Effector function of leucocytes from susceptible and resistant mice against distinct isolates of Candida albicans

Neutrophils and macrophages were generated in vitro from mice that display either high or low tissue susceptibilities to Candida albicans infection and their ability to phagocytose and kill three isolates of the yeast with different virulence characteristics was evaluated. In the absence of opsonization, phagocytosis by BALB/c and CBA/CaH neutrophils was comparable, but the killing was very poor. Opsonization with normal serum slightly decreased phagocytosis, but it had markedly different effects on killing, either enhancing or inhibiting candidacidal activity, depending on the combination of yeast isolate and mouse strain. In contrast, BALB/c macrophages showed high levels of phagocytosis and killing of both unopsonized yeasts and opsonized yeasts; whereas killing of unopsonized yeasts by CBA/CaH macrophages was poor, it was markedly enhanced by opsonization.     

Cryptococcus neoformans is resistant to surfactant protein A mediated host defense mechanisms

Initiation of a protective immune response to infection by the pathogenic fungus Cryptococcus neoformans is mediated in part by host factors that promote interactions between immune cells and C. neoformans yeast. Surfactant protein A (SP-A) contributes positively to pulmonary host defenses against a variety of bacteria, viruses, and fungi in part by promoting the recognition and phagocytosis of these pathogens by alveolar macrophages. In the present study we investigated the role of SP-A as a mediator of host defense against the pulmonary pathogen, C. neoformans. Previous studies have shown that SP-A binds to acapsular and minimally encapsulated strains of C. neoformans. Using in vitro binding assays we confirmed that SP-A does not directly bind to a fully encapsulated strain of C. neoformans (H99). However, we observed that when C. neoformans was incubated in bronchoalveolar fluid, SP-A binding was detected, suggesting that another alveolar host factor may enable SP-A binding. Indeed, we discovered that SP-A binds encapsulated C. neoformans via a previously unknown IgG dependent mechanism. The consequence of this interaction was the inhibition of IgG-mediated phagocytosis of C. neoformans by alveolar macrophages. Therefore, to assess the contribution of SP-A to the pulmonary host defenses we compared in vivo infections using SP-A null mice (SP-A-/-) and wild-type mice in an intranasal infection model. We found that the immune response assessed by cellular counts, TNFalpha cytokine production, and fungal burden in lungs and bronchoalveolar lavage fluids during early stages of infection were equivalent. Furthermore, the survival outcome of C. neoformans infection was equivalent in SP-A-/- and wild-type mice. Our results suggest that unlike a variety of bacteria, viruses, and other fungi, progression of disease with an inhalational challenge of C. neoformans does not appear to be negatively or positively affected by SP-A mediated mechanisms of pulmonary host defense.     

Failure of Trypanosoma cruzi to trigger the respiratory burst of activated macrophages. Mechanism for immune evasion and importance of oxygen-independent killing

Activated macrophages are thought to kill Trypanosoma cruzi, which lack catalase, by the generation of hydrogen peroxide. We investigated triggering of the respiratory burst of activated macrophages induced by phagocytosis of virulent T. cruzi, bloodform trypomastigotes, amastigotes obtained from spleens, and tissue culture organisms; and of relatively nonvirulent epimastigotes. All stages of T. cruzi prompted the release of less than 10% of hydrogen peroxide released by activated macrophages when stimulated with PMA or Candida. Superoxide anion production was not stimulated by PMA or Candida in activated macrophages nor was there a significant qualitative reduction of nitroblue tetrazolium induced by ingestion of virulent T. cruzi. Opsonization of T. cruzi with specific antibody did not promote the release of hydrogen peroxide or the reduction of nitroblue tetrazolium. Similar results were observed with activated spleen macrophages. Incubation of activated macrophages with catalase, catalase and superoxide dismutase, sodium benzoate with or without catalase, and respiratory burst-exhausting PMA failed to inhibit the killing of T. cruzi in vitro. These results indicate that 1) virulent opsonized or unopsonized T. cruzi do not trigger a respiratory burst by activated macrophages and 2) oxygen-independent killing of T. cruzi is of prime importance.     

Experimental and calculated parameters on particle phagocytosis by alveolar macrophages.

Phagocytosis of three types of fluorescein-labeled test particles by rat alveolar macrophages (AM) were studied: spherical silica (3.2 microm), heat-killed Candida albicans (3.8 microm), and heat-killed Cryptococcus neoformans (6.1 microm) opsonized with specific IgG. These particles should attach to scavenger, mannose, and Fc receptors, respectively. Both control AM and AM pretreated for 20 h with interferon-gamma (12.5 or 50 U/ml) were studied. The sum of the number of attached and ingested particles per AM (accumulated attachment) was used as a measure of the attachment process, and the number of ingested particles per AM divided by the accumulated attachment (ingested fraction) was used as a measure of the ingestion process. The average ingestion time (IT), which is also a measure of the ingestion process, was calculated from the experimental data. The ingestion process was independent of the attachment process. IT increased with the time of observation. This is explained by the fact that IT determined from observation times shorter than the whole distribution of IT for a certain particle results in a shorter IT than the real average IT. C. albicans (mannose receptor) had the fastest ingestion process, C. neoformans opsonized with specific IgG (Fc receptor) had ingestion that was nearly as fast, and the silica particles (scavenger receptors) had the slowest ingestion process. Treatment with interferon-gamma markedly impaired the attachment process for all three types of particles (and three types of receptors) but clearly impaired the ingestion process only for silica particles (scavenger receptors).     

C1q enhances the phagocytosis of Cryptococcus neoformans blastospores by human monocytes

We investigated whether C1q, a subunit of the first component of C, could modulate human peripheral blood monocyte-mediated phagocytosis of Cryptococcus neoformans (CN). Adherence of monocytes to C1q-coated surfaces induced a significant enhancement of ingestion of CN blastospores that had been opsonized with specific anticapsular IgG (IgG-CN). Additionally, C1q enhanced the monocyte-mediated phagocytosis of CN opsonized with C (CN-absorbed, nonimmune, normal human serum; C-CN). Ingestion of IgG- and C-CN by control and C1q-stimulated monocytes was maximal by 1 h of incubation. The monocyte-mediated enhancement of phagocytosis caused by C1q was paralleled by a proportionate increase in fungicidal activity, an effect which was maximal by 3 h of incubation. Human serum albumin-adherent, control monocytes exhibited only a low level of killing after 3 h of incubation. C1q enhancement was blocked by preincubation of the surfaces with a goat, polyclonal F(ab')2 anti-C1q. This study describes a new cellular function for the cell surface C1q receptor: the enhancement of phagocytosis of a pathogenic organism by monocytes.     

Catch me if you can: phagocytosis and killing avoidance by Cryptococcus neoformans

After inhalation of infectious particles, Cryptococcus neoformans resides in the alveolar spaces, where it can survive and replicate in the extracellular environment. This yeast has developed different mechanisms to avoid internalization by phagocytic cells, the main one being a polysaccharide capsule around the cell body, which inhibits the uptake of the yeast by macrophages. In addition, capsule-independent mechanisms have also been described, such as the production of antiphagocytic proteins. Despite these mechanisms, phagocytosis can occur in the presence of opsonins, and once C. neoformans is internalized, multiple outcomes are possible, including pathogen killing or intracellular replication and escape from macrophages. For this reason, C. neoformans is considered a facultative intracellular pathogen. As alveolar macrophages are the first component of the host immune system to confront C. neoformans, the outcome of this interaction could determine the degree of infection, producing either a severe disseminated disease or a latency state. In this review, we will tackle the complexity of the interaction between C. neoformans and macrophages, including the phagocytic avoidance mechanisms and all the possible outcomes that have been described for this interaction. Finally, we will discuss the consequences of the different outcomes for the type of infection produced in the host.     

Cryptococcus neoformans: in vivo protection of mice by pretreatment with pyran copolymer

Synthetic polyanions have been shown to alter host resistance to infection. The anticryptococcal effect of pyran copolymer was assessed in vivo and in vitro. Pretreatment with pyran copolymer significantly extended mean survival in mice lethally infected with Cryptococcus neoformans when compared to untreated animals (p < 0.01). The anticryptococcal effect of peritoneal exudate cells (PEC) elicited by 10% thioglycollate or pyran copolymer (25 mg/kg) was assessed in vitro. Initial percent phagocytosis of both encapsulated and non-encapsulated isolates of C. neoformans was greatest in the pyran elicited PEC. Significant killing of C. neoformans in vitro was observed only in pyran-activated PEC cultures combined with non-encapsulated cells of C. neoformans, although pyran PEC did inhibit initial growth of phagocytized encapsulated yeast cells. The protection of pyran copolymer pretreated mice from infection with C. neoformans, but the absence of significant killing of encapsulated yeast in vitro suggest a complex mechanism of host defense which may involve an activation of the reticuloendothelial system by pyran copolymer.     

Selective down-regulation of alveolar macrophage oxidative response to opsonin-independent phagocytosis

We have compared the oxidative response of alveolar macrophages (AM) during opsonin-dependent and independent phagocytosis by using multiparameter flow cytometry. The respiratory burst of AM during phagocytosis was quantitated by the intracellular oxidation of the nonfluorescent precursors dichlorofluorescin diacetate (DCFH) or hydroethidine (HE, a reduced precursor of ethidium) to their fluorescent (oxidized) counterparts. After loading freshly isolated normal hamster AM with DCFH or HE, red or green fluorescent beads, respectively, were added to the shaking cell suspensions. Ingestion of opsonized particles by AM caused a marked increase in oxidation of both DCFH and HE proportional to the number of beads ingested. In contrast, uptake of one to three unopsonized particles per cell led to inhibition of oxidative activity compared to control cells incubated without particles. AM ingesting four or more unopsonized particles showed some increase in oxidative metabolism, but far less than that with identical numbers of particles in opsonin-dependent ingestion. Similar results were obtained using fluorescent labeled staphylococcal bacteria. Using three-color flow cytometry to study cells ingesting both types of particles, cells first ingesting unopsonized beads were also found to have an inhibited oxidative response to subsequently ingested opsonized particles. The mitochondrial poison antimycin inhibited most of the intracellular oxidative response to either type of phagocytosis. The remaining antimycin-insensitive, membrane derived respiratory burst of AM was also substantially diminished after phagocytosis of unopsonized particles vs similar numbers of opsonized particles. The greatly increased mitochondrial respiration in AM during phagocytosis of opsonized particles may be related to bactericidal mechanisms. Killing of ingested Staphylococcus by AM was markedly impaired in the presence of antimycin. The results suggest that AM may ingest the numerous, unopsonized inert particles that are inhaled without generation of potentially toxic oxygen metabolites, while retaining the capacity to undergo a respiratory burst after ingesting opsonized particles and bacteria. The mechanism(s) for this distinct response may include generation of an inhibitor of intracellular oxidative metabolism.     

Influenza virus-induced immune complexes suppress alveolar macrophage phagocytosis

Immune complexes in the lungs are capable of inducing adverse responses. Herein we have detailed the formation of immune complexes in the lungs of influenza virus-infected mice and examined their effect on alveolar macrophage defenses. On days 3, 7, 10, 15, and 30 after aerosol infection with influenza A/PR8/34 virus, the acellular pulmonary lavage fluid was tested for viral antigen, specific viral antibody, and immune complexes by immunoassays. Whereas peak viral antigen (day 3) diminished to undetectable levels by day 10, specific viral antibody remained at a low concentration until day 10, after which it rapidly increased. Immune complex concentrations increased through day 7, peaked at day 10, and gradually returned to the control level by day 30. These data demonstrate that immune complexes of detectable size are induced by influenza virus infection during the interface between antigen excess and antibody excess conditions. Since alveolar macrophages are the pivotal phagocytic defense cells in the lung, the ability of normal alveolar macrophages to ingest opsonized erythrocytes was quantitated in the presence of immune complexes from lavage fluid. Immune complexes from day 10 virus-infected lungs caused a dose-dependent suppression of antibody-mediated phagocytosis to 30% of control values. In contrast, although these immune complexes also markedly decreased the phagocytosis of antibody-coated yeast cells, they did not significantly impair the antibody-independent ingestion of unopsonized yeast cells by macrophages. the suppressive effects of immune complexes on alveolar macrophages may, in part, explain the phagocytic dysfunction that occurs 7 to 10 days after influenza virus pneumonia.     

Treatment of experimental Cryptococcus neoformans infection in newborn rabbits by airway instillation of specific antibody and surfactant

Cryptococcosis in AIDS patients has a slow response to antifungal chemotherapy, and passive antibody treatment has thus been considered as an adjunct. Polyclonal anticryptococcal IgG dissolved in a suspension of modified natural surfactant was given intratracheally to near-term rabbits. Killing of Cryptococcus neoformans within the lungs was determined by counting the colony forming units (cfu). After 5 h a significant decrease in cfu was observed in rabbits treated with the IgG-surfactant mixture compared with control animals receiving saline. In conjunction with conventional therapy, the combined treatment of IgG-surfactant given by bronchoscopy might be used in high-risk patients to enhance killing of the yeast within the lungs.     

Trypanosoma lewisi-induced immunosuppression: the effects on alveolar macrophage activities against Cryptococcus neoformans

The immunosuppressive effect of Trypanosoma lewisi infection on alveolar macrophage (AM) activities against Cryptococcus neoformans was studied in an animal model. Two groups of rats were treated with T. lewisi and killed after 4 (4d-rats) and 7 days (7d-rats), respectively. A third group not given T. lewisi, served as control. AM were challenged in vitro with C. neoformans. Phagocytosis was assessed with a fluorescence method. Superoxide anion production was evaluated with the nitroblue tetrazolium (NBT) test. The survival of cryptococci was estimated by counting colony-forming units. The numbers of detached AM from culture plates were determined using a Bürker chamber. The NBT response, adhesion to plate surface and killing activity, but not the phagocytosis of AM from 4d-rats were significantly impaired compared to control or 7d-rats. Thus, T. lewisi causes transitory immunosuppressive effects on AM activities. This rapid T. lewisi immunosuppression model may be useful to study new approaches to anticryptococcal therapy.     

Interaction of mycoplasmas and phagocytes

Aspects of the interaction of certain mycoplasmas with macrophages and neutrophils in vivo and in vitro have been studied using two systems, one involving M. pulmonis in mice and the other involving M. bovis with bovine leucocytes. Studies with M. pulmonis indicated that the disappearance of viable organisms from the peritoneal cavity was not enhanced in SPF mice in which a peritoneal exudate rich in neutrophils had been induced. However, viable M. pulmonis organisms disappeared more rapidly from the peritoneal cavities with exudates containing increased numbers of macrophages. Experiments in vitro studied the opsonic effect of bovine IgG isotypes for bovine neutrophils and alveolar macrophages. Both IgG1 and IgG2 promoted killing of M. bovis by alveolar macrophages but IgG2 was more effective than IgG1 at promoting mycoplasma killing by neutrophils. Further studies in vitro indicated that certain bovine mycoplasma could inhibit killing of Escherichia coli by bovine neutrophils.     

NK cells eliminate Cryptococcus neoformans by potentiating the fungicidal activity of macrophages rather than by directly killing them upon stimulation with IL-12 and IL-18

In the present study, we examined whether natural killer (NK) cells have direct fungicidal activity against Cryptococcus neoformans. Splenic NK cells were obtained from SCID mice and stimulated with a combination of interleukin (IL)-12 and IL-18 in flat culture plates or round tubes. They were then or at the same time cultured with the yeast cells and the number of viable yeast cells was examined. We could not detect direct fungicidal activity by NK cells under any culture condition, although they produced a large amount of IFN-gamma and exerted marked cytotoxic activity against YAC-1 cells. On the other hand, NK cells significantly potentiated the nitric oxide-mediated cryptococcocidal activity of thioglycolate-elicited peritoneal macrophages obtained from SCID mice upon stimulation with IL-12 and IL-18. The culture supernatants of NK cells stimulated with IL-12 and IL-18 provided similar results when used in place of NK cells. The induction of macrophage anticryptococcal activity by NK cells and NK cell culture supernatants were both mediated by IFN-gamma because the specific mAb almost completely abrogated such effect. Considered collectively, our results suggested that NK cells may play a regulatory role in potentiating macrophage-mediated fungicidal mechanisms in host resistance to infection with C. neoformans rather than exerting a direct killing activity against the fungal pathogen.     

Challenge of Drosophila melanogaster with Cryptococcus neoformans and role of the innate immune response

We found that the ingestion of Cryptococcus neoformans by Drosophila melanogaster resulted in the death of the fly but that the ingestion of Saccharomyces cerevisiae or the nonpathogenic Cryptococcus kuetzingii or Cryptococcus laurentii did not. The C. neoformans protein kinase A and RAS signal transduction pathways, previously shown to be involved in virulence in mammals, also played a role in killing DROSOPHILA: Mutation of the Toll immune response pathway, the predominant antifungal pathway of the fly, did not play a role in Drosophila defense following ingestion of the yeast. However, the Toll pathway was necessary for the clearance of C. neoformans introduced directly into the hemolymph of D. melanogaster and for the survival of systemically infected flies.     

The Effect of Prednisolone on Canine Neutrophil Function: In Vivo and in Vitro Studies

The in vivo effect of a therapeutic dose of prednisolone on canine neutrophil adherence, random migration, Chemotaxis, phagocytosis of IgG and C3b opsonized yeast cells, chemiluminescence, Fc- and CR3-receptor expression was investigated. Prednisolone was also added in vitro to neutrophils as isolated cells and in whole blood. In the in vivo study, prednisolone increased the IgG mediated ingestion of yeast cells and the number of activated neutrophils in the phagocytosis assay, while flow cytometric investigation of the IgG-receptor FcγRIII with a monoclonal antibody showed similar expression before, during and after treatment. Prednisolone also increased the ingestion of C3b-opsonized yeast cells, while the expression of CR3-receptors (CD11b CD18) measured by flow cytometry was unchanged. Chemiluminescence and the chemotactic response towards zymosan activated serum were increased, while adherence to nylon wool was decreased. The in vitro studies revealed that prednisolone had no or a dampening effect on neutrophils in cell suspensions. Adherence as well as IgG mediated ingestion was decreased at the highest prednisolone concentration (800 ng/ml) in whole blood. The present study suggests that the part of the antiinflammatory effect of corticosteroids mediated through their influence on neutrophils, besides reduced adherence, may be exerted by increased clearance of microorganisms and IgG-complexes through an elevated functional capacity.     

Fc- and complement-receptor activation stimulates cell cycle progression of macrophage cells from G1 to S

Phagocytosis of microorganisms by macrophages is an important host defense mechanism. While studying the phagocytosis of the human pathogenic fungus Cryptococcus neoformans, we noted that macrophage-like J774 cells with ingested fungal cells had frequent mitotic figures. By analyzing the relative proportion of phagocytic cells as a function of cell cycle phase, we observed an increase in S phase cells after Fc-mediated phagocytosis of polystyrene beads, live or heat-killed C. neoformans. This result was confirmed by increased nuclear BrdU incorporation after Fc-mediated phagocytosis. The induced progression to S phase was observed after both Fc- and complement-mediated phagocytosis of live yeasts. Fc-mediated stimulation of cell division did not require ingestion, because it could be triggered by incubating cells in IgG1-coated plates. Phagocytosis-mediated stimulation of replication was confirmed in vitro using primary bone marrow macrophages and in vivo for peritoneal macrophages. We conclude that phagocytosis of microbes or inert particles can stimulate macrophages to enter S phase and commence cell division. This observation suggests a potential mechanism for increasing the number of effector cells after microbial ingestion, but can also promote the spread of infection.     

Monocyte responses to Candida albicans are enhanced by antibody in cooperation with antibody-independent pathogen recognition

Although most individuals are colonized with Candida albicans, only patients with insufficient or nonfunctional phagocytes develop life-threatening C. albicans disease. Because recognition of bacterial pathogens through phagocyte receptors for IgG (FcgammaR) is known to augment phagocyte responses, we postulated that antibody opsonization would enhance monocyte damage to C. albicans and subsequent tumor necrosis factor-alpha (TNF-alpha) production. After exposure to the human monocytic cell line THP-1, opsonized yeast showed an 89% decrease in metabolic activity, compared with 40% for unopsonized yeast (P<0.05). Culture supernatants contained 1316 pg mL(-1) of TNF-alpha after monocytes were exposed to opsonized yeast vs. 341 pg mL(-1) for unopsonized yeast (P=0.003). Similar results were obtained using peripheral blood mononuclear cells. Antibody opsonization of C. albicans germ tubes enhanced TNF-alpha production but did not affect organism damage. Antibody-dependent and antibody-independent factors were found to act synergistically to increase TNF-alpha production. ERK activation was important for both antibody-dependent and antibody-independent stimulation of TNF-alpha production, but not for monocyte-mediated organism damage. These data suggest that FcgammaR cooperates positively with antibody-independent recognition mechanisms in what may be a novel link between innate and adaptive immunity to C. albicans.     

Human alveolar macrophage FcR-mediated cytotoxicity. Heteroantibody- versus conventional antibody-mediated target cell lysis

Human alveolar macrophage have three distinct receptors for IgG: FcRI, FcRII, and FcRIII. In order to compare the ability of these receptors to mediate target cell lysis, three different assay systems were examined. First, we studied lysis of chicken E (CE) opsonized with heteroantibodies, which are synthetic antibodies composed of Fab fragments with anti-FcR activity covalently linked to Fab fragments with anti-CE activity. We found alveolar macrophage readily lysed heteroantibody-opsonized CE via each of the three FcR classes (FcRI, 20 +/- 5%; FcRII, 27 +/- 7%; and FcRIII, 13 +/- 13%, p less than 0.05). Non-FcR-dependent lysis of anti-beta 2-microglobulin x anti-CE heteroantibody-opsonized CE was not detected. Second, lysis of hybridoma cell lines bearing anti-FcR antibodies on their cell surface was examined to assess killing of "tumor-like" target cells. Whereas peripheral blood monocytes and lymphocytes were able to lyse hybridoma cell lines bearing surface anti-FcR mAb, alveolar macrophages were not. Third, activity of alveolar macrophage FcR was examined in a conventional antibody-dependent cellular cytotoxicity assay by using O+ (R1,R2) human RBC opsonized with human anti-D and anti-CD serum as target cells. We found lysis of anti-D and anti-CD opsonized human RBC was mediated exclusively via FcRI. No activity of FcRII or FcRIII was detected in these latter assays even if performed under conditions that impair FcRI activity. Thus, all three FcR present on alveolar macrophage mediate lysis of heteroantibody-opsonized CE; in contrast, with the use of a conventional antibody-dependent cellular cytotoxicity assay, only FcRI activity was detected. We were unable to demonstrate lysis of anti-FcR-bearing hybridoma cell lines by alveolar macrophages.     

Quantitative analysis of phagocytosis of Cryptococcus neoformans by adherent phagocytic cells by fluorescence multi-well plate reader

Macrophages and monocytes are adherent phagocytic cells which play an important role in host defence against the yeast-like fungus Cryptococcus neoformans. Before, phagocytosis by adherent phagocytes could only be measured by means of microscopy or by a radioactive assay, which both have obvious disadvantages. We have developed a new, rapid and objective method to measure phagocytosis of C. neoformans by adherent phagocytes (e.g. alveolar macrophages) using a fluorescence multi-well plate reader. This method allows us to discriminate accurately between adherence and internalisation of C. neoformans by macrophages during long term incubation. In addition, the method was used to study the role of the mannose receptor in phagocytosis of the acapsular yeast in the absence of serum by human monocyte-derived macrophages (MDM).     

Human pulmonary surfactant protein (SP-A), a protein structurally homologous to C1q, can enhance FcR- and CR1-mediated phagocytosis

C1q, a subunit of the first component (C1) of the classical complement pathway, and the pulmonary surfactant protein SP-A are structurally homologous molecules, each having an extended collagen-like domain contiguous with a non-collagenous domain. It is the collagen-like region of C1q that binds to mononuclear phagocytes and mediates the enhancement of phagocytosis of opsonized particles by these cells. Because SP-A enhances the endocytosis of phospholipids by alveolar type II cells and alveolar macrophages, we examined whether these two molecules were functionally interchangeable. The phagocytosis of sheep erythrocytes opsonized with IgG or with IgM and complement was enhanced by the adherence of monocytes or macrophages, respectively, to SP-A. The enhanced response was dependent on the concentration of SP-A used for coating the surfaces, similar to that seen when monocytes were adhered to C1q-coated surfaces. Both the percentage of cells ingesting the opsonized targets and the number of targets ingested per cell increased with increasing concentrations of SP-A. No such enhancement was seen with cells adhered to albumin, iron-saturated transferrin, or uncoated surfaces. However, SP-A did not substitute for C1q in the formation of hemolytically active C1. C1q did not stimulate lipid uptake by alveolar type II cells or alveolar macrophages and had only a slight inhibitory effect on the binding of SP-A to alveolar type II cells. Thus, these results suggested that a function which requires interactions of both the collagenous and the non-collagenous regions (i.e. initiation of the classic complement cascade) could not be mimicked by a protein sharing structural macromolecular similarity but lacking sequence homology in the non-collagen-like region. However, SP-A could substitute for C1q in stimulating a function previously shown to be mediated by the collagen-like domains of the C1q molecule.