Pierson J. Van Alten

Steroid sex hormones regulate the release of tumor necrosis factor by macrophages

Generally, females have been found to have a heightened immune response and a concomitantly higher incidence of autoimmune diseases compared to males. We have used male rat peritoneal macrophages (M phi) to study the effect of female sex hormones on tumor necrosis factor (TNF) release. The amount of TNF released by macrophages (M phi) exposed to 10(-2) and 10(-3) ng/ml of 17 beta-estradiol (E2) (35.1 +/- 7.3 and 23.2 +/- 2.5 units/ml, respectively) was significantly (P < 0.05; n = 9) greater than that released by untreated M phi. Progesterone (P) also significantly (P < 0.05; n = 8) stimulated a maximal TNF release (24.4 +/- 2.8 units/ml TNF) at 10(-2) ng/ml. On the other hand, the amount of TNF released by M phi exposed to E2 or P at concentrations greater than 10(-1) or less than 10(-4) ng/ml was significantly (P < 0.05) reduced compared to untreated controls. In contrast, testosterone did not significantly affect TNF release at any concentration. Within the physiological range of E2 and P concentrations, TNF release from M phi is finely regulated and dramatically affected by relatively small changes in hormone concentrations.

Steroid Sex Hormones and Macrophage Function: Modulation of Reactive Oxygen Intermediates and Nitrite Release

PROBLEM: In general, females have a more active immune response than do males. The effects of female sex hormones on lymphocytes have been studied extensively but their effects on macrophages are poorly understood.

Steroid Sex Hormones and Macrophage Function: Regulation of Chemiluminescence and Phagocytosis

PROBLEM: Female sex hormones modulate a variety of humoral and cell‐mediated immunologic functions. In this study, the effects of estrogen, progesterone, and testosterone on the chemiluminescence (CL) response and phagocytic ability of male rat peritoneal macrophages (Mφ) were examined.

Definition of fibronectin-mediated uptake of gelatinized latex by liver slices and macrophages

These studies show that both liver slices and macrophages carried out fibronectin concentration-dependent uptake of 125I-labeled gelatin-coated latex (test latex). Lack of phagocytosis of test latex by liver slices was shown directly by electron microscopy and indirectly by trypsin treatment, which caused the release of all test latex taken up in response to fibronectin. Inhibitors of phagocytosis did not alter this uptake. On the other hand, trypsin released only a portion of test latex from macrophages. Inhibitors of phagocytosis did not effect the released radioactive particles from macrophages but greatly reduced the trypsin-resistant radioactivity, taken as representing phagocytized particles. Opsonization of test latex with fibronectin did not require heparin but its association with liver slices occurred only in the presence of heparin. Macrophages, however, readily bound and internalized the opsonized test latex and heparin only potentiated these reactions. Gelatin competed with test latex for fibronectin for opsonization, but did not inhibit binding and phagocytosis of fibronectin-test latex complexes. Finally, soluble fibronectin-gelatin complexes did not compete for binding and phagocytosis of fibronectin-test latex complexes. Thus, fibronectin concentrated on the surface of latex is preferred for interaction with the fibronectin receptor of macrophages. Gelatin, however, was not essential for this reaction, because fibronectin directly coupled to latex was also readily taken up.

Natural antibody to sheep erythrocytes in bursectomized chickens.

Summary Chickens surgically bursectomized 2 days before hatching or on the day of hatching were assayed at 4 to 6 wk of age for their ability to react with sheep erythrocytes. Bursectomized and normal, unoperated chickens were tested using immunocytoadherence technique, hemolytic plaque, serum hemagglutinin and hemolysin assays. These tests showed that both groups of bursectomized chickens possessed “naturally” occurring rosette forming and hemolytic plaque producing spleen cells in numbers comparable to those of normal chickens. The serum hemagglutinin and hemolysin titers also were of the same magnitude in bursectomized and normal chickens. From our results we suggest that bursectomy may deplete a population of antigen-reactive cells which are necessary for the propagation of a normal response to antigenic stimulation at 4 or more wk of age.

Mercuric Bromphenol Blue Staining of Precipitin Lines in Agar

Lines formed by antibody-organ antigen reactions are stained particularly well by a modification utilizing the mercuric bromphenol blue (MBB) mixture of Mazia et al. (Biol. Bull., 104: 57-67, 1953). The agar covered slides are placed overnight in 0.85% NaCI at 4 C, followed by washing for 2 hr in 0.85% NaCI at 25 C. They are then rinsed for 10 min in distilled water, and dried overnight at 37 C. The precipitin lines are fixed by immersing the slides for 25 min in 95% alcohol, followed by 5 min hydration in distilled water. They are stained for 25 min in MBB mixture (HgCI2, 10 gm; bromphenol blue, 0.1 gm; 95% ethanol, 100 ml). Excess stain is removed by immersing in acidified alcohol (95% ethanol, 98 ml; glacial acetic acid, 2 ml). Finally, the slides are passed through alcohol and xylene, and resin-mounted under coverslips.

Defects in monocyte chemotaxis in patients with neoplastic disease

The immuno-surveillance system is generally believed to detect and deter the spread of neoplastic disease. As major effecters of this complex system, infiltrating macrophages or monocytes accumulate at tumor sites and are thought to participate directly in tumor destruction [26, 381. Macrophage migration plays an essential role in such cell-mediated immune responses and in inflammatory reactions [65]. Studies of experimental tumor models have shown that macrophage migration is reduced in tumor-bearing animals [ 34, 41, 481. The migration of macrophages into sites of inflammation and macrophage accumulation around tumors are mediated primarily by soluble and surface-adherent chemical stimuli [40,60]. Soluble factors have been used for in vitro assays of directional locomotion, i.e., chemotaxis, to evaluate leukocyte function in tumor patients. Chemotaxis assays have employed primarily lymphokines (lymphocyte derived chemotactic factors) or complement fragments (C5a) as attractants for human monocytes [9, 44, 601. In addition, we have recently used a third group of chemoattractants, the synthetic formylpeptides, to study the migration of tumor patient neutrophils (PMN) and monocytes [68]. Chemotaxis of PMN from tumor patients is generally normal but a deficiency is clearly evident in the chemotaxis of monocytes from patients with malignant disease. In this review, we discuss the methodology used for leukocyte chemotaxis studies, summarize the data currently available on the chemotaxis of tumor patient leukocytes, and assess the biological and clinical significance of the leukotactic defect seen in these patients.

Antigenic changes in developing hamster brain using antisera to myelinated and unmyelinated brain.

Abstract Saline-perfused brain tissue of either adult or 5-day neonatal hamsters, homogenized in saline solution and complete Freunds adjuvant, was injected into a separate series of Polish rabbits. “Fresh”, perfused adult brain and perfused adult brain which had undergone alternate freezings and thawings were compared for use in antisera production and in precipitin reactions. Five tissues other than brain were utilized for observing cross-reactions and for inhibition studies. Sera from rabbits injected with perfused fresh adult brains reacted negligibly in agar with perfused adult and 5-day old (unmyelinated) brain. Sera from rabbits injected with perfused, sequentially frozen-thawed adult brain or perfused, fresh 5-day old brain formed a series of precipitin lines in agar with adult frozen-thawed or fresh 5-day old brain. Antisera to 5-day brain always was stronger. Cross-reactions and inhibition tests with other tissues indicate that the 5-day brain antiserum is organ-specific whereas adult brain antiserum is not. These studies indicate that in nervous tissue the antigenic sites have become masked during the process of myelination to such a degree that good humoral antibody production is inhibited. Freeze-thawing tends to “unmask” adult brain antigen so that it reacts immunologically more like unmyelinated brain. Our observations also suggest that erroneous conclusions can result from studies on the development of antigenicity in nervous tissue which use only adult brain antisera as a level for reference.