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F H Bach

Publications and source records attributed to F H Bach.

At least 235 records · Page 13Linked to original sources
PubMed1988

Molecular studies of a rare DR2/LD-5a/DQw3 HLA class II haplotype. Multiple genetic mechanisms in the generation of polymorphic HLA class II genes.

A cDNA library was constructed from a homozygous B lymphoblastoid cell line (REM) obtained from an individual of a long isolated American Indian tribe, the Warao. The REM cell line expresses serologically defined determinants, DR2 and DQw3, and the T lymphocyte-defined (Dw/LD) specificity, LD-5a. T cells can recognize differences between FJO (a DR2/DQw1 cell line that expresses the Dw specificity MN2) and REM for both DR and DQ molecules. cNDA clones encoding the polymorphic DR beta 1-, DR beta 2-, DQ beta-, and DQ alpha-chains were sequenced and compared with other DR and DQ gene sequences. The DR beta 1-sequence of REM is identical to the DR beta 1-sequence of FJO; the DR beta 2-sequence is also identical to that of FJO except for one amino acid difference at position 67 in the polymorphic first domain (Leu in REM, Phe in FJO) due to a single point mutation. The DQ beta-sequence is identical to that of DR4/DQw3 haplotype; the DQ alpha is different from the DQ alpha of DR4/DQw3 haplotype and identical to the DQ alpha of both the DR3/DQw2 haplotype of a Raji cell line and the DR5/DQw3 haplotype in deduced amino acid sequence. Taken together, these findings suggest that: 1) a single amino acid difference (position 67) in the third hypervariable region of the first domain of the DR beta 2-chain in the DR2 haplotype is apparently sufficient for stimulating T cell responses; 2) the DQw3 serologic specificity may be defined mainly by the DQ beta-rather than DQ alpha-chain; and 3) multiple genetic events have probably occurred to generate the rarely found REM (DR2/LD-5a/DQw3) haplotype.

Amino Acid Sequence↗
PubMed1988

Augmentation of cell number and LAK activity in peripheral blood mononuclear cells activated with anti-CD3 and interleukin-2. Preliminary results in children with acute lymphocytic leukemia and neuroblastoma.

A wide variety of human cancers currently have no effective treatment and are potential targets for lymphokine-activated killer (LAK) cellular immunotherapy. Relapsed acute lymphocytic leukemia (ALL) and neuroblastoma are two of the major therapeutic challenges in pediatric oncology today. However, one problem which makes LAK immunotherapy in children particularly difficult is obtaining the large numbers of cells required. Present adult therapeutic LAK protocols have utilized short-term (5 day) cultures of interleukin-2 (IL2)-activated cells which are initially obtained from leukopheresis. Since routine use of this procedure in small children is not practical, we have investigated a different approach to obtain increased cell numbers by activation of peripheral blood mononuclear cells with OKT3, a mitogenic anti-CD3 monoclonal antibody, and IL2. Cell growth and LAK activity in OKT3 + IL2-activated cultures were compared to cultures activated with IL2 alone in 2 children with relapsed ALL and 2 children with stage IV neuroblastoma. OKT3 + IL2-activated cultures had marked increases in cell number: after 14 days the OKT3 + IL2-activated cultures yielded an approximately 500-fold increase in cell number compared to a 7-fold increase for cultures activated with IL2 alone. In vitro 51Cr release assays were used to estimate LAK activity of the cultures at 7 and 14 days. When tested against HL60, a natural killer (NK)-resistant tumor cell line, not only were total cytolytic units greatly increased in OKT3 + IL2-stimulated cultures by lytic activity on a per cell basis (lytic units/1 x 10(6) cells) had also markedly increased on day 14 of culture. Phenotypic analysis demonstrated that 80% to 90% of cells in OKT3 + IL2-stimulated cultures were CD3 + T cells. Variable low percentages of CD16 + NK cells were seen in these cultures. In summary, OKT3 + IL2 activation resulted in a large increase in cell yield and the development of high level LAK activity using peripheral blood mononuclear cells from children with cancer. This approach may facilitate the utilization of increased cell numbers in future adoptive immunotherapy protocols, especially in pediatric patients.

Adjuvants, Immunologic↗
PubMed1988

Large granular lymphocyte proliferation: an analysis of T-cell receptor gene arrangement and expression and the effect of in vitro culture with inducing agents.

The status of the T cell receptor beta and gamma genes in natural killer (NK) cells was investigated in two patients with a marked expansion of CD2+, CD3- NK cells. Both genes were found to be in the germline state. The T alpha and complete T beta gene transcripts were not detected, but a 1.0-kilobase T beta gene transcript could be demonstrated at low levels in freshly isolated cells and at a much higher level in interleukin-2 (IL-2)-cultured cells. The transcript coding for the delta chain of the CD3 complex was also absent. These cells were cultured in IL-2 with or without the addition of the differentiation-inducing agents: retinoic acid, N,N-hexamethylene bisacetamide, and sodium butyrate. The cultured cells retained their NK activity except in culture with sodium butyrate at greater than or equal to 1 mmol/L. Expression of CD3 or other T cell surface markers by the NK cells was not observed in these cultures. Either CD2+, CD3- NK cells are derived from a non-T lineage, or they have diverged from the T cell lineage earlier than the stage of T gamma gene rearrangement and CD3 delta chain expression; they are refractory to many induction signals in undergoing further T cell differentiation.

Acetamides↗
PubMed1988

Oligonucleotide probing. Applications to HLA typing.

Human leukocyte antigen typing is currently being done by both serologic and cellular techniques. It is this methodology that forms the basis for organ transplantation, since HLA antigens are the major proteins involved in graft rejection. Matching organ graft donors with recipients at these polymorphic HLA loci clearly increases graft survival in most transplant systems. However, the serologic and cellular techniques have shortcomings, including the problem of standardizing antisera, the need for technologists with expertise in T-cell cloning, and the failure to work successfully with some cells (eg, leukemic blasts). We have begun to develop a more rapid, sensitive typing method using molecular biological techniques to tissue-type cells. Using oligonucleotide probes, we can now identify HLA-D region alleles (which may differ by only a single nucleotide) that can only be differentiated by a highly technical, lengthy cellular technique. Oligonucleotide probing not only provides the basis for a rapid method of HLA typing, which in all likelihood will become generally applicable as additional sequence information becomes available, but also allows definition of a gene transcript from an individual locus rather than the phenotypic results provided by current HLA typing techniques.

Alleles↗
PubMed1987

Stepwise activation of T cells. Role of the calcium ionophore A23187.

The calcium ionophore A23187, at a concentration of 1 microgram/ml, is able to stimulate proliferation of freshly isolated peripheral blood lymphocytes, CD4+-enriched cells, or CD8+-enriched cells as measured by [3H]thymidine incorporation. This proliferation is accompanied by an increase in interleukin 2 (IL-2) receptor expression but not by a detectable up-regulation in (IL-2) production or the development of cytotoxicity. Proliferation can be blocked by anti-CD3, CD4, or CD8 monoclonal antibodies, but not by anti-Tac. If CD8+-enriched cells are activated for 3 days with A23187 and the blasts present on day 3 are sorted and returned to culture, they rapidly develop cytolytic activity in the presence of recombinant IL-2 but not recombinant interferon-gamma. CD4+ enriched cells, after activation with A23187, do not become cytotoxic in the presence of either recombinant IL-2 or recombinant interferon-gamma. These findings permit study of the stepwise maturation of T cells in this alternative pathway by using "minimal signals" that do not, by themselves and as used in these studies, stimulate precursor Tc to mature to full effector cytotoxic function. These findings are consistent with the model that A23187 drives T cells only part way along a pathway of maturation and that an additional second signal must be given to effect maturation of cytotoxic status.

Antibodies, Monoclonal↗
PubMed1987

Current concepts: immunology. Transplantation immunology.

The MHC is of overwhelming importance in determining the fate of an allograft. Over the past 10 years, our understanding of the serology of this locus, its role in cellular immunity, its biochemistry, and most recently, its molecular biology has increased enormously. Genes for all the known loci, both Class I and Class II, have been cloned and sequenced, and their evolution and function are therefore now more amenable to investigation at the molecular level. Concomitantly, an understanding of the different populations of immune cells involved in the recognition of the antigens encoded by these loci has permitted a more precise immunologic description of the rejection response. It is now clear that recognition of both Class I and Class II allogeneic differences by both helper and cytotoxic T cells has an important role in graft rejection. The goal of transplantation biologists is to take advantage of this increased understanding in order to design new techniques for specifically modifying the immune response to MHC antigens and achieving specific tolerance. Some exciting approaches are already being studied, including specific modifications of both the antigen and the receptor arms of the immune response. It is hoped that these approaches will have new clinical applications in the near future.

Animals↗
PubMed1987

HLA class II restriction of T helper cell response to cytomegalovirus (CMV). I. Immunogenetic control of restriction.

All three HLA class II families (DR, DQ, and DP) are involved in restriction of helper T cell (Th) recognition of nominal antigens including CMV. Only limited studies have been described previously to determine whether restricting determinants of DR and especially DQ are subtypic to the serologically defined DR and DQ specificities, and to what extent restricting determinants are associated with Dw specificities defined in alloresponses. In the present report, we describe a large number of CMV-specific Th clones derived from two different individuals who are seropositive for CMV. Clones were classified as being DR-, DQ-, or DP-reactive based on blocking with monoclonal antibodies. DR- and DQ-restricted clones were then examined in panel studies using antigen-presenting cells (APC) expressing the Dw subtype of the restricting DR-DQ haplotype, as well as APC expressing different Dw subtypes associated with the serologically defined specificity. Unrelated specificities were also included. Our findings show that not only for DR but for DQ as well, the primary restricting determinants appear to be subtypic to the serologically defined antigen; furthermore, subtype restriction for both DR and DQ is very closely associated with single Dw specificities. In several cases in which cross-reactivity among restricting Dw specificities was observed in association with a given DR or DQ haplotype, a molecular basis could be suggested to explain the cross-reacting determinants. A small minority of the clones appeared to be CMV specific, but was restricted by a determinant(s) that is either monomorphic or minimally polymorphic.

Antigen-Presenting Cells↗
PubMed1987

cDNA cloning and sequencing reveals that the electrophoretically constant DR beta 2 molecules, as well as the variable DR beta 1 molecules, from HLA-DR2 subtypes have different amino acid sequences including a hypervariable region for a functionally important epitope.

Two-dimensional gel electrophoresis (2D-PAGE) of DR molecules from three different Dw subtypes (Dw2, Dw12, and FJO) of the HLA-DR2 haplotype reveals that at least two DR beta genes are expressed. Protein mixing experiments demonstrate that one of the two expressed DR beta molecules is electrophoretically variable (referred to as DR beta 1), and the other (DR beta 2) migrates constantly among DR2 subtypes. We have constructed cDNA libraries from Dw12 and FJO homozygous typing cells (HTC DHO for Dw12 and HTC FJO for FJO) and isolated DR beta cDNA clones. Four of these clones (FJO-13, DHO-8, FJO-6, and DHO-7) were sequenced, and the deduced amino acid sequences were compared with each other and with two published amino acid sequences for the DR beta molecules derived from a DR2-Dw2HTC. Prediction of the migration patterns on 2D-PAGE from the amino acid sequences of these and other DR beta molecules allows the tentative designation of the two full-length cDNA (DHO-8 and FJO-13) as coding for DR beta 2 molecules and the other two cDNA (DHO-7 and FJO-6) for DR beta 1 molecules. Amino acid sequence comparisons also show that the constantly migrating DR beta 2 molecules, as well as the electrophoretically variable DR beta 1 molecules, from Dw2, Dw12, and FJO have different primary amino acid sequences, including a clustered difference in the third hypervariable region of the polymorphic first domain.

Amino Acid Sequence↗
PubMed1987

Long-term growth of lymphokine-activated killer (LAK) cells: role of anti-CD3, beta-IL 1, interferon-gamma and -beta.

Peripheral blood lymphocytes (PBL) cultured in interleukin 2 (IL 2)-containing medium in conventional tissue culture develop the ability to lyse fresh tumor cells; such cells are referred to as lymphokine-activated killer (LAK) cells. LAK activity peaks by day 5 of culture and declines rapidly thereafter. We studied culture conditions and signals that allow for long-term culture and expansion of cells with LAK activity. By culturing cells at relatively low densities and regularly replenishing medium and recombinant IL 2 (r-IL 2), LAK function is significantly higher as compared with short-term cultures, and remains present for at least 21 days while cell numbers undergo an average 100-fold expansion. By activating these cultures with anti-CD3 (OKT3) monoclonal antibody and r-IL 2, an approximately 1000-fold expansion in the cell number is obtained with maintenance of comparable levels of LAK activity. The exogenous addition of beta interleukin 1 (beta-IL 1), interferon-beta (IFN-beta) or interferon-gamma (IFN-gamma) can augment the lytic activity of cell populations expanded by anti-CD3 plus r-IL 2. These approaches may enable the in vitro generation from individual donors of much greater numbers of LAK cells for adoptive immunotherapy than can now be obtained with the 3 to 5 day in vitro culture systems.

Antibodies, Monoclonal↗
PubMed1987

Role of CD2 and IL 1 beta in T cell responsiveness to IL 2.

We have evaluated the effects of an anti-CD2 monoclonal antibody (moAb), Leu-5b, on inhibition of lymphocyte responsiveness. Given previous reports that anti-CD2 moAb prevent production of interleukin 2 (IL 2), as well as up-regulation of IL 2 receptor, we chose a test system in which lymphocytes were activated by exogenously added recombinant IL 2 (rIL 2). Our studies thus allowed us to evaluate other possible mechanisms by which anti-CD2 moAb could act. We found that Leu-5b not only blocked the proliferative response of human peripheral blood leukocytes (PBL) to IL 2, but also the development of cytotoxicity as assayed in lectin-dependent cytotoxicity (LDCC). Addition of recombinant interleukin 1 beta (rIL 1 beta) reconstituted the proliferative response in the presence of Leu-5b but did not allow development of cytotoxicity. Our findings are consistent with the concept that Leu-5b delivers a signal to IL 2-responsive cells that prevents their subsequent proliferation and development of cytotoxicity, but that only the former event, i.e., proliferation, is reconstituted by the addition of IL 1 beta.

Antibodies, Monoclonal↗
PubMed1987

HLA: its role in pathogenesis of disease.

The complexity of the HLA system, the major histocompatibility complex in man, is reviewed with regard both to the isotypic complexity (the number of loci) as well as the polymorphism of the individual loci. Different methods used to define polymorphism, including serological analysis and T lymphocyte response analysis, are considered. The possible role of HLA antigens in association with a variety of diseases is discussed.

Antibody Specificity↗
PubMed1987

Determination of DR beta 1 alleles of DR4/Dw subtypes by oligonucleotide probing.

The extensive Dw polymorphism of HLA-D has, to date, been defined using cumbersome and lengthy techniques involving T-cell proliferative reactions. These techniques have not necessarily provided information about the genotype at any single locus of HLA-D because the products of more than a single locus of HLA-D (e.g., DR beta 1 and DR beta 2 of the DR2 haplotypes and in some cases DQ) can contribute to T-cell proliferation and thus to assignment of Dw phenotype. We have recently reported that DR beta 1 alleles relating to the Dw polymorphism can vary by only a single, or a very few, nucleotide differences; the products of this locus are recognized in allo- and restricted responses. Given the importance of defining this polymorphism, we have used specific oligonucleotides as hybridization probes to differentiate between these very closely related alleles. This approach provides the basis for a rapid method for HLA-D typing which, as additional sequence information becomes available, will likely be generally applicable. In addition, oligonucleotide probing allows definition of a gene transcript from an individual locus of HLA-D rather than the phenotype results provided by Dw typing.

Alleles↗
PubMed1987

T-cell clonal analysis of HLA-DR2 haplotypes.

Individuals carrying the serologically-defined DR2 haplotypes can be further subdivided by utilizing T cell response to define several HLA-Dw/LD clusters: Dw2, Dw12, LD-5a, and a related group composed of FJO, AZH, and MN2. We have used cytotoxic T cell clones generated against these DR2 haplotypes to define the DR and DQ associated Dw/LD polymorphisms. Clones were categorized as DR, DQ, DP, or class I-directed based on monoclonal antibody (MoAb) inhibition studies. We analyzed the distribution of the determinants detected by these clones based on analysis with panels of cells. Some DR-directed clones only lysed cells positive for the Dw subtype of the sensitizing cell. A few DR-directed clones lysed most DR2 positive cells. Other DR-directed clones detected determinants shared by cells of different subtypes: determinants shared by Dw2 and Dw12 positive cells and determinants shared by MN2, FJO, AZH, and LD-5a positive cells. The majority of the DQ-directed clones only lysed cells positive for the sensitizing HLA-Dw subtype. A group of clones primed against an LD-5a HTC, whose cytolytic activity was not blocked by any monoclonal antibodies tested, lysed some, but not all, DR2 positive cells as well as Dw1 and Dw10 positive cells. The results suggest the detection of determinants on these molecules which may be shared by cells of specific Dw subgroups and determinants shared by most cells of the sensitizing serological specificity. The basis of this observed polymorphism and influence on the allogeneic response are discussed.

Antibodies, Monoclonal↗
PubMed1987

Comparison of DR beta 1 alleles from diabetic and normal individuals.

Insulin-dependent diabetes (IDD) is positively associated with HLA-D proteins. A critical question is whether or not sequence differences within the HLA-D coding region are the same or different in diabetics and normal individuals of the same haplotype. We have isolated both DR beta 1 alleles from a Dw4/LD MN2 cDNA library and compared them to DR beta 1 genes isolated from normal individuals of the same Dw phenotype. We found no nucleotide differences in the coding region between the normal and diabetic alleles of DR beta 1 suggesting to us that DNA differences other than the DR beta 1 coding region may account for the observed association of HLA-D and diabetes.

Alleles↗