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At least 37 records · Page 2Linked to original sources
PubMed2005

Early detection of dominant Env-specific and subdominant Gag-specific CD8+ lymphocytes in equine infectious anemia virus-infected horses using major histocompatibility complex class I/peptide tetrameric complexes.

Cytotoxic T lymphocytes (CTL) are critical for control of lentiviruses, including equine infectious anemia virus (EIAV). Measurement of equine CTL responses has relied on chromium-release assays, which do not allow accurate quantitation. Recently, the equine MHC class I molecule 7-6, associated with the ELA-A1 haplotype, was shown to present both the Gag-GW12 and Env-RW12 EIAV CTL epitopes. In this study, 7-6/Gag-GW12 and 7-6/Env-RW12 MHC class I/peptide tetrameric complexes were constructed and used to analyze Gag-GW12- and Env-RW12-specific CTL responses in two EIAV-infected horses (A2164 and A2171). Gag-GW12 and Env-RW12 tetramer-positive CD8+ cells were identified in nonstimulated peripheral blood mononuclear cells as early as 14 days post-EIAV inoculation, and frequencies of tetramer-positive cells ranged from 0.4% to 6.7% of nonstimulated peripheral blood CD8+ cells during the 127-day study period. Although both horses terminated the initial viremic peak, only horse A2171 effectively controlled viral load. Neutralizing antibody was present during the initial control of viral load in both horses, but the ability to maintain control correlated with Gag-GW12-specific CD8+ cells in A2171. Despite Env-RW12 dominance, Env-RW12 escape viral variants were identified in both horses and there was no correlation between Env-RW12-specific CD8+ cells and control of viral load. Although Gag-GW12 CTL escape did not occur, a Gag-GW12 epitope variant arose in A2164 that was recognized less efficiently than the original epitope. These data indicate that tetramers are useful for identification and quantitation of CTL responses in horses, and suggest that the observed control of EIAV replication and clinical disease was associated with sustained CTL recognition of Gag-specific epitopes.

Amino Acid Sequence↗
PubMed1995

Detection by polymerase chain reaction of BCR/ABL transcripts in myeloproliferative diseases at time of diagnosis and for monitoring chronic myelogenous leukaemia patients after bone marrow transplantation.

The Philadelphia chromosome t(9;22)(q34;q11) is a cytogenetic marker for chronic myelogenous leukaemia (CML), and is also present in some acute leukaemias. The translocation in CML gives rise to two BCR/ABL chimeric transcripts (b3a2 and b2a2) encoding a 210-kD tyrosine kinase protein. These leukaemia-specific transcripts can be detected easily by the reverse transcriptase polymerase chain reaction (PCR). PCR has improved the possibility of detecting minimal residual leukaemia cells in Ph-positive patients, especially after bone marrow transplantation (BMT). With PCR, we looked for BCR/ABL transcripts in 30 patients with CML and 4 with essential thrombocythaemia at time of diagnosis, finding a significant difference in the platelet counts of CML patients carrying b3a2 or b2a2 transcripts. The BCR/ABL transcript was monitored by PCR in 6 CML patients after BMT. The usefulness of PCR in clinical practice at time of diagnosis, and the biological and clinical significance of positive/negative PCR results, in patients with transplants, are discussed.

Adult↗
PubMed1994

The effect of TCR V beta 8 peptide protection and therapy on T cell populations isolated from the spinal cords of Lewis rats with experimental autoimmune encephalomyelitis.

Vaccination or treatment of Lewis rats with TCR V beta 8 peptides can prevent or reverse the clinical signs of experimental autoimmune encephalomyelitis (EAE) which is mediated predominantly by V beta 8.2+ CD4+/CD45R lo T cells. However, rats protected or treated with V beta 8 peptides still developed histological lesions in the spinal cord (SC), even though they remained clinically well. We sought to discern phenotypic changes characteristic of these SC infiltrating lymphocytes. In particular, we focused on whether the immunoregulatory mechanism induced by TCR peptides caused a reduction of V beta 8.2+ T cells, or induced changes in CD45R lo or hi/CD4+ subpopulations that have been associated respectively with EAE induction or recovery. In the V beta 8 peptide vaccinated rats there was a dramatic decrease in the number of V beta 8.2+ T cells isolated from the SC early in disease. During the recovery phase, however, the number of V beta 8.2+ SC T cells was similar in protected and control groups; in contrast, there was striking reduction in the number and size of CD45R hi/CD4+ T cells in the protected animals. In rats treated with V beta 8.2 peptide, no changes were observed in the number of SC V beta 8.2+ T cells or expression of V beta 8.2 message, but similar to vaccinated rats, there was a marked decrease in the number of CD45R hi/CD4+ T cells. These data suggest that vaccination with TCR peptides prevented the initial influx of encephalitogenic V beta 8.2+ T cells into the central nervous system (CNS), whereas treatment appeared to inactivate V beta 8.2+ T cells already present in the CNS. In both cases, TCR peptide-induced inhibition of the encephalitogenic T cells apparently preempted the need for CD45R hi/CD4+ T cells that may normally be necessary to resolve the disease.

Amino Acid Sequence↗
PubMed1996

Chronic myeloid leukemia of thrombocythemic onset: a CML subtype with distinct hematological and molecular features?

Among 84 consecutive patients with chronic phase Ph-positive chronic myeloid leukemia (CML) who were investigated for the hybrid BCR/ABL mRNA, in six cases (7%) the disease mimicked essential thrombocythemia (ET) at presentation, because of marked thrombocytosis (platelet counts ranging from 1003 x 10(9)/l to 2800 x 10(9)/l) and moderate leukocytosis (WBC counts from 10 x 10(9)/l to 19 x 10(9)/l). At initial examination, four of the six patients showed a few (4-9%) immature myeloid cells in the peripheral blood, and two had blood basophilia. All six patients later developed increasing leukocytosis, and two subsequently died from blast crisis and accelerated CML, respectively. In the overall series, 38 patients (45%) expressed the b2a2 type of BCR/ABL mRNA, and 46 (55%) either the b3a2 or both mRNAs. By contrast, only one of the six patients with CML of thrombocythemic onset expressed the b2a2 mRNA vs five with either b3a2 (n = 4) or both mRNA types (n = 1). The above results, in conjunction with similar data from a few previously published cases, suggest an association between the above form of CML and b3a2 type of BCR/ABL transcript.

Adolescent↗
PubMed2004

Human T cell lymphotropic virus type I (HTLV-I)-specific CD4+ T cells: immunodominance hierarchy and preferential infection with HTLV-I.

CD4(+) T cells predominate in early lesions in the CNS in the inflammatory disease human lymphotropic T cell virus type I (HTLV-I)-associated myelopathy/tropical spastic paraparesis (HAM/TSP), but the pathogenesis of the disease remains unclear and the HTLV-I-specific CD4(+) T cell response has been little studied. We quantified the IFN-gamma-producing HTLV-I-specific CD4(+) T cells, in patients with HAM/TSP and in asymptomatic carriers with high proviral load, to test two hypotheses: that HAM/TSP patients and asymptomatic HTLV-I carriers with a similar proviral load differ in the immunodominance hierarchy or the total frequency of specific CD4(+) T cells, and that HTLV-I-specific CD4(+) T cells are preferentially infected with HTLV-I. The strongest CD4(+) T cell response in both HAM/TSP patients and asymptomatic carriers was specific to Env. This contrasts with the immunodominance of Tax in the HTLV-I-specific CD8(+) T cell response. The median frequency of HTLV-I-specific IFN-gamma(+) CD4(+) T cells was 25-fold greater in patients with HAM/TSP (p = 0.0023, Mann-Whitney) than in asymptomatic HTLV-I carriers with a similar proviral load. Furthermore, the frequency of CD4(+) T cells infected with HTLV-I (expressing Tax protein) was significantly greater (p = 0.0152, Mann-Whitney) among HTLV-I-specific cells than CMV-specific cells. These data were confirmed by quantitative PCR for HTLV-I DNA. We conclude that the high frequency of specific CD4(+) T cells was associated with the disease HAM/TSP, and did not simply reflect the higher proviral load that is usually found in HAM/TSP patients. Finally, we conclude that HTLV-I-specific CD4(+) T cells are preferentially infected with HTLV-I.

Amino Acid Sequence↗
PubMed2005

Selective escape from CD8+ T-cell responses represents a major driving force of human immunodeficiency virus type 1 (HIV-1) sequence diversity and reveals constraints on HIV-1 evolution.

The sequence diversity of human immunodeficiency virus type 1 (HIV-1) represents a major obstacle to the development of an effective vaccine, yet the forces impacting the evolution of this pathogen remain unclear. To address this issue we assessed the relationship between genome-wide viral evolution and adaptive CD8+ T-cell responses in four clade B virus-infected patients studied longitudinally for as long as 5 years after acute infection. Of the 98 amino acid mutations identified in nonenvelope antigens, 53% were associated with detectable CD8+ T-cell responses, indicative of positive selective immune pressures. An additional 18% of amino acid mutations represented substitutions toward common clade B consensus sequence residues, nine of which were strongly associated with HLA class I alleles not expressed by the subjects and thus indicative of reversions of transmitted CD8 escape mutations. Thus, nearly two-thirds of all mutations were attributable to CD8+ T-cell selective pressures. A closer examination of CD8 escape mutations in additional persons with chronic disease indicated that not only did immune pressures frequently result in selection of identical amino acid substitutions in mutating epitopes, but mutating residues also correlated with highly polymorphic sites in both clade B and C viruses. These data indicate a dominant role for cellular immune selective pressures in driving both individual and global HIV-1 evolution. The stereotypic nature of acquired mutations provides support for biochemical constraints limiting HIV-1 evolution and for the impact of CD8 escape mutations on viral fitness.

Acute Disease↗
PubMed1993

The relationship between the type of bcr-abl hybrid messenger RNA and thrombopoiesis in Philadelphia-positive chronic myelogenous leukemia.

Formation of the Philadelphia (Ph1) chromosome, which contains the hybrid bcr-abl gene, is thought to be the initial event in chronic myelogenous leukemia (CML). The positions of the breakpoint within the breakpoint cluster region (bcr) on the bcr-abl gene in 22 chronic-phase cases of Ph1-positive CML were determined using conventional Southern blots, and the splicing pattern were also determined the species of the fused bcr-abl mRNA in 79 CML cases using the polymerase chain-reaction procedure (RT-PCR). The location of the breakpoint within the bcr locus was assigned to one of five zones. Breakpoints in zones 1 and 2 were grouped as 5', and those in zones 3, 4 and 5 as 3'. Nine patients had 5' breakpoints and 13 patients had 3' breakpoints. The platelet counts of 3' patients were significantly higher than those of 5' patients (1395 vs 274 x 10(9)/L; p < 0.03). The megakaryocyte counts from bone marrow histological sections in 3' patients (n = 12) and 5' patients (n = 7) were 63.4/mm2 and 19.5/mm2, with a significant difference at p < 0.006. The mean number of megakaryocyte progenitor cells assayed by in vitro cloning was 128.3/2 x 10(5) bone marrow cells for 3' patients (n = 7) compared with 46.3 for 5' patients (n = 4). Using the RT-PCR technique, the bcr exon 2/abl exon 2 fused mRNA (b2-a2) was detected in 18 patients, the bcr exon 3/abl exon 2 fused mRNA (b3-a2) was detected in 45 patients, and both types of mRNA were detected in 16 patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗
PubMed1996

Time course analysis of alpha+ beta+ T cell clones during normal pregnancy.

During normal pregnancy, the fetus continues to mature inside the uterus without rejection. Inherited paternal antigens could be targeted by the maternal immune system. These reactions are believed to play a role in a number of habitual abortions. However, the precise maternal mechanisms preventing fetal tissue rejection are not well understood. Maternal T cells should recognize fetal antigens, so it is conceivable that antigen-specific T cell response to fetal antigens would occur by proliferation and accumulation of certain T cell clones in the pregnant mother. To elucidate the maternal immune response to the fetus we investigated the clonality of expanded T cells in peripheral blood lymphocytes in ten normal pregnant women. We employed reverse transcriptase-polymerase chain reaction for T cell receptor beta chain gene and subsequently analyzed the PCR product by single-strand conformation polymorphism analysis. A large number of distinctly expanded T cell clones were detected during pregnancy. These accumulations were observed as early as the ninth to tenth week post-conception and reached a maximum during the second trimester, suggesting the existence of dynamic antigen-specific T cell responses in the pregnant mother. However, after the 30th week of gestation, nearly all expanded T cell clones disappeared before parturition and the degree of clonality reached almost normal levels. Our results clearly indicate the existence of dynamic maternal T cell responses during pregnancy.

Adult↗
PubMed2004

A novel splice-site mutation in the common gamma chain (gammac) gene IL2RG results in X-linked severe combined immunodeficiency with an atypical NK+ phenotype.

Mutations in the gene encoding the common gamma chain (gammac) of interleukin receptors 2, 4, 7, 9, 15 and 21 result in X-linked severe combined immunodeficiency (SCID-X1). Classically, this disease is characterised by an absence of T and NK cells, and near normal numbers of functionally deficient B cells (B(+), T(-), NK(-) phenotype). Atypical phenotypes have also been described, but relatively little is known about the mechanisms by which the underlying mutations impair gammac-dependent interleukin receptor signalling to produce these disease variants. Here we describe a novel splice-site mutation resulting in the presence of near normal numbers of functionally deficient NK cells (B(+), T(-), NK(+) phenotype), in a SCID-X1 infant who was subsequently treated by gene therapy. The mutation, c.468+3A>C affecting the third base of intron 3 in the IL2RG gene, was shown to result in the production of two aberrantly spliced gammac mRNA species and reduction of correctly spliced message to trace levels, consistent with failure to detect gammac on the surface of B and NK cells by FACS analysis. The infant's genotype-phenotype correlation supports the hypothesis that interleukin 15 receptor-mediated signalling is preferentially retained as the amount of cell surface gammac becomes limiting. The possible implications for immunological reconstitution following gene therapy are also discussed.

Base Sequence↗
PubMed1995

Thrombopoietin, the c-mpl ligand, is a major regulator of platelet production.

Hematopoiesis is regulated by a family of glycoproteins, the hematopoietic growth factors. Although the cytokines that influence the late stages of granulopoiesis (granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor) or erythropoiesis (erythropoietin) have been identified, characterized, and cloned, and have been in clinical use since that late 1980's, the cytokine that stimulates thrombopoiesis had remained elusive. By using strategies based on the c-mpl receptor, several groups have recently succeeded in purifying and cloning thrombopoietin [1-6]. This study outlines the cloning and biological characterization in vitro and in vivo of thrombopoietin, a major regulator of platelet production.

Amino Acid Sequence↗
PubMed2001

Development of a novel helicase assay using electrochemiluminescence.

DNA helicases are ubiquitous enzymes involved in DNA replication, recombination, and repair. These enzymes use the energy of ATP to unwind duplex DNA. A common approach to measure DNA helicase activity utilizes electrophoresis to visualize product formation in a gel. The present study develops a rapid helicase assay based on electrochemiluminescence. The assay is adaptable to rapid high-throughput screening for chemical inhibitors. The assay is applied to Escherichia coli DnaB. Three other technologies have also been applied to DnaB and are compared to results of the electrochemiluminescence-based assay.

Base Sequence↗
PubMed1995

Comparisons of T cell receptor (TCR) V beta repertoires of lamina propria and peripheral blood lymphocytes with respect to frequency and oligoclonality.

The TCR repertoires of CD4+ and CD8+ lamina propria and peripheral blood lymphocytes (PBL) were compared with respect to V beta frequencies and oligoclonality. Disease-free colon specimens and paired peripheral blood samples were obtained from eight adult patients undergoing surgical resections for colorectal carcinoma. Mononuclear cell were isolated from the lamina propria and peripheral blood and separated into CD4+ and CD8+ cells by immunomagnetic adsorbtion. Analysis of V beta frequencies by qPCR revealed that PBL and lamina propria lymphocytes (LPL) from the same individual have very different repertoires, especially within the CD8+ population. Furthermore, CD8+, but not CD4+, LPL display extensive oligoclonality, similar to that which has previously been reported for CD8+ PBL. However, the oligoclonal receptors observed in CD8+ LPL are, in general, distinct from those observed in CD8+ PBL, and differ for each individual. These observations indicate that the TCR repertoires of LPL are as diverse as PBL, and suggest that LPL and PBL are normally exposed to different sets of antigens. In addition, these observations provide a baseline for examining the effects of various disease states and environmental insults on the LPL repertoire.

Base Sequence↗
PubMed1993

HIV-1 and HCV co-infected patients: detection of active viral expression using a nested polymerase chain reaction.

The aim of the present study was to determine, in a population of 70 HIV-1 infected patients with antibodies to HCV, the percentages of individuals with an active replication of HIV-1, HCV or both. During a one year follow-up of these patients at different stages of disease, blood samples were regularly collected for determination of transaminase, beta 2 microglobulin and CD4+ lymphocytes. Total RNAs were extracted from the sera, retrotranscribed with MoMuLV reverse transcriptase and nested PCR assays were carried out separately with sets of primers homologous to the 5' non-translated region of HCV and in HIV-1 gag. The amplified products were subjected to electrophoresis and observed under u.v. illumination after staining with ethidium bromide. For some samples, the identity of the amplified products was confirmed by Southern blotting by hybridization with enzyme-labelled probes. A total of 57% of the patients were found to produce HIV-1 RNA and 62% HCV RNA, while 34% produced both. HIV-1 RNA production was correlated with beta 2 microglobulin and CD4+ levels; active replication of HCV was correlated with hepatitis but not with CD4+ levels nor with HIV-1 RNA synthesis.

Base Sequence↗
PubMed1990

Graded cytopathogenicity of the human immunodeficiency virus (HIV) in the course of HIV infection.

To determine whether the biological variability of HIV-1 has any clinical significance, the highly variable cytopathogenicity of 153 HIV-1 strains, isolated from 119 hemophiliacs, was related to the number of CD4+ lymphocytes present in the patient's blood at the time of virus isolation. It was shown that the cytopathogenicity of the HIV-1 isolates was inversely correlated with the number of CD4+ lymphocytes. The highest CD4+ cell number were observed in 34 latently infected patients characterized by HIV seropositivity, failure of virus isolation, and detection of viral DNA by the polymerase chain reaction. Cytopathogenicity of the HIV-1 isolates was a reliable prognostic marker and correlated well with other less-sensitive prognostic parameters, including the detection of infectious virus and p24 antigen in the plasma, and the decline of p24 antibody in the serum. The results suggest that the viral isolates - if not subjected to extensive passage - represent in vivo variants selected from a heterogeneous viral population according to the particular immunological conditions of the host.

Base Sequence↗
PubMed1991

Patterns of antibody reactivity to selected human immunodeficiency virus type 1 (HIV-1) gp160 epitopes infected individuals grouped according to CD4+ cell levels.

We examined sera from 160 HIV-infected individuals for antibodies reactive to HIV-1 gp160 epitopes defined by seven synthetic peptides. Seropositive individuals were placed into three groups based upon levels of circulating CD4+ cells. These groups consisted of individuals with (1) more than 400 CD4+ cells, (2) 200-400 CD4+ cells, and (3) fewer than 200 CD4+ cells/mm3. The percentage of sera containing antibodies reactive with two immunodominant gp160 epitopes (a.a. 304-321 and 600-611) was unchanged between groups, regardless of CD4 cell numbers. The percentage of sera containing antibodies reactive with weakly immunogenic gp160 epitopes, such as those defined by peptides 425-448 and 846-860, declined in the groups as CD4 values decreased. Our results suggest that the patterns of antibody reactivity to gp160 epitopes change as CD4 levels decline. A narrowing of the humoral immune response to epitopes on the envelope of HIV-1 appears to occur with disease progression.

Acquired Immunodeficiency Syndrome↗
PubMed1992

Absence of selection of HIV-1 variants in vivo based on transcription/transactivation during progression to AIDS.

The activity of the human immunodeficiency virus type 1 (HIV-1) transactivation protagonists tat and TAR has been analyzed from sequential primary material. The sequences were amplified from uncultured peripheral blood mononuclear cells. Despite fluctuations within the tat and TAR quasispecies there was no obvious selection for a variant encoding more powerful transactivation components either in vivo or ex vivo, indicating that this system is not exploited during disease progression. The basal levels of the natural promoters were, depending on the cell line, two- to fourfold higher than that of the reference promoter, itself derived from ex vivo adapted HIV-1 Lai.

Acquired Immunodeficiency Syndrome↗
PubMed1992

Frequency of T cells specific for myelin basic protein and myelin proteolipid protein in blood and cerebrospinal fluid in multiple sclerosis.

T cell sensitization to two myelin components, myelin basic protein (MBP) and myelin proteolipid protein (PLP), may be important to the pathogenesis of multiple sclerosis (MS). Using the limiting dilution assay, we demonstrated that the blood of MS patients had an increased frequency of MBP-reactive T cells compared with normal subjects and patients with other neurological diseases (OND) and rheumatoid arthritis. There was no difference in T cell frequency to a synthetic peptide, PLP139-151, or Herpes simplex virus. Within cerebrospinal fluid (CSF), 37% of IL-2/IL-4-reactive T cell isolates from MS patients responded either to MBP or PLP139-151 while only 5% of similar isolates from OND patients responded to these myelin antigens. The mean relative frequency of MBP-reactive T cells within CSF from MS patients was significantly higher than that of OND patients (22 x 10(-5) cells versus 1 x 10(-5) cells) and was similar to that of MBP reactive T cells within the central nervous system of rats with experimental autoimmune encephalomyelitis. These results lend new support to the hypothesis that myelin-reactive T cells mediate disease in MS.

Amino Acid Sequence↗
PubMed2005

Experimental inoculation of beagle dogs with Ehrlichia species detected from Ixodes ovatus.

Three beagle dogs were inoculated with mice spleen/liver homogenate infected with Ehrlichia species detected from Ixodes ovatus (EIO) and one dog was used as a control. All three infected dogs did not show clinical signs of disease except for mild pyrexia throughout the 41-day study period. Splenomegaly was observed from Day 7 post-inoculation (p.i.) in two of the dogs. Hematological and biochemical abnormalities included mild thrombocytopenia, hypoproteinaemia, hypoalbuminaemia and increased C-reactive protein values. One of the dogs' splenic aspirate sample was PCR-positive for Ehrlichia Day 7 p.i. and another dogs' blood and bone marrow aspirate sample was PCR-positive Day 41 p.i. Sequence analysis of the PCR products showed 100% homology with the 16SrRNA partial gene sequence of Ehrlichia sp. HF565. Antibody titers to EIO were observed in all three experimentally infected dogs starting from the first week p.i. and cross-reactivity with Ehrlichia canis was detectable in one of the dogs starting Day 7 p.i. These data suggest that infection of dogs with EIO is possible, though is probably of low pathogenic importance. Cross-reactivity of EIO infected dog serum with E. canis raises the likelihood of false E. canis seropositive dogs.

Animals↗