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D P Stites

Publications and source records attributed to D P Stites.

At least 19 recordsLinked to original sources
PubMed1992

Neutrophil function screening in patients with chronic granulomatous disease by a flow cytometric method.

Neutrophils from patients with chronic granulomatous disease (CGD) fail to produce a significant oxidative burst following stimulation. We have evaluated the use of flow cytometry and the dye 2',7'-dichlorofluorescein diacetate (DCF) for routine screening for deficiencies of neutrophil oxidative burst. A range for DCF fluorescence for phorbol myristate acetate stimulated and non-stimulated neutrophils was established based on data from 52 healthy adults. Samples from three patients with suspected neutrophil dysfunction, three patients with X-linked CGD, and one patient with autosomal recessive (AR) CGD were evaluated with both the DCF assay and the quantitative nitroblue tetrazolium dye reduction (NBT) test. For the DCF test, the ratio of mean fluorescence intensity of stimulated to non-stimulated neutrophils was less than 5 for CGD patients and from 16 to greater than 50 for healthy individuals. With the DCF test, two populations of neutrophils could be identified in samples from four carriers of X-linked CGD, although two carriers of AR CGD had NBT and DCF results in the normal range. Our data suggest the DCF test is a sensitive and convenient method for detecting CGD.

Adult↗
PubMed1992

A phase I study of HGP-30, a 30 amino acid subunit of the human immunodeficiency virus (HIV) p17 synthetic peptide analogue sub-unit vaccine in seronegative subjects.

HGP-30-KLH vaccine in alum at doses of 10, 25, 50, and 100 micrograms/kg administered intramuscularly at weeks 0, 4, and 10 appear well-tolerated clinically. Local pain at the injection site, appears to be the main clinical toxicity. Laboratory parameters are not affected by administration of the vaccine candidate except for perhaps mild urinalysis abnormalities at the highest dose. This vaccine candidate has no apparent immunotoxicity and does not appear to affect lymphocyte populations or T-cell functional studies. Low levels and transient antibodies develop in a minority of subjects early after immunization with the vaccine candidate. These responses were observed in the lowest dose range. Higher doses, and longer follow-up will be needed to confirm this observation. T-cell proliferative responses to KLH and KLH-HGP-30 are consistent and may not be dose dependent, but the proliferative responses are variable and more data need to be accumulated. Preliminary, there appears to be an HGP-30-induced CTL response of HGP-30-coated EBV-transformed autologous B cell lines. This study was approved under an IND for the California Department of Health Services' Food and Drug Branch. They have provided excellent support and regulatory guidelines for this project. Future work will extend and confirm these initial observations.

AIDS Vaccines↗
PubMed1991

Epitopes of human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins recognized by antibodies in the sera of HIV-1-infected individuals.

Sera from human immunodeficiency virus (HIV)-infected study subjects and controls were analyzed by enzyme-linked immunosorbent assay using 10 synthetic peptides to identify epitopes of HIV envelope glycoproteins (ENVgp) that were recognized by antibodies. Two epitopes of HIV ENVgp, ENVP466 (amino acids 466-481) and ENVP497 (amino acids 497-509), were recognized by antibodies in the sera of most HIV-infected individuals. The frequency of individuals with detectable serum antibodies to these two epitopes was not associated with the stage of HIV disease. Purified antibodies to ENV497 had only very weak neutralizing activity against infectious HIV. These data suggest that a particular dominant type of antibody response to HIV's ENVgp has minimal protective effects. These and other studies to identify and stimulate immune responses to selected epitopes of HIV antigens may be useful in the design of vaccines to prevent or treat HIV infections.

Acquired Immunodeficiency Syndrome↗
PubMed1991

Effects of 0.60 PPM nitrogen dioxide on circulating and bronchoalveolar lavage lymphocyte phenotypes in healthy subjects.

Nitrogen dioxide (NO2) is a common oxidant air pollutant. Animal studies have suggested that NO2 exposure causes a decrease in the numbers of some splenic lymphocyte subtypes and impairs lymphocyte-dependent immune responses. To investigate whether ambient levels of NO2 alter circulating and bronchoalveolar lavage fluid (BALF) human lymphocytes, we studied five healthy nonsmoking adult volunteers. In each subject, blood and bronchoalveolar lavage fluid was obtained and then, more than 2 weeks later, volunteers were exposured to 0.60 ppm NO2 for 2 hr with intermittent light to moderate exercise on 4 separate days within a 6-day period. We measured standard tests of pulmonary function (airway resistance, thoracic gas volume, maximal expiratory flow) and had the subjects rate the severity of respiratory symptoms before and after each NO2 exposure. Circulating and BALF lymphocytes were labeled with fluorochrome-conjugated monoclonal antibodies to human lymphocyte antigens and a flow cytometer was used to count lymphocyte subtypes. Neither any single day's exposure nor all four exposures caused a change in symptoms or in the results of tests of pulmonary function. The total number of circulating lymphocytes obtained after NO2 exposure was slightly greater than at baseline (1792 +/- 544 vs 1598 +/- 549 cells/mm3 at baseline; P = not significant) but the proportions of lymphocyte subtypes did not differ. In the BALF obtained after NO2 exposure and in the baseline state, the total number of lymphocytes and the percentages of T cells (CD 3), B cells (CD 20), T cytotoxic-suppressor cells (CD 8), T helper-inducer cells (CD 4), and large granular lymphocytes (CD 57) also did not differ after NO2 exposure. A slightly but significantly greater proportion of natural killer cells (CD 16) was found in the BALF obtained after NO2 exposure (7.2 +/- 3.1 vs 4.2 +/- 2.4% of total lymphocytes). We conclude that repeated exposures of healthy nonsmoking adults to 0.60 ppm NO2 are not associated with clinically significant symptoms, changes in airway caliber, or alterations in circulating and BALF lymphocyte subtypes. We suggest that brief, daily exposures to NO2 at levels higher than those achieved in urban atmosphere are unlikely to provoke acute respiratory impairment in healthy, nonsmoking adults.

Adult↗
PubMed1991

AUR Memorial Award 1991. Immunogenicity of gadolinium-based contrast agents for magnetic resonance imaging. Induction and characterization of antibodies in animals.

To evaluate the immunogenic potential of gadolinium-based magnetic resonance imaging (MRI) contrast agents, Sprague-Dawley rats were sensitized with gadolinium diethylenetriamine pentaacetic acid (Gd-DTPA) dimeglumine and with Gd-DTPA covalently linked to either human serum albumin, dextran, or polylysine. IgG antibodies directed against Gd-DTPA were detected in immune sera by an enzyme-linked immunosorbent assay (ELISA), and were confirmed by competitive inhibition of antibody binding using free Gd-DTPA dimeglumine. Antiserum induced by immunization with human serum albumin-(Gd-DTPA) was characterized by a monophasic competition curve with 50% inhibition (IC50) = 5.5 x 10(-4) M when Gd-DTPA dimeglumine was used as both the well-coating and the displacing agent in a competition ELISA. Antiserum induced by Gd-DTPA dimeglumine alone was characterized by a biphasic competition curve with IC50 = 6.5 x 10(-7) M and 7.9 x 10(-4) M. Antisera obtained after exposure to either dextran-(Gd-DTPA) or polylysine-(Gd-DTPA) were of insufficient titer for characterization. The detection of antibodies specific for Gd-DTPA suggests in vivo protein binding with formation of hapten-carrier conjugates. This hypothesis is supported by increased relaxivity values observed when Gd-DTPA dimeglumine is incubated in serum rather than in water. Gd-DTPA dimeglumine and albumin-(Gd-DTPA) are immunogenic in rats under idealized experimental conditions. Additional studies will be necessary to determine the potential for immunologic response in humans to gadolinium chelates under conditions of exposure inherent in clinical use.

Animals↗
PubMed1990

Human spleen and peripheral blood lymphocytes activated by interleukin-2 have similar phenotypic and functional characteristics.

Phenotypic markers and cytotoxic function were monitored in cultures of normal human mononuclear cells obtained from peripheral blood or spleen and stimulated by recombinant interleukin-2 (IL-2; 1,500 U/ml). Fresh spleen cells contained less than 5% natural killer (NK) cells (CD3-NKH1+), which increased about threefold after activation with IL-2. In both spleen and peripheral blood cultures, T cells with the NKH1 marker showed the highest relative increment among all cell types studied. Lymphokine-activated killer (LAK) cells from peripheral blood and spleen displayed very similar cytotoxic activity against K562, Daudi, and COLO carcinoma cell lines. Killing of the three targets peaked at 7 days of culture. Antibody-dependent cell cytotoxicity against a B-cell line was mediated by both circulating and splenic LAK cells from 2 to 14 days of culture. Cell sorting experiments showed that K562 targets were killed by both CD5+NKH1+ and CD5-NKH1+ cells whereas Daudi targets are only killed by CD5-NKH1+ activated NK cells from both spleen and peripheral blood. In summary, human spleen LAK cells have similar phenotypic and functional properties to circulating LAK cells, and they may be used for adoptive immunotherapy of human cancer.

Antigens, Differentiation↗
PubMed1989

CD8+ T lymphocyte control of HIV replication in cultured CD4+ cells varies among infected individuals.

Production of the human immunodeficiency virus (HIV) by cultured peripheral blood mononuclear cells (PMC) from many seropositive individuals is inhibited by the presence of CD8+ T lymphocytes. In a study of 10 subjects, high levels of virus replication could be detected in cultures of purified CD4+ cells, but not in unseparated PMC. Addition of highly purified, autologous CD8+ cells to the enriched CD4+ cells resulted in a dose-dependent inhibition of HIV growth and revealed that for some individuals, even low numbers of CD8+ cells can prevent replication of the virus. The data also indicated that culturing enriched CD4+ cells could greatly enhance detection of infectious virus in blood specimens and demonstrated that the CD4+ molecule is expressed on infected T cells isolated directly from the peripheral blood.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Development of a microsphere-based fluorescent immunoassay and its comparison to an enzyme immunoassay for the detection of antibodies to three antigen preparations from Candida albicans.

A sensitive assay for the simultaneous detection of multiple serum antibodies by flow cytometry was developed. Polystyrene microspheres of 5, 7 and 9.3 micron in diameter were used as solid supports for the attachment of three different antigen preparations from Candida albicans. These antigens were a whole cell extract; a cytoplasmic protein extract and a cell wall polysaccharide. Microsphere-associated fluorescence was quantitated by flow cytometry, with the different sized microspheres analyzed separately using electronic volume gating. This procedure allowed for different antigen-coated microspheres with discrete sizes to be analyzed independently for immunofluorescence. The assay detected antibody levels in human serum at dilutions up to 10(-6) and provided complete discrimination, using all three antigen preparations, between antibody levels seen in healthy subjects and those seen in patients suspected of having a systemic Candida infection. A standard enzyme immunoassay (EIA) failed to provide complete discrimination between healthy subjects and patient samples: at least 17% of patient values fell within the healthy subject range using all three antigen preparations. The microsphere assay which allowed for the simultaneous detection of multiple antibodies, has increased dynamic range over EIA and provides for better discrimination of patients from healthy subjects in comparison to EIA. Precise quantitation of antibodies is possible and the rapid analysis of thousands of microspheres markedly enhances the statistical accuracy of the assay. We suggest this assay is likely to have many other important applications in immunologic testing.

Antibodies, Fungal↗
PubMed1989

Immunophenotyping in a multicenter study: the Transfusion Safety Study experience.

The Transfusion Safety Study (TSS) is a cooperative investigation of factors that determine the occurrence of and modify the expression of transfusion-transmitted infections. A major component of its data is derived from lymphocyte immunophenotyping using a large panel of monoclonal antibodies and two-color flow cytometric analysis. The multicenter longitudinal character of TSS necessitates a uniformity of instrumentation, reagents, and protocols, as well as an intensive quality control program. The baseline assessment of a cohort of males 10 years of age and over with congenital clotting disorders (CCD) exemplifies the approach and some of the flow cytometry results. A comparison of anti-HIV-1 positive and negative subjects shows that more of the loss of T4+ cells was attributable to a decrease in the T4+4B4+ subset than the T4+2H4+ subset. There was an overall increase in CD8 cells, with a significant increase in the I2+T8+ and Leu7+T8+ cells, but a fall in NKH.1+T8+ cells. Monocytes, MO2+I2+ cells, increased. In CCD patients under the age of 10, both anti-HIV-1 positive and negative, there were absolute elevations in immunocytes, including CD4. There was also a distinctly different distribution of CD4 subsets. The suppressor inducer subset, 2H4+T4+, was increased relative to the helper inducer subset, 4B4+T4+, in the younger subjects.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Lymphocyte subset analysis to predict progression to AIDS in a cohort of homosexual men in San Francisco.

A group of 10 leukocyte and lymphocyte subsets were measured by simultaneous dual immunofluorescence and flow cytometry in a group of homosexual men from the San Francisco General cohort. Absolute numbers and percentages of lymphocytes were determined in 30 individuals who progressed to AIDS and 29 who did not over a 44-month period at annual intervals. At entry into the study, all subjects were asymptomatic, HIV seropositive, and had multiple changes in lymphocyte subsets compared to HIV-negative controls. In progressors, large changes occurred from the first visit to the last visit before progression in both absolute numbers and percentages of CD4 cells. The percentage of HLA-DR-bearing CD8 cells also increased. We utilized a proportional hazards model to assign a predictive value for progression to AIDS to lymphocyte subsets in both univariate and multivariate tests. Increased DR-positive CD8 cells, decreased CD4 cells, and increased CD8-positive, Leu 7-positive cells independently predicted progression to AIDS at P less than 0.006 (relative hazard 5.8-4.0). In a multivariate model, the most useful tests were either increased numbers or percentages of DR-positive CD8 cells. These data suggest parsimonious approaches to following HIV-positive individuals and further support the possibility of autoreactive T cells in the pathogenesis of HIV-associated diseases.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Lymphocyte proliferative responses to human immunodeficiency virus antigens in vitro.

All HIV seronegative (HIV Ab-) and most HIV seropositive (HIV Ab+) individuals' lymphocytes failed to proliferate in primary cultures in response to purified HIV or to recombinant envelope and core antigens of HIV, even in the presence of recombinant interleukin 2 (rIL-2). Most HIV Ab- and HIV Ab+ individuals' lymphocytes, however, could proliferate or be induced by rIL-2 to proliferate in response to lysates of Escherichia coli or Saccharomyces cerevisiae. These findings indicate selective defects in lymphocyte proliferative responses to HIV antigens before the development of AIDS in which lymphocytes are unable to proliferate in response to any antigens. These defects in cell-mediated immune responses to HIV antigens are likely to play an important role in the pathobiology of HIV infections. Although intact HIV or glycosylated gp120 envelope protein of HIV are involved in these defects, a non-glycosylated recombinant form of the HIV gp120 envelope (ENV2-3) and p25 core proteins did not inhibit antigen- or mitogen-driven lymphocyte proliferation.

AIDS-Related Complex↗
PubMed1989

Stress reduction training changed number of sexual partners but not immune function in men with HIV.

We tested the impact of stress management training on sexual behavior and immune functioning in 64 gay men infected with human immunodeficiency virus (HIV). Subjects randomized to the stress management group met for eight two-hour sessions and one all day retreat to learn systematic relaxation, health behavior change, and stress management skills. Compared to those randomized to a wait list control, treatment subjects reported significantly fewer sexual partners in the prior month at post-test (1.10 vs 2.29 for controls). There were no differences between groups in lymphocyte numbers and function.

Adult↗
PubMed1988

Simultaneous detection of antibodies to cytomegalovirus and herpes simplex virus by using flow cytometry and a microsphere-based fluorescence immunoassay.

A sensitive assay for the simultaneous detection of anti-cytomegalovirus and anti-herpes simplex virus antibodies was developed. Two different sizes of polystyrene microspheres were coated with purified viral antigens. Human antiviral antibodies were detected with a biotin-streptavidin amplification procedure with phycoerythrin as the fluorescent label. Microsphere-associated fluorescence was quantitated with a flow cytometer. Sixteen percent of samples initially scored as seronegative for cytomegalovirus and 35% of samples initially scored as seronegative for herpes simplex virus by conventional assays were clearly found positive by the microsphere technique. This flow cytometric assay can simultaneously detect several specific antibodies at levels which are below the sensitivity of standard assays. The dynamic range of this assay is at least sixfold greater than that of enzyme immunoassays. This technique is amenable to numerous serologic assays and could greatly expand the clinical laboratory applications of flow cytometry.

Antibodies, Viral↗
PubMed1988

Transcription and replication of human immunodeficiency virus-1 in B lymphocytes in vitro.

We and others have shown that HIV-1 transcription and replication in vitro are increased by the binding of transcriptional enhancer DNA sequences in the HIV-1 long terminal repeat (LTR) to a cellular protein designated Nuclear Factor-kappa B (NF-kappa B), a trans-acting transcription factor which is present in activated but not in resting T cells. Because NF-kappa B is also expressed in resting B cells we suggested that B cells might provide an optimal intracellular environment for HIV-1 transcription. Using a transient transfection assay, we show that the HIV-1 LTR is indeed more efficiently transcribed in the B cell line Raji than in the T cell line Jurkat, and that this effect maps to the transcriptional enhancer (NF-kappa B binding site) of the LTR. Furthermore, we show that Raji cells which have been rendered CD4-positive by gene transfection are susceptible to HIV-1 infection, and that viral gene expression and replication are more efficient in these CD4-positive B cells than in CD4-positive T cells. These findings demonstrate the crucial role of the transcriptional enhancer in regulating HIV-1 expression and, since receptors for HIV-1 are expressed by some Epstein-Barr virus (EBV)-positive B cells, they also suggest that HIV-1 replication could occur in EBV-positive B cells in vivo.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1987

The effects of human immunodeficiency virus recombinant envelope glycoprotein on immune cell functions in vitro.

The effect of human immunodeficiency virus (HIV) recombinant envelope glycoprotein 120 (rgp 120) on the functions of peripheral blood mononuclear cells (PBMC) in vitro was investigated. The results demonstrate that rgp 120 used at concentrations less than 1 microgram/ml has no significant effects on PBMC function in vitro. However, the addition of 1-20 micrograms/ml of rgp 120 significantly inhibits the tetanus toxoid-induced PBMC proliferative response in a dose-related manner as determined by [3H]thymidine incorporation. The data also show that rgp 120 (5 micrograms/ml) causes up to 70% reduction in the number of immunoglobulin G-secreting cells in pokeweed mitogen-stimulated PBMC cultures. Further, rgp 120 can selectively interact with the CD4a epitope of the CD4 helper cell membrane receptor. These results indicate that microgram per milliliter levels of rgp 120 can depress certain immune functions in vitro. The significance of these findings to the pathogenesis of immunodeficiency in HIV infection remains to be determined.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1986

CD8+ lymphocytes can control HIV infection in vitro by suppressing virus replication.

Lymphocytes bearing the CD8 marker were shown to suppress replication of human immunodeficiency virus (HIV) in peripheral blood mononuclear cells. The effect was dose-dependent and most apparent with autologous lymphocytes; it did not appear to be mediated by a cytotoxic response. This suppression of HIV replication could be demonstrated by the addition of CD8+ cells at the initiation of virus production as well as after several weeks of virus replication by cultured cells. The observations suggest a potential approach to therapy in which autologous CD8 lymphocytes could be administered to individuals to inhibit HIV replication and perhaps progression of disease.

Acquired Immunodeficiency Syndrome↗