B. Wilbrink

Immune reactivity in SL2 lymphoma-bearing mice compared with SL2-immunized mice

SummaryWe have studied the rather paradoxical phenomenon of the growth of an antigenic tumor in an immunocomponent host. This phenomenon was studied by comparing (a) the lymphocyte reactivity and (b) the macrophage cytotoxicity, during SL2 growth in DBA/2 mice (SL2-bearing mice) and in DBA/2 mice immunized against SL2 tumor cells (SL2-immune mice). Immune mice rejected a challenge of tumor cells. The immune T-lymphocytes rendered macrophages cytotoxic (arming) and were able to transfer tumor resistance to naive animals. Nonimmunized mice did not reject a challenge of SL2 cells. In these tumor-bearing mice various forms of immune reactivity were tested. Lymphocytes with the capacity to arm macrophages could not be found in the lymphoid organs. However, lymphocytes isolated from the tissue directly surrounding the subcutaneous SL2 tumor could arm macrophages in vitro.Shortly after subcutaneous tumor grafting cytotoxic macrophages were found in the peritoneal cavity. In the serum macrophage arming factors were detected that rendered macrophages cytotoxic in vitro. This cytotoxicity of the peritoneal macrophages and the presence of macrophage arming factors in the serum showed a similar biphasic pattern. The first phase of cytotoxicity between day 3 and 8 after tumor grafting was tumor (SL2) specific. The second phase from day 12 and onwards was not tumor specific. During the first 4 days after SL2 grafting the DBA/2 mice expressed a specific concomitant immunity to a second tumor graft. Then 7 or more days after grafting the first SL2 tumor, the concomitant immunity was nonspecific as the growth of a second SL2 tumor graft and a L5178Y (DBA/2) tumor graft were inhibited. In addition, the immune suppressive activity of serum and lymphocytes was tested. Neither serum nor lymphocytes from SL2-bearing mice suppressed the macrophage arming capacity of SL2 immune lymphocytes. Lymphocytes from tumor-bearing mice did not inhibit the capacity of SL2-immune lymphocytes to transfer resistance to naive animals. On the contrary, lymphocytes obtained from SL2-bearing mice 14 days after SL2 grafting transfered tumor resistance in a Winn-type assay. These data suggest that the growth of an antigenic tumor is due to the inability of the immune system to mount an effective antitumor effector cell population during tumor growth, rather than an immune suppression of the antitumor reactivity, as a limited immune reactivity could be detected in tumor-bearing mice, whereas immune suppression could not be detected.

Matrix depletion of young and old human articular cartilage by cultured autologous synovium fragments: a chondrocyte-independent effect

SummaryHuman articular cartilage of different ages was cultured for 8 days and proteoglycan (PG) release into the medium was measured. Retinol and synovial co-culture increased the PG release of cartilage of all ages. The effect of retinol was dose-dependent. Synovium increased also the PG release of dead cartilage, whereas retinol did not. The increased PG release by synovial co-culture is therefore mainly the result of synovial enzymes acting directly on the matrix rather than of a factor inducing chondrocyte-mediated breakdown.

In vitro influence of ketoprofen on the proteoglycan metabolism of human normal and osteoarthritis cartilage

We investigated the influences of ketoprofen on the proteoglycan (PG) turnover of human articular cartilage explants in three groups: normal young with a high basal PG synthesis, normal adult and osteoarthritis cartilage, both with a low basal PG synthesis. Ketoprofen had no influence on the mean PG synthesis rate of normal adult and OA cartilage after 4 days of culture nor on the cartilage PG content after 8 days of culture. There was no relation between the histological grade of OA and effects of ketoprofen. In normal young cartilage ketoprofen induced an increase of the PG synthesis rate when added to the culture in a concentration of 10−4M. No correlation existed between the effect of ketoprofen and the basal PG synthesis of normal cartilage.

Antigen‐Activated T Cells Inhibit Cartilage Proteoglycan Synthesis Independently of T‐Cell Proliferation

Previously we have shown that blood mononuclear cells (MNC) obtained from patients with rheumatoid arthritis (RA) have the capacity to induce depletion of proteoglycans (PG) in human cartilage explants. This was observed especially after stimulating MNC with mycobacterial antigens, rather than with the miiogen Concanavalin A (Con A). We have now co‐cultured cartilage explants in the presence of T‐cell clone A2b obtained from the rat model of adjuvant arthritis (A A). We show that inhibition of the cartilage PG synthesis is a consequence of antigenspecific T‐cell activation and that it is mediated by a humoral factor. This seems to be a cytokine rather than an enzyme. Moreover, at the level of polyclonally responding T cells. inhibition of PG synthesis due to T‐cell activation by mycobacterial antigens was shown to depend on prior mycobacterial immunization. Arthritogenic T‐cell clone A2b also showed PG synthesis inhibitory effects when co‐cultured with cartilage alone. The inhibitory activity was shown to be unrelated to the degree of T‐cell proliferation. We conclude that antigen‐specific T‐cell activation may be one of the initiating events leading to cartilage damage in arthritic processes. The measurement of T‐cell‐mediated PG synthesis inhibition may be a more sensitive and relevant assay for the detection of pathogenic T cells than T‐cell proliferation.

Initial immunochemical characterization of specific macrophage-arming factor

SummaryThis paper describes the initial immunochemical characterization of specific macrophage-arming factor (SMAF). SMAF is an antigen-specific factor that is released by (sensitized) T lymphocytes after contact with the specific antigen. It renders macrophages specifically cytotoxic. The specificity is dependent on the tumor-mouse combination. In allogeneic systems the specificity is H-2-directed, whereas in the syngeneic systems the specificity is tumor-specific. SMAF has a molecular mass of 65–85 kDa (established by gel filtration). By affinity chromatography SMAF could not be adsorbed with anti-(κ + λ light chain) immunoglobulins or anti-IgG from SMAF-containing supernatants. SMAF could be adsorbed with the monoclonal antibody 14–30 (directed against specific T-cell factors), and could be eluted from columns containing the latter. Furthermore, SMAF could also be adsorbed with and eluted from affinity chromatography columns to which specific tumor cell membranes or KCl extracts of these tumor cell membranes were coupled. Other tumor cell membranes could not adsorb SMAF. Together these data show that SMAF is not an antibody but a T-cell factor with an antigen-specific recognition site.

Differential responses of old human cartilage explants to synovial- and mononuclear-cell factors

SummaryTo investigate mechanisms of cartilage destruction that may apply to rheumatoid arthritis, young and old human, and young porcine, articular cartilage was cultured for 8 days and the effects on proteoglycan (PG) metabolism of normal synovium supernatant (NSS), rheumatoid synovial fluid (RFL), and blood mononuclear cell supernatant (MCF) were studied. The effects were chondrocyte-mediated. An inverse correlation was found between baseline net PG synthesis and the effect of NSS on PG synthesis. Responses of young (porcine and human) cartilage were similar. In young cartilage the three agents induced PG depletion by suppression of net PG synthesis. In old cartilage NSS and RFL induced PG depletion, whereas MCF did not. In cartilage of low baseline net PG sysnthesis, NSS and MCF stimulated both PG release and PG synthesis; NSS stimulated predominantly PG release, and MCF predominantly PG synthesis. In conclusion, young and old human cartilage differ in the quality of their in vitro response to potentially catabolic factors. This may be due to the difference in baseline net PG synthesis. Synovial extracts differ from mononuclear-cell supernatants in their effects on old cartilage. It is suggested that this is caused by the presence, in different relative amounts, of factors that influence either PG synthesis or PG release.

Influence of rheumatoid synovial fluid and cells on proteoglycans in human cartilage explants. Modulation by piroxicam.

SummaryWe have studied the effects of cell-free rheumatoid synovial fluid (RASF) and the conditioned medium (CM) from these cells on the proteoglycans (PGs) of normal human cartilage and the influence which piroxicam might have on these processes. Both RASF and the CM from RASF cells enhanced the PG release from the cartilage explants. The effects of the above mentioned fluids on the cartilage PG content depended on the metabolic state of the cartilage i.e. correlated inversely with the PG synthesis. Whether this was due to the presence of anabolic and catabolic factors in these fluids is discussed. Piroxicam had no adverse effect on the PGs of human cartilage in vitro. Piroxicam prevented the cartilage PG depletion when it was induced by the CM from RASF cells.

Degradation of human cartilage by cytokines in vitro.

past history of ischaemic heart disease and was taking ibuprofen, digoxin, and frusemide. On examination a large fluctuant swelling of the shoulder extending posteriorly over the right deltoid and scapula was seen. The shoulder was cool but swollen and tender with limited movement. An x ray examination of the joint initially seemed to support the clinical diagnosis of Milwaukee shoulder with areas of abnormal lucency and sclerosis in the humeral head. Attempted aspiration of the joint was unsuccessful but a sample from the scapular swelling contained frank pus. On microscopy

Tumor growth stimulatory macrophages induced by a subclinical bacterial infection in vivo.

Nonelicited peritoneal macrophages obtained from normal mice from our animal house unexpectedly expressed a strong tumor growth stimulatory effect in vitro. Macrophages expressing this stimulatory effect had an aberrant morphology compared to the morphology of normal macrophages as observed by electron microscopy.