Delayed hypersensitivity in vitro: its mediation by cell-free substances formed by lymphoid cell-antigen interaction.

Abstract

It has long been thought that delayed hypersensitivity is mediated by cells or cell-associated substances. In an attempt to investigate the mechanism of delayed hypersensitivity, in vitro studies using the inhibition of cell migration by specific antigen as an assay have been carried out. In this in vitro system, it has been shown that peritoneal exudate cells taken from guinea pigs exhibiting delayed hypersensitivity and placed in capillary tubes are inhibited from migration by specific antigen." 2 Moreover, when mixed populations of normal and sensitive cells were prepared, it was observed that if as few as 2.5 per cent of the cells in a population were from a sensitive animal, the whole population (97.5% normal cells) would be inhibited by antigen.3The results of recent experiments suggest that the lymphocyte is the specifically sensitive cell in this system. It was found that lymph node cells (approximately 95% lymphocytes) obtained from sensitive guinea pigs will, when mixed with a population of normal peritoneal exudate cells, cause the whole population to be inhibited by specific antigen.4 It is of note that such sensitive lymphoid cells, when assayed alone in culture, are not themselves inhibited from migrating by antigen.Experiments were initiated in an attempt to determine the manner by which these sensitive lymphoid cells achieved their effect. The results, here reported, demonstrate that, following incubation of sensitive lymphoid cells with specific antigen for 24 hr, a nondialyzable substance is detected in the cell-free supernatants which inhibits the migration of normal peritoneal cells.Materials and Methods.-Sensitization: The antigens used were ovalbumin (Worthington) and o-chlorobenzoyl chloride conjugated to bovine gamma globulin (OCBC-BGG) by the method of Benacerraf and Levine,5 kindly supplied by Dr. Y. Borel. Guinea pigs of the Hartley strain weighing 500-800 gm were sensitized with the appropriate antigen diluted in saline and emulsified in an equal volume of complete adjuvant (Difco H37Ra). Each animal received a total dose of 100 ,ug of antigen distributed into the four footpads, 0.1 ml per footpad.Tissue culture media: The basic tissue culture media used throughout were minimal essential media, Eagle 12-125 (Microbiological Associates, Bethesda, Md.) containing 15% normal guinea pig serum and 85 units of penicillin and 85 jAg of streptomycin/ml.Preparation of lymph node cell suspensions: Twelve to twenty one days after sensitization axillary, inguinal and popliteal lymph nodes were obtained aseptically from guinea pigs which had been anesthetized with ether and exsanguinated by cardiac puncture. The nodes were diced into tissue culture medium, teased gently with mouse-toothed forceps, and the resulting cell suspension was pipetted into centrifuge tubes. The suspension was allowed to stand for 4 min so that the tissue fragments settled by gravity. The supernatant was then removed to a fresh tube. After three such settlings the cell suspensions were essentially free of tissue fragments. ...

ASSAY OF HUMAN LYMPHOKINES IN VITRO. EVIDENCE FOR A MIGRATION STIMULATION FACTOR (MStF) WHICH INTERFERES WITH THE MACROPHAGE MIGRATION INHIBITION ASSAY

“…as yet, tmeharaeterized and apparently immunologically non-speeific migration stimtilatiort factors which act in vitro. As was shown in David's (1966) results, some ot the non-specific migration stimulation could be attribttted to variations between the protein concentration of "control" and "test" cultures. It remains to be shown whether other enhancing factors such as that described by Jatiis and Bach (1970), wliich potentiated antigen-induced lymphocyte transformation, also interfere with the MMI test.…”

“…In some studies (Svejcar and Johanovsky, 1961;David, 1966), a non-specific migration stimulation was occasionally obscr\'ed when leucocvte.s were cultured with antigens to which they were not sensitized. Other workers (Federlin, Maini, Russell and Dumonde, 1971) have suggested that migration stimulation occurs when sensitized leucocytes are challenged in vitro with snb-optimal doses of antigen.…”

Promoter region polymorphism of macrophage migration inhibitory factor is strong risk factor for young onset of extensive alopecia areata

“…MIF was originally identified as a lymphokine that concentrates macrophages at inflammatory loci, is a potent activator of macrophages in vivo and is considered to play an important role in cell-mediated immunity. 3,10 Since the molecular cloning of MIF, 11 this cytokine has been reevaluated as a proinflammatory cytokine and pituitaryderived hormone that potentiates endotoxemia. 12,13 It has been shown that T cells and macrophages secrete MIF in response to various proinflammatory stimuli such as endotoxins.…”

MIF loss impairs Myc-induced lymphomagenesis

“…24 Other retroviral vectors included those encoding CDK4, E2F1, and E2F4. For viral infections, 10 5 MEFs plated onto a 6-cm dish were incubated overnight with an appropriate amount of the corresponding retroviral stock. Cells were analyzed for the corresponding protein levels 3-4 days postinfection.…”

“…[1][2][3][4] MIF was originally identified as a product of activated T cells. 5 However, subsequent work showed that MIF is almost ubiquitously expressed in various tissues, including neural, epithelial, and hematopoietic lineage cells. [6][7][8][9] Studies using neutralizing antibodies implicated MIF in the development of pathologies of acute and chronic inflammation such as allograft rejection, wound repair, glomerulonephritis, inflammatory arthritis, and septic shock.…”