An economic model for a novel viral influenza diagnostic.

The authors present a model that tests the economic value of a new diagnostic test that can identify type A and B influenza. Compared with traditional treatment without trying to objectively differentiate viral from bacterial infection, substantial cost savings may be achieved if diagnostic testing is appropriately utilized in a comprehensive influenza management program.

Ocular disease in children with vertically acquired human immunodeficiency virus infection.

BACKGROUND: Clinical reports suggest that ocular disease in infants and children vertically infected with human immunodeficiency virus (HIV) is different from that in adults. Pediatric patients with acquired immunodeficiency syndrome (AIDS) and HIV infection are being treated more aggressively and are living longer, but current literature on the incidence of AIDS-related ocular disease in vertically acquired HIV infection is limited. METHODS: Thirty-three children with culture-positive, vertically acquired HIV infections were prospectively followed with ophthalmic examinations between September 1991 and August 1996 at the University of Massachusetts. Patients were categorized as having symptomatic or asymptomatic HIV disease according to the U.S. Centers for Disease Control and Prevention guidelines. Absolute CD4 counts and other measures of immune function were obtained. RESULTS: The average length of follow-up was 30 months, and the average number of ophthalmic examinations per patient was 4.8. Ten patients developed ophthalmic findings. Nine of 18 (50%) patients with symptomatic AIDS disease developed ophthalmic findings. One of 15 asymptomatic HIV-infected patients developed ocular findings. Two patients with absolute CD4 counts less than 10 developed cytomegalovirus retinitis. CONCLUSIONS: These results suggest that AIDS-related ophthalmic disease is less common in vertically infected children than in adult AIDS patients. It also supports intensified clinical surveillance for cytomegalovirus retinitis in children with end-stage disease and very low CD4 counts.

Use of Epstein-Barr virus-transformed, autologous B-lymphoblastoid cells as antigen-presenting cells for establishment and maintenance of dengue virus-specific, human cytotoxic T lymphocyte clones.

We have been maintaining dengue virus specific CD8+ cytoxic T lymphocyte (CTL) clones by repeated stimulation using autologous peripheral blood mononuclear cells (PBMC) as antigen presenting cells (APCs). In the present study, Epstein-Barr virus (EBV)-transformed autologous lymphoblastoid cell lines (LCL) were compared with autologous PBMC as APCs for long term culture of a dengue virus-specific, HLA class I-restricted CD8+ CTL clone CB2.8. We substituted autologous LCL for autologous PBMC and maintained CB2.8 for several months. CB2.8 cultured using LCL as APCs maintained antigen specific cytolytic activity. No demonstrable difference in the specificity or in the level of cytolytic activity against a panel of target cells was noted between the CB2.8 maintained with LCL and those maintained with PBMC. Lysis of the target cells was blocked by the anti-HLA-class I antibody indicating that HLA class I-restriction was also maintained. We then compared autologous LCL with autologous PBMC in the establishment of CD4 + CTL clones from the PBMC of a dengue-1 immune donor. Dengue 1-specific clones were derived from limiting dilution cultures using either type of APCs. Similar numbers of dengue virus-specific CD4+ CTL clones were established using LCL or PBMC as APCs. These results indicate that autologous LCL act as APCs for long term culture of virus-specific CTL clones and represent a cost effective alternative to repeated collection of PBMC.

Modulation of the functions of dengue virus-specific human CD8+ cytotoxic T cell clone by IL-2, IL-7 and IFN gamma.

Lymphokines play an important role in immune responses to viruses by modulating functions of T lymphocytes. In the present study, we examined the effects of interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-6 (IL-6), interleukin-7 (IL-7), and interferon gamma (IFN gamma) on proliferation, cytotoxic activity and lymphokine production of a dengue virus-specific CD8+ human cytotoxic T lymphocyte (CTL) clone. IL-2 and IL-7 induced proliferation of the CD8+ CTL clone in the presence or absence of specific antigen, while IFN gamma suppressed proliferation of the clone. IL-7 and IFN gamma augmented dengue virus-specific cytotoxic activity without inducing non-specific cytotoxic activity, and IL-2 induced non-specific cytotoxic activity. IL-2 induced IFN gamma production by the CD8+ CTL clone. IL-4 and IL-6 did not modulate the functions of the CD8+ CTL clone in these experimental conditions. These results suggest that functions of dengue virus-specific CD8+ CTL are modulated by IL-2, IL-7 and IFN gamma, and that IL-7 is a lymphokine useful to induce growth and to maintain specific cytotoxic activity of CD8+ CTL clones in vitro.

Dengue virus-specific, HLA-B35-restricted, human CD8+ cytotoxic T lymphocyte (CTL) clones. Recognition of NS3 amino acids 500 to 508 by CTL clones of two different serotype specificities.

Dengue virus infections are a major cause of morbidity and mortality in tropical and subtropical areas of the world. We analyzed dengue virus-specific CD8+ CD4- CTL at the clonal level to further understand the role of CD8+ CTL in dengue virus infections. Dengue virus-specific CD8+ CTL clones were established from lymphocytes of a dengue 4-immune adult. Three patterns of dengue serotype specificities were identified: 1) specific for dengue 4, 2) cross-reactive for dengue 2 and dengue 4 (subcomplex-specific); and 3) cross-reactive for all four dengue virus serotypes. Three dengue 4-specific clones and one dengue 2/dengue 4 cross-reactive clone were further analyzed. All four of the clones were HLA-B35 restricted and recognized NS3. The epitopes were mapped to amino acids (aa) 483 to 618 of NS3. The epitope was then defined by using synthetic peptides. Three dengue 4-specific clones and one dengue 2/dengue 4 cross-reactive clone recognized the same peptide (TPEGIIPTL) encompassing aa 500 to 508 of dengue 4 NS3. The peptide encompassing aa 500-508 of dengue 2 NS3 was recognized by a dengue 2/dengue 4 cross-reactive clone but was not recognized by the dengue 4-specific clones. Dengue 4-specific and dengue 2/dengue 4 cross-reactive clones used different TCR. These results indicate that CD8+ CTL clones that use different TCR and demonstrate two distinct serotype specificities recognize the same 9-mer peptide in the context of HLA-B35.

Recognition of envelope protein by dengue virus serotype-specific human CD4+ CD8- cytotoxic T-cell clones.

We analyzed dengue virus-specific CD4+ CD8- cytotoxic T lymphocytes (CTL) at the clonal level to further understand their role in dengue virus infections. Stimulation of peripheral blood mononuclear cells from two dengue virus type 4 (D4V)-immune donors with live D4V or noninfectious D4V antigen generated 17 HLA class II-restricted CD4+ CTL capable of specific lysis of dengue virus antigen-treated autologous lymphoblastoid cell lines. Thirteen clones were D4V specific, three clones were cross-reactive for D2V and D4V, and one clone was cross-reactive for D1V, D3V, and D4V. Antigen recognition by six D4V-specific clones and three D2V- and D4V-cross-reactive clones was restricted by HLA-DR7. Five D4V-specific CD4+ CTL clones lysed autologous lymphoblastoid cell lines infected with a dengue virus-vaccinia virus recombinant containing the E gene of D4V, whereas three serotype-cross-reactive CTL clones did not. These results indicate that E-specific clones are serotype specific and HLA-DR7 restricted in these two donors and suggest that a common epitope on E protein may be recognized. E protein-specific CD4+ CTL may be important mediators of virus clearance especially during reinfection with the same serotype as that in primary infection by providing help for virus-specific antibody production and lysis of virus-infected cells.

Immunopathologic mechanisms of dengue hemorrhagic fever and dengue shock syndrome.

Dengue virus infections are a major cause of morbidity and mortality in tropical and subtropical areas of the world. The immunopathological mechanisms that result in severe complications of dengue virus infection, i.e. dengue hemorrhagic fever (DHF), are important to determine. Primary dengue virus infections induce serotype-specific and serotype-cross-reactive, CD4+ and CD8+ memory cytotoxic T lymphocytes (CTL). In secondary infections with a virus of a different serotype from that which caused primary infections, the presence of cross-reactive non-neutralizing antibodies results in an increased number of infected monocytes by dengue virus--antibody complexes. This in turn results in marked activation of serotype cross-reactive CD4+ and CD8+ memory CTL. We hypothesize that the rapid release of cytokines and chemical mediators caused by T cell activation and by CTL-mediated lysis of dengue virus-infected monocytes triggers the plasma leakage and hemorrhage that occurs in DHF.

Definition of an HLA-DPw2-restricted epitope on NS3, recognized by a dengue virus serotype-cross-reactive human CD4+ CD8- cytotoxic T-cell clone.

We previously reported that the clone JK34 was cross-reactive for dengue virus types 1, 2, 3, and 4 and recognized NS3 (I. Kurane, M. A. Brinton, A. L. Samson, and F. A. Ennis, J. Virol. 65:1823-1828, 1991). In the present experiments, we defined the epitope at the amino acid level, with 93 15-mer overlapping peptides which cover the entire NS3. A peptide 4 which contains amino acids 251 to 265 of NS3 sensitized the autologous B lymphoblastoid cell line (LCL) to the lysis by JK34. The smallest peptide recognized by JK34 was a 10-mer peptide which contains amino acids 255 to 264 (EIVDLMCHAT). A monoclonal antibody to HLA-DP inhibited the lysis of epitope peptide-pulsed autologous LCL by JK34. Genotypic typing revealed that the HLA-DP of this donor is DPA1*01, DPB1*0201, which is serologically defined as HLA-DPw2. JK34 lysed peptide 4-pulsed allogeneic LCL which carried HLA-DPw2. These results indicate that HLA-DPw2 is the restriction allele for recognition of this epitope by JK34.

Thyroid growth immunoglobulins in feline hyperthyroidism.

Feline hyperthyroidism bears a strong clinical and pathologic resemblance to toxic nodular goiter in humans. To evaluate whether the observed thyroid growth might be due to circulating thyroid antibodies, as has been postulated in humans, we studied the effect of purified immunoglobulin (Ig) G preparations on a rat thyroid follicular (FRTL-5) cell line. When compared with control, hyperthyroid cat IgG caused significantly increased [3H]-thymidine (Tdr) incorporation into DNA (p less than 0.02) and stimulated cellular proliferation 15-fold. Stimulation of 3H-Tdr incorporation tended to be biphasic and could be inhibited completely by a potent, specific TSH receptor blocking antibody. Hyperthyroid cat IgG also significantly inhibited 125I-bTSH binding to porcine thyroid membranes, an effect that could be reproduced using electrophoretically pure IgG and normal cat thyroid membranes. Unlike its effect on growth, hyperthyroid cat IgG did not stimulate intracellular cAMP, and there was no correlation between thyroid function in vivo and IgG growth-promoting activity in vitro. These data suggest that elevated titers of thyroid growth IgGs, probably acting through the TSH receptor, are present in feline hyperthyroidism and may play a role in goiter formation. Unlike growth, the thyroid hyperfunction observed is not IgG dependent. Further study of feline hyperthyroidism may contribute important insights into human nodular goiter and into the mediation of thyroid growth in general.

Human immune responses to dengue viruses.

Dengue fever (DF) and dengue hemorrhagic fever (DHF)/dengue shock syndrome (DSS) are major public health problems in many areas of the world. We are analyzing the human immune responses to dengue viruses, in order to understand the mechanism of recovery from dengue virus infections and the pathogenesis of DHF/DSS. Human natural killer (NK) cells lyse dengue virus-infected cells to a greater degree than uninfected cells. Antibodies to dengue viruses augment the lysis of dengue virus-infected cells by NK cells. Dengue virus-infected monocytes produce high levels of interferon alpha (IFN alpha). DR+ lymphocytes also produce high levels of IFN alpha after contact with dengue virus-infected monocytes. The IFN alpha produced protects uninfected monocytes from dengue virus infection. These results suggest that NK cells and IFN alpha may play an important role in controlling primary dengue virus infection. Dengue virus-specific CD4+CD8(-)T lymphocytes and CD4(-)CD8+T lymphocytes are present in the peripheral blood mononuclear cell population from donors who were infected with dengue virus. Most of CD4+T lymphocytes are dengue serotype-crossreactive. They lyse dengue virus-infected autologous cells in an HLA class II-restricted fashion, and produce interferon gamma (IFN gamma). IFN gamma augments dengue virus infection of monocytic cells in the presence of antidengue virus antibodies by increasing the number of Fc gamma receptors. Dengue virus-specific CD8+T lymphocytes lyse dengue virus-infected autologous cells in an HLA class I-restricted fashion. These CD8+T lymphocytes are also dengue serotype-crossreactive.(ABSTRACT TRUNCATED AT 250 WORDS)