T-cell hybridoma bearing heavy chain variable region determinants producing (T,G)-A--L-specific helper factor

Thymus-derived lymphocytes (T cells) exert their regulatory effect (help or suppression) on the antibody production by B cells either by direct cell to cell interaction or by soluble mediators or factors. The low frequency of specific T cells, the heterogeneity of their responses and their relatively short life span have hampered the molecular characterization of the antigen recognition unit of T cells, and its structure is largely unknown. The lymphocyte hybridization technique, which has been found very useful for the production of B-cell hybridomas secreting specific monoclonal antibodies, has also been used for the generation of homogeneous and stable T-cell hybridomas with unlimited growth potential. So far the only specific effector function demonstrated in the established T hybridomas is the property to generate a factor(s) which suppresses antibody responses. We now describe the establishment of hybrid lines which exhibit characteristic T-cell markers. One of the hybridomas (denoted R-9) releases into the culture supernatant factor(s) with helper activity specific to the synthetic polypeptide (T,G)-A--L and bears surface determinants of the immunoglobulin heavy chain variable region (VH). Such hybrid cell lines are of great value for studies on the nature of the T-cell receptor.     

I-J as an idiotype of the recognition component of antigen-specific suppressor T-cell factor

The I-J determinant of membrane glycoprotein is known to be expressed exclusively on suppressor T cells (TS), which have a crucial role in the regulation of immune responses. I-J also comprises part of the soluble factor (TSF) with suppressor activity which is secreted from TS. Gene-mapping experiments have indicated that the I-J gene lies between the I-A and I-E subregions of the mouse major histocompatibility complex (MHC) and is defined by the H-2 congeneic pair, that is, B10.A(3R) and B10.A(5R). In fact, antibodies raised in the reciprocal combinations of B10.A(3R) and B10.A(5R) define the I-Jb and I-Jk alleles, and are able to detect the I-J determinants on TS and TSF. Biochemical and functional analyses, using I-J-positive TS clones and hybridomas, have demonstrated that monoclonal anti-I-J antibodies precipitate I-Jk or I-Jb with a relative molecular mass of 25,000-28,000 (25-28K) and that the I-J+ molecule mediates the restriction specificity of TSF in association with an antigen-binding protein (45K). However, molecular genetic studies on the I-J gene reveal no genetic difference between B10.A(3R) and B10.A(5R) and also that there is no room to accommodate a gene encoding I-J in the expected I region. These discrepancies between the molecular genetic and serological/functional data require explanation. Here we demonstrate that TS and TSF expressing I-J of the host type were produced by fully allogeneic bone marrow cells of donor origin in chimaeric mice, when the chimaeras received the host antigen-presenting cells (APC) at the time of immunization. The results show that APC are necessary for the activation and clonal expansion of TS and also support the notion that I-J is an idiotypic determinant of the recognition component of TS and TSF.     

Functional and molecular organisation of an antigen-specific suppressor factor from a T-cell hybridoma

Thymus-dependent (T) lymphocytes have been shown to have antigen specificity. The antigen receptor on T lymphocytes, in contrast to that on B lymphocytes, does not appear to be of the conventional immunoglobulin (Ig) type. Studies on the antigen-specific factors derived from helper and suppressor T cells (Ts) demonstrated that they possess determinants with antigen binding affinity and products of genes in the H-2 complex (MHC). Furthermore, antibodies against the variable region of Ig heavy chains or idiotypes have been shown to react with T-cell antigen receptors as well as antigen-specific helper and suppressor T-cell factors (TsF). It is, therefore, conceivable that at least two gene products are involved in the structural entity of these receptors: one each coded for by genes in either. To establish the molecular nature of the recognition component of T cells we have used homogeneous TsF from a T-cell hybridoma with a specific function. We report here that the antigen binding and I-J coded molecules on TsF are independently synthesised in the cytoplasm, and are secreted as an associated form of the two molecules; this association is required for antigen-specific suppression of antibody response.     

Qa-1 restricted recognition of foreign antigen by a gamma delta T-cell hybridoma

Distinct T-lymphocyte subsets recognize antigens in conjunction with different classes of major histocompatibility complex (MHC) glycoproteins using the T-cell receptor (TCR), a disulphide-linked heterodimer associated with the CD3 complex on the cell surface. In general, class I and class II MHC products provide a context for the recognition of foreign antigens by CD8+ and CD4+ T cells, respectively. This recognition seems to be largely dependent on alpha beta TCR heterodimers, whereas the function of the second gamma delta TCR, present on a minor subpopulation of cells, is still unknown. In the mouse, the existence of six cell-surface MHC class I products (K, D, L, Qa-1, Qa-2 and Tla) has been firmly established by serological, biochemical and genetic evidence. So far, only the most polymorphic of them, K, D and L ('classical' class I) have been reported as restriction elements for T-cell recognition of foreign antigens. The function of the relatively invariant Qa and Tla molecules remains unknown. We have made a T-helper cell hybridoma clone (DGT3) that recognizes synthetic copolymer poly(Glu50Tyr50) in the context of Qa-1 cell surface product, and has a CD4-CD8- phenotype. Our studies indicate that DGT3 cells express the gamma delta TCR on the cell surface, implicating its role in Qa-1-restricted antigen recognition. This is the first evidence that T cells can recognize foreign antigen in association with self Qa product, confirming that Qa molecules not only topologically, but also functionally, belong to the MHC.     

Expression and function of CD4 in a murine T-cell hybridoma

The CD4 (T4) antigen was originally described as a phenotypic marker specific for helper T cells, and has recently been shown to be the receptor for the human immunodeficiency virus (HIV). Functional studies using monoclonal antibodies directed at CD4 and major histocompatibility complex (MHC) class II molecules led to the suggestion that CD4 binds to the MHC class II molecules expressed on stimulator cells, enhancing T-cell responsiveness by increasing the avidity of T cell-stimulator cell interaction and/or by transmitting a positive intracellular signal. But recent evidence that antibodies to CD4 inhibit T-cell responsiveness in the absence of any putative ligand for CD4 has been interpreted as suggesting that antibody-mediated inhibition may involve the transmission of a negative signal via the CD4 molecule instead. We have infected a murine T-cell hybridoma that produces interleukin 2 (IL-2) in response to human class II HLA-DR antigens with a retroviral vector containing CD4 cDNA. The resulting CD4-expressing hybridoma cell lines produce 6- to 20-fold more IL-2 in response to HLA-DR antigens than control cell lines. Furthermore, when antigen levels are suboptimal, the response of the cell lines is entirely CD4-dependent. The data presented here clearly demonstrate that CD4 can enhance T-cell responsiveness and may be crucial in the response to suboptimal levels of antigen.     

Dissociation of phosphoinositide hydrolysis and Ca2+ fluxes from the biological responses of a T-cell hybridoma

T lymphocytes can be activated in a variety of ways, including occupancy of the T cell antigen receptor (TCR) complex or cross-linking of certain cell-surface molecules with antibody. Two of the earliest events seen after stimulation are the hydrolysis of phosphatidylinositol bisphosphate to inositol trisphosphate (Ins P3) and 1,2-diacylglycerol (DAG), and an increase in the concentration of intracellular Ca2+ ([Ca2+]i). Later, the cell secretes lymphokines and expresses lymphokine receptors. It has been postulated that the products of the hydrolysis of phosphatidylinositols (Ptd Ins) and fluctuations in [Ca2+]i are critical 'second messengers', transmitting the signals for the initiation of the later events. We have examined the relationship between these second messengers and the secretion of IL-2 in a murine T cell variant whose missing TCR complex had been reconstituted by gene transfer. Surprisingly, although the IL-2 responses of the transfectant could not be distinguished from the original line expressing the same TCR, Ptd Ins hydrolysis and the increase in [Ca2+]i were substantially reduced or absent in the reconstituted cell. It is therefore possible to dissociate these early biochemical changes from a late biological response, raising questions about the putative causal relationship of these events.     

高品质轴承钢LF--VD过程非金属夹杂物演变规律

对国内一钢厂EAF→LF→VD→CC工艺生产的高品质GCr15轴承钢进行系统取样,针对DS类非金属夹杂物随机性强的特点,利用能够进行大面积试样检测的ASPEX自动扫描电镜分析统计钢中非金属夹杂物的成分、尺寸、数量等信息。研究发现：GCr15轴承钢冶炼过程中非金属夹杂物主要为MgO- Al2 O3- CaO复合夹杂物和MnS,同时有少量的SiO2- Al2 O3和MgO-Al2 O3复合夹杂物;夹杂物尺寸主要集中在3~8μm,并有少量DS类夹杂物出现且尺寸范围波动很大,最大可以达70μm以上,形貌为圆形或近似圆形;VD有较强的去除夹杂物功能,经过VD真空精炼,夹杂物中CaO含量有增加趋势;吊包至铸坯过程,夹杂物成分向Al2 O3含量增多的区域移动,最终轴承钢铸坯中夹杂物成分位于高Al2 O3含量(≥80%),少量MgO (<20%)和低CaO(<5%)的区域;DS夹杂物的生成和去除具有较强的随机性。