Studies on the regulation of lymphocyte reactivity by normal and activated macrophages.

To determine the potential role of macrophages as regulators of the immune response, the effect of mouse peritoneal macrophages on transforming mouse spleen lymphocytes was investigated. Mitogen and antigen stimulated lymphocyte transformation, as measured by DNA synthesis, was enhanced by all concentrations of normal macrophages tested, but only by low concentrations of activated macrophages. High concentrations of activated macrophages markedly inhibited lymphocyte transformation. This inhibition occurred whether lymphocyte DNA synthesis was measured by incorporation of [³H]TdR or of ³²P. Activated macrophages cultured with lymphocytes within 4 hr of being removed from the peritoneal cavity inhibited lymphocyte transformation. When activated macrophages were cultured alone for 24 or more hours before addition of lymphocytes, enhancement of transformation was noted. Once lymphocytes were exposed to activated macrophages, they could not be induced to undergo transformation in the presence of Con A. Whereas heat-killed activated macrophages, which appeared intact morphologically, lost their capacity to inhibit lymphocyte transformation, macrophages treated with mitomycin C to inhibit DNA synthesis retained this capacity. Syngeneic and allogeneic macrophages had similar inhibitory ability. Supernatants from cultures of many cell types (including normal or activated macrophages, lymphocytes, lymphocytes plus macrophages, and L cells) inhibited [³H]TdR incorporation by both mitogen stimulated lymphocytes and tumor cells. These studies demonstrate the capacity of macrophages to regulate lymphocyte transformation in vitro and suggest a role for these cells as regulators of cell-mediated immunity in vivo.     

Effect of tumor cell culture media and sera from tumor hosts on spreading,phagocytosis, and antitumor cytotoxicity of C. parvum-activated murine macrophages

Summary We investigated whether the media of tumor cell cultures and sera from tumor-bearing hosts exert inhibitory effects upon macrophage spreading, phagocytosis, and cytotoxicity. Peritoneal macrophages from normal and Corynebacterium parvum-treated C3Hf/Bu mice were incubated in media from a syngeneic mammary carcinoma, allogeneic Ehrlich ascites carcinoma, and human malignant melanoma or cervical carcinoma cell cultures, or in the serum of hosts bearing these tumors. Such media and sera inhibited the ability of both normal and C. parvum-activated macrophages to spread on glass surfaces and to ingest latex particles. In contrast, they did not interfere with the in vitro destruction of tumorigenic L929 cells by C. parvum-activated macrophages. Media of murine embryo fibroblasts and a human benign tumor and sera from healthy mice or humans did not, however, inhibit either of the macrophage functions tested.     

Effects of immune lymphocyte products and serum antibody on the multiplication of Toxoplasma in murine peritoneal macrophages.

In vitro studies on cell-mediated immunity against Toxoplasma infection were carried out by estimating the ability of antigenically stimulated lymphocytes. Splenic lymphocytes from normal mice and from hyper-immunized mice were cultured in the presence or absence of Toxoplasma lysate antigen. The cell-free supernatant fluids from the lymphocyte cultures were assassed for their ability to alter the functional capacities of normal macrophages. When glycogen-induced peritoneal macrophages were incubated with the culture supernatant from immune lymphocytes reacting with specific angigen, the intracellular multiplication of the organisms was inhibited remarkably. In contrast, the addition of antitoxoplasma antibody to culture medium had no effect on the enhancement of phagocytic activity of culture macrophages. However, when extra-cellular organisms were exposed to fresh or heat-inactivated immune serum just before infection of the monolayers, the intracellular multiplication of Toxoplasmas was inhibited significantly by either activated or normal macrophages.     

Cytotoxic lymphocytes induced by soluble factors derived from the medium of leukocyte cultures.

Studies were undertaken to evaluate the cytotoxic capacity of human peripheral blood lymphocytes activated by either supernatants (CFM) derived from lymphocyte cultures or lymphocytes treated for 60 min at 45 degrees C. The effect of the addition of heat-treated cells on the cytotoxic activity of CFM-induced effector cells was also studied. CFM from either unmixed or mixed cultures of lymphocytes was capable of activating cytotoxic effector cells. These effector cells could kill any allogeneic target cells but failed to effect cytotoxicity on the target cells autologous to the responding cells. Both the heat-treated cells and CFM from cultures of these cells also activated lymphocytes to cytotoxic effector cells having specific receptors for nonself antigens. The question of whether heat-treated cells activate cytotoxic cells by themselves or through secreted soluble factor cannot yet be clearly answered. The findings of the present investigation suggest that expression of cytotoxicity induced in MLC is not necessarily restricted to the target cells syngeneic to the stimulator cells, but can be extended to any allogeneic target cells by the indirect effect of soluble factor secreted from stimulated cells that causes a polyclonal activation of cytotoxic precursors in the responding cell populations. The present findings also emphasize the need for caution in the use of heat-treated lymphocytes as innocent-bystander cells in MLC to provide additional cytotoxic specificities in the responder cells, since heat-treated cells alone can activate lymphocytes to cytotoxic effector cells that kill any allogeneic target cells.     

Induction of macrophage-mediated cytolysis of neoplastic cells by mycoplasmas

Unexpected cytolysis was encountered when nonactivated murine peritoneal macrophages were cultured with [³H]TdR-prelabeled syngeneic or allogeneic tumor cells at a 10:1 ratio. The level of specific cytolysis reached 70% within 48 hr of cocultivation. Similar killing was observed whether the macrophages were derived from untreated, thioglycollate-treated, or germ-free mice. Cytolytic activity was also demonstrated when bone marrow-derived or peritoneal macrophages from 9- and 5-day in vitro cultures, respectively, were employed rather than freshly harvested peritoneal macrophages. Thus, the macrophage-mediated killing was neither the result of in vivo preactivation nor a consequence of the presence of lymphocytes in the assay. Moreover, macrophages derived from different strains caused similar effects. Our study revealed that the neoplastic target cell cultures susceptible to cytolysis by nonactivated macrophages were contaminated with mycoplasma. A mycoplasma was isolated from the supernatant of a culture of the A9HT fibrosarcoma line, identified as Mycoplasma orale, and cultivated. Addition of viable mycoplasma from that isolate to mixed cultures of thioglycollate-elicited macrophages and [³H]TdR-prelabeled mycoplasma-free target cells resulted in specific cytolysis of transformed A9 cells, but not of normal mouse fibroblasts. The level of macrophage-dependent cytolysis correlated with the number of viable mycoplasma cells added and was higher than that attained by activation with LPS at optimal concentration. Similar specific cytolysis was observed with heat-killed mycoplasmas. Our results demonstrate that mycoplasmas may cause selective macrophage-mediated cytolysis of neoplastic but not of normal target cells, perhaps via activation of the macrophages. It is suggested that undetected infection of experimental systems by mycoplasmas may account for some reports on lysis of neoplastic cells by nonactivated macrophages.     

The role of tumor-derived cytokines on the immune system of mice bearing a mammary adenocarcinoma. I. Induction of regulatory macrophages in normal mice by the in vivo administration of rGM-CSF

Using a dimethylbenzanthracene-induced immunogenic nonmetastatic murine mammary adenocarcinoma in BALB/c mice, our previous work has shown that splenocytes from tumor bearers have reduced responses to both mitogens and Ag including tumor-associated Ag. NK and cytotoxic T cell activities are also reduced in splenocytes of tumor bearers. Mac-1+2+ macrophages induced in mammary tumor bearers are capable of down-regulating lymphocyte responses to mitogens and tumor-associated Ag by cell to cell contact interaction and increased PGE2 production. We have found that the tumor constitutively releases a granulocyte-macrophage (GM)-CSF-like factor in vivo and in vitro, which may be responsible for the systemic increase in cells of the macrophage lineage in tumor-bearing mice. A tumor cell line established from the in vivo tumor expresses and releases GM-CSF as shown by Northern and Western blot analyses. Daily i.p. injections for 3 wk of 10,000 U of rGM-CSF into normal mice induces hemopoietic and immunologic alterations similar to those observed in tumor bearers. Mac-1+ and/or Mac-2+ macrophages can also be detected in the spleens and bone marrow of the mice treated with rGM-CSF. Additionally, splenocytes from rGM-CSF-treated mice have reduced responses to mitogens and their peritoneal exudate cells can cause in vitro down-regulation of proliferative responses of lymphocytes from normal mice. The suppression can be partially reversed by the addition of indomethacin to the cultures suggesting that PGE2 may contribute to the effect. rGM-CSF enhances the in vitro release of PGE2 by the spleen, bone marrow, and peritoneal cells of normal mice. These data indicate that the high levels of GM-CSF constitutively produced by the tumor may be responsible for the hemopoietic changes and immunologic alterations observed in tumor-bearing mice.     

In vitro studies on transplantation immunity. II. The migration inhibition in homograft reactions in guinea pigs

The migration of peritoneal exudate cells from guinea pigs exhibiting transplantation immunity is inhibited in the presence of donor antigens. This inhibition of migration is demonstrable whether the donor transplantation antigens are presented in the form of viable cells (peritoneal exudate cells) or as particulate subcellular antigens (spleen microsomes). A greater degree of inhibition was observed when transplantation immunity was induced with lymphoid cells in Freud's adjuvant compared to sensitization with orthotopic skin grafts. There was no inhibition of migration in mixtures of normal allogeneic cells or when peritoneal cells from guinea pigs exhibiting tuberculin hypersensitivity were mixed with similar cells from normal animals. Finally, supernatants from cultures of sensitive lymphocytes plus donor antigens inhibited the migration of normal peritoneal cells indicating the presence of migration inhibitory factor (MIF) activity.     

Staphylococcal enterotoxin B induced release of macrophage migration inhibition factor from normal lymphocytes

Staphylococcal enterotoxin B (SEB), a potent lymphocyte mitogen, inhibits migration of peritoneal exudate cells from most guinea pigs but does not inhibit migration of purified macrophages. Experiments were designed to test the ability of highly purified SEB to induce normal lymphocytes to release migration inhibition factor (MIF). Supernatants of lymph node lymphocytes cultured with SEB inhibited the migration of purified macrophages, indicating the release of a migration inhibition factor. Mitomycin-C blocked the SEB-induced release of MIF. SEB-induced MIF localized in the albumin fraction on Sephadex G-200 chromatography. Antibody to SEB specifically blocked the inhibitory effect of SEB on migration of normal guinea pig peritoneal exudate cells.     

Activation of peritoneal macrophages by lysophosphatidylcholine

Lysophosphatidylcholine (lyso-PC), a product of inflammation induced by infectious and other agents, is able to stimulate mouse peritoneal macrophages to ingest target cells coated with IgG but not IgM regardless of the presence of complement. In vitro treatment of mouse resident peritoneal macrophages (adherent cells) alone with lyso-PC stimulated spreading activity but did not enhance ingestion activity of macrophages. However, when mixed cultures of adherent and nonadherent (lymphocytes) cells were treated with lyso-PC, macrophage ingestion activity of IgG-coated target cells (i.e., via Fc-mediated ingestion) was markedly enhanced. Analysis of lyso-PC activation process of macrophages for ingestion activity suggests that nonadherent (lymphocytes) cells are required for the induction of the manifestation of ingestion capacity. This requirement was also met by addition of untreated nonadherent cells to treated adherent cells. Thus, the activation mechanism of macrophages by lyso-PC for ingestion requires contribution of lymphocytes to promote enhanced ingestion activity. Since lyso-PC is a metabolite of a representative membrane phospholipid, we propose that lyso-PC and other lysophospholipids are mediators for activation of macrophages regardless of the type of inflammation-causative agent.     

Generation of specific helper cells and suppressor cells in vitro for the IgE and IgG antibody responses.

Normal splenic lymphocytes from BDF1 mice were cultured on ovalbumin (OA)-bearing syngeneic peritoneal adherent cells for 5 days and their subsequent helper function was tested by an adoptive transfer technique. Lymphocytes harvested from the culture were mixed with DNP-KLH-primed spleen cells and transferred into irradiated syngeneic mice followed by challenge with DNP-OA. The results showed that the cultured lymphocytes has helper function for both IgE and IgG anti-DNP antibody responses. Depletion of mast cells and T cells in the peritoneal adherent cell preparations did not affect the generation of helper cells in the culture. The helper function of the cultured lymphocytes was abolished by the treatment with anti-theta antiserum and complement and was specific for ovalbumin. The OA-specific helper T cells were generated in vitro by the culture of a T cell-rich fraction of normal spleen on T cell-depleted OA-bearing peritoneal cells. If the normal splenic lymphocytes or T cell-rich fraction were cultured with 10 mug/ml of OA in the absence of macrophages, cultured lymphocytes lacked helper function. The transfer of splenic lymphocytes or splenic T cells cultured with soluble OA to normal non-irradiated mice, however, suppressed both IgG and IgE antibody responses of the recipients to subsequent immunization with DNP-OA. The suppressor cells were sensitive to anti-theta antiserum and complement and their activity was specific for OA. The cultured cells transferred into normal mice did not suppress anti-hapten antibody response to DNP-KLH. Normal lymphocytes cultured on OA-bearing macrophages and had helper function in adoptive transfer experiments failed to suppress antibody response of non-irradiated recipients to DNP-OA. The results indicate that OA-bearing macrophages primed T cells and generated helper T cells, whereas the culture of normal lymphocytes with soluble OA in the absence of macrophages generated suppressor T cells.     

Mechanism and specificity of macrophage-mediated cytotoxity.

The cytotoxic effect of macrophages derived from alloimmunized mice (immune macrophages) was found to be immunologically specific. The immune macrophages killed only target macrophages carrying the alloantigens used for immunization in mixed macrophage cultures (MMC) under optimal conditions of contact between effector and target cells. T-sensitized lymphocytes, but not B cells, were capable of arming nonimmune macrophages and conferring upon them cytotoxic activity; the arming factor, which seemed to be a T mediator or T-cell receptor (membrane component) was removable by trypsin. Frequent rinsing or addition of hydrocortisone significantly decreased the cytotoxicity of the MMC. Pretreatment of peritoneal cells with anti-θ antisera and complement markedly decreased immune macrophage cytotoxic activity. It is suggested that the presence of a very small number of T-sensitized lymphocytes is required for strong cytotoxic activity to be manifested by the macrophages.     

Tumor therapy of neoplastic diseases with tumor cells and neuraminidase

The therapeutic effect of i.d. injection of tumor cells mixed with Vibrio cholerae neuraminidase (VCN) on tumor progression in dogs with spontaneous mammary tumors was investigated. The i.d. injections were performed in a chessboard-like manner: different numbers (10(5), 10(6), 10(7), and 10(8) of mitomycin-treated autologous tumor cells (M-TC) were each mixed with different amounts (10, 50, and 100 mU) of VCN. These different mixtures were injected i.d. at different sites in dogs on the day of resection of a part of multiple tumors.     

A unique antigen expressed on myeloid cells and acute leukemia blast cells defined by a monoclonal antibody

A murine hybridoma-derived monoclonal antibody, PM-81, was obtained from a fusion of cells of the NS-1 myeloma cell line with cells from a mouse immunized with the HL-60 promyelocytic leukemia cell line. This cytotoxic IgM monoclonal antibody was specific for myeloid cells. Employing indirect immunofluorescence and flow cytometry, we determined that this antibody reacts strongly with normal human granulocytes, eosinophils, and monocytes but not lymphocytes (including phytohemagglutinin-activated lymphocytes), null cells, red blood cells, or platelets. Moreover, the PM-81 antibody reacts with leukemia cells from 19 of 22 patients with acute myelocytic leukemia of all FAB subclasses, three of three patients with common acute lymphocytic leukemia, four of four patients with chronic myelocytic leukemia (CML) in myeloid blast crisis (terminal transferase (TdT)-negative) but did not react with cells from two patients with CML in lymphoid blast crisis (TdT-positive) or five patients with chronic lymphocytic leukemia. The myeloid cell lines HL-60, K562, KG-1, and U937 were all reactive with PM-81. The lymphoid lines CCRF-CEM and Daudi did not express PM-81 but HSB-2 was positive. The PM-81 antigen was absent on myeloid and erythroid progenitor cells as determined by their insusceptibility to complement-dependent lysis. In addition, only PM-81-unreactive cells were capable of colony formation. Furthermore, the PM-81 antibody does not appear to induce modulation of the antigen to which it binds. Thus, this monoclonal antibody appears to fulfill several criteria for clinical utility in the diagnosis and treatment of both acute myelocytic and acute lymphocytic leukemia.     

Interaction of rat peritoneal macrophages with homologous, sialidase-treated lymphocytes in vitro

The interaction in vitro between rat peritoneal macrophages and homologous, sialidase-treated lymphocytes was investigated. Lymphocytes were isolated from blood, thymus, and spleen on a density gradient. Total sialic acids obtained by acid hydrolysis were 10 nmol/10(8) lymphocytes, composed of 29% N-acetyl-neuraminic acid and 71% N-glycoloylneuraminic acid. Sialidase treatment released maximally 33% of membrane sialic acids. Lymphocytes were bound to peritoneal macrophages to an extent which increased in parallel with the amount of sialic acids released, whereas binding of untreated lymphocytes was not significant. This interaction was inhibited by free galactose and substances containing terminal galactose residues. Asialoorosomucoid with its oligoantennary sugar chains proved to be a 10(5) times more potent inhibitor of the interaction than lactose. The addition of homologous serum had no influence on binding. Electron microscopy revealed that vital lymphocytes were tightly bound to macrophages and only damaged lymphocytes appeared to be phagocytozed. The experiments demonstrate that the interaction between rat peritoneal macrophages and sialidase-treated lymphocytes is mediated by a macrophage receptor specific for galactose. This sugar is demasked on the surface of lymphocytes after the removal of terminal sialic acids. The role of this mechanism in cell recognition, elimination and homing of lymphocytes is discussed.     

Gamma interferon blocks gammaherpesvirus reactivation from latency in a cell type-specific manner

Gammaherpesviruses are important pathogens whose lifelong survival in the host depends critically on their capacity to establish and reactivate from latency, processes regulated by both viral genes and the host immune response. Previous work has demonstrated that gamma interferon (IFN-gamma) is a key regulator of chronic infection with murine gammaherpesvirus 68 (gammaHV68), a virus that establishes latent infection in B lymphocytes, macrophages, and dendritic cells. In mice deficient in IFN-gamma or the IFN-gamma receptor, gammaHV68 gene expression is altered during chronic infection, and peritoneal cells explanted from these mice reactivate more efficiently ex vivo than cells derived from wild-type mice. Furthermore, treatment with IFN-gamma inhibits reactivation of gammaHV68 from latently infected wild-type peritoneal cells, and depletion of IFN-gamma from wild-type mice increases the efficiency of reactivation of explanted peritoneal cells. These profound effects of IFN-gamma on chronic gammaHV68 latency and reactivation raise the question of which cells respond to IFN-gamma to control chronic gammaHV68 infection. Here, we show that IFN-gamma inhibited reactivation of peritoneal cells and spleen cells harvested from mice lacking B lymphocytes, but not wild-type spleen cells, suggesting that IFN-gamma may inhibit reactivation in a cell type-specific manner. To directly test this hypothesis, we expressed the diphtheria toxin receptor specifically on either B lymphocytes or macrophages and used diphtheria toxin treatment to deplete these specific cells in vivo and in vitro after establishing latency. We demonstrate that macrophages, but not B cells, are responsive to IFN-gamma-mediated suppression of gammaHV68 reactivation. These data indicate that the regulation of gammaherpesvirus latency by IFN-gamma is cell type specific and raise the possibility that cell type-specific immune deficiency may alter latency in distinct and important ways.     

Studies of latent cytomegalovirus infection: the macrophage as a virus-harboring cell

During chronic infection of mice with mouse cytomegalovirus (MCMV), the virus was isolated from various tissues by cocultivation with allogeneic mouse embryonic fibroblasts (MEF). Infectious virus was recovered from over 15% of the pancreases, salivary glands, kidneys, lacrimal glands, and spleens. When activated macrophages were obtained by intraperitoneal injection of peptone into mice infected 3 months earlier, they harbored MCMV. Macrophages or lymphocytes were infected with MCMV in vitro and injected into normal mice intravenously. The peritoneal cavities of these mice were then stimulated by peptone injection 3 months after the transfer, and peritoneal or splenic macrophages and lymphocytes were cocultured with allogeneic MEF. MCMV was recovered from the peritoneal and splenic macrophages and not from the lymphocytes.     

Mobility of surface proteins on normal rat macrophages and on a "macrophagelike" rat tumor

Peritoneal macrophages endocytosed their histocompatibility antigens (RT1), Fc receptors (FcR), and concanavalin A (Con A) receptors after cross-linking by ligands, but did not cap these membrane proteins. The 323N cell, a "macrophage like" tumor cell, under identical conditions capped its surface proteins. Experiments measuring fluorescence recovery after photobleaching showed that the mobile fraction of RT1 was significantly greater in 323N cells than in normal peritoneal macrophages. Presumably, the membrane proteins of 323N are not as tethered to the cytoskeleton, or, if so, are in a nexus that is not the same as that which occurs between membrane proteins of normal macrophages and the cytoskeleton. The mobility of RT1 on normal lymphocytes was also different from that of macrophages. These observations suggest that the movement of membrane molecules is determined by cell type and is regulated by the cytoskeleton which varies in structure and function from cell type to cell type.     

Macrophage-dependent stimulation of T cell-depleted spleen cells by Clostridium difficile toxin A and calcium ionophore

Clostridium difficile toxin A causes severe intestinal inflammation and fluid secretion in rabbit ileum and is chemotactic for neutrophils in vitro. The mechanism of intestinal injury produced by toxin A appears to involve direct epithelial cell damage as well as recruitment of an inflammatory cell response. The current study was undertaken to determine if toxin A can directly stimulate a proliferative response in lymphocytes. Highly purified toxin A, in the presence of the calcium ionophore, ionomycin, stimulated substantial [3H]thymidine incorporation by murine splenic lymphocytes, which was maximal at 10(-9) M toxin A and 800 ng/ml ionomycin. Removal of T cells with anti-Thy-1.2 antibody plus complement had no effect on the proliferative response induced by toxin A. However, [3H]thymidine incorporation in response to toxin A was significantly inhibited (P less than 0.001) by the removal of macrophages from splenocyte suspensions and was restored by the addition of peritoneal macrophages or cell-free supernatant from toxin A-treated macrophage cultures. Analysis of the toxin A-treated macrophage supernatants showed high levels of IL-1, but not IL-2 or IL-4. The combination of recombinant IL-1 plus ionomycin was found to stimulate [3H]thymidine incorporation by T cell-depleted splenic lymphocytes. These results suggest that toxin A stimulates the release of IL-1, and possibly other factors, from macrophages which can costimulate murine B lymphocytes.     

IL-10 inhibits cytokine production by activated macrophages

IL-10 inhibits the ability of macrophage but not B cell APC to stimulate cytokine synthesis by Th1 T cell clones. In this study we have examined the direct effects of IL-10 on both macrophage cell lines and normal peritoneal macrophages. LPS (or LPS and IFN-gamma)-induced production of IL-1, IL-6, and TNF-alpha proteins was significantly inhibited by IL-10 in two macrophage cell lines. Furthermore, IL-10 appears to be a more potent inhibitor of monokine synthesis than IL-4 when added at similar concentrations. LPS or LPS- and IFN-gamma-induced expression of IL-1 alpha, IL-6, or TNF-alpha mRNA was also inhibited by IL-10 as shown by semiquantitative polymerase chain reaction or Northern blot analysis. Inhibition of LPS-induced IL-6 secretion by IL-10 was less marked in FACS-purified peritoneal macrophages than in the macrophage cell lines. However, IL-6 production by peritoneal macrophages was enhanced by addition of anti-IL-10 antibodies, implying the presence in these cultures of endogenous IL-10, which results in an intrinsic reduction of monokine synthesis after LPS activation. Consistent with this proposal, LPS-stimulated peritoneal macrophages were shown to directly produce IL-10 detectable by ELISA. Furthermore, IFN-gamma was found to enhance IL-6 production by LPS-stimulated peritoneal macrophages, and this could be explained by its suppression of IL-10 production by this same population of cells. In addition to its effects on monokine synthesis, IL-10 also induces a significant change in morphology in IFN-gamma-stimulated peritoneal macrophages. The potent action of IL-10 on the macrophage, particularly at the level of monokine production, supports an important role for this cytokine not only in the regulation of T cell responses but also in acute inflammatory responses.     

Adhesion,phagocytosis and cell surface energy. The binding of fixed human erythrocytes to rat macrophages and polymethylpentene

Fixed human erythrocytes were used as model particles for the study of adhesion and phagocytosis by rat peritoneal macrophages. Erythrocytes were fixed with various concentrations of glutaraldehyde or tannic acid, or were treated with neuraminidase. Adhesion and phagocytosis of these cells were measured. In addition, the surface energy of these erythrocytes and macrophages was estimated by the contact angle technique. Free energies of adhesion, based on the cell surface energies, were correlated with both adhesion and phagocytosis.