摘要:

The monoclonal antibody MoAb‐VκIIIb binds a cross‐reactive idiotopic (CRI) determinant on light (L) chains encoded by the VκIIIb subgroup A27a (Humkv325) gene segment. The aim of this study was to localize the MoAb‐VκIIIb CRI. Mutational analyses involving region exchanges between a CRI‐positive VκIIIb chain and a CRI‐negative Vκ1 chain indicate that the MoAb‐VκIIIb CRI is located in framework region (FR) 3 of A27a (Humkv325) encoded L chains. CRI‐positive kappa chains unpaired with a heavy (H) chain are reactive with MoAb‐VκIIIb, indicating that the CRI is located on the kappa chain alone without involvement of H chain residues. Combinatorial antibodies composed of non‐parental L and H chain pairings are reactive with MoAb‐VκIIIb only when the L chain is A27a (Humkv325) encoded. The CRI, therefore, is not readily perturbed by H chain interactions. When the FR3 from a CRI‐positive kappa chain replaced the FR3 in a CRI‐negative lambda chain, the determinant was no longer detectable with MoAb‐VκIIIb. It is possible, therefore, to exchange regions between kappa chains from different families and retain the CRI structure, however the determinant is lost when placed in a more foreign background such as a lambda chain. These data more precisely define the interaction between MoAb‐VκIIIb and its CRI, and indicate that there are limits within which antibody FRs can be shuffled and

ISSN:0300-9475    

DOI:10.1046/j.1365-3083.1996.d01-319.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Apoptosis of normal thymocytes was shown to be triggered by several mechanisms (e.g. glucocorticoids, γ‐irradiation). In the present study the authors report on thymocyte apoptosis that is induced by thymic epithelial cells. The thymocytes undergo a massive apoptotic death within 24 h of cocultivation with thymic epithelial cell monolayers derived from primary cultures (PTEC) or from a thymic epithelial cell line (TEC). Non‐thymic monolayers were inactive. Apoptosis induction in this experimental model requires direct contact between the thymocytes and the thymic epithelial monolayer and can be blocked by anti‐CD2 and anti‐LFA‐1 antibodies. The immature CD3−/+dullCD4+CD8+thymocytes were the cells which undergo apoptosis. The fact that the authors are dealing with a massive apoptotic process of immature cells in the absence of exogenous antigen suggests that it involves the nonselected thymocytes. The apoptotic pathway selected by thymocytes following their culturing on TEC involves p53 expression. Indeed it was found that TEC‐induced apoptosis, led to the accumulation of p53 protein that preceded the step of DNA fragmentation in freshly isolated thymocytes as well as in a glucocorticoid resistant thymoma cell line. Since glucocorticoid‐induced thymocyte apoptosis is p53‐independent, glucocorticoids are conceivably not involved in TEC‐induced thymocyte death. Thein vitroexperimental model presented here may reflect the physiological sequence of events leading to thymocy

ISSN:0300-9475    

DOI:10.1046/j.1365-3083.1996.d01-312.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

The influence of the transcription factor Oct2A on V(D)J recombination was investigated. It was found that  co‐transfection of Oct2A together with an artificial recombination substrate increased the rate of V(D)J recombination by a factor of 3.4 in the fibroblast cell line L4 which had been stably transfected with genomic  DNA containing the rag‐1 and rag‐2 genes and their flanking regulatory sequences. This effect, however, was not observed in the pre‐B‐cell line 38B9. The effect of Oct2A depends on the presence of its transactivation domains indicating that Oct2A increases the transcription of genes that increase the rate of V(D)J recombination. This effect can be reversed by additional co‐transfection of the glucocorticoid receptor. In contrast to L4 cells Oct2A and the glucocorticoid receptor had no effect in 38B9 cells. From these results one may conclude that the effect of Oct2A is masked in 38B9 cells by endogenous Oct2A and other  redundant pre‐B‐cell‐specific transcription factors and that it can only be seen i

ISSN:0300-9475    

DOI:10.1046/j.1365-3083.1996.d01-316.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

The skin interfaces directly with the external environment that contains innumerable infectious agents. Therefore, an appropriate and rapid immunologic response is required to preserve internal homeostasis. An essential feature of the ‘skin immuno system' (SIS) is the presence of substantial numbers of T cells in normal skin. The T‐cell receptor repertoire from normal human breast skin was analysed quantitatively and qualitatively by using PCR amplification of reverse transcribed RNA, T‐cell receptor BV3 and BV14 gene usage was increased in skin T lymphocytes in all individuals tested (n = 8) compared to peripheral blood CD4+and CD8+T lymphocytes from the same individuals. The T‐cell receptor junctional diversity analysed by high resolution gel electrophoresis showed skin T‐cell BV3 and BV14 gene usage to be predominantly polyclonal. Superantigen stimulation of T cells in human skin is considered a likely explanation of the pr

ISSN:0300-9475    

DOI:10.1046/j.1365-3083.1996.d01-313.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Allergy to fish is common in Northern Europe. Variable reactions to different fish species are usually experienced among fish allergic patients. The allergens of cod fish and particularly the major allergen parvalbumin β (Gadus callarias) have been extensively studied in Norway. In the present communication, the white muscle parvalbumin was similarly found to be a major allergen in Atlantic salmon (Salmo salar,Sal s1 ). A purified salmon parvalbumin was obtained by anion exchange chromatography, gel filtration chromatography (GFC) and high‐performance liquid chromatography (HPLC) of the muscle extracts. The antigenicity and allergenicity of salmon parvalbumin were confirmed using various immunologic and electrophoretic techniques. The protein is representative for several isoallergens judged by the amino acid (AA) sequence variance at certain sites in the AA sequence of CNBr cleavage peptides. Using sera from patients with cod and salmon allergySal s1was demonstrated to be the major allergen of Atlantic salmon, as judged by RAST‐ and ELISA‐inhibitions and crossed radioimmunoelectrophoresis (CRIE) techniques. The protein was also demonstrated to be antigenic by the use of polyclonal cod and salmon antibodies in IgG ELISA and immunoelectrophoretic methods. Cloning of parvalbumin cDNA from Atlantic salmon was performed based on an alignment of parvalbumin AA sequences from other species. A probe was generated by PCR and used for screening a salmon muscle cDNA‐library. Subcloning and sequencing of two hybridizing clones revealed transcripts from two different parvalbumin genes. The translated sequences of both clones belong to the β‐lineage of parvalbumins and include the entire

ISSN:0300-9475    

DOI:10.1046/j.1365-3083.1996.d01-314.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

IFN‐α production in Sendai virus‐stimulated human buffy coat cultures could readily be demonstrated in individual cells at a protein level by the use of a novel immuno‐enzymatic staining procedure. A distinctive rounded, juxtanuclear staining pattern was generated in producer cells by the accumulation of the intracellularly synthesized IFN‐α in the Golgi stacks. The technology is based upon acquiring a video image of stained monolayers of cells, viewed in a microscope by a colour camera, which then transfers binary images directly into a computer‐controlled operating system. The characteristic appearance of the immunocytochemical staining enabled a computerized image‐analysis system to measure IFN‐α producing cells based on defined criteria set for morphology, intensity, colour and size. The automated system could accurately and reproducibly register a range of 0.1–7.0% of the total cell population as IFN‐α producing cells during the kinetic studies of the response. Congruent results were obtained with manual microscopy and image analysis concerning the assessment of the incidence of IFN‐α producing cells in the total cell populations. All IFN‐α producing cells expressed surface HLA‐DR molecules and 95% of these cells belonged to the myelomonocytic lineage. The image analysis system provided, in contrast to conventional microscopy, an opportunity to assess and document differences of signal intensity and cell size of individual IFN‐α producin

ISSN:0300-9475    

DOI:10.1046/j.1365-3083.1996.d01-317.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

Lymphocyte–endothelium interactions are pivotal steps in mediating inflammatory responses. The authors have analysed the influence of ultraviolet B (UVB) irradiation on intercellular adhesion molecule (ICAM)‐1 expression on cells of the human microvascular endothelial cell line (HMEC)‐1 and the intracellular signalling pathways involved. Flow cytometry revealed dose‐dependent ICAM‐1 up‐regulation with maximum induced expression 24 h after sublethal UVB irradiation of 10 mJ/cm2. While anti‐tumour necrosis factor (TNF)‐α antibodies or recombinant human interleukin (IL)‐10 did not influence this response, anti‐interferon (IFN)‐γ antibodies blocked the UVB‐induced ICAM‐1 up‐regulation. Significant induction of intracellular/membrane‐bound IFN‐γ was measured as early as 6 h post‐UVB. Since previous work has shown a differential role of protein kinase C (PKC) in cytokine induced ICAM‐1 expression, the effect of a selective  bisindolylmaleimide‐derived PKC‐inhibitor (GF109203X) was studied. Ultraviolet B‐induced ICAM‐1 up‐regulation was effectively blocked by the PKC‐inhibitor, whereas a PKA‐inhibitor was ineffective. Moreover, immunofluorescence analysis showed a radiation‐induced membrane translocation of PKC‐α, indicative of enzyme activation, in HMEC‐1 cells already 30 min post‐UVB. The functional relevance of the UVB‐induced ICAM‐1 expression and involvement of PKC in this process was demonstrated in an adhesion assay with peripheral blood mononuclear cells. In conclusion, UVB‐induced ICAM‐1 expression on human endothelial cells involves PKC‐dependent pathways and can be prevented by a PKC‐inhibitor. The use of PKC‐inhibitors as additive modulators in immune reactions may bear clinical potential. The me

ISSN:0300-9475    

DOI:10.1046/j.1365-3083.1996.d01-324.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

A major problem in the intensive care unit nowadays is the development of multiple organ dysfunction syndrome (MODS), a cumulative sequence of progressive deterioration of organ functions. While the pathogenic pathways of MODS remain to be elucidated, it is assumed that cells of the host defence system, especially the macrophages, are altered in their function. During the development of MODS it is assumed that macrophages are overactivated and that an exaggerated inflammatory response may contribute to its pathogenesis. In order to gain insight into the alterations of the functional status of the macrophage during the development of MODS, a series of macrophage functions was measured in the subsequent phases of zymosan induced generalized inflammation in mice. Male C57BL/6 mice received a single dose of zymosan intraperitoneally and groups of animals were killed after 2, 5, 8, and 12 days. Peritoneal macrophages were collected forin vitroassessment of the ADCC, the production of superoxide (O−2) and nitric oxide (NO), and complement mediated phagocytosis and intracellular killing ofStaphylococcus aureus. A single intraperitoneal injection with zymosan resulted in a three‐phase illness. During the third phase the animals developed MODS‐like symptoms. Peritoneal cells from control animals produced very low to non‐detectable amounts of O−2and NO, and the cytotoxic activity was also low. During the development of MODS, from day 7 onwards, the ability to produce O−2and NO‐2became strongly elevated, as did the cytotoxic activity. These findings are in parallel with the development of MODS whereas the phagocytic and killing capacity remained essentially unaltered. The changes found could be detrimental for the organism, thus possibly contributing to the onset and develo

ISSN:0300-9475    

DOI:10.1046/j.1365-3083.1996.d01-315.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

The authors have previously reported on the ability of A60, an immunodominant antigenic complex of Mycobacterium bovis BCG, to prevent cancer development in mice challenged with EMT 6 tumour cells. Such effect proved to rely on neoplastic cell lysis by cytolytic T lymphocytes and activated macrophages. The involvement of cytokines in triggering the immune response leading to tumour rejection is analysed in the present work. The synthesis of IL‐2, IFN‐α and TNF‐α was strongly increased in A60‐primed mice. Cancer development depressed the blood levels of these three cytokines. In vitro cultures of lymphocytes from lymph nodes and blood of A60‐primed mice produced higher levels of these cytokines in the presence of A60, as  compared to cultures lacking A60. Such effect was inhibited by co‐incubation of lymphocytes with EMT  6  tumour cells. In vitro cultures of macrophages yielded higher levels of TNF‐α in the presence of A60 and co‐incubation of these cells with EMT 6 tumour cells also inhibited TNF‐α production. The enhanced synthesis of IL‐2 and IFN‐α, which promote activation of cytolytic T lymphocytes and macrophages, accounts for the increased tumour cell lysis inducedin vivoby A60. The A60‐promoted synthesis of TNF‐α is partly responsible for the latter effect. The inhibitory action of EMT‐6 tumour cells on cytokine synthesis is a powerful mechanism of t

ISSN:0300-9475    

DOI:10.1046/j.1365-3083.1996.d01-318.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY

摘要:

It has been previously demonstrated that the sea star axial organ is a primitive immune organ. Phagocytic, lymphoid‐like cells have been characterized with properties similar to those of vertebrates. There is also evidence for an invertebrate cytokine network because IL‐1 and TNF‐like activities are clearly demonstrable. In addition, the authors have previously described preliminary evidence for IL‐2‐like activity in the sea star. In the present report, the authors obtained evidence for the presence of IL‐1‐ and IL‐2‐like molecules on axial organ cells. More interestingly, the results suggested that sea star cells express structures similar to human receptors for IL‐1, I

ISSN:0300-9475    

DOI:10.1046/j.1365-3083.1996.d01-322.x

出版商:Blackwell Science Ltd    

年代:1996

数据来源: WILEY