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Biomedical subjects

C D Gregory

Publications and source records attributed to C D Gregory.

70 records · Page 4Linked to original sources
PubMed1988

Different patterns of Epstein-Barr virus gene expression and of cytotoxic T-cell recognition in B-cell lines infected with transforming (B95.8) or nontransforming (P3HR1) virus strains.

Epstein-Barr virus (EBV)-negative Burkitt's lymphoma (BL) cell lines have been converted to EBV genome positivity by in vitro infection with the transforming EBV strain B95.8 and with the nontransforming mutant strain P3HR1, which has a deletion in the gene encoding the nuclear antigen EBNA2. These B95.8- and P3HR1-converted lines have been compared for their patterns of expression of EBV latent genes (i.e., those viral genes constitutively expressed in all EBV-transformed lines of normal B-cell origin) and for their recognition by EBV-specific cytotoxic T lymphocytes (CTLs), in an effort to identify which latent gene products provide target antigens for the T-cell response. B95.8-converted lines on several different EBV-negative BL-cell backgrounds all showed detectable expression of the nuclear antigens EBNA1, EBNA2, and EBNA3 and of the latent membrane protein (LMP); such converts were also clearly recognized by EBV-specific CTL preparations with restriction through selected human leukocyte antigen (HLA) class I antigens on the target cell surface. The corresponding P3HR1-converted lines (lacking an EBNA2 gene) expressed EBNA1 and EBNA3 but, surprisingly, showed no detectable LMP; furthermore, these converts were not recognized by EBV-specific CTLs. Such differences in T-cell recognition were not due to any differences in expression of the relevant HLA-restricting determinants between the two types of convert, as shown by binding of specific monoclonal antibodies and by the susceptibility of both B95.8 and P3HR1 converts to allospecific CTLs directed against these same HLA molecules. The results suggest that in the normal infectious cycle, EBNA2 may be required for subsequent expression of LMP and that both EBNA2 and LMP (but not EBNA1 or EBNA3) may provide target antigens for the EBV-specific T-cell response.

Antigens, Viral↗
PubMed1987

Identification of a subset of normal B cells with a Burkitt's lymphoma (BL)-like phenotype.

Fresh biopsy cells from cases of Burkitt's lymphoma (BL) display a homogeneous cell surface phenotype. The cells were found to be reactive with the pan B cell marker B1, and consistently co-expressed the BL-associated glycolipid antigen, BLA, and the common acute lymphoblastic leukemia antigen, CALLA, but lacked the B cell "activation" antigens characteristically expressed on EB virus-transformed normal B cells. Microscopic and cell sorter analysis of cells isolated from a series of fresh normal tonsils have identified a subpopulation of normal B cells carrying the same cell surface markers. That BLA and CALLA could be co-expressed on individual B cells was demonstrated by two-color immunofluorescence (IF) of tonsils in suspension, and immunoperoxidase (IP) staining of serial tonsil sections. These BLA+, CALLA+, "activation" antigen- cells were further characterized as B1+, sIgM+, sIgD-, C3d/EB virus receptor+ and were susceptible to virus-induced transformation in vitro. IF studies on Percoll-fractionated tonsillar cell populations and direct examination of IP-stained tonsil semi-thin sections indicated that the BLA+, CALLA+ cells were localized in germinal centers. Their morphological characteristics matched those of BL cells, and their location within germinal centers was consistent both with the known phenotype of germinal center tonsillar B cells and with the description of BL as a proliferation of centroblasts. We suggest that this population of tonsillar germinal center B cells provides the normal counterpart of BL tumor cells.

Antibodies, Monoclonal↗
PubMed1987

Epstein-Barr virus-transformed human precursor B cell lines: altered growth phenotype of lines with germ-line or rearranged but nonexpressed heavy chain genes.

A series of lymphoblastoid cell lines (LCLs) have been established by in vitro infection of fetal bone marrow and fetal liver cells with Epstein-Barr virus (EBV). While most lines showed the usual mature B cell phenotype, a small proportion were cytoplasmic and surface immunoglobulin (Ig) heavy and light chain negative. Analysis of gene rearrangements indicated that the Ig- lines were either germ-line or nonproductively rearranged when probed for JH and were in germ-line configuration for C chi; no mu or chi mRNA could be detected in such cells. Precursor B cell lines were indistinguishable from their normal Ig+ counterparts in their expression of a wide variety of cell surface markers including "activation" antigens usually associated with the lymphoblastoid state; even the single LCL showing germ-line heavy and light chain genes expressed B lineage-specific cell surface antigens. However, the Ig- lines were distinct from their Ig+ counterparts in three important respects: (a) they grew much more slowly and achieved lower saturation densities, (b) they showed unusually high proportions (8-16%) of cells in EBV-productive cycle, and (c) they contained unusually high proportions (up to 40%) of cells expressing free joining (J) chain. These results suggest that precursor B cells differ in their response to the growth-transforming effects of EBV such that the virus-cell interaction in precursor B cell lines is inherently less stable than in conventional LCL. In particular there may be a greater movement of cells out of cycle and along the B cell maturation pathway. It is possible that such movement leads in individual cells either to virus replication or to a "sterile" plasmacytoid differentiation with J chain expression in the absence of Ig synthesis.

Antibody Formation↗
PubMed1987

Differences in B cell growth phenotype reflect novel patterns of Epstein-Barr virus latent gene expression in Burkitt's lymphoma cells.

Recently established Epstein-Barr virus (EBV)-positive Burkitt's lymphoma (BL) cell lines, carrying chromosomal translocations indicative of their malignant origin, have been monitored for their degree of in vitro progression towards a more 'lymphoblastoid' cell surface phenotype and growth pattern, and for their expression of three EBV latent gene products which are constitutively present in all virus-transformed normal lymphoblastoid cell lines (LCLs). BL cell lines which stably retained the original tumour biopsy phenotype on serial passage were all positive for the nuclear antigen EBNA 1 but did not express detectable amounts of two other 'transforming' proteins, EBNA 2 and the latent membrane protein (LMP). This novel pattern of EBV gene expression was also observed on direct analysis of BL biopsy tissue. All three viral proteins became detectable, however, in BL cell lines which had progressed towards a more LCL-like phenotype in vitro. This work establishes a link between B cell phenotype and the accompanying pattern of EBV latent gene expression, and identifies a novel type of EBV:cell interaction which may be unique to BL cells.

Antibodies, Monoclonal↗
PubMed1987

Viruses and cancer risks: outgrowth of Epstein-Barr virus-positive Burkitt's lymphoma in the immune host.

This work examines ways in which Epstein-Barr (EB) virus-positive Burkitt's lymphoma (BL) cells achieve outgrowth in vivo in the face of prevailing EB virus-specific cytotoxic T-cell surveillance. Earlier work has shown that some, but not all, BL cell lines in vitro are insensitive to virus-specific T-cell cytolysis and the present study identifies two mechanisms whereby the tumour cells might evade detection. First, BL-cell lines which stably retain the original tumour cell phenotype on serial passage in vitro show very low expression of two cell adhesion-related molecules, LFA-1 and ICAM 1, and are negative for a third such molecule, LFA-3; these molecules are thought to play a crucial role in the non antigen-dependent phase of effector: target cell conjugation which precedes antigen-specific recognition and target cell lysis. Secondly, those same BL cell lines display an unusually restricted pattern of EB virus latent gene expression with at least two potentially important target proteins for the T-cell response, namely EBNA 2 and LMP, not detectably expressed.

Animals↗
PubMed1987

Phenotypic heterogeneity and recycling capacity of natural killer cells in normal human pregnancy.

Natural killer (NK) cells have the ability to kill a variety of target cell types and the possibility that such cells could mount an effective attack on the developing fetus has not been discounted. The present study extends previous work showing that maternal NK reactivity against K562 target cells (TC) is reduced during pregnancy. Here we demonstrate using cytotoxicity assays at both the population and single cell level that, although depressed in number, maternal NK cells exhibiting the capacity to kill K562 TC are as lytically active in their ability to recycle and destroy multiple TC as NK cells from non-pregnant females. Moreover, two colour immunofluorescence analysis of the NK cell-associated markers Leu-7 and Leu-11b indicates that, in addition to a reduction in the absolute number of TC conjugate-forming cells, pregnant females present in their peripheral blood a larger proportion of TC-binding Leu-7+11- cells. These cells may be lytically immature. Small changes in NK cell profile and activity in maternal peripheral blood may be indicative of much more significant changes at the feto-maternal interface. It is, however, clear that pregnant females retain a population of highly active NK cells, thus minimising the possibility of immunocompromise.

Adult↗
PubMed1987

Cytotoxic activity and phenotypic analysis of natural killer cells in early normal human pregnancy.

Peripheral blood lymphocytes taken from healthy women planning a pregnancy and then at various intervals up to the 16th week of pregnancy were assayed for natural killer (NK) cell activity against K562 target cells both in a 51Cr-release assay and in a single cell cytotoxicity assay. Results indicated a depression in NK activity from the earliest stages of pregnancy. The target binding capacity of the effector cells remained unimpaired up to 16 weeks, but a significant reduction in the post-binding lytic potential was observed, which parallelled the drop in cytotoxicity as assayed by the 51Cr-release method. The ability of individual effector cells to recycle and kill multiple targets remained essentially unimpaired. Analysis of lymphocyte subpopulations using the monoclonal antibodies anti-Leu-7 and anti-Leu-11b, which recognize NK cell-associated antigens, showed a significant reduction in the proportion of the mature, lytically active Leu-7-11+ cells capable of both binding and lysing K562 target cells. The suggestion that the depression in cytotoxicity may be associated with the reduction in the Leu-7-11+ subpopulation is supported by the high correlation levels observed between the proportion of Leu-7-11+ cells and target cell lysis.

Antibodies, Monoclonal↗
PubMed1987

Epstein-Barr virus nuclear antigen 2 specifically induces expression of the B-cell activation antigen CD23.

Epstein-Barr virus (EBV) infection of EBV-negative Burkitt lymphoma (BL) cells induces some changes similar to those seen in normal B lymphocytes that have been growth transformed by EBV. The role of individual EBV genes in this process was evaluated by introducing each of the viral genes that are normally expressed in EBV growth-transformed and latently infected lymphoblasts into an EBV-negative BL cell line, using recombinant retrovirus-mediated transfer. Clones of cells were derived that stably express the EBV nuclear antigen 1 (EBNA-1), EBNA-2, EBNA-3, EBNA-leader protein, or EBV latent membrane protein (LMP). These were compared with control clones infected with the retrovirus vector. All 10 clones converted to EBNA-2 expression differed from control clones or clones expressing other EBV proteins by growth in tight clumps and by markedly increased expression of one particular surface marker of B-cell activation, CD23. Other activation antigens were unaffected by EBNA-2 expression, as were markers already expressed on the parent BL cell line, including BL markers (cALLA and BLA), proliferation markers (transferrin receptor and BK19.9), and cell adhesion-related molecules (LFA-1 and LFA-3). Increased CD23 expression in cells expressing EBNA-2 was apparent from monoclonal anti-CD23 antibody binding to the cell surface, from immunoprecipitation of the 45-kDa and 90-kDa CD23 proteins with monoclonal antibody, and from RNA blots probed with labeled CD23 DNA. The results indicate that EBNA-2 is a specific direct or indirect trans-activator of CD23. This establishes a link between an EBV gene and cell gene expression. Since CD23 has been implicated in the transduction of B-cell growth signals, its specific induction by EBNA-2 could be important in EBV induction of B-lymphocyte transformation.

Antigens, Viral↗
PubMed1986

Burkitt-like lymphoma in an English child: characterisation of tumour biopsy cells and of the derived tumour cell line.

An eight year old English boy presented with an abdominal undifferentiated 'Burkitt-like' lymphoma. Lymphoma cells from ascitic fluid were cultured on a human embryo fibroblast feeder layer and, after a short lag period, a cell line (DH-BL) was established which, like the original tumour, was both negative for the Epstein-Barr nuclear antigen (EBNA) and expressed a monoclonal pattern of surface immunoglobulin (alpha lambda). DH-BL also possessed the Burkitt-related 8:14 chromosome translocation in all metaphases analysed; no other chromosomal abnormalities were present. The cell surface phenotype of the original biopsy cells and the cultured tumour cells in early passage were investigated using a panel of monoclonal antibodies to B lineage-associated antigens. These antibodies had recently been used to characterise African 'endemic' Burkitt's lymphoma (BL) biopsy cells and their derived cell lines. The cell surface phenotype of this English EBNA negative Burkitt-like lymphoma biopsy was indistinguishable from that previously shown by biopsies of EBNA positive endemic BLs. It therefore appears that both the endemic and sporadic forms of BL, as illustrated by this case, may be derived from the same subset of progenitor cells.

Abdominal Neoplasms↗
PubMed1986

Endemic Burkitt's lymphoma: phenotypic analysis of tumor biopsy cells and of derived tumor cell lines.

Tumor cells from 10 patients with Epstein-Barr virus-positive endemic Burkitt's lymphoma (BL) have been examined for cell surface phenotype, both at the biopsy stage and during BL cell line outgrowth in vitro, the cultures being followed for up to 150 passages. In all 10 cases, the biopsy cells showed coexpression of the common acute lymphoblastic leukemia antigen (CALLA) and of the BL-associated glycolipid antigen (BLA) with no accompanying expression of several "lymphoblastoid" cell surface markers defined by selected monoclonal antibodies. During cell line establishment and in vitro passage, the individual BL cell lines showed different degrees of progression toward a more "lymphoblastoid" cell surface phenotype, some even losing CALLA and BLA expression while retaining the chromosomal translocations indicative of their malignant origin. This differential capacity for phenotypic progression in vitro explains much, if not all, of the heterogeneity of the BL cell phenotype apparent from many previous studies with panels of long-established lines. Such heterogeneity in vitro belies the true homogeneity of the tumor cell phenotype in vivo.

Antibodies, Monoclonal↗
PubMed1985

Natural killer cells in normal pregnancy: analysis using monoclonal antibodies and single-cell cytotoxicity assays.

Peripheral blood lymphocytes (nylon wool non-adherent) from healthy pregnant women and normal non-pregnant females were tested for natural killer (NK) cell-mediated cytotoxicity against K562 target cells both by 51Cr-release assay and single-cell cytotoxicity assay in agarose. The results indicated depression of NK cytotoxicity in pregnancy due to a decrease in the proportion of target-binding lymphocytes as well as a reduction in the lytic capacity of target-bound cells. The ability of active pregnancy-associated NK lymphocytes to recycle appeared to be unimpaired. Analysis of lymphocyte populations with monoclonal antibodies recognizing NK cell-associated antigens showed that the number of Leu-11+ lymphocytes was reduced in pregnancy. Enumeration of Leu-7+ cells and correlation of NK cell subpopulation data with cytotoxicity assay data suggest that pregnancy is associated with a reduction in the number of mature NK cells and probably also an inhibition of post-binding lytic activity.

Adult↗
PubMed1985

Cytotoxic reactivity of human natural killer (NK) cells during normal pregnancy: a longitudinal study.

A longitudinal analysis of natural killer (NK) cell-mediated cytotoxicity of maternal peripheral blood lymphocytes was carried out at various stages (16-36 weeks) of normal human pregnancy, within 1 week following delivery and up to 40 weeks post-partum. NK cell-mediated lysis of K562 target cells (TC) in short term 51Cr-release assays was significantly depressed throughout pregnancy, returning to control levels 9-40 weeks post-partum. Natural cytotoxicity of unseparated maternal peripheral blood was also substantially depressed at all stages of pregnancy and, in addition, remained impaired in the late post-partum period. Longitudinal enumeration of Leu 3a+ and Leu 2a+ lymphocytes indicated an absence of helper/suppressor T-cell imbalance during pregnancy, and analysis of Leu 7+ cells showed no difference in population sizes between control and pregnancy groups. Comparison of blood and lymphocyte cytotoxicity in control and pregnancy samples suggested a complex regulation of NK reactivity during pregnancy. The potential role in vivo of both plasma-associated and cellular regulatory elements is discussed.

Adolescent↗
PubMed1984

Quantitative ultrastructure of cytolytic lymphocytes mediating allograft rejection in the mouse. I. Cellular alterations in T lymphocytes during specific target cell lysis.

A quantitative ultrastructural analysis of cytolytic T lymphocytes (CTL) is presented which allows both the distinction of these cells from normal T lymphocytes and permits the demonstration of ultrastructural alterations of putative CTL following interaction with target cells (TC). Alloreactive CTL were generated in C57BL/10 mice receiving intraperitoneal fibroblastic allografts and target-binding splenic lymphocytes (TBSL) were concentrated by specific immunoadsorption on fibroblast monolayers. TBSL were subjected to ultrastructural quantification either at the onset of TC interaction or following 30 or 60 min incubation at 37 degrees C. By means of simple stereological relationships it was shown that, in comparison with normal, non-cytolytic splenic T lymphocytes, TBSL were slightly larger cells, displaying around 60% more cytoplasm, a similarly-sized nucleus and approximately triple the volume of Golgi apparatus. During the first 30 min of interaction with TC, the target binding surface of the TBSL plasma membrane decreased in area. This change was accompanied by a polarization of the TBSL towards the target. Incubation of lymphocytes with TC for a further 30 min resulted in a general polarization of lymphocytic cellular constituents away from the TC. These results were only attainable by objective quantitative analysis and are discussed in relation to possible mechanisms of CTL-mediated lysis.

Animals↗
PubMed1984

Large agranular lymphocytes: early non-specific effector cells in allograft rejection in the mouse.

The cytolytic reactivity and ultrastructure of centrally-reactive and allograft-infiltrating lymphocyte populations was investigated in a murine peritoneal allograft system. Animals sensitized with a single intraperitoneal dose of allogeneic L929 fibroblasts generated a population of splenic cytolytic T cells maximally reactive 10 days after immunization. Sensitized splenic lymphocytes, isolated by immunoadsorption on L929 monolayers, were ultrastructurally classified as mature small lymphocytes. At the graft site, cytolytic non-T lymphoid cells displaying the ability to kill K562 target cells, were demonstrable between 4 and 6 days after sensitization. Six-day peritoneal lymphocyte populations were found to contain both cytolytic T cells (L929 killers) and highly reactive K562 killers. Immunoadsorption and cold target competition studies indicated that the K562 killer cells were able to recognize both K562 and L929 targets. K562 target-binding cells appeared to be ultrastructurally immature and were designated 'large agranular lymphocytes'. The role of cytolytic non-T cells in rejecting allografts is discussed.

Animals↗
PubMed1979

On the problem of counting and sizing mitochondria: a general reappraisal based on ultrastructural studies of mammalian lymphocytes.

The authors ask the question whether the parameters "numbers" and "volume" are suitable for the morphometric analysis of mitochondria. In several types of cell, irregularity of mitochondrial shape makes it technically difficult, if not impossible, to obtain reliable stereological estimates of mean organelle volume or number per unit volume. Of more fundamental concern is whether number of mitochondria per cell is of any real value as a structural correlate of respiratory potential and hence as a measure of cell function. Alternative parameters might serve better for this purpose. Though the problem is illustrated by reference to quantitative studies of lymphocytes, it is also pertinent to the investigation of many other cell types.

Animals↗