David L. Rosenstreich

The role of macrophages in the acute-phase response: SAA inducer is closely related to lymphocyte activating factor and endogenous pyrogen

Abstract An increase in the concentration of the acute-phase reactant, serum amyloid A (SAA), following endotoxin treatment, is a consequence of the action of lipopolysaccharide (LPS) on macrophages to produce a monokine, the SAA inducer, which in turn, triggers SAA synthesis by hepatocytes. We have found that murine SAA inducer is closely related, if not identical, to murine lymphocyte activating factor (LAF), otherwise known as Interleukin 1 (IL 1). Furthermore, both rabbit endogenous pryrogen (EP), which is believed to be identical to LAF (IL 1), and human LAF (IL 1), induced elevated SAA concentrations in C3H/HeJ mice. Antiserum previously shown to block both pyrogenic and thymocyte proliferating activities of the species of rabbit EP exhibiting an isoelectric point of pH 7.3 (EP 7), also blocked the SAA inducing activity of EP7. Phenylglyoxal treatment of highly purified murine LAF (IL 1) abrogated both thymocyte proliferating activity and the SAA inducing activity. These studies support and extend previous reports suggesting that within 2 hr of an inflammatory stimulus, macrophages produce a monokine that acts systemically to alter body temperature, activate T cells, and induce hepatic protein synthesis of acute-phase reactants.

Phorbol myristic acetate stimulates LAF production by the macrophage cell line, P388D

Abstract Phorbol myristic acetate markedly stimulated LAF production by the murine macrophage cell line, P388D 1 . This effect was both time and concentration dependent. Other analogs, such as phorbol didecanoate and to a lesser extent, phorbol dibenzoate also enhanced LAF production, but the parent compound, phorbol, was inactive. The PMA induced supernatant LAF had a molecular weight of 16,000 daltons and exhibited charge heterogenity on DEAE cellulose. Since PMA is a small compound of defined structure which induces the production of large amounts of LAF it should be a useful tool with which to probe the biology and biochemistry of LAF.

The distribution of histocompatibility antigens on T and B cells in the guinea pig

SUMMARY The distribution of histocompatibility (H) antigens on different lymphocyte populations in the guinea pig was examined with alloantisera prepared by cross immunizing strain 2 and strain 13 animals. These alloantisera killed only 30-40% of lymph node cells, and those cells that remain alive after killing by the alloantisera are markedly depleted in immunoglobulin-bearing B lymphocytes. Absorption studies of the anti-2 serum demonstrated that the alloantigen content of lymph node cells was 7-8 times that of thymocytes, while the alloantigen content of L2C leukemia cells, a pure population of malignant B cells, was 25-30 times that of thymocytes. Furthermore, pure B lymph node cells, prepared by treating lymph node cells with an anti-T cell serum and C, were more efficient than normal lymph node cells in the absorption of the alloantisera. Finally, those lymph node cells which remain alive after treatment with the alloantisera and C failed to respond to endotoxin, a B cell mitogen, whereas these same cells responded normally to phytohemagglutinin, a T cell mitogen. These results demonstrate an increased content of H antigen on the B lymphocyte compared to the T lymphocyte. Since H antigens play a role in the cooperation between B and T cells, information as to the distribution of H antigens on these cells may have important implications for cell to cell interaction in the immune response.

7 – The Role of Macrophages in the Activation of T and B Lymphocytes in Vitro

Publisher Summary This chapter discusses the mechanisms of action of macrophages, the role of macrophages in antigen-induced lymphocyte proliferative responses in vitro, macrophage dependence of antigen-induced T lymphocyte proliferation in vitro, macrophage dependence of B lymphocytes for proliferation induced by thymus-dependent antigens, and mechanisms of macrophage action in antigen-induced proliferation of T and B lymphocytes. It has been observed that syngeneic macrophages and lymphocytes interact more effectively than allogeneic cells, indicating that something more than antigen presentation is involved in the interaction. Using inbred guinea pigs and antigens, which were under strict genetic control, Rosenthal and Shevach found that there had to be histocompatability between macrophages and lymphocytes to obtain lymphocyte activation. This observation supports the idea that direct macrophage–lymphocyte contact is required for activation. However, it also suggests that there are more sophisticated requirements for activation beyond a passive presentation of antigen by macrophages so that the lymphocyte must recognize additional determinants on the macrophage surface in addition to antigen.

Analysis of the systemic corticosteroid sensitivity of patients with primary open-angle glaucoma.

Patients with primary open-angle glaucoma have an ocular and systemic sensitivity to corticosteroids. We adapted a cellular assay that used peripheral blood lymphocytes to detect this corticosteroid sensitivity in vitro in a microtiter assay. It reduced the time, cost, and amount of blood required to examine a patient. We examined ten subjects on three separate days and demonstrated that the reliability of one 50% inhibitory concentration was about 76%. We then studied 25 patients with primary open-angle glaucoma and 25 control subjects using this in vitro assay. The patients with primary open-angle glaucoma were significantly more sensitive to corticosteroids than the control subjects (P less than .001).

Resistance of C3H/HeJ Mice to Lethal Challenge with Herpes Simplex Virus

Summary In vitro replication of herpes simplex virus (HSV) in murine spleen cells requires simultaneous cell stimulation with a B-cell mitogen such as lipopolysaccharide (LPS). As expected, spleen cells of LPS-unresponsive C3H/HeJ mice did not support HSV replication in LPS-pretreated cultures, while spleen cells from closely related but LPS-responsive C3HeB/FeJ did. More importantly, the C3H/HeJ strain was found to be intrinsically resistant to HSV infection in vivo. After intraperitoneal (ip) inoculation, HSV was 50-120 times more virulent for C3HeB/FeJ mice than for the C3H/HeJ strain. This resistance appeared to be due to a failure of HSV to replicate in C3H/HeJ peritoneal cells, since after ip infection with HSV, recovery of virus was higher and more consistent from peritoneal exudate cells of C3HeB/FeJ mice than from C3H/HeJ mice. In addition, no difference in lethality was observed between these two strains after a direct intracerebral inoculation of HSV. This observation that LPS-unresponsive mice are intrinsically resistant to lethal HSV infection, coupled with the LPS requirement for HSV replication in vitro, suggests an important but as yet unexplained link between host sensitivity to HSV and to LPS.

Location of theSas-1 locus on mouse chromosome 1

Using three sets of recombinant inbred strains (BXD, BXH, and BXJ), we found the locus controlling an antigenic substance (Sas}-1) in murine serum to be closely linked to the Chromosome-1 marker,Dip-1. This linkage was confirmed by an analysis of backcross linkage. The BXD and backcross data suggest that the gene order isId-1-Dip-1-Sas-1-Mls. Data from the three sets of RI strains and the 32 backcross mice lead to the estimate that the recombination frequency betweenDip-1 andSas-1 is 0.030 ±0.015.

Cellular Sources of Lymphokines

Publisher Summary This chapter discusses the cellular sources of lymphokines. Lymphokines consist of a large, heterogeneous family of molecules, all of which in various ways regulate the immune response. The chapter reviews the lymphokines produced by lymphoid cells, that is, B- and T-lymphocytes, macrophages, and lymphoid cell lines. Lymphokines are the products of activated lymphoid cells. For the most part, both T- and B-lymphocytes are capable of producing lymphokines with similar functional properties. Lymphokine production is generally as a result of cell activation. The process of activation of lymphokine production is similar to the activation that results in cell proliferation or antibody production. The mechanisms by which B-lymphocytes are activated to produce lymphokines differ from those of T-cells. Lymphokines have been detected in the culture supernatants of both B- and T-, normal and malignant, and human and animal cell lines. There are at least three major determinants in the process of lymphocyte activation: (1) the nature of the signaling agent, (2) the T-cell requirement for B-cell activation by certain antigens, and (3) the requirement for macrophages as accessory cells.

Signal requirements for lymphocyte activation: Role of a T-cell growth factor produced by guinea pig peritoneal exudate lymphocytes

Abstract Peritoneal exudate lymphocytes (PEL) from immunized guinea pigs, when pulsed with antigen, rapidly release a T-cell stimulatory factor (TSF). TSF nonspecifically enhances the proliferation of purified guinea pig T cells in the presence of another signal such as PMA, PHA, or Con A. On a per cell basis, antigen-pulsed PEL produce about 14 times more activity than similarly stimulated lymph node lymphocytes. Several lines of evidence support the view that TSF is the guinea pig equivalent of TCGF (IL-2). TSF containing supernatants have IL-2 activity when assayed on the IL-2-dependent CT6 cell line. When TSF containing supernatants were absorbed with CT6 cells, there was a significant decrease of both TSF activity as assessed on guinea pig T cells as well as IL-2 activity as assessed by the CT6 assay. Additionally, partially purified human and mouse IL-2 have TSF activity, while the macrophage product, IL-1, has no TSF activity. After chromatography on a S-200 column, TSF activity and IL-2 activity coelute at an apparent molecular weight of 19,000.

The Lymphocyte Uropod: A Specialized Surface Site for Immunologic Recognition

Functional specialization of the cell surface of motile protozoa such as the Paramecium and amoeba are generally recognized (Ambrose and Forrester, 1968; Wolpert and Gin-gell, 1968). Less well appreciated is the existence of differentiated surface membranes on mammalian leukocytes. In this laboratory we have been interested in a modification of the lymphocyte surface called the uropod, an area consisting of microvillus projections of cell membrane adjacent to the golgi-associated cell pole. The cytoplasm contained in the uropod is rich in microtubules, mitochondria, endoplasmic reticulum, and numerous endocytic vesicles, and is quite distinct from the pseudopod region which contains few such subcellular organelles. Moreover, the uropod is not engaged in cell motility as is the pseudopod. The lymphocyte surface membrane functions in immunologic recognition, cell cooperation, and cell-mediated cytotoxicity in an as yet undefined manner. Circumstantial evidence suggests that the uropod may represent the site of such interactions.