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We have previously shown that tumour cells may release soluble factors which inhibit the division of a variety of cells. These factors had marked immunosuppressive activity in vitro as shown by their ability to Inhibit lymphocyte mitogenic responses to lectins. allogeneic lymphocytes and pokeweed mitogen-induced immunoglobulin production. It was postulated that these factors may play an important role in determining the outcome of tumour host interactions in humans and be the source of immunosuppressive factors in the circulation of patients with cancer. In the present study, characterization of these mitogenic inhibitory factors released into the supernatants of cultured tumour cells was carried out by a number of sequential separation procedures involving affinity chromatography on wheat-germ lectin (WGL). gel filtration on PL-agarose and electrophoresis in polyacrylamide gels in the presence of sodium dodecyl sulphate (PAGE-SDS). The factors were identified throughout the separation procedure using inhibition of phytohaemagglutinin (PHA) stimulation of normal lymphocytes as an index of inhibitory function. By these techniques the inhibitory activity was shown to be associated with glycoproteins containing N-acetyl-β-D-glucosamine (NAcGlu) and having molecular weights (mol. wt) of approximately 140.000 and over 200,000 daltons. A further small mol. wt peak of 70,000 daltons was observed on PAGE-SDS. pi values of the fractions were 7.4 and 4.2. The activity was destroyed by heating, low pH and repeated freeze-thawing but was resistant to proteolytic enzymes, deoxyribonuclease, ribonuclease and neuraminidase. The active fractions were not associated with proteolytic activity and the inhibition of mitogenic responses was irreversible in nature. These results form the basis of studies to detect this factor in biological fluids and further evaluate its role in the tumour host relationship. [ABSTRACT FROM AUTHOR]

A number of previous reports have described the presence of factors which inhibit the response of lymphoid cells to phytohaemagglutinin. The present study describes an inhibitor of cell division synthesized and released directly from cultured tumour cells which appears to have similar immunosuppressive effects in vitro. This factor(s) was detected in a wide variety of cultured tumour cells and some cultures of normal foetal tissue. It inhibited the mitogenic response of lymphocytes to PHA and also the spontaneous cell division of a variety of cultured cells including the cells producing the factor. Pulse cytophotometry showed that cells were inhibited in the G1 phase. Production of the factor increased with time and was related to the number of cells in culture. Its synthesis was inhibited by cycloheximide. DNA and RNA synthesis was not required for its production. The factor appeared to have a high degree of biological activity m terms of the number of cultured cells required for production and in regard to the number of cells inhibited. The biological significance of this factor in vivo is unknown but in vitro it was shown to inhibit immunoglobulin and mitogen-induced responses which suggests it may play an important role in suppression of the immune response against tumour cells in the host. [ABSTRACT FROM AUTHOR]

Blood from patients with hairy cell leukaemia (HCL) was used to investigate subpopulations of the non-specific activated lymphocyte killer (ALK) cells generated in mixed lymphocyte culture (MLC). Mononuclear cells from four of live patients with HCL had decreased natural killer (NK) cell activity and., of these, all showed a decrease in ALK cell activity against K-562 target cells (NK like cells), but three had normal levels of ALK cells able to kill melanoma cells. One patient who had received splenectomy had normal NK cell activity, and in MLC his cells generated normal levels of both subpopulations of ALK cells. The results support the hypothesis of separate subsets of ALK cells: NK like cells with NK cell precursors and which preferentially lyse the K-562 target cell and a second separate population which arises independent of NK cell precursors and which has strong cytolytic activity against an NK insensitive melanoma cell line. [ABSTRACT FROM AUTHOR]

The action of levamisole on immunoglobulin production was investigated in pokeweed mitogen-stimulated cultures of B and T cells. Immunoglobulin production was measured by nephelometric methods using specific antisera to IgA, IgG and IgM. Levamisole was shown to have no effect on B lymphocytes but pretreatment of T lymphocytes with levamisole resulted in an increase in immunoglobulin production which was maximal at 10-6 M. The latter increase appeared to be due to an effect on suppressor T cells in that pretreatment of T cells depleted of suppressor cells (by irradiation or removal of T cells with receptors with IgG) did not result in an increase in immunoglobulin production. Similar effects of levamisole were demonstrated against suppressor cells induced by concanavalin A in vitro and against spontaneous suppressor cell activity in vivo. The effect of levamisole and irradiation on suppressor cell activity against particular classes of immunoglobulin varied between individuals. Suppressor cell activity against IgA production alone was demonstrated in six of 17 subjects whereas in the remainder, suppressor cell activity was either not detected or inhibited production of all three immunoglobulin classes. The implications of this action of levamisole for its effects in vivo are unknown, but the results appear consistent with many of the observed effects of the drug. Selection of patients with diseases associated with high suppressor cell activity may lead to more effective use of the drug. [ABSTRACT FROM AUTHOR]

Mixed lymphocyte cultures were set up between blood mononuclear cells and irradiated autologous or allogeneic B lymphoblasts infected with Epstein Barr virus. The resulting cytotoxic effector cells were cloned and tested for activity against the stimulating B lymphoblast, K562 and melanoma targets. Specific clones which killed only the stimulating B lymphoblasts (cytotoxic T lymphocytes: CTL) were re-cloned and the subclones tested for cytolysis of B lymphoblasts and melanoma cells. Of seven primary CTL clones generated in allogeneic culture, 308 subclones developed the ability to kill melanoma cells and none retained specific CTL function. In the autologous system, 180 subclones were derived from three specific primary clones: of these. 13 (7%) retained specific function 29(16%) were able to kill both B lymphoblasts and melanoma cells, and 93 (52%) killed only the melanoma target. Testing of random clones demonstrated that whereas both B lymphoblast killing (CTL function) and melanoma cell killing (anomalous killer; AK function) were blocked by a monoclonal antibody to LFA-1, only CTL function was blocked by anti-T3 or anti-T8 antibodies. The factor(s) causing the progression of CTL to AK cells are discussed. These data thus demonstrate that the majority of CTL are capable of mediating AK cell function and are thus potentially suitable for passive immunotherapy. [ABSTRACT FROM AUTHOR]

Synopsis Objective To examine the potential of non-animal collagens as a new option for cosmetic applications. Methods Non-animal collagens from three species, Streptococcus pyogenes, Solibacter usitatus and Methylobacterium sp 4-46, have been expressed as recombinant proteins in Escherichia coli using a cold-shock, p Cold, expression system. The proteins were purified using either metal affinity chromatography or a simple process based on precipitation and proteolytic digestion of impurities, which is suitable for large-scale production. Samples were examined using a range of analytical procedures. Results Analyses by gel electrophoresis and mass spectrometry were used to examine the purity and integrity of the products. Circular dichroism spectroscopy showed stabilities around 38°C, and calculated p I values were from 5.4 to 8.6. UV-visible light spectroscopy showed the clarity of collagen solutions. The collagens were soluble at low ionic strength between p H 5 and p H 8, but were less soluble under more acidic conditions. At lower p H, the insoluble material was well dispersed and did not form the fibrous associations and aggregates found with animal collagens. The materials were shown to be non-cytotoxic to cells in culture. Conclusions These novel, non-animal collagens may be potential alternatives to animal collagens for inclusion in cosmetic formulations. [ABSTRACT FROM AUTHOR]

Collagen is an important component for cosmetic formulation, where it is an effective natural humectant with high substantivity. Commercial collagen preparations have a wide range of properties. In the present study, various techniques have been used to examine three distinct commercial collagens that illustrate the range of properties that are available. The usefulness of the various techniques for assessing collagen quality and batch-to-batch variation is discussed. The results indicate that there are several simple, cheap and effective methods such as gel electrophoresis that provide excellent information on collagen quality. The appropriate selection of tests allows informed decisions on the choice of which collagen preparation to use in providing the desired functionality and shelf life of a formulation. [ABSTRACT FROM AUTHOR]

Objective To evaluate the biological response to two urethane-based adhesives used to repair full thickness meniscal wounds created in the partially vascularised (red-white) zone. Design An ovine bilateral meniscal defect model was used to evaluate the initial biological response of the meniscal cartilage and synovium over a 1-month period. A 10-mm full-thickness defect was created in the medial meniscus of each femorotibial joint. The defects were either left untreated or repaired using the urethane-based adhesives. Synovial fluid, synovial membrane and the meniscal cartilages were retrieved at necropsy for cytological and histological assessment. Results The ovine model proved to be a suitable system for examining meniscal repair. Untreated defects showed no tissue apposition or cellular healing response, whereas all eight defects repaired with the two urethane-based adhesive formulations showed signs of repair and tissue regeneration with indications of cell infiltration and new collagen deposition in and around the polymer. No adverse cellular response to the adhesives was observed in the meniscal defect or in the synovial membrane and fluid. Conclusion Trauma to the knee commonly results in tears to the meniscal cartilage, with the majority of these occurring in the partially vascularised (red-white) or non-vascularised (white) zones of the meniscus. Repair, and subsequent healing, of these tears is poor because of the reduced vascularity and limited surgical access. The present data indicate that an ovine model is a suitable system for examining meniscal repair, and that development of urethane-based adhesives offers a strategy that may be clinically effective for the treatment of these injuries. [ABSTRACT FROM AUTHOR]

Lymphocytes from six Chediak Higashi (CH) patients were markedly depressed in their ability to lyse tumour cell targets in both 51Cr release and single cell cytotoxicity assays. The frequency of lymphocytes bearing the OKM1 marker and the frequency of T3+, T4+, T8+, la-, Mol+, Mo2+ and BI+ cells was normal among sheep erythrocyte rosetting (E+ ) and non-resetting (E-) peripheral blood leucocytes analysed by flow cytofluorography. Cells expressing the NK shared markers. OKMI mac-1, FcR, and the characteristic large granular lymphocyte (LGL) morphology of NK cells were also present in normal numbers in the highly enriched NK fraction separated on Percoll density gradients. This fraction did not contain detectable numbers of cells expressing the Mo2 marker of human monocytes. Therefore most of the cells stained by monoclonal OKMI and mac-1 in this fraction are likely NK cells, rather than monocytes. and we conclude that the size of the NK pool in CH patients is probably normal. The capacity of CH lymphocytes to recognize anti bind to tumour cells was also normal as was the subsequent burst at oxygen intermediates produced by the NK cells in a chemiluminenscence assay We have shown elsewhere that O2 generation is directly involved in activating subsequent steps in the NK cytolytic pathway. These results suggest that NK cells in CH patients are present in normal frequency but are blocked at some post-recognition, post-activation step in the cytolytic pathway subsequent to the burst of oxygen intermediates but preceding the lethal hit. [ABSTRACT FROM AUTHOR]