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Biomedical subjects

L K Borysiewicz

Publications and source records attributed to L K Borysiewicz.

At least 37 records · Page 2Linked to original sources
PubMed1998

Vaccinia virus-induced changes in cytokine-regulated acute phase plasma protein synthesis by hepatoma cells.

Human hepatoma cell line, HepG2, has been infected with vaccinia virus and synthesis of plasma proteins was determined by electroimmunoassay and corresponding mRNA's measured by Northern blotting. The inhibitory effect of the virus was dose- and time-dependent. Electrophoretic mobility shift assay revealed a decrease in C/EBP binding activities in nuclear extracts isolated from the infected hepatoma cells. Supershift analysis of the C/EBP isoforms showed alpha and beta subunit involvement in DNA binding. The treatment of the cells with interleukin-1, interleukin-6, and dexamethasone at the initial stage of infection appears to delay the virally induced inhibition of host cell protein synthesis. Thus, possible "protective" role of the acute phase cytokines in viral infection is proposed.

Acute-Phase Proteins↗
PubMed1997

Human papillomavirus-specific cytotoxic T lymphocytes in patients with cervical intraepithelial neoplasia grade III.

Human papillomaviruses (HPVs), especially types 16 and 18, are strongly associated with the development of cervical intraepithelial neoplasia (CIN) and invasive carcinoma. The HPV E6 and E7 proteins are expressed constitutively in the majority of CIN lesions and carcinomas. Therefore, they are targets for the immune response against HPV and candidates for active immunotherapy. We have previously detected HPV-specific CTLs from the peripheral blood mononuclear cells from a cervical cancer patient following immunization with a recombinant vaccinia using in vitro restimulation with adenovirus recombinants expressing HPV16 or HPV18 E6/E7 fusion proteins. In this study, we used a similar protocol to determine the prevalence of CTL responses against HPV16 and HPV18 E6/E7 in the peripheral blood mononuclear cells from 10 CIN III and 10 normal subjects. HPV-specific CTL responses were detected in 6 of 10 CIN III subjects. These CTL lines recognized HPV16 E6/E7 proteins presented by at least three MHC class 1 HLA alleles and by HPV-transformed CaSki cells. No HPV-specific CTLs were detected in normal subjects. This study demonstrates the presence of naturally occurring HPV-specific memory CTLs in a majority of CIN III patients and provides an approach for further study of their role in modulating cervical malignancy.

Adult↗
PubMed1997

Infiltration of cervical cancer tissue with human papillomavirus-specific cytotoxic T-lymphocytes.

CTLs specific for high-risk human papillomaviruses (HPVs) have recently been found in the peripheral blood of cervical cancer patients. Although cell-mediated immunity is thought to be important in the control of HPV infection, the functional relevance and site of activation of HPV-specific CTLs are unclear. We identified HLA-A*0201-restricted HPV-16 E7 peptide-specific CTLs in the peripheral blood (four of five patients), draining lymph nodes (three of four patients) and tumors (one of three patients) of cervical cancer patients. In four of four cancer patients, the frequency of CTLs specific for a recombinant vaccinia virus expressing HPV-16 and -18 E6/E7 gene products was found to be higher in tumors and lymph nodes compared with that of peripheral blood. HPV-specific CTLs were not identified in any of seven healthy controls, but primary responses could be generated by peptide-pulsed dendritic cells (four of four controls). In a non-HLA-A*0201 subject with invasive carcinoma, other HLA alleles also presented HPV antigens. This is the first demonstration that virus-specific CTLs infiltrate the virus-associated tumor, where they may play an important role in restricting disease progression.

Adult↗
PubMed1997

Detection and biochemical characteristics of the receptor for complexes of soluble CD14 and bacterial lipopolysaccharide.

Soluble CD14 (sCD14) has been found to bind LPS and mediate LPS activation of several cell types. It has been postulated that sCD14-LPS complexes induce cell responses by interacting with a cell surface structure, which, in turn, triggers cell activation. There has been no biochemical evidence, however, for a direct interaction of sCD14 with a cell surface structure, and the putative receptor has not been identified. To rigorously test this hypothesis, we studied the interaction of human rsCD14 with cells in the absence of serum and in the presence and the absence of LPS. We found 1) there was specific and saturable binding of 125I-sCD14, indicative of a typical receptor-ligand interaction, to several cell types, including endothelial cells, epithelial cells, astrocytes, and human monocytes; 2) specific binding to all the cell types and IL-6 induction in membrane-bound CD14 (mCD14)-negative cells occurred only when both sCD14 and LPS were present; 3) competitive displacement experiments of 125I-sCD14 binding to astrocytes and Scatchard plots revealed a binding of high affinity (Kd = 3.3 +/- 0.4 nM) and approximately 25,000 single class binding sites/cell; 4) the steady state for the association of 125I-sCD14 was obtained after 180-200 min; 5) chemical cross-linking experiments revealed the association of sCD14 with a binding structure of approximately 216 kDa; 6) binding of 125I-sCD14 to CD14-expressing cell transfectants was about 50% lower than that to nontransfected cells. Maximal binding, however, was recovered after removing mCD14, suggesting that the sCD14-LPS receptor may also interact with mCD14. These results provide direct biochemical evidence for the existence of a cell surface signal-mediating binding structure for LPS-bearing sCD14 and suggest that this structure may represent the signaling unit of the postulated multimeric LPS receptor in mCD14-bearing cells.

Animals↗
PubMed1997

CD8high+ (CD57+) T cells in patients with rheumatoid arthritis.

OBJECTIVE: To investigate the development and T cell receptor (TCR) usage of CD8+, CD57+ T cells in rheumatoid arthritis (RA) patients. METHODS: Three-color flow cytometry using monoclonal antibodies (MAb) to CD8, CD57 and different TCR V beta gene products. RESULTS: The proportion of CD8+ T cells expressing CD57 (CD57/CD8) was significantly higher in RA patients compared with age-matched controls. Expanded TCR V beta populations were more frequent, and were found in both RA patient-derived CD8high+ (CD57+) and CD8+, CD57- populations. TCR V beta 5+ and TCR V beta 13+ expansions were present at high frequency (5 of 26 and 7 of 26, respectively). TCR V beta expansions in CD8high+ (CD57+) lymphocytes from RA patients were significantly larger than those in age-matched controls (expansion index 2.38 +/- 0.28, n = 41 and 1.63 +/- 0.09, n = 32, respectively), and were stable over time. CONCLUSION: RA leads to an increase in the frequency of expanded CD8+ T cell subsets expressing selected TCR, due to expansion of TCR V beta + populations in CD8high+ (CD57+) T cells. Their restricted TCR usage suggests potential specificity for RA antigens and, therefore, a potential role in the pathogenesis of RA.

Aged↗
PubMed1997

Consequences of live poliovirus vaccine administration in chronic fatigue syndrome.

The effect of live oral polio virus vaccination on chronic fatigue syndrome (CFS) patients was examined in a double-blind study. CFS patients were allocated randomly to placebo (N = 7) or vaccine (N = 7) conditions. All controls subjects received the vaccine (9). Vaccine administration was not associated with clinical exacerbation of CFS. However, objective responses to the vaccine revealed differences between patients and controls: increased poliovirus isolation, earlier peak proliferative responses, lower T-cell subsets on certain days post vaccination and a trend for reduced gamma-interferon in the CFS-vaccine group. Polio vaccination was not found to be clinically contraindicated in CFS patients, however, there was evidence of altered immune reactivity and virus clearance.

Adult↗
PubMed1997

Development of a model for cytomegalovirus infection of oligodendrocytes.

The well-characterized human oligodendroglioma (HOG) cell line, cells of which resemble immature oligodendrocytes, was used to investigate the level of permissiveness to human cytomegalovirus (CMV) infection. Expression of CMV genes was incomplete following exposure of HOG cells to CMV, in contrast with results observed with the astroglioma cell line U373-MG (used as a positive control). However, treatment with phorbol 12-myristate 13-acetate (PMA) or with dibutryl cAMP (dbcAMP) plus the phosphatase inhibitor 1-isobutyl-3,3-methyl xanthine (IBMX) rendered the HOG cells fully permissive to CMV; down-regulation of HLA class I and production of virions were only observed under these conditions. In contrast to the findings seen with the HOG cell line, treatment of U373-MG cells with dbcAMP/IBMX or PMA did not interfere with CMV-induced down-regulation of HLA class I. However, these chemical stimulators reduced virus production in U373-MG cells by 30 and 70%, respectively.

Animals↗
PubMed1996

A recombinant vaccinia virus encoding human papillomavirus types 16 and 18, E6 and E7 proteins as immunotherapy for cervical cancer.

BACKGROUND: Human papillomavirus (HPV) infection, especially with type 16 or 18, is associated with cervical cancer. Two HPV proteins, E6 and E7, are consistently expressed in tumour cells. The objectives of the study were to examine the clinical and environmental safety and immunogenicity in the first clinical trial of a live recombinant vaccinia virus expressing the E6 and E7 proteins of HPV 16 and 18 (TA-HPV). METHODS: The study was an open label phase I/II trial in eight patients with late stage cervical cancer. The patients were vaccinated with a single dose of TA-HPV and kept in strict isolation to monitor local and systemic side-effects, environmental spread, and anti-E6/E7 immune responses. FINDINGS: Vaccination resulted in no significant clinical side-effects and there was no environmental contamination by live TA-HPV. Each patient mounted an antivaccinia antibody response and three of the eight patients developed an HPV-specific antibody response that could be ascribed to the vaccination. HPV-specific cytotoxic T lymphocytes, the effector mechanism most likely to be of therapeutic benefit, were detected in one of three evaluable patients. INTERPRETATION: Further studies to investigate the use ot TA-HPV for immunotherapy of cervical cancer are warranted.

Adult↗
PubMed1996

Cytotoxic T cell immunity to human cytomegalovirus glycoprotein B.

Human cytomegalovirus (HCMV) is associated with significant morbidity and mortality following immunosuppression and in pregnancy. HCMV infection may be accompanied by acute disease but persists asymptomatically. Cytotoxic T lymphocytes (CTL) appear to be an important immune effector mechanism in maintaining the normal host-virus equilibrium. Glycoprotein B may be an important target for future subunit vaccines as it has been found to elicit both neutralising antibody and CTL responses. We therefore studied the ability of normal asymptomatic HCMV-seropositive individuals and women throughout pregnancy to determine the presence of HCMV and gB-specific CTL responses. CTL effector cells were induced by stimulation of peripheral blood mononuclear cells (PBMC) with AD169 HCMV-infected cells and gB-specific CTL were identified using chromium labeled, vac.gB-infected cells. In 7 HCMV-seropositive individuals, HCMV-specific CTL were identified. Three of the 7 individuals which lysed HCMV-infected cells lysed vac.gB-infected B cells. However, vac.gB-infected autologous fibroblasts, which only present MHC class I, were not killed. Using MHC class I single allele targets, no specific lytic response was observed, suggesting a MHC class II restricted CTL response. Flow cytometric analysis showed the gB-specific effector cell phenotype to be CD3+, CD4+, CD8-. In conclusion, a gB-specific CTL lytic response was identified in seropositive individuals which in most cases was MHC class II-restricted.

Adult↗
PubMed1996

A malignant astrocytoma in a patient with AIDS: a possible association between astrocytomas and HIV infection.

We describe a malignant astrocytoma in a patient with AIDS confirmed at autopsy. The object of this report is to draw attention to a further example of an unusual tumour associated with HIV infection. This is the tenth case in the world literature and it is possible that this tumour is represented with a higher than expected frequency in this group of the population. This case and others add malignant astrocytoma to the differential diagnosis of an intracranial space occupying lesion in an AIDS patient.

Acquired Immunodeficiency Syndrome↗
PubMed1996

Construction and characterisation of a recombinant vaccinia virus expressing human papillomavirus proteins for immunotherapy of cervical cancer.

The presence and consistent expression of the genes encoding the human papillomavirus (HPV) E6 and E7 proteins in the great majority of cervical tumours presents the opportunity for an immunotherapeutic approach for control of the disease. This report describes the construction and characterisation of a recombinant vaccinia virus designed to express modified forms of the E6 and E7 proteins from HPV16 and HPV18, the viruses most commonly associated with cervical cancer. The recombinant virus (designated TA-HPV) was based on the Wyeth vaccine strain of vaccinia, and was shown to express the desired gene products. Studies in mice indicated that the recombinant virus was less neurovirulent than the parental virus and was capable of inducing an HPV-specific CTL response. This pre-clinical evaluation has provided a basis for the initiation of human trials in cervical cancer patients.

Amino Acid Sequence↗
PubMed1995

CD8highCD57+ T lymphocytes in normal, healthy individuals are oligoclonal and respond to human cytomegalovirus.

CD8+CD57+ lymphocytes from normal peripheral blood are divided into T cells expressing high levels of CD8 and NK cells expressing low levels of surface CD8. Increased numbers of CD8highCD57+ T cells correlate with previous exposure to human cytomegalovirus (HCMV) infection, but no virus-specific function or function for CD8highCD57+ cells has been recorded. We have studied the TCR repertoire and responses of the CD8highCD57+ population induced by virus-infected fibroblasts in healthy individuals. Using three-color flow cytometry of PBL, we detected restricted TCRBV usage in the CD8highCD57+ subset of 11/15 subjects, compared with 1/15 in the CD8+, CD57- subset. The results of anchored PCR and sequencing also showed oligoclonality of TCR; 40-70% of CD8highCD57+ lymphocytes (10-20% of total CD8+ T cells) were derived from single clones. Such expansions were stable with time and detected in one subject over a 2-yr period. Functionally, CD8highCD57+ lymphocytes proliferated strongly to HCMV-, but not HSV-, VZV-, or influenza-infected fibroblasts in an MHC-unrestricted manner in vitro, including preferential augmentation of particular in vivo oligoclonally expanded subpopulations. HCMV-specific MHC-restricted CTL were detected, but limiting dilution analysis showed that these were a minority (< 10%) and not the oligoclonal subsets. In contrast, depletion of oligoclonally expanded CD8highCD57+ subpopulations, resulted in the increase of HCMV-specific CTL, suggesting functional heterogeneity in these cells.

Adult↗
PubMed1995

Human HLA-A0201-restricted cytotoxic T lymphocyte recognition of influenza A is dominated by T cells bearing the V beta 17 gene segment.

The major histocompatibility complex class I-restricted cytotoxic T lymphocyte (CTL) response is important in the clearance of viral infections in humans. After influenza A infection, a peptide from the matrix protein, M58-66, is presented in the context of the MHC allele HLA-A0201 and the resulting CTL response is detectable in most HLA-A0201 subjects. An initial study suggested that M58-66-specific CTL clones show conserved T cell receptor (TCR) alpha and beta gene segments. We have addressed the significance of this observation by determining the expression of V beta 17 during the development of M58-66-specific CTL lines in 21 unrelated HLA-A0201 subjects, and analyzing TCR usage by M58-66-specific CTL clones. TCR V beta 17 was the dominant V beta segment used and CD8 V beta 17 expansion correlated with M58-66-specific lysis. Limiting dilution analysis from five subjects showed the M58-66 CTL precursor frequency to vary between 1/54,000 and less than 1/250,000, and that up to 85% of the matrix peptide (M58-66)-specific CTL used the V beta 17 gene segment. The M58-66 specific CTL response was dependent on previous viral exposure and specific V beta 17 expansion, as it was not found in cord blood, despite a readily expandable V beta 17+ CD8+ T cell subpopulation. Sequence analysis of 38 M58-66-specific V beta 17 transcripts from 13 subjects revealed extensive conservation in the CDR3 region including conservation of an arginine-serine motif. To test the dependence of this CTL response on the V beta 17 gene segment, peripheral blood lymphocytes were depleted of CD8+ TCR V beta 17+ cells, before the generation of M58-66-specific CTL. In most cases such depletion blocked or severely reduced the generation of the M58-66-specific response, and under limiting dilution conditions could abolish M58-66-specific CTL precursors. These studies reveal the dependence of this natural human immune response on a particular TCR gene segment.

Alleles↗
PubMed1995

Gene therapy and viral vaccination: the interface.

Live viral vaccines have had a major impact on the incidence of acute virus infections world-wide. Virus infections recognised as future vaccine targets will require a modified approach based on a detailed understanding of the immunobiology of specific infections combined with the application of new technologies designed to generate specific and appropriate protective immunity. A similar vector technology directed at in vivo gene delivery is currently being exploited both for gene therapy and vaccination. The induction of an immune response to an expressed transgene represents a potential hazard for a gene therapy protocol but is the object of a vaccine strategy. In vivo gene delivery using replication-competent or replication-deficient viral vector systems and by direct transfer of naked DNA can generate an effective humoral, secretory and cell-mediated immune response to expressed transgenes.

Adenoviridae↗
PubMed1995

The role of CD8+, CD57+ cells in human cytomegalovirus and other viral infections.

Peripheral blood lymphocytes expressing CD8 and CD57 determinants are a small (1-15%) subset in healthy humans. CD8+, CD57+ peripheral blood lymphocytes may be divided by the level of CD8 expression, into CD8+high (CD57+) T-cells and CD8+low (CD57+) natural killer (NK) cells. CD8+high (CD57+) T-cell numbers are increased in human cytomegalovirus (HCMV)-seropositive subjects, and there is substantial evidence that HCMV is integral in the development of this subset in health and disease. Furthermore, the CD8+high (CD57+) subset is clonally derived, expressing a limited range of T-cell receptors, and are therefore likely to have restricted antigen specificity. Functionally, CD8+low(CD57+) cells exhibit NK activity, while CD8+high(CD57+) T-cells from healthy subjects mediate contact-dependent suppression in several in vitro systems including: (i) pokeweed mitogen-induced proliferation and immunoglobulin synthesis, and (ii) generation of antiviral MHC-restricted cytotoxic T-lymphocytes. This is distinct from the nonspecific, soluble factor-mediated suppression exhibited from a phenotypically similar subset in human immunodeficiency virus (HIV) and bone marrow transplant recipients. This suggests an important immunoregulatory, suppressive role for CD8+high(CD57+) T-cells that may be potentiated by HCMV and altered in diseases associated with higher numbers of this subset including HIV, allograft recipients and rheumatoid arthritis.

Biomarkers↗
PubMed1994

CD8high (CD57+) T cells in normal, healthy individuals specifically suppress the generation of cytotoxic T lymphocytes to Epstein-Barr virus-transformed B cell lines.

We have previously identified two subsets of CD8+, CD57+ lymphocytes in normal peripheral blood: i) T cells expressing high levels [CD8high(CD57+)] and ii) natural killer cells expressing low levels of surface CD8[CD8low(CD57+)]. We investigated the cytotoxic and suppressive function of CD8high(CD57+) T lymphocytes from normal, healthy individuals using standard chromium-release assays and limiting dilution analysis. In normal, healthy subjects, this cell subset suppressed the generation of cytotoxic T lymphocytes (CTL) to autologous, Epstein-Barr virus (EBV)-transformed B cell lines (BCL). Depletion of CD8high(CD57+) T lymphocytes from peripheral blood mononuclear cells (PBMC) resulted in a three- to sevenfold rise in CTL precursor frequency to autologous EBV-transformed BCL, but not allogeneic PBMC or BCL by LDA. Replacement of CD8high(CD57+) T lymphocytes in limiting dilution cultures led to the dose-dependent suppression of EBV-specific, but not allogeneic, CTL generation. Supernatant from CD8high(CD57+) T lymphocytes cultured with autologous BCL did not exhibit suppression, suggesting that soluble factors were not responsible. As CD8high(CD57+) T lymphocytes did not, themselves, exhibit cytotoxicity against autologous BCL, removal of BCL stimulator cells in co-culture was not the mechanism of suppression. Furthermore, while the CD8high(CD57+) T lymphocytes from healthy subjects suppressed the generation of CTL to autologous BCL, they did not suppress the cytotoxic activity of established mixed lymphocyte reactions or peptide-specific CTL clones, as has been reported in bone marrow transplant recipients and human immunodeficiency virus patients. This suggests that CD8high(CD57+) T lymphocytes from healthy subjects suppress the generation of, rather than killing by, CTL in a contact-dependent manner. To our knowledge, this is the first identification of a phenotypically distinct subset of human CD8+ T cells that can suppress generation of antigen-specific major histocompatibility complex class I-restricted CTL.

Adult↗