Biomedical subjects

D M Koelle

Publications and source records attributed to D M Koelle.

At least 37 records · Page 2Linked to original sources

PubMed1998

Structural basis of specificity and degeneracy of T cell recognition: pluriallelic restriction of T cell responses to a peptide antigen involves both specific and promiscuous interactions between the T cell receptor, peptide, and HLA-DR.

TCR engagement of peptide-MHC class II ligands involves specific contacts between the TCR and residues on both the MHC and peptide molecules. We have used molecular modeling and assays of peptide binding and T cell function to characterize these interactions for a CD4+ Th1 cell clone, ESL4.34, which recognizes a peptide epitope of the herpes simplex type 2 virus virion protein, VP16 393-405, in the context of several HLA-DR alleles. This clone responded to VP16 393-405 in proliferation and cytotoxicity assays when presented by DRB1*0402, DRB1*1102, and DRB1*1301, which share a common amino acid sequence, ILEDE, at residues 67-71 in the alpha-helical portion of the DRbeta polypeptide, but not when presented by other DR4, DR11, and DR13 alleles that are negative for this sequence. Using a panel of APCs expressing DR4 molecules that were mutagenized in vitro at individual residues within this shared epitope and using peptide analogues with single amino acid substitutions of predicted MHC and TCR contact residues, a unit of recognition was identified dependent on DRbeta residues 67-71 and relative position 4 (P4) of the VP16 393-405 peptide. The interactions of this portion of the peptide-DR ligand with the ESL4.34 TCR support a structural model for MHC-biased recognition in some Ag-specific and alloreactive T cell responses and suggest a possible mechanism for autoreactive T cell selection in rheumatoid arthritis.

Alleles↗

PubMed1998

Clearance of HSV-2 from recurrent genital lesions correlates with infiltration of HSV-specific cytotoxic T lymphocytes.

The mechanisms involved in host clearance of symptomatic mucocutaneous herpes simplex virus (HSV) infection are unclear. We studied the functional properties of bulk cultures of skin-infiltrating lymphocytes from normal skin and serial biopsies of recurrent genital HSV-2 lesions, and compared HSV-specific and NK responses with viral clearance. HSV-specific CD4+ or CD8+ T cells were rarely detected in lymphocytes cultured from normal skin. The total lymphocyte count and HSV-specific and NK-like effector cell activities were markedly higher in cultures derived from lesional skin. HSV-specific CD4+ proliferative responses and NK-like cytotoxic responses were present at all stages of herpetic lesions, including biopsies early in the disease course. In contrast, cytotoxic T lymphocyte activity was generally low among cells derived from early culture-positive lesions, and increased during lesion evolution. Viral clearance from the lesion site was associated with a high level of local cytolytic activity towards HSV-infected cells. The phenotypes of cells with HSV-specific cytotoxic responses varied between patients, having CD4+ and CD8+ components. Immunotherapeutic approaches to HSV should be directed at improving in vivo cytolytic activity to HSV.

Adult↗

PubMed1998

Reactivities of human sera with human herpesvirus-8-infected BCBL-1 cells and identification of HHV-8-specific proteins and glycoproteins and the encoding cDNAs.

The reactivates of human sera with uninduced and phorbol ester (TPA)-induced human herpesvirus-8 (HHV-8)-infected BCBL-1 cells were examined by immunofluorescence assay (IFA) and by radioimmunoprecipitation reactions (RIP). The seroprevalence of HHV-8 infections is low in the United States general population and only low levels of HHV-8 antibodies were detected in the seropositive sera. In contrast, high levels of antibodies against HHV-8 lytic and latent antigens were detected by IFA in the sera from HIV+ Kaposi's sarcoma (KS)-positive individuals. These sera recognized several proteins and glycoproteins from BCBL-1 cells in RIP reactions. Two types of antibody responses were detected in the sera from HIV+ KS- homosexual men. In majority of the sera with and without detectable HHV-8 DNA in the peripheral blood mononuclear cells (PBMC), significantly low levels of HHV-8 antibodies were detected by IFA. These sera recognized only a subset of HHV-8 proteins and glycoproteins in RIP reactions. In contrast, in a subgroup of sera from HIV+ KS- homosexual men, higher levels of IFA antibodies against HHV-8 lytic and latent antigens were detected. These sera also recognized several viral proteins and glycoproteins in RIP reactions. These results suggest that antibody response profiles to HHV-8 infection vary significantly and serologic assays to detect antibody responses to a panel of both lytic and latent antibodies may be required for maximum sensitivity. Screening of a cDNA library from TPA-induced BCBL-1 cells with an HIV+ KS+ serum identified cDNAs encoding 12 HHV-8 proteins. Further characterization of these HHV-8 proteins would define the HHV-8 antigens useful for seroepidemiological studies and in discriminating lytic, latent, past, and/or reactivation infections.

Acquired Immunodeficiency Syndrome↗

PubMed1998

Famciclovir for the suppression of symptomatic and asymptomatic herpes simplex virus reactivation in HIV-infected persons. A double-blind, placebo-controlled trial.

BACKGROUND: Herpes simplex virus (HSV) infection is one of the most common opportunistic infections in HIV-infected persons. However, most documentation of the effectiveness of antiviral therapy in reducing HSV reactivation is anecdotal. OBJECTIVE: To evaluate the quantitative effect of antiviral therapy on the frequency of HSV reactivation in HIV-infected persons. DESIGN: Double-blind, placebo-controlled, crossover trial. SETTING: Research clinic at a university hospital. PATIENTS: 48 persons (45 men and 3 women) who were HIV positive and HSV seropositive. INTERVENTION: Patients were randomly assigned to receive famciclovir, 500 mg orally twice daily, or placebo for 8 weeks. They then crossed over to receive the other regimen after a 1-week washout period. MEASUREMENTS: Patients obtained daily cultures of their perirectal, urethral, oral, and genital areas and kept dairy records of signs and symptoms of genital and oral-labial herpes. RESULTS: The median CD4 cell count at study entry was 384 cells/mm3. In the intention-to-treat analysis of the first study period, HSV was isolated on 122 of 1114 (11%) placebo days compared with 9 of 1071 (1%) famciclovir days (relative risk, 0.15; P < 0.001). For patients who completed the crossover, the median difference in days with symptoms between placebo and famciclovir was 13.8% of days and the median difference in days on which HSV was isolated was 5.4% of days (P < 0.001 for both). Percentage of days with HSV-2 shedding was reduced from 9.7% to 1.3%. Breakthrough reactivations that occurred while patients were receiving famciclovir were infrequent, short, and often asymptomatic, HSV-2 isolates from these reactivations were susceptible to penciclovir in vitro. CONCLUSIONS: Antiviral chemotherapy with famciclovir results in clinically and statistically significant reductions in the symptoms associated with HSV infection and the symptomatic and asymptomatic shedding of HSV among HIV-positive persons.

2-Aminopurine↗

PubMed1998

Tipping the scales of herpes simplex virus reactivation: the important responses are local.

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CD4-Positive T-Lymphocytes↗

PubMed1998

Chronic vulvar ulceration in an immunocompetent woman due to acyclovir-resistant, thymidine kinase-deficient herpes simplex virus.

A 34-year-old healthy woman presented with a 15-month history of persistent, nonhealing vulvar ulcerations due to herpes simplex virus (HSV) type 2. Extensive dermatologic workup and serial skin biopsies failed to reveal an underlying vulvar dermatosis or autoimmune bullous disorder. Virologic studies revealed resistance to acyclovir in vitro due to deficiency in thymidine kinase activity. Serum antibody to human immunodeficiency virus was negative on two occasions, separated by 1 year. Immunologic evaluation showed normal HSV-specific proliferative and CD8 cytotoxic T lymphocyte responses as well as normal NK cell function. Vulvar lesions failed to heal in association with trials of topical trifluorothymidine and oral valacyclovir but resolved completely with the application of 1% foscarnet cream. No recurrence of HSV has been observed in 24 months of follow-up to date.

Acyclovir↗

PubMed1998

Herpes simplex virus type 1 renders infected cells resistant to cytotoxic T-lymphocyte-induced apoptosis.

Many viruses interfere with apoptosis of infected cells, presumably preventing cellular apoptosis as a direct response to viral infection. Since cytotoxic T lymphocytes (CTL) induce apoptosis of infected cells as part of the "lethal hit," inhibition of apoptosis could represent an effective immune evasion strategy. We report here herpes simplex virus type 1 (HSV-1) interference with CTL-induced apoptosis of infected cells and show that HSV-1 inhibits the nuclear manifestations of apoptosis but not the membrane changes. The HL-60 cell line (human promyelocytic leukemia) undergoes apoptosis in response to many stimuli, including incubation with ethanol. After HSV-1 infection (strains E115 and 17+), ethanol-treated cells did not produce oligonucleosomal DNA fragments characteristic of apoptosis, as assayed by gel electrophoresis and enzyme-linked immunosorbent assay. Inhibition was detected 2 h after infection and increased over time. Importantly, HSV-1-infected cells were resistant to apoptosis induced by antigen-specific CD4+ CTL, despite the fact that CTL recognition and degranulation in response to infected targets remained intact. Unlike HSV-1, HSV-2 (strains 333 and HG52) did not inhibit DNA fragmentation. In contrast to the inhibition of DNA fragmentation by HSV-1, none of the HSV-1 or -2 strains interfered with the ethanol-induced exposure of surface phosphatidylserine characteristic of apoptosis, as determined by annexin V binding. These results demonstrate that genes of HSV-1 inhibit the nuclear manifestations of apoptosis but not the membrane manifestations, suggesting that these may be mediated via separate pathways. They also suggest that HSV-1 inhibition of CTL-induced apoptosis may be an important mechanism of immune evasion.

Apoptosis↗

PubMed1998

Human herpesvirus 8 in the prostate glands of men with Kaposi's sarcoma.

The epidemiology of human immunodeficiency virus (HIV)-associated Kaposi's sarcoma (KS) resembles that of a sexually transmitted pathogen. However, human herpesvirus 8 (HHV-8), the proposed cause of KS, is found in semen only infrequently and at low titers. To determine whether HHV-8 was present in the urogenital tract, transrectal ultrasound-guided prostate biopsies were obtained from six men with KS (five with concurrent HIV infection) and four without KS (three with concurrent HIV) and assayed for HHV-8 by PCR. Nine of the 10 men were seropositive for HHV-8. Five of nine HHV-8-seropositive men had HHV-8 DNA detected in prostate tissue by solution-based PCR. All five currently had KS or had it previously. In two subjects, prostate tissue was the only identified source of HHV-8. In situ PCR on serial sections of prostate indicated that HHV-8 infection was localized to discrete areas of the prostate. When detected, HHV-8 DNA was present in the nuclei of >90% of the glandular epithelial cells. In situ hybridization for HHV-8 mRNA revealed that between 1 and 5% of cells harboring HHV-8 DNA expressed viral transcripts associated with HHV-8 replication (T1.1 transcript), while >90% expressed gene products associated with viral latency (T0.7 transcript). Intermittent replication of HHV-8 in the prostate and subsequent shedding of virus in semen may be crucial factors for determining whether HHV-8 can be transmitted through sexual activity.

Adult↗

PubMed1998

Recognition of herpes simplex virus type 2 tegument proteins by CD4 T cells infiltrating human genital herpes lesions.

The local cellular immune response to herpes simplex virus (HSV) is important in the control of recurrent HSV infection. The antiviral functions of infiltrating CD4-bearing T cells may include cytotoxicity, inhibition of viral growth, lymphokine secretion, and support of humoral and CD8 responses. The antigens recognized by many HSV-specific CD4 T cells localizing to genital HSV-2 lesions are unknown. T cells recognizing antigens encoded within map units 0. 67 to 0.73 of HSV DNA are frequently recovered from herpetic lesions. Expression cloning with this region of DNA now shows that tegument protein VP22 and the viral dUTPase, encoded by genes UL49 and UL50, respectively, are T-cell antigens. Separate epitopes in VP22 were defined for T-cell clones from each of three patients. Reactivity with the tegument protein encoded by UL21 was identified for an additional patient. Three new epitopes were identified in VP16, a tegument protein associated with VP22. Some tegument-specific CD4 T-cell clones exhibited cytotoxic activity against HSV-infected cells. These results suggest that herpes simplex tegument proteins are processed for antigen presentation in vivo and are possible candidate compounds for herpes simplex vaccines.

Amino Acid Sequence↗

PubMed1997

Severe genital herpes infections in HIV-infected individuals with impaired herpes simplex virus-specific CD8+ cytotoxic T lymphocyte responses.

The specific mechanisms underlying the varied susceptibility of HIV-infected (HIV+) individuals to opportunistic infections (OI) are still incompletely understood. One hypothesis is that quantitative differences in specific T cell responses to a colonizing organism determine the development of an AIDS-defining OI. We evaluated this hypothesis for herpes simplex virus (HSV) infection, a common OI in HIV+ patients. Using limiting dilution analyses, the frequency of HSV-specific CD8+ cytotoxic T lymphocyte precursors (pCTL) and proliferative precursors were quantitated in peripheral blood mononuclear cells from 20 patients coinfected with HIV and HSV-2. The frequency of HSV-specific CD8+ pCTL in HSV+HIV+ individuals was significantly lower than in HSV+HIV- individuals (1 in 77,000 vs. 1 in 6,000, P = .0005) and was not different than in HSV-HIV- individuals (1 in 100,000, P = .24). HIV+ patients who suffered more severe genital herpes recurrences had significantly lower HSV-specific CD8+ pCTL frequencies than those patients with mild recurrences (1 in 170,000 vs. 1 in 26,000, P = .03). In contrast, no significant difference was seen in proliferative precursor frequencies between those patients with mild vs. severe genital herpes (1 in 3,800 vs. 1 in 6,600, P > .5). Quantitative differences in pCTL frequency to HSV appear to be the most important host factor influencing the frequency and severity of HSV reactivation in HIV+ patients. Studies to reconstitute such immunity, especially in people with acyclovir-resistant HSV, appear warranted.

CD8-Positive T-Lymphocytes↗

PubMed1997

Preferential presentation of herpes simplex virus T-cell antigen by HLA DQA10501/DQB10201 in comparison to HLA DQA10201/DQB10201.

The HLA DQA1 locus is polymorphic. Haplotypes containing HLA DQA1*0501, but not HLA DQA1*0201, together with HLA DQB1*0201 are associated with Grave's disease and celiac sprue. In this report, we demonstrate a functional correlate of DQA1 polymorphism. T cells infiltrating a herpes simplex virus (HSV) lesion from a HLA DQ 2,7 individual yielded a virus-specific CD4+ clone restricted by DQ2. Presentation of viral peptide and protein segregated with DQA1 allele, because cell lines bearing DQA1*0501/DQB1*0201 heterodimers presented antigen in proliferation and cytotoxicity assays much more efficiently than cell lines bearing DQA1*0201/DQB1*0201. Binding of viral peptide to cell lines bearing DQA1*0201, in comparison to DQA1*0501, was only moderately reduced and may not explain this effect. Truncation and substitution analyses of peptide binding and T-cell activation were performed to determine which viral peptide residues contacting TCR might therefore be presented in an altered conformation by DQA1*0201/DQB1*0201. Residues 432, 435, 437, 438, and 440 (position P1, P4, P6, P7, and P9) contributed to DQ2 binding, whereas residues 431, 433, 434, and 436 (positions P 1, P2, P3, and P5) contributed to TCR contact. Differential presentation of peptide by HLA DQ2 heterodimers varying at the DQA1 locus may have relevance to host defense and the pathogenesis of HLA DQ2-associated autoimmune diseases.

Alleles↗

PubMed1997

Frequent detection of Kaposi's sarcoma-associated herpesvirus (human herpesvirus 8) DNA in saliva of human immunodeficiency virus-infected men: clinical and immunologic correlates.

The prevalence, quantity, temporal pattern, and clinical and immunologic correlates of shedding of Kaposi's sarcoma (KS)-associated herpesvirus (KSHV; or human herpesvirus [HHV]-8) DNA in saliva were studied. KSHV DNA was detected in saliva from 18 (75%) of 24 human immunodeficiency virus (HIV)-positive patients with KS and from 1 of 1 HIV-negative patient with KS, 3 (15%) of 20 HIV-positive patients without KS, and none of 24 controls. KSHV DNA levels ranged from 10(2.4) to 10(6) copies/mL and were lower than levels for Epstein-Barr virus but comparable to those for HHV-6. Detection of KSHV DNA in saliva was not associated with oral KS or decreased peripheral blood CD4 cell counts. KSHV DNA was not detected in semen. Resistance of KSHV DNA from saliva to DNase treatment was consistent with the presence of virions. These data suggest that KSHV can replicate in the oropharynx and that salivary contact could contribute to KSHV transmission.

Antibodies, Viral↗

PubMed1997

Transmissible Kaposi's sarcoma-associated herpesvirus (human herpesvirus 8) in saliva of men with a history of Kaposi's sarcoma.

We have evaluated the physical state and infectious nature of Kaposi's sarcoma-associated herpesvirus (KSHV) in the saliva of nine persons with past or current Kaposi's sarcoma (KS). KSHV DNA in saliva had the physical characteristics of DNA present in virions. Inoculation of 293 cells with cell-free saliva fluid resulted in the persistence of KSHV DNA in culture for at least 13 passages of the cells. The addition of tetradecanoyl phorbol acetate to KSHV-infected 293 cells led to increased viral DNA. Two virus-specific RNAs were detected by reverse transcriptase PCR in 293 cells infected with cell-free saliva fluid and in cells present in saliva from subjects with KSHV salivary shedding. These results indicate that infectious KSHV can be present in saliva of patients with KS.

Cell Line, Transformed↗

PubMed1996

HLA-DQB1 codon 57 is critical for peptide binding and recognition.

The association of specific HLA-DQ alleles with autoimmunity is correlated with discrete polymorphisms in the HLA-DQ sequence that are localized within sites suitable for peptide recognition. The polymorphism at residue 57 of the DQB1 polypeptide is of particular interest since it may play a major structural role in the formation of a salt bridge structure at one end of the peptide-binding cleft of the DQ molecules. This polymorphism at residue 57 is a recurrent feature of HLA-DQ evolution, occurring in multiple distinct allelic families, which implies a functional selection for maintaining variation at this position in the class II molecule. We directly tested the amino acid polymorphism at this site as a determinant for peptide binding and for antigen-specific T cell stimulation. We found that a single Ala-->Asp amino acid 57 substitution in an HLA-DQ3.2 molecule regulated binding of an HSV-2 VP-16-derived peptide. A complementary single-residue substitution in the peptide abolished its binding to DQ3.2 and converted it to a peptide that can bind to DQ3.1 and DQ3.3 Asp-57-positive MHC molecules. These binding studies were paralleled by specific T cell recognition of the class II-peptide complex, in which the substituted peptide abolished T cell reactivity, which was directed to the DQ3.2-peptide complex, whereas the same T cell clone recognized the substituted peptide presented by DQ3.3, a class II restriction element differing from DQ3.2 only at residue 57. This structural and functional complementarity for residue 57 and a specific peptide residue identifies this interaction as a key controlling determinant of restricted recognition in HLA-DQ-specific immune response.

Alanine↗

PubMed1996

Primary and secondary syphilis lesions contain mRNA for Th1 cytokines.

Phagocytosis of Treponema pallidum by cytokine-activated macrophages aids bacterial clearance and lesion resolution in early syphilis. To investigate the cytokine profiles of cells infiltrating primary and secondary syphilis lesions, reverse transcription and polymerase chain reaction (RT-PCR) were used to detect cytokine mRNA in 13 lesion biopsies. Both primary and secondary lesions contained mRNA encoding interleukin (IL)-2, interferon-gamma (IFN-gamma), IL-12p40, and IL-10. In contrast to a lesion from a patient with recurrent herpes simplex virus type 2, no message for IL-4 could be detected in any of the syphilis lesions, and 10 of 13 had no mRNA for IL-5 or IL-13. These findings are consistent with a Th1-predominant local cellular response activating macrophages and support the hypothesis that IFN-gamma-activated macrophages are primary effectors in treponeme clearance.

Base Sequence↗

PubMed1996

High frequency of CD8+ cytotoxic T-lymphocyte precursors specific for herpes simplex viruses in persons with genital herpes.

Herpes simplex virus (HSV)-specific CD8+ cytotoxic T lymphocytes (CTL) have rarely been detected in humans, presumably because of virus-induced mechanisms that downregulate major histocompatibility complex class I expression. We have developed a method that has allowed us to consistently demonstrate HSV-specific CD8+ precursor CTL (pCTL) from HSV type 1- and 2-seropositive persons. Major histocompatibility complex-restricted HSV-specific CD8+ pCTL were found in 10 consecutively tested HSV type 1- and 2-seropositive subjects at frequencies ranging from 1 in 21,000 to 1 in 300 (median, 1 in 6,000) versus a pCTL frequency of 1 in 100,000 in HSV-seronegative donors. These results suggest that CD8+ CTL are important effector cells in resolving HSV lesions.

Herpes Genitalis↗

PubMed1994

Direct recovery of herpes simplex virus (HSV)-specific T lymphocyte clones from recurrent genital HSV-2 lesions.

T lymphocytes were recovered from recurrent vesicular and ulcerative herpes simplex virus type 2 (HSV-2) lesions from 4 patients and cloned without exogenous secondary antigenic stimulation. Resultant cultures were screened for antigen-specific proliferative and cytotoxic responses. Of the 39 HSV-specific clones recovered, all were CD3-positive; 38 were CD4-positive and 1 was CD8-positive. Of the T cell clones recovered directly from lesions, 6%-10% had HSV-specific proliferative responses, in contrast to a peripheral blood precursor frequency of HSV-specific CD4 cells of approximately 1/10(3) to 1/10(4) as measured by limiting dilution assays. CD4+ clones studied restricted by HLA-DR, -DP, and -DQ were detected and both type-common and type-specific CD4+ T cell clones were recovered. HSV-specific T cells localize to recurrent human genital HSV-2 lesions and can be obtained directly from early lesions. The ability to isolate HSV-specific T cell clones from lesions allows the study of antigenic specificities and functional properties of tissue-resident antigen-specific T lymphocytes.

Cell Division↗

PubMed1994

Antigenic specificities of human CD4+ T-cell clones recovered from recurrent genital herpes simplex virus type 2 lesions.

Lesions resulting from recurrent genital herpes simplex virus (HSV) infection are characterized by infiltration of CD4+ lymphocytes. We have investigated the antigenic specificity of 47 HSV-specific CD4+ T-cell clones recovered from the HSV-2 buttock and thigh lesions of five patients. Clones with proliferative responses to recombinant truncated glycoprotein B (gB) or gD of HSV-2 or purified natural gC of HSV-2 comprised a minority of the total number of HSV-specific clones isolated from lesions. The gC2- and gD2-specific CD4+ clones had cytotoxic activity. The approximate locations of the HSV-2 genes encoding HSV-2 type-specific CD4+ antigens have been determined by using HSV-1 x HSV-2 intertypic recombinant virus and include the approximate map regions 0.30 to 0.46, 0.59 to 0.67, 0.67 to 0.73, and 0.82 to 1.0 units. The antigenic specificity of an HLA DQ2-restricted, HSV-2 type-specific T-cell clone was mapped to amino acids 425 to 444 of VP16 of HSV-2 by sequential use of an intertypic recombinant virus containing VP16 of HSV-2 in an HSV-1 background, recombinant VP16 fusion proteins, and synthetic peptides. Each of the remaining four patients also yielded at least one type-specific T-cell clone reactive with an HSV-2 epitope mapping to approximately 0.67 to 0.73 map units. The antigenic specificities of lesion-derived CD4+ T-cell clones are quite diverse and include at least 10 epitopes. Human T-cell clones reactive with gC and VP16 are reported here for the first time.

Amino Acid Sequence↗