IntroductionA profound impairment of immune functions occurs in individuals infected with human immunodeficiency virus type 1 (HIV-1). Both the cellular and the humoral arms of the immune system are unable to control the infection, which ultimately results in severe exhaustion of several lymphocyte functions and increased susceptibility to secondary and opportunistic infections. Major immunologic defects occur in the B-cell compartment. 1 Polyclonal B-cell activation is demonstrated by hypergammaglobulinemia and spontaneous antibodies' (Abs) production by cultured peripheral lymphocytes 2,3 ; additional signs of B-cell abnormality are the high incidence of B-cell tumors 4 and the deregulated expression of several surface molecules like Fas, Fas ligand (FasL), CD5, CD21, and CD27. 5-8 B-cell hyperactivity is also accompanied by functional defects since humoral immune responses following immunization are severely impaired in HIV-1-infected subjects and B lymphocytes from patients are poorly responsive to in vitro stimulation. [9][10][11] Several mechanisms may account for the B-cell abnormalities in HIV-1 infection. A direct effect of virus replication or viral proteins on B-cell function has been shown 12 and sustained by the observation that polyclonal B-cell activation is strongly reduced following effective antiretroviral treatment. 13-15 HIV-driven unbalanced production of several cytokines like tumor necrosis factor ␣ (TNF-␣), interleukin 6 (IL-6), IL-10, and IL-15 has also been involved in B-cell dysfunctions. [16][17][18] Defective T-cell help may account for B-cell unresponsiveness to T-cell-dependent antigens. 19,20 The defect of B cells in HIV-1 infection appears, however, to be intrinsic since it begins early during infection preceding functional and quantitative defects in T-helper activity and cannot be restored by allogenic normal CD4 ϩ T cells in vitro. 3,21 The mechanisms inducing hypergammaglobulinemia in HIV-1 infection are only partially known. Activation driven by CD4 ϩ T cells, monocytes, and natural killer (NK) cells through CD40-CD40 ligand (CD40L) interaction and an inappropriate cytokine supply may have a relevant role in inducing abnormal differentiation of B cells. 17,22 In addition, HIV-1 itself may directly affect B-cell activation and dysfunction, inducing the appearance of a subset of CD21 Ϫ B cells which have been proposed to contribute to increased antibody production. 23,24 A recent work by Hunziker et al 25 has suggested that naive B cells represent an important source of hypergammaglobulinemia and autoantibody production in chronic viral infections.Because of the lack of protective humoral immunity, HIV-1-infected individuals receive vaccination against several pathogens. However, many studies have reported an impaired humoral immune response in most of the patients after vaccination. [9][10][11]26,27 From the Microbiology and Tumor Biology Center, Karolinska Institutet, and the Gay Men's Health Clinic, The Soder Hospital, Stockholm, Sweden; the Swedish Institute for Infectious...

Circulating memory B cells are severely reduced in the peripheral blood of HIV-1-infected patients. We investigated whether dysfunctional serologic memory to non-HIV antigens is related to disease progression by evaluating the frequency of memory B cells, plasma IgG, plasma levels of antibodies to measles, and Streptococcus pneumoniae, and enumerating measles-specific antibody-secreting cells in patients with primary, chronic, and long-term nonprogressive HIV-1 infection. We also evaluated the in vitro production of IgM and IgG antibodies against measles and S pneumoniae antigens following polyclonal activation of peripheral blood mononuclear cells (PBMCs) from patients. The percentage of memory B cells correlated with CD4 ؉ T-cell counts in patients, thus representing a marker of disease progression. While patients with primary and chronic infection had severe defects in serologic memory, long-term nonprogressors had memory B-cell frequency and levels of antigen-specific antibodies comparable with controls. We also evaluated the effect of antiretroviral therapy on these serologic memory defects and found that antiretroviral therapy did not restore serologic memory in primary or in chronic infection. We suggest that HIV infection impairs maintenance of long-term serologic immunity to HIV-1-unrelated antigens and this defect is initi- IntroductionThe ability to maintain an intact memory B-cell compartment is an essential component of the immune response to (re-) infections. 1 Maintenance of serologic memory is carried out by plasma cells and memory B cells [1][2][3] ; memory B cells play an essential role in the maintenance of antibody (Ab) levels by rapidly generating secondary immune responses upon reinfection or antigenic stimulation. 4 One of the most deleterious effects of HIV-1 infection is B-lymphocyte hyperactivation, which manifests as hypergammaglobulinemia, increased expression of activation markers, high spontaneous Ab production in vitro, and increased incidence of B-cell lymphomas. 5 Paradoxically, HIV-1-infected persons, especially those in advanced stages of disease, also have impaired humoral immune response to vaccination, and their B cells respond poorly to in vitro stimulation by common mitogens such as SAC and PWM. 5 Earlier studies suggest that naive and memory B cells differentially contribute to B-cell dysfunctions in HIV-1 infection. [6][7][8][9][10] Circulating memory B cells in peripheral blood from patients with chronic HIV-1 infection (CHI) are severely reduced and die by apoptosis. 6,11 Serum Abs against measles, tetanus toxoid, and HIV-1 antigens are significantly reduced in patients with low memory B cells, indicating that this phenotypic alteration may severely affect memory B-cell functions. 10 Recently, we reported that during primary HIV-1 infection (PHI), memory B cells are phenotypically and functionally altered though not significantly reduced in number. 12 These alterations were only partially recovered upon antiretroviral therapy (ART), suggesting that PHI sets the stage ...

Proton pumps like the vacuolar-type H + ATPase (V-ATPase) are involved in the control of cellular pH in normal and tumor cells. Treatment with proton pump inhibitors (PPI) induces sensitization of cancer cells to chemotherapeutics via modifications of cellular pH gradients. It is also known that low pH is the most suitable condition for a full PPI activation. Here, we tested whether PPI treatment in unbuffered culture conditions could affect survival and proliferation of human B-cell tumors. First, we showed that PPI treatment increased the sensitivity to vinblastine of a pre-B acute lymphoblastic leukemia (ALL) cell line. PPI, per se, induced a dose-dependent inhibition of proliferation of tumor B cells, which was associated with a dose-and time-dependent apoptotic-like cytotoxicity in B-cell lines and leukemic cells from patients with pre-B ALL. The effect of PPI was mediated by a very early production of reactive oxygen species (ROS), that preceded alkalinization of lysosomal pH, lysosomal membrane permeabilization, and cytosol acidification, suggesting an early destabilization of the acidic vesicular compartment. Lysosomal alterations were followed by mitochondrial membrane depolarization, release of cytochrome c, chromatin condensation, and caspase activation. However, inhibition of caspase activity did not affect PPI-induced cell death, whereas specific inhibition of ROS by an antioxidant (N-acetylcysteine) significantly delayed cell death and protected both lysosomal and mitochondrial membranes. The proapoptotic activity of PPI was consistent with a clear inhibition of tumor growth following PPI treatment of B-cell lymphoma in severe combined immunodeficient mice. This study further supports the importance of acidity and pH gradients in tumor cell homeostasis and suggests new therapeutic approaches for human B-cell tumors based on PPI. [Cancer Res 2007;67(11):5408-17]

We constructed a single-chain Fv antibody library that permits human complementarity-determining region (CDR) gene fragments of any germline to be incorporated combinatorially into the appropriate positions of the variable-region frameworks VH-DP47 and VL-DPL3. A library of 2 x 109 independent transformants was screened against haptens, peptides, carbohydrates, and proteins, and the selected antibody fragments exhibited dissociation constants in the subnanomolar range. The antibody genes in this library were built on a single master framework into which diverse CDRs were allowed to recombine. These CDRs were sampled from in vivo-processed gene sequences, thus potentially optimizing the levels of correctly folded and functional molecules, and resulting in a molecule exhibiting a lower computed immunogenicity compared to naive immunoglobulins. Using the modularized assembly process to incorporate foreign sequences into an immunoglobulin scaffold, it is possible to vary as many as six CDRs at the same time, creating genetic and functional variation in antibody molecules.

The development of the human immune system is a continuous process where both accelerated and retarded development is deleterious.

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ABSTRACT. Objective. To evaluate viral vaccination immunity and booster responses in children treated successfully for acute lymphoblastic leukemia by chemotherapy and to study the response to treatment of antibody-producing plasma cells that are important for persistence of humoral immunity.Methods. Forty-three children who were in continuous first remission for a median of 5 years (range: 2-12 years) were studied. Before the leukemia was diagnosed, all children had been immunized against measles, mumps, and rubella according to the Swedish National immunization program. We analyzed levels of serum antibodies against measles and rubella by enzyme immunoassays. Avidity tests for measles antibodies were concomitantly performed by enzyme-linked immunosorbent assay for measles virus immunoglobulin G detection. The proportion of plasma cells in bone marrow was studied by flow cytometry at different times during treatment and follow-up. Children who lacked protective levels of antibodies to vaccination antigens were reimmunized. Serum was collected 3 months after immunization to assess vaccination responses.Results. After completion of the treatment, only 26 of the 43 children (60%) were found to be immune against measles and 31 (72%) against rubella. The proportion of bone marrow plasma cells decreased during treatment but returned to normal after 6 months. Revaccination caused both primary and secondary immune responses. Six of the 14 children without immunity failed to achieve protective levels of specific antibodies against measles and 3 against rubella.Conclusions. Our finding of loss of antibodies against measles and rubella in children treated with intensive chemotherapy suggests that reimmunization of these patients is necessary after completion of the treatment. To determine reimmunization schedules for children treated with chemotherapy, vaccination responses need to be studied further. A n increasing number of children survive leukemia as a result of improved and more intense chemotherapy. Other factors that influence outcome are improved supportive care including platelet transfusions, treatment with growth factors such as granulocyte colony-stimulating factor, and prophylactic antibiotic treatment. 1,2 Few studies have focused on potential long-term immunologic consequences of chemotherapy in survivors of childhood leukemia. Short-term (Ͻ2 years) effects of chemotherapy on immune function have previously been documented in children who were treated for malignancies, including acute lymphoblastic leukemia (ALL). 3,4 In those children, severe Band T-cell depletion results in clinical complications related to immune incompetence, 5,6 although the total B-and T-cell counts resolve quantitatively 6 months to 1 year after cessation of therapy. [7][8][9][10] In earlier studies, children who were treated with chemotherapy had lower levels of antibodies against common viral vaccination antigens such as measles, mumps, rubella, and polio. 11 The clinical implications, if any, of this finding are not completely und...

IntroductionHIV-1 infection is associated with extensive B-cell abnormalities, manifested by phenotypic alterations and polyclonal B-cell activation, increased frequencies of B-cell malignancies, hypergammaglobulinemia, as well as poor antigen-specific immune responses to recall and de novo antigens. [1][2][3][4][5] In secondary lymphoid tissue, HIV-1 infection induces follicular hyperplasia and alterations in the architecture of germinal center (GC) 6 and splenic marginal zones. 7 Defects in the B-cell compartment become overt already during primary HIV-1 infection 8 as measured by a decline of B-cell number, increased expression of activation, and apoptosis markers. 9 The mechanisms by which HIV-1 impairs humoral immunity may be the result of intrinsic B-cell defects and/or a lack of functional dialogue between B and T cells in secondary lymphoid organs.Lymphocyte migration and recirculation between the periphery and lymphoid tissue are critical for effective immunity and are in part regulated by chemokine receptors on lymphocytes together with the expression of their respective ligands (chemokines) in different tissue compartments. 10,11 In recent years, increasing attention has been given to the potential role of viruses to interfere with chemokine receptor expression, binding, and signaling. [12][13][14] In this respect, HIV-1 has been extensively studied because the virus uses CXC chemokine receptor 4 (CXCR4) and CC chemokine receptor 5 (CCR5) as coreceptors for entry into target cells, in addition to the main receptor, the CD4 molecule. 15 However, the expression of chemokine receptors in the context of B-cell trafficking is still poorly studied in chronic HIV-1 infection.The chemokine receptor CXCR4 is broadly expressed on a majority of B cells in the bone marrow (BM), as well as in the periphery, and plays an important role for early B-cell development 16,17 and plasma cell homing to the BM. 18,19 On the other hand, CXCR5 is expressed by mature B cells and contributes to the recruitment of naive B cells into the lymph nodes 20 where the GC reaction occurs with class switch, somatic hypermutation, and affinity maturation. The microanatomic organization of GCs into light and dark zones has been attributed to the expression of CXCR4 and CXCR5. 21,22 B cells also express a moderate amount of CCR7, which contributes to the migration within the lymph node. 23 In the present study, we examined the cell surface expression of chemokine receptors CXCR4, CXCR5, and CCR7 on B cells isolated from the blood of HIV-1-infected patients because these receptors mediate important events of B-cell homing to lymphoid tissue. 20,[23][24][25][26] Using gene expression profiling of chemokine receptors and chemokines, we found a high level of CXC chemokine ligand 13 (CXCL13) mRNA in B cells from HIV-1-infected patients compared with controls; in addition, these cells secreted a high level of the CXCL13 protein after in vitro activation. Histopathology studies performed in lymphoid tissues revealed the presence of CXCL13 ϩ B cel...