as they possess the potential for suppression that is both antigen-specific and permanent. The experiments herein identified a novel antagonistic relationship between the type-2 cytokines interleukin-5 (IL-5) and IL-4 regarding their effects on FasL+ B cell function. Treating murine B cells with IL-5 expanded a population of B cells with potent killing activity against CD4+ T cells and that secreted the anti-inflammatory cytokine IL-10. In contrast, treatment with IL-4 inhibited both of these regulatory mechanisms in B cells. Therefore, drugs that activate pathways downstream of the IL-5 receptor or inhibit those downstream of the IL-4 receptor may lead to novel areas of drug discovery for the treatment of immune-mediated disorders. Although FasL+ B cells are rare in vivo, lymphoblastoid cell lines (LCLs) generated by transformation of human B cells with Epstein-Barr virus (EBV) showed robust expression of intracellular FasL, suggesting that the EBV latency program in transformed B cells drives the production of FasL. LCLs also secreted MHCII+FasL+ exosomes that induced antigen specific apoptosis in CD4+ T cells in two independent experimental designs. LCLs consequently represent a realistic source for immuosuppressive exosomes for therapeutic use in the treatment of human disease. This study has therefore set the groundwork for future investigations that may one day lead to powerful and novel therapeutic strategies.PHDImmunologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/102347/1/mklinker_1.pd

The Grizzly, September 25, 1981

Greaseband Tonight • Campus Welcome • Fridge Fee Unfrozen • Deutsch und Deutschland Heute: German Professor Co-authors Text • Public Speaking Exemption Exam • Books Sought by Ursinus Friends • Red Cross Bloodmobile at Helfferich Hall • Career Planning and Placement Office • Dessert Held in Union • Fast Food Service Losing Convenience • ProTheatre: Canterbury Tales Presented • Transplanted Texan: Nobody Expects the Moral Majority • School Bands Looking for Musicians • WRUC: Back on the Air? • First Coffeehouse Sparkles With Talent • Late Mail for Off-Campus Houses • [Reprinted Articles About the Greaseband] • Bear\u27s Booters Kick Off Season • Business as Usual for Cross-Country • Bears Drop 10-3 Decision to Western Maryland • Davis Leads Hockey Over Widenerhttps://digitalcommons.ursinus.edu/grizzlynews/1061/thumbnail.jp

The per-protocol effect of immediate versus deferred antiretroviral therapy initiation

The START trial found a lower risk of a composite clinical outcome in HIV-positive individuals assigned to immediate initiation of antiretroviral therapy (ART) compared with those assigned to deferred initiation. However, 30% of those assigned to deferred initiation started ART earlier than the protocol specified. To supplement the published intention-to-treat effect estimates, here we estimate the per-protocol effect of immediate versus deferred ART initiation in START

Circulating microRNAs in sera correlate with soluble biomarkers of immune activation but do not predict mortality in ART treated individuals with HIV-1 infection: A case control study

Introduction: The use of anti-retroviral therapy (ART) has dramatically reduced HIV-1 associated morbidity and mortality. However, HIV-1 infected individuals have increased rates of morbidity and mortality compared to the non-HIV-1 infected population and this appears to be related to end-organ diseases collectively referred to as Serious Non-AIDS Events (SNAEs). Circulating miRNAs are reported as promising biomarkers for a number of human disease conditions including those that constitute SNAEs. Our study sought to investigate the potential of selected miRNAs in predicting mortality in HIV-1 infected ART treated individuals. Materials and Methods: A set of miRNAs was chosen based on published associations with human disease conditions that constitute SNAEs. This case: control study compared 126 cases (individuals who died whilst on therapy), and 247 matched controls (individuals who remained alive). Cases and controls were ART treated participants of two pivotal HIV-1 trials. The relative abundance of each miRNA in serum was measured, by RTqPCR. Associations with mortality (all-cause, cardiovascular and malignancy) were assessed by logistic regression analysis. Correlations between miRNAs and CD4+ T cell count, hs-CRP, IL-6 and D-dimer were also assessed. Results: None of the selected miRNAs was associated with all-cause, cardiovascular or malignancy mortality. The levels of three miRNAs (miRs -21, -122 and -200a) correlated with IL-6 while miR-21 also correlated with D-dimer. Additionally, the abundance of miRs -31, -150 and -223, correlated with baseline CD4+ T cell count while the same three miRNAs plus miR- 145 correlated with nadir CD4+ T cell count. Discussion: No associations with mortality were found with any circulating miRNA studied. These results cast doubt onto the effectiveness of circulating miRNA as early predictors of mortality or the major underlying diseases that contribute to mortality in participants treated for HIV-1 infection

Development and Validation of a Risk Score for Chronic Kidney Disease in HIV Infection Using Prospective Cohort Data from the D:A:D Study

Ristola M. on työryhmien DAD Study Grp ; Royal Free Hosp Clin Cohort ; INSIGHT Study Grp ; SMART Study Grp ; ESPRIT Study Grp jäsen.Background Chronic kidney disease (CKD) is a major health issue for HIV-positive individuals, associated with increased morbidity and mortality. Development and implementation of a risk score model for CKD would allow comparison of the risks and benefits of adding potentially nephrotoxic antiretrovirals to a treatment regimen and would identify those at greatest risk of CKD. The aims of this study were to develop a simple, externally validated, and widely applicable long-term risk score model for CKD in HIV-positive individuals that can guide decision making in clinical practice. Methods and Findings A total of 17,954 HIV-positive individuals from the Data Collection on Adverse Events of Anti-HIV Drugs (D:A:D) study with >= 3 estimated glomerular filtration rate (eGFR) values after 1 January 2004 were included. Baseline was defined as the first eGFR > 60 ml/min/1.73 m2 after 1 January 2004; individuals with exposure to tenofovir, atazanavir, atazanavir/ritonavir, lopinavir/ritonavir, other boosted protease inhibitors before baseline were excluded. CKD was defined as confirmed (>3 mo apart) eGFR In the D:A:D study, 641 individuals developed CKD during 103,185 person-years of follow-up (PYFU; incidence 6.2/1,000 PYFU, 95% CI 5.7-6.7; median follow-up 6.1 y, range 0.3-9.1 y). Older age, intravenous drug use, hepatitis C coinfection, lower baseline eGFR, female gender, lower CD4 count nadir, hypertension, diabetes, and cardiovascular disease (CVD) predicted CKD. The adjusted incidence rate ratios of these nine categorical variables were scaled and summed to create the risk score. The median risk score at baseline was -2 (interquartile range -4 to 2). There was a 1: 393 chance of developing CKD in the next 5 y in the low risk group (risk score = 5, 505 events), respectively. Number needed to harm (NNTH) at 5 y when starting unboosted atazanavir or lopinavir/ritonavir among those with a low risk score was 1,702 (95% CI 1,166-3,367); NNTH was 202 (95% CI 159-278) and 21 (95% CI 19-23), respectively, for those with a medium and high risk score. NNTH was 739 (95% CI 506-1462), 88 (95% CI 69-121), and 9 (95% CI 8-10) for those with a low, medium, and high risk score, respectively, starting tenofovir, atazanavir/ritonavir, or another boosted protease inhibitor. The Royal Free Hospital Clinic Cohort included 2,548 individuals, of whom 94 individuals developed CKD (3.7%) during 18,376 PYFU (median follow-up 7.4 y, range 0.3-12.7 y). Of 2,013 individuals included from the SMART/ESPRIT control arms, 32 individuals developed CKD (1.6%) during 8,452 PYFU (median follow-up 4.1 y, range 0.6-8.1 y). External validation showed that the risk score predicted well in these cohorts. Limitations of this study included limited data on race and no information on proteinuria. Conclusions Both traditional and HIV-related risk factors were predictive of CKD. These factors were used to develop a risk score for CKD in HIV infection, externally validated, that has direct clinical relevance for patients and clinicians to weigh the benefits of certain antiretrovirals against the risk of CKD and to identify those at greatest risk of CKD.Peer reviewe

Human B cell-derived lymphoblastoid cell lines constitutively produce Fas ligand and secrete MHCII+FasL+ killer exosomes

Immune suppression mediated by exosomes is an emerging concept with potentially immense utility for immunotherapy in a variety of inflammatory contexts, including allogeneic transplantation. Exosomes containing the apoptosis-inducing molecule Fas ligand (FasL) have demonstrated efficacy in inhibiting antigen-specific immune responses upon adoptive transfer in animal models. We report here that a very high frequency of human B cell-derived lymphoblastoid cell lines (LCLs) constitutively produce MHCII+FasL+ exosomes that can induce apoptosis in CD4+ T cells. All LCLs tested for this study (>20 independent cell lines) showed robust expression of FasL, but had no detectable FasL on the cell surface. Given this intracellular sequestration, we hypothesized that FasL in LCLs was retained in the secretory lysosome and secreted via exosomes. Indeed, we found both MHCII and FasL proteins present in LCL-derived exosomes, and using a bead-based exosome capture assay demonstrated the presence of MHCII+FasL+ exosomes among those secreted by LCLs. Using two independent experimental approaches, we demonstrated that LCL-derived exosomes were capable of inducing antigen-specific apoptosis in autologous CD4+ T cells. These results suggest that LCL-derived exosomes may present a realistic source of immunosuppressive exosomes that could reduce or eliminate T cell-mediated responses against donor-derived antigens in transplant recipients

Interleukin-5 supports the expansion of fas ligand-expressing killer B cells that induce antigen-specific apoptosis of CD4(+) T cells and secrete interleukin-10.

Beyond their critical role in humoral immunity, B lymphocytes can employ a variety of immunomodulatory mechanisms including expression of the apoptosis-inducing molecule Fas ligand (FasL; CD178). Here, we extensively characterized the surface phenotype of FasL(+) killer B cells, showing they are enriched in the IgM(high)CD5(+)CD1d(high) B cell subset previously reported to contain a higher frequency of B cells producing interleukin-10 (IL-10). A rare population of B cells expressing IL-10 was present among FasL(+) B cells, but most FasL(+) B cells did not produce IL-10. We also identify interleukin-5 (IL-5) as a novel inducer of killer B cell function. Constitutively FasL(+) B cells expressed higher levels of the IL-5 receptor, and treating B cells with IL-5 and CD40L resulted in the expansion of a B cell population enriched for FasL(+) cells. B cells stimulated with IL-5 and CD40L were potent inducers of apoptosis in activated primary CD4(+) T cells, and this killing function was antigen-specific and dependent upon FasL. IL-5 also enhanced IL-10 secretion in B cells stimulated with CD40L. Taken together these findings elucidate the relationship of FasL(+) B cells and IL-10-producing B cells and demonstrate that IL-5 can induce or enhance both killer B cell activity and IL-10 secretion in B cells. Finally, we found that the killer B cell activity induced by IL-5 was completely blocked by IL-4, suggesting the existence of a previously unknown antagonistic relationship between these type-2 cytokines in modulating the activity of killer B cells. Targeting this IL-5/IL-4 signaling axis may therefore represent a novel area of drug discovery in inflammatory disorders

The majority of FasL⁺ splenocytes are B cells.

<p>Splenocytes from naïve mice were stained for surface expression of FasL and the cellular composition of FasL⁻ and FasL⁺ subsets was examined by flow cytometry. (<b>A</b>) Representative dot plots of splenocytes stained with a T cell marker (CD3) and three independent B cell markers (B220, CD19, and IgM). (<b>B</b>) The frequency (mean ± SEM) of cells staining positive for the markers presented in (A) among FasL⁺ splenocytes. Data are representative of more than 5 independent experiments with at least three animals per experiment.</p

IL-5 enhances IL-10 secretion from CD40L-stimulated B cells.

<p>CD19⁺ B cells were isolated by MACS from naïve mice and cultured for five days with CD40L-expressing fibroblasts in the presence or absence of IL-5. B cells were then harvested, washed in PBS, and cultured overnight at equivalent cell concentrations (7.5×10⁵ viable cells/mL) in the presence or absence of stimulation with PMA and ionomycin. IL-10 in the culture supernatants was measured by ELISA. (<b>A</b>) IL-10 secretion from B cells after overnight culture with no further stimulation. (<b>B</b>) IL-10 secretion from B cells after overnight culture with PMA and ionomycin. Data are representative of at least three independent experiments (mean ± SEM). ***<i>p</i><0.001.</p