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Alan L Rothman

Publications and source records attributed to Alan L Rothman.

At least 37 records · Page 2Linked to original sources
PubMed2004

Identification and analysis for cross-reactivity among hantaviruses of H-2b-restricted cytotoxic T-lymphocyte epitopes in Sin Nombre virus nucleocapsid protein.

Sin Nombre virus (SNV) causes hantavirus pulmonary syndrome (HPS), with a high rate of mortality in humans who are infected by the transmission of virus from the natural rodent host. In humans, cytotoxic T lymphocytes (CTL) specific for SNV appear to play an important role in the pathogenicity of HPS. There is a correlation between the frequencies of SNV-specific CTLs and the severity of HPS disease. In order to create a mouse model to study the role of SNV-specific T cells in vivo, T cell responses to SNV nucleocapsid (N) protein in B6.PL Thy1(a)/Cy mice (H-2(b)) immunized with plasmid DNA or recombinant vaccinia virus expressing SNV N protein were examined. Four peptides, NC94-101, NC175-189, NC217-231 and NC331-345, were recognized by CD8(+) T cells in CTL and ELISPOT assays in SNV N-immunized mice. Interestingly, two of these epitopes are located in the central region of the SNV N protein, where several human CD8(+) T-cell epitopes have been defined in Puumala virus and SNV. CTL lines specific for these four epitopes were cross-reactive to corresponding Puumala virus peptides, but only one of them was cross-reactive to Hantaan virus peptides. These results will enable the analysis of the roles of CTL in immunopathology of HPS in experimental mouse models of HPS.

Amino Acid Sequence↗
PubMed2004

Dengue: defining protective versus pathologic immunity.

Dengue is an expanding public health problem, and an effective vaccine remains elusive. This review discusses how the significant influence of sequential infection with different dengue virus serotypes on the severity of disease can be viewed in terms of beneficial and detrimental effects of heterologous immunity. A more complete understanding of these effects is likely to be critical for predicting optimal vaccine-induced immune responses.

Dengue↗
PubMed2003

A blunted blood plasmacytoid dendritic cell response to an acute systemic viral infection is associated with increased disease severity.

At least two distinct human dendritic cell (DC) subsets are produced in the bone marrow and circulate in the peripheral blood-precursor myeloid DCs (pre-mDCs) and plasmacytoid DCs (PDCs). Both lineages of DCs are instrumental in antiviral innate immunity and shaping Th1 adaptive immune responses. PDCs are the most potent IFN-alpha-producing cells to viral pathogens. Dengue, an acute flavivirus disease, provides a model to study DC responses to a self-limited human viral infection. We analyzed circulating DC subsets in a prospective study of children with dengue across a broad range of illness severities: healthy controls; mild, nondengue, presumed viral infections; moderately ill dengue fever; and, the most severe form of illness, dengue hemorrhagic fever. We also examined PDC responses in monkeys with asymptomatic dengue viremia and to dengue virus exposure in vitro. The absolute number and frequency of circulating pre-mDCs early in acute viral illness decreased as illness severity increased. Depressed pre-mDC blood levels appeared to be part of the typical innate immune response to acute viral infection. The frequency of circulating PDCs trended upward and the absolute number of circulating PDCs remained stable early in moderately ill children with dengue fever, mild other, nondengue, febrile illness, and monkeys with asymptomatic dengue viremia. However, there was an early decrease in circulating PDC levels in children who subsequently developed dengue hemorrhagic fever. A blunted blood PDC response to dengue virus infection was associated with higher viremia levels, and was part of an altered innate immune response and pathogenetic cascade leading to severe disease.

Acute Disease↗
PubMed2003

Quantitation of CD8+ T cell responses to newly identified HLA-A*0201-restricted T cell epitopes conserved among vaccinia and variola (smallpox) viruses.

Immunization with vaccinia virus resulted in long-lasting protection against smallpox and was the approach used to eliminate natural smallpox infections worldwide. Due to the concern about the potential use of smallpox virus as a bioweapon, smallpox vaccination is currently being reintroduced. Severe complications from vaccination were associated with congenital or acquired T cell deficiencies, but not with congenital agammaglobulinemia, suggesting the importance of T cell immunity in recovery from infection. In this report, we identified two CD8+ T cell epitopes restricted by the most common human major histocompatibility complex (MHC) class I allele, HLA-A*0201. Both epitopes are highly conserved in vaccinia and variola viruses. The frequency of vaccinia-specific CD8+ T cell responses to these epitopes measured by interferon (IFN)-gamma enzyme-linked immunospot (ELISPOT) assay and HLA/peptide tetramer staining peaked 2 wk after primary immunization and then declined, but were still detectable 1 to 3 yr after primary immunization. 2 wk after immunization, IFN-gamma-producing cells specific to these two epitopes were 14% of total vaccinia virus-specific IFN-gamma-producing cells in one donor, 35% in the second donor, and 6% in the third donor. This information will be useful for studies of human T cell memory and for the design and analyses of the immunogenicity of experimental vaccinia vaccines.

CD8-Positive T-Lymphocytes↗
PubMed2003

Immunology and immunopathogenesis of dengue disease.

The pathophysiological basis of severe dengue disease (i.e., dengue hemorrhagic fever [DHF]), appears to be multifactorial, involving complex interactions among viral factors, host genetics, and the immunologic background of the host, principally prior exposure to dengue virus. Analysis of these processes has been limited to observational studies of naturally infected humans because there have not been useful animal models of dengue disease. Substantial evidence points to dengue virus-reactive T cells as a critical effector in the development of DHF. We are beginning to define the critical elements of T-cell epitope specificity and functional responses that contribute to DHF. Additional studies in well-characterized patient cohorts from different geographic regions will be needed to advance this research and guide new approaches to prevention and treatment of DHF.

Animals↗
PubMed2003

Dengue virus induces novel changes in gene expression of human umbilical vein endothelial cells.

Endothelial cells are permissive to dengue virus (DV) infection in vitro, although their importance as targets of DV infection in vivo remains a subject of debate. To analyze the virus-host interaction, we studied the effect of DV infection on gene expression in human umbilical vein endothelial cells (HUVECs) by using differential display reverse transcription-PCR (DD-RTPCR), quantitative RT-PCR, and Affymetrix oligonucleotide microarrays. DD identified eight differentially expressed cDNAs, including inhibitor of apoptosis-1, 2'-5' oligoadenylate synthetase (OAS), a 2'-5' OAS-like (OASL) gene, galectin-9, myxovirus protein A (MxA), regulator of G-protein signaling, endothelial and smooth muscle cell-derived neuropilin-like protein, and phospholipid scramblase 1. Microarray analysis of 22,000 human genes confirmed these findings and identified an additional 269 genes that were induced and 126 that were repressed more than fourfold after DV infection. Broad functional responses that were activated included the stress, defense, immune, cell adhesion, wounding, inflammatory, and antiviral pathways. These changes in gene expression were seen after infection of HUVECs with either laboratory-adapted virus or with virus isolated directly from plasma of DV-infected patients. Tumor necrosis factor alpha, OASL, and MxA and h-IAP1 genes were induced within the first 8 to 12 h after infection, suggesting a direct effect of DV infection. These global analyses of DV effects on cellular gene expression identify potentially novel mechanisms involved in dengue disease manifestations such as hemostatic disturbance.

Cells, Cultured↗
PubMed2002

High circulating levels of the dengue virus nonstructural protein NS1 early in dengue illness correlate with the development of dengue hemorrhagic fever.

Infection with any 1 of 4 dengue viruses produces a spectrum of clinical illness ranging from a mild undifferentiated febrile illness to dengue fever (DF) to dengue hemorrhagic fever (DHF), a potentially life-threatening disease. The morbidity and mortality of DHF can be reduced by early hospitalization and careful supportive care. To determine its usefulness as a predictor of DHF, plasma levels of the secreted dengue virus nonstructural protein NS1 (sNS1) were measured daily in 32 children with dengue-2 virus infections participating in a prospective, hospital-based study. Free sNS1 levels in plasma correlated with viremia levels and were higher in patients with DHF than in those with DF. An elevated free sNS1 level (> or =600 ng/mL) within 72 h of illness onset identified patients at risk for developing DHF.

Adolescent↗
PubMed2002

Epidemiology of inapparent and symptomatic acute dengue virus infection: a prospective study of primary school children in Kamphaeng Phet, Thailand.

Dengue viruses are a major cause of morbidity in tropical and subtropical regions of the world. Knowledge about the epidemiology and host determinants of inapparent and severe dengue virus infections is limited. In this paper, the authors report findings from the first 3 years of a prospective study of dengue virus transmission and disease severity conducted in a cohort of 2,119 elementary school children in northern Thailand. A total of 717,106 person-school days were observed from 1998 to 2000. The incidence of inapparent and of symptomatic dengue virus infection was 4.3% and 3.6% in 1998, 3.2% and 3.3% in 1999, and 1.4% and 0.8% in 2000, respectively. Symptomatic dengue virus infection was responsible for 3.2%, 7.1%, and 1.1% of acute-illness school absences in 1998, 1999, and 2000, respectively. The early symptom complex of acute dengue virus infection is protean and difficult to distinguish from other causes of febrile childhood illnesses. The authors' results illustrate the spatial and temporal diversity of dengue virus infection and the burden of dengue disease in schoolchildren in Thailand. Their findings increase understanding of dengue virus transmission and disease severity in a well-defined cohort population and offer a study design in which to test the efficacy of potential dengue vaccines.

Acute Disease↗
PubMed2002

Spatial and temporal circulation of dengue virus serotypes: a prospective study of primary school children in Kamphaeng Phet, Thailand.

Dengue virus occurs as four distinct serotypes, each of which causes epidemics throughout the tropical and subtropical regions of the world. Few studies have examined co-circulation of multiple dengue virus serotypes in a well-defined cohort population over time and their capacity to produce severe dengue disease. In this paper, the authors report the details and findings of the first 3 years (1998-2000) of an ongoing prospective study of dengue virus transmission and disease severity in a cohort of children in northern Thailand. A total of 108 dengue virus isolates were obtained from 167 acute dengue virus infections; 23% were DEN-1, 35% were DEN-2, 41% were DEN-3, and 1% were DEN-4. Despite the proximity of the schools, there was marked spatial and temporal clustering of transmission of each dengue serotype. Serotype-specific antibody levels prior to the dengue transmission season were not predictive of the incidence of dengue virus infections or the predominant serotype transmitted at individual schools. All dengue serotypes produced severe dengue illness, although DEN-3 produced more severe symptoms than the other dengue serotypes. The authors' findings emphasize the complexity of dengue serotype-specific virus transmission and severe dengue disease and have important implications for dengue control and vaccine development.

Acute Disease↗
PubMed2002

T cell responses to an HLA-B*07-restricted epitope on the dengue NS3 protein correlate with disease severity.

Dengue hemorrhagic fever (DHF), the severe manifestation of dengue virus (DV) infection characterized by plasma leakage, is more common in secondary DV infections in previously infected individuals and is associated with high levels of immune activation. To determine the Ag specificity of this immune response, we studied the response to an HLA-B*07-restricted T cell epitope, residues 221-232 of the DV NS3 protein, in 10 HLA-B*07(+) Thai children who were studied during and after acute DV infections. Peptide-specific T cells were detected in 9 of 10 subjects. The frequency of peptide-specific T cells was higher in subjects who had experienced DHF than in those who had experienced DF. We also detected peptide-specific T cells in PBMC obtained at the time of the acute DV infection in 2 of 5 subjects. These data suggest that the NS3 (221-232) epitope is an important target of CD8(+) T cells in secondary DV infection and that the activation and expansion of DV-specific T cells is greater in subjects with DHF than in those with dengue fever. These findings support the hypothesis that activation of DV-specific CD8(+) T cells plays an important role in the pathogenesis of DHF.

Adolescent↗
PubMed2002

Dengue-specific T cell responses in peripheral blood mononuclear cells obtained prior to secondary dengue virus infections in Thai schoolchildren.

Children who experience secondary dengue virus (DV) infections are at increased risk for dengue hemorrhagic fever. To study the effect of preexisting T cell responses to DV on the severity of secondary virus infection, peripheral blood mononuclear cells (PBMC) from 10 subsequently hospitalized and 12 nonhospitalized Thai schoolchildren were stimulated with inactivated dengue antigens, and proliferation of interferon (IFN)-gamma or tumor necrosis factor (TNF)-alpha responses of the preinfection PBMC were measured. Proliferation responses were observed in 11 subjects, and IFN-gamma responses were seen in 12 subjects, 6 of whom showed broad serotype cross-reactive IFN-gamma responses. TNF-alpha responses were detected exclusively in 4 hospitalized subjects. Four PBMC samples that showed neither proliferation nor cytokine responses to any dengue antigen were from nonhospitalized subjects. This study, thought to be the first to investigate T cell responses to DV in preinfection PBMC, suggests that the pattern of preexisting T cell responses influences the risk for severe disease.

Animals↗
PubMed2002

Primary induction of human CD8+ cytotoxic T lymphocytes and interferon-gamma-producing T cells after smallpox vaccination.

This study measured the ability of a standard smallpox vaccine, given by scarification (by bifurcated needle), to induce primary human vaccinia virus-specific cytotoxic and interferon (IFN)-gamma-producing T lymphocyte responses. Because protection against smallpox may be mediated in part by T cell memory responses induced by vaccination, an analysis of the induction of primary human cytotoxic T lymphocytes (CTL) and IFN-gamma-producing T cell responses was performed. Although smallpox is no longer an epidemic threat under natural conditions, vaccination is still recommended for persons working with vaccinia viruses in the laboratory and for those who may be at risk from the potential use of smallpox virus as a bioterrorism agent. The results demonstrate that smallpox vaccine given by bifurcated needle induces strong vaccinia virus-specific CD8(+) CTL and IFN-gamma-producing T cell responses and provide baseline information useful for planning the immunologic assessment of future smallpox vaccines.

Adult↗
PubMed2002

Differing influences of virus burden and immune activation on disease severity in secondary dengue-3 virus infections.

Dengue hemorrhagic fever (DHF), the most severe form of illness following infection with a dengue virus, is characterized by plasma leakage, thrombocytopenia, and hepatic inflammation. The interrelationships among virus burden, immune activation, and development of DHF were examined in 54 children with secondary dengue-3 virus infections participating in a prospective, hospital-based study. DHF was associated with higher mean plasma viremia early in illness and earlier peak plasma interferon-gamma levels. Maximum plasma viremia levels correlated with the degree of plasma leakage and thrombocytopenia. Maximum plasma levels of interleukin (IL)-10 and soluble tumor necrosis factor receptor-II correlated with the degree of thrombocytopenia, independently of viremia levels. Hepatic transaminase elevation correlated with plasma soluble IL-2 receptor levels and not with viremia levels. Quantitative differences in virus burden and host immune responses, and the timing of type 1 cytokine responses, have differing influences on the severity of disease manifestations during secondary dengue-3 virus infections.

Antigens, CD↗
PubMed2002

Dose-related effects of smallpox vaccine.

BACKGROUND: We conducted a double-blind, randomized trial of three dilutions of vaccinia virus vaccine in previously unimmunized adults in order to assess the clinical success rates, humoral responses, and virus-specific activity of cytotoxic T cells and interferon-gamma-producing T cells. METHODS: Sixty healthy adults were inoculated intradermally by bifurcated needle with undiluted vaccine (dose, 10(7.8) plaque-forming units [pfu] per milliliter), a 1:10 dilution (dose, 10(6.5) pfu per milliliter), or a 1:100 dilution (dose, 10(5.0) pfu per milliliter); there were 20 subjects in each group. The subjects were monitored with respect to vesicle formation (an indicator of successful vaccination), the viral titer at the time of peak lesion formation, antiviral antibodies, and cellular immune responses. RESULTS: A vaccinia vesicle developed in 19 of the 20 subjects who received undiluted vaccine (95 percent), 14 of the 20 who received the 1:10 dilution (70 percent), and 3 of the 20 who received the 1:100 dilution (15 percent). One month after vaccination, 34 of 36 subjects with vesicles had antibody responses, as compared with only 1 of 24 subjects without clinical evidence of vaccinia virus replication. Vigorous cytotoxic T-cell and interferon-gamma responses occurred in 94 percent of subjects with vesicles, and a cytotoxic T-cell response occurred in only one subject without a vesicle. CONCLUSIONS: The vaccinia virus vaccine (which was produced in 1982 or earlier) still has substantial potency when administered by a bifurcated needle to previously unvaccinated adults. Diluting the vaccine reduces the rate of successful vaccination. The development of vesicular skin lesions after vaccination correlates with the induction of the antibody and T-cell responses that are considered essential for clearing vaccinia virus infections.

Adolescent↗
PubMed2002

Human cytotoxic T lymphocyte responses to live attenuated 17D yellow fever vaccine: identification of HLA-B35-restricted CTL epitopes on nonstructural proteins NS1, NS2b, NS3, and the structural protein E.

Yellow fever virus (YFV) is a re-emerging problem despite the existence of an effective live-attenuated vaccine. The induction of YFV-neutralizing antibodies undoubtedly contributes to vaccine efficacy, but T lymphocyte responses to YFV likely play a role in long-term efficacy. We studied the T lymphocyte responses to YFV in four vaccinees. Proliferation and cytolytic responses to YFV were demonstrated in all subjects. We isolated 13 YFV-specific CD8(+) CTL lines that recognized epitopes on the E, NS1, NS2b, and NS3 proteins; eight CTL lines were HLA-B35-restricted. YFV-specific T cell responses were detectable by IFN gamma ELISPOT assays 14 days postvaccination, with T cell frequencies sustained for up to 19 months. To our knowledge, this is the first report of human T lymphocyte responses following YFV vaccination. These results indicate that the live 17D YFV vaccine induced CD8(+) T cell responses directed against at least four different HLA-B35-restricted YFV epitopes.

Alleles↗
PubMed2002

Cytokine gene expression and protein production in peripheral blood mononuclear cells of children with acute dengue virus infections.

Plasma leakage in dengue hemorrhagic fever (DHF) is associated with elevated plasma levels of cytokines. To define further the contribution of immune activation to DHF and the source of cytokines, we analyzed the production of cytokines in peripheral blood mononuclear cells (PBMC) obtained from children with dengue, using RT-PCR and immunostaining. Tumor necrosis factor-alpha (TNF-alpha) and TNF-beta expression was detected in all samples by PCR and in < 50% of samples by immunostaining. Interferon-gamma (IFN-gamma) expression was detected in < 50% of samples by either method. Interleukin-2 (IL-2) and IL-4 expression was detected in a few samples by immunostaining but was not detectable by PCR. We found greater expression of TNF-alpha and IL-4 in DHF than in dengue fever or other (non-dengue) febrile illnesses. These results support the model of immunopathogenesis of DHF. However, low levels of cytokine expression in PBMC suggest that cellular activation in tissues may contribute to high serum cytokine levels in DHF.

Acute Disease↗
PubMed2002

Increased production of interleukin-8 in primary human monocytes and in human epithelial and endothelial cell lines after dengue virus challenge.

The more severe form of dengue virus infection, dengue hemorrhagic fever, is characterized by plasma leakage and derangements in hemostasis. As elevated interleukin-8 (IL-8) levels have been observed in sera from patients with more severe disease manifestations, a study was initiated to look at the effect of dengue virus infection in vitro on proinflammatory cytokine secretion and expression. A significant increase in IL-8 levels in the culture supernatant of primary human monocytes infected with dengue 2 virus (D2V) New Guinea C (NGC) was found by enzyme-linked immunosorbent assay. Additionally, by reverse transcriptase PCR, the mRNA was also augmented. Among the proinflammatory cytokines and their mRNAs measured (IL-6, IL-1 beta, IL-8, and tumor necrosis factor alpha), IL-8 showed the greatest change following D2V infection. Similarly, two cell lines, 293T (a human epithelial cell line) and ECV304 (an endothelial cell line), were permissive to D2V NGC and responded to the infection by increasing the synthesis of IL-8. Nuclear factor kappa B (NF-kappa B) and nuclear factor IL-6 (NFIL-6) are primary mediators of IL-8 expression. We studied the transcriptional regulation of IL-8 in the ECV304 and 293T cell lines and found that the induction of IL-8 gene expression involved the activation of NF-kappa B (P = 0.001) and, to a lesser extent, the activation of NFIL-6 in ECV304 cells only. We next observed by the chromatin immunoprecipitation procedure in vivo acetylation of core histones bound to the IL-8 promoter after D2V infection. IL-8 produced by infected monocytes and also IL-8 that may be produced by endothelial or other epithelial cells is associated with the hyperacetylation of histones bound to the IL-8 promoter in addition to the activation of transcription by NF-kappa B. We hypothesize that the overall increase in IL-8 synthesis observed in this in vitro study may play a role in the pathogenesis of the plasma leakage seen in dengue hemorrhagic fever and dengue shock syndrome.

Animals↗