Evolving strategies for diagnosing human immunodeficiency virus infection.
Explore the source record for details and available documents.
Clinical Laboratory Techniques↗
Biomedical subjects
K A Nguyen
Publications and source records attributed to K A Nguyen.
The PPAR activator docosahexaenoic acid prevents acetaminophen hepatotoxicity in male CD-1 mice.
Acetaminophen (APAP)-induced hepatocellular necrosis can be prevented by treatment with peroxisome proliferators. This protection is associated with lowered protein arylation and glutathione depletion in mice. Peroxisome proliferators have been shown to activate nuclear receptors. These receptors, termed peroxisome proliferator activated receptors (PPARs), can also be activated by free fatty acids. This study was designed to determine if treatment with the PPAR activator docosahexaenoic acid (DHA) would also lower APAP toxicity. Male CD-1 mice received 250 mg DHA/kg or 500 mg clofibrate (CFB)/kg, i.p., for 5 d. Controls received corn oil vehicle, i.p. After overnight fasting, mice received 800 mg APAP/kg, p.o. At 24 h after APAP, hepatotoxicity was evident in control mice by elevated plasma sorbitol dehydrogenase activity (SDH) and histologic evidence of hepatic degeneration and necrosis. As expected, CFB pretreatment significantly decreased this. Similarly, DHA protected against APAP-induced hepatotoxicity at 24 h after challenge. However, treatment with DHA did not increase hepatic glutathione prior to APAP, as previously shown with CFB. Interestingly, DHA did not increase palmitoyl coenzyme A (CoA) oxidase activity or other biochemical parameters associated with peroxisome proliferation after 5 d of treatment at 250 mg/kg. No significant alterations in microsomal APAP glucuronidation or cytochrome P-450-mediated bioactivation were detected either. Collectively, these results show that DHA also prevents APAP-induced hepatotoxicity at 24 h after challenge. However, the association between resistance against APAP-induced liver injury, PPAR activation, and peroxisome proliferation is not clearly understood.
Acetaminophen↗
The 86-kD subunit of Ku autoantigen mediates homotypic and heterotypic adhesion of multiple myeloma cells.
Previous studies have shown that triggering multiple myeloma (MM) cells via CD40 induces IL-6-mediated autocrine growth as well as increased expression of cell surface adhesion molecules including CD11a, CD11b, CD11c, and CD18. In this study, we generated the 5E2 mAb which targets an antigen that is induced upon CD40 ligand (CD40L) activation of MM cells. Immunofluorescence, immunoprecipitation, and protein sequencing studies identified the target antigen of 5E2 mAb as the 86-kD subunit of the Ku autoantigen. We demonstrate that increased cell surface expression of Ku on CD40L-treated cells is due to migration of Ku from the cytoplasm to the cell surface membrane. Moreover, cell surface Ku on CD40L-treated MM cells mediates homotypic adhesion of tumor cells, as well as heterotypic adhesion of tumor cells to bone marrow stromal cells and to human fibronectin; and 5E2 mAb abrogates IL-6 secretion triggered by tumor cell adherence to bone marrow stromal cells. These data suggest that CD40L treatment induces a shift of Ku from the cytoplasm to the cell surface, and are the first to show that Ku functions as an adhesion molecule. They further suggest that cell surface Ku may play a role in both autocrine and paracrine IL-6-mediated MM cell growth and survival.
3T3 Cells↗
Self-reactive B cells in nonautoimmune and autoimmune mice.
The defining feature of autoimmune disease is the presence of specific autoreactive lymphocytes. Systemic lupus erythematosus (SLE), for example, is characterized by a discrete set of antibodies directed to nuclear antigens; these include autoantibodies to DNA and snRNPs that are diagnostic for SLE. The murine model of SLE, the MRL-lpr/lpr mouse, likewise, has a similar autoantibody profile. To understand how SLE-associated autoantibodies are regulated in healthy individuals and to identify mechanisms underlying their expression in autoimmunity, we have developed a transgenic (tg) model system using multiple sets of tgs. The development of B cells bearing these tgs has been studied in BALB/c and MRL-lpr/lpr autoimmune backgrounds, and the relative fates of anti-ssDNA and anti-dsDNA tg B cells when they are a part of a diverse as well as monoclonal B cell repertoire have been evaluated.
Animals↗
CD28/B7 costimulation: a review.
The current model of T cell activation requires two signals. The first signal is specific, requiring T cell receptor recognition and binding to MHC/Antigen presented by an antigen-presenting cell. The second signal is nonspecific, resulting from the binding of B7 ligand on the antigen-presenting cell with its receptor, CD28, on the T cell. If both signals are provided, the T cell will proliferate and secrete cytokines. Recently, it has been shown that CTLA4, another receptor for B7 that is upregulated following T cell after activation, can deliver an inhibitory signal, downregulating T cell proliferation. The B7 family of ligands has two family members, B7-1 and B7-2. They both bind to CD28 and CTLA4, but they differ in their binding affinity, structure, and temporal expression. Considerable research has been done on the CD28/B7 costimulatory pathway. Different ways of manipulating this pathway could provide insights into the mechanism and treatment of opposing pathological states. Blocking the CD28/B7 pathway could result in immunosuppression, with implications for the treatment of autoimmune diseases, organ transplantation, and graft vs. host disease. Activating the CD28/B7 pathway could be useful for including the immune system to recognize and eliminate tumors that evade the immune system. Finally, the CD28/B7 pathway could be involved with maintaining immune tolerance, as recent studies suggest the preferential binding of the B7-CTLA4 pathway results in the down-regulation of the responding T cells. Thus, the B7/CD28/CTLA4 pathway has the ability to both positively and negatively regulate immune responses.
Abatacept↗
Characterization of anti-single-stranded DNA B cells in a non-autoimmune background.
Anti-DNA Abs are prevalent in the serum of systemic lupus erythematosus (SLE) patients and in the MRL-lpr/lpr mouse model of SLE, but are generally absent in normal individuals. We have studied the regulation of anti-ssDNA B cells in a non-autoimmune (BALB/c) background by using Ig transgenes (Tgs) encoding anti-DNA Abs. In one case, they are present with other non-DNA-binding B cells (the VH3H9 Tg with endogenous light chains); in the other, they are present as an essentially monospecific population (VH3H9/Vkappa8). We have previously observed that serum anti-ssDNA levels in these Tg mice were no higher than those of non-Tg mice, despite the fact that anti-ssDNA B cells dominate the peripheral B cell repertoire. These results suggested that the anti-ssDNA Tg B cells present are functionally inactivated. In this paper, we isolate B cells from VH3H9/Vkappa8 Tg mice to show that this is indeed the case and go on to further define this state. We demonstrate that VH3H9/Vkappa8 Tg B cells have diminished Ig secretion in response to both T-independent and T-dependent stimuli compared with non-Tg controls. VH3H9/Vkappa8 Tg B cells also show suboptimal proliferation in response to anti-IgM F(ab)'2 fragments and LPS, and are phenotypically distinct in expressing decreased total surface Ig. Despite their functional defects, however, VH3H9/Vkappa8 Tg B cells have an in vivo turnover rate comparable to non-Tg B cells, suggesting that they are long lived.
Animals↗
Persistence of functionally compromised anti-double-stranded DNA B cells in the periphery of non-autoimmune mice.
Both anti-single-stranded (ss) and anti-double-stranded (ds) DNA antibodies are associated with the autoimmune disease systemic lupus erythematosus (SLE), but only anti-dsDNA antibodies are considered one of the diagnostic criteria. Using Ig transgenes coding for anti-DNA we have determined the fate of anti-dsDNA B cells in a non-autoimmune environment. In a Rag-2 wild-type background, B cells expressing the anti-dsDNA Ig transgenes are present in the spleen but dsDNA specificity is disrupted due to expression of endogenous L chains. In a Rag-2-deficient background where co-expression of endogenous Ig is blocked, splenic B cells expressing only the anti-dsDNA transgene Ig are present, indicating that endogenous Ig expression is not required for bone marrow export. The anti-dsDNA B cells that persist are profoundly crippled in that they are unable to proliferate to lipopolysaccharide or anti-Ig stimulation. Furthermore, these anti-dsDNA Ig transgene B cells show a decreased lifespan relative to non-transgene BALB/c B cells. Persistence of anti-dsDNA B cells in the periphery of non-autoimmune mice raises the possibility that their appearance in the context of SLE is due to their reactivation by T cell help.
Amino Acid Sequence↗
B cell selection and allelic exclusion of an anti-DNA Ig transgene in MRL-lpr/lpr mice.
We have used an Ig transgene (VH3H9) that increases the frequency of anti-DNA autoantibodies to address whether the production of antinuclear Abs in systemic lupus erythematosus is the consequence of a breakdown of B cell tolerance. We have shown that nonautoimmune mice regulate anti-DNA B cells, and that lupus-prone MRL-lpr/lpr mice are defective in this regulation. Here we show that a subset of anti-DNA B cells, namely those that stain nuclei in a homogeneous fashion, not only fail to be deleted in MRL-lpr/lpr mice, but undergo preferential clonal expansion. In addition, we describe a surprising finding: the VH3H9 transgene is less efficient at inhibiting endogenous heavy chain gene rearrangement on the autoimmune-prone MRL-lpr/lpr genetic background than on the nonautoimmune BALB/c background.
Amino Acid Sequence↗
Breakdown of B cell tolerance in a mouse model of systemic lupus erythematosus.
Anti-DNA antibodies, specifically those that stain nuclei in a homogenous nuclear (HN) fashion, are diagnostic of systemic lupus erythematosus (SLE) and the MRL-lpr/lpr SLE murine model. We have used a heavy chain transgene that increases the frequency of anti-HN antibodies to address whether their production in SLE is the consequence of a defect in B cell tolerance. Anti-HN B cells were undetectable in nonautoimmune-prone transgenic mice, but in MRL-lpr/lpr transgenic mice their Ig was evident in the sera and they were readily retrievable as hybridomas. We conclude that nonautoimmune animals actively delete anti-HN-specific B cells, and that MRL-lpr/lpr mice are defective in this process possibly because of the lpr defect in the fas gene.
Amino Acid Sequence↗
Fas receptor expression on B-lineage cells.
Mice homozygous for the lpr mutation have B and T cell defects and develop autoantibodies, suggesting that lpr plays a role in their genesis. The lpr defect has been identified as a mutation in the apoptosis-associated Fas receptor (FasR) gene. To begin to define the role of FasR in B cells, we have surveyed FasR expression on B-lineage cells from early progenitors in the bone marrow through their maturation in the periphery. Contrary to some reports, we found that FasR is expressed on B cells at all stages of their development and is highest on germinal center B cells. FasR is not expressed on lpr!lpr-derived cells. These data are consistent with the idea that lpr/lpr mice have an intrinsic B cell defect that may be manifested in developing as well as peripheral B cells. An unexpected finding is that B-1 (CD5) B cells do not constitutively express FasR: FasR becomes detectable on B-1 B cells only after activation.
Animals↗
Anatomy of autoantibody production: dominant localization of antibody-producing cells to T cell zones in Fas-deficient mice.
The goal of this study was to examine the in vivo site of autoantibody production in normal and autoimmune-prone mice. B cells were identified in tissue sections with IgM- and IgG2a-specific riboprobes that readily distinguished resting cells from antibody-forming cells (AFC). In normal mice, the few identifiable IgG2a-secreting cells were found in the red pulp. By contrast, in Ipr mice exceedingly high numbers of IgG2a and autoantibody-producing cells were found deep within the T cell-rich periarteriolar lymphoid sheaths (PALS). This unusual anatomic location of autoantibody-secreting B cells is unique to Fas dysregulated strains, since IgG2-producing cells in MRL/+ and (SWR x NZB)F1 mice were found predominantly in the red pulp or outer PALS, similar to normal mice. Furthermore, analysis of spleens from Ipr and non-Ipr anti-DNA immunoglobulin transgenic mice revealed dramatic accumulation of Tg+ cells in the inner PALS only in Ipr mice. These data suggest that in the absence of Fas, autoreactive B cells accumulate in T cell-rich zones, and this anatomic feature may contribute to autoantibody production.
Animals↗
Increased expression of major histocompatibility complex antigens on lymphocytes from aged mice.
Many studies have reported age-related changes in immune responses that could be due to alterations in lymphoid cell numbers or functions. Here we report the results of studies using immunofluorescent staining and in vitro assays of cellular function to compare the expression of cell surface antigens on lymphocytes from mice up to 2 years of age. No significant changes were observed in the frequencies of spleen cells bearing class I or class II major histocompatibility complex (MHC) antigens, surface immunoglobulin, or Thy-1, Ly-1, Ly-2, or L3T4 antigens. However, the densities (per cell) of both class I and class II MHC antigens were increased significantly on cells from aged as compared to young mice, whereas the densities of the other cell surface antigens studied were unchanged or slightly decreased. The increased levels of MHC antigen expression in old relative to young mice were shown to be functionally significant regarding immunological stimulation. These data suggest that T-cell clones silent in young individuals may be activated in comparable situations in older animals, leading to immunological alterations perhaps including increased autoreactivity.
Aging↗
Secretion of hydrogen peroxide by phagocytic cells from bovine non-lactating mammary glands.
Phagocytic cells from non-lactating bovine mammary glands have the capacity to secrete hydrogen peroxide when exposed to the soluble membrane stimulant phorbol myristate acetate (PMA). Unfractionated cell suspensions, containing mainly neutrophils and macrophages, and cell monolayers enriched for macrophages secreted hydrogen peroxide. A correlation was observed between the amount of hydrogen peroxide secreted, the antibacterial activity of the cells and the number of neutrophils present in the cell suspensions. Pre-exposure of cells to PMA significantly impaired their antibacterial activity against Staphylococcus aureus suggesting the importance of oxygen metabolism in the bactericidal capacity of these cells.
Animals↗
Phagocytic and bactericidal properties of bovine macrophages from non-lactating mammary glands.
Macrophages were isolated from the mammary glands of non-lactating (dry) cows and their ability to phagocytose and kill staphylococci in vitro assessed. Normal bovine serum enhanced the uptake of staphylococci and was required for optimal killing in the bactericidal test. Dry gland secretion interfered with uptake. Secretions taken progressively into the dry period became more inhibitory. The phagocytic ability of macrophages was significantly less than that of neutrophils present in the same gland preparation when tested in the presence of dry gland secretion. A marked variation in the antibacterial activity of macrophages from different cows was noted.
Animals↗