Depletion Of C3 Research Articles

Specificity of an Anti-Murine B Cell Serum

An antiserum (ABS) specific for murine B cells was prepared in rabbits by immunizationwith lymph node cells from nude mice as reported previously (1). To make the antiserum specific for antibody-forming cell precursors (AFCP) extensive absorption was required with mouse serum, erythrocytes and thymus cells. Even after ABS was no longer cytotoxic for thymus cells, further absorption with thymus cells was necessary to make it ineffective for plaque-forming cells (PFC). The correlation between cells bearing C3- or Fc-receptors and the cells bearing antigen(s) towhich ABS reacts was studied. In the residual spleen cells treated with ABS and complement, there were some EA 8 and EA mC m rosette-forming cells. There were also some spleen cells reacting with ABS even after depletion of C3- and/or Fc-receptor-bearing cells. These results show that the antigen(s) to which ABS reacts could be a marker on B cells different from C3- or Fc-receptors. Spleen cells reacting with ABS were compared with cells bearing C3- or Fc-receptorsvarious days after birth. The cells sensitive to ABS increased in spleens earlier after birth than those with C3- or Fc-receptors.

Role of complement in lethal bacterial lipopolysaccharide-induced hypotensive and coagulative changes.

The effect of C3 depletion on the multiple pathophysiological changes produced by a lethal dose of Serratia marcescens lipopolysaccharide (LPS) was evaluated. The injection of this LPS into rabbits resulted in biphasic hypotensive changes and thrombocytopenia. These changes were characterized by an acute, transient decrease occurring within minutes after injection followed by a second more gradual decrease beginning 30 to 60 min post-LPS. Prior depletion of C3, with the anticomplementary protein from cobra venom (CoF), did not alter the extent of either the gradual hypotensive and platelet changes or the coagulative and metabolic changes when normal and C3-depleted rabbits were compared. Importantly, the lethal effects of S. marcescens LPS were not reduced by prior depletion of C3. Only the immediate, reversible thrombocytopenia and hypotension were abrogated by C3 depletion.

Complement Depletion Following Envenomation by Russell's Viper and Echis Carinatus (Saw-Scaled Viper) in the Rhesus Monkey *

Significant depletion of C3 was observed following administration of lethal and sublethal doses of Russell's viper and Echis carinatus venom to rhesus monkeys. A sequential study carried out over a period of 3 weeks following sublethal envenomation showed a steady rise in the C3 level from 48 hours onwards. By the 20th day the C3 values had attained the pre-envenomation levels. The extent of depletion of C3 was similar in both Russell's viper and E. carinatus envenomation. Because of the close immunological similarity of monkey and man, the rise in C3 which corresponded with recovery of the animals in this experimental model could serve as a useful parameter of recovery in clinical cases of snake poisoning.

Trypanosoma musculi infections in normocomplementemic, C5-deficient, and C3-depleted mice.

The role of complement in host resistance to infection with Trypanosoma musculi was studied in normal, C5-deficient, and C3-depleted mice. Infections in normocomplementemic strains (CBA and B10.D2/n) were generally similar to those in strains genetically deficient in C5 (A and B10.D2/o). There were no differences in inhibition of reproduction, duration of infection, persistence of parasites in the kidneys, or resistance to reinfection. However, peak parasitemias in B10.D2/o mice were slightly greater than in B10.D2/n mice. In addition, B10.D2/o mice had slightly decreased serum levels of C1 early in the course of infection and of C3 early during the elimination of adult forms. These components were unchanged or increased in infections of B10.D2/n. Depletion of C3 and late-acting components in B10.D2/n mice by treatment with cobra venom factor during the reproductive stage of infection resulted in an increase of reproductive forms before the apparent development of ablastic immunity as well as slightly greater peak parasitemias when compared with those of untreated controls. Cobra venom factor treatment of B10.D2/o mice during the reproductive stage did not alter the course of infection. Cobra venom factor treatment of C3H mice during the adult stage prolonged infections by interfering with parasite elimination. It is concluded that complement-mediated lysis is not involved in control of T. musculi. It is not clear whether a C3-dependent function such as phagocytosis may facilitate elimination of the parasites. The major difference in degree of parasitemias among the various strains of mice studied is due to genetic factors rather than the levels of C3, C5, or late-acting complement components.

Hemodialysis leukopenia. Pulmonary vascular leukostasis resulting from complement activation by dialyzer cellophane membranes.

Acute leukopenia occurs in all patients during the first hour of hemodialysis with cellophanemembrane equipment. This transient cytopenia specifically involves granulocytes and monocytes, cells which share plasma membrane reactivity towards activated complement components. The present studies document that complement is activated during exposure of plasma to dialyzer cellophane, and that upon reinfusion of this plasma into the venous circulation, granulocyte and monocyte entrapment in the pulmonary vasculature is induced. During early dialysis, conversion of both C3 and factor B can be demonstrated in plasma as it leaves the dialyzer. Moreover, simple incubation of human plasma with dialyzer cellophane causes conversion of C3 and factor B, accompanied by depletion of total hemolytic complement and C3 but sparing of hemolytic C1. Reinfusion of autologous, cellophane-incubated plasma into rabbits produces selective granulocytopenia and monocytopenia identical to that seen in dialyzed patients. Lungs from such animals reveal striking pulmonary vessel engorgement with granulocytes. The activated complement component(s) responsible for leukostasis has an approximate molecular weight of 7,000-20,000 daltons. Since it is generated in C2-deficient plasma and is associated with factor B conversion, it is suggested that activation of complement by dialysis is predominantly through the altermative pathway.

Complement in experimental Trypanosoma lewisi infection of rats.

The role of complement in host resistance to infection with Trypanosoma lewisi was studied in normal, C4-deficient, and C3-depleted rats. Complement levels were measured in normal rats throughout the course of infection. A drastic reduction of total complement and C4 hemolytic activities occurred, and C3 levels measured immunochemically were decreased. Although total complement and C4 levels were regularly reduced to less than 10% of preinfection levels regardless of parasite numbers, the degree of C3 consumption correlated with the parasitemia. C3 levels varied from 100% of preinfection in rats with light infections to 35% in animals with heavy parasitemias. Recovery to normal levels followed trypanosome elimination from the peripheral blood. The infection had no significant effect on C6 hemolytic activity. Parasitemias and C3 levels in C4-deficient rats did not differ from those of normocomplementemic controls. Depletion of C3 and late-acting components by cobra venom factor during the reproductive or adult stages of infection did not alter the parasitemias. In addition, T. lewisi and immune serum caused complement activation in vitro, which could be inhibited with ethylene glycol-bis-(beta-aminoethyl ether)N,N'-tetraacetic acid or ethylenediaminetetraacetic acid. It is concluded that T. lewisi infection in rats result in activation of the classical complement pathway with extensive consumption of the early-acting components, as well as a low degree of activation of the alternative pathway. However, complement does not appear to play a major role in the control and termination of the infection.

The Complement Profile in Babesiosis

Rats have been infected with Babesia rodhaini, a protozoan agent that induces immune complex disease. Assays for complement have revealed depletion of C2, C3, C4, C5, and whole complement in the course of infection. No evidence of depletion in the alternative (properdin) pathway was found. These findings are consistent with the conclusion that the classical complement pathway is activated during the course of the protozoan infection.

Necrosis, Haemorrhage and Complement Depletion Following Bites by the Spitting Cobra (<italic>Naja nigricollis</italic>)

The Spitting Cobra, Naja nigricollis, is widely and densely distributed in Africa. Fourteen patients with proven N. nigricollis bites, who were seen in the savanna region of Nigeria, did not exhibit the neurological signs, such as cranial nerve lesions and respiratory paralysis, expected following Elapid poisoning. All had local swelling, in eight cases involving the entire limb, and ten developed local tissue necrosis. Spontaneous haemorrhage was detected in three cases and was the probable cause of death in one of them; the other death in this series was unexplained. Haematological abnormalities included prolonged clot lysis anf failure of clot retraction due to a platelet defect. There was no specific deficit in clotting factors and a delayed rise in fibrin degradation products was attributed to extensive tissue damage at the site of the bit. Most patients showed depletion of complement component C3 and glycine-rich beta-glycoprotein (GBG), suggesting activation of the alternative pathway of complement fixation. There was evidence of hepatocellular damage in two out of six patients investigated. There was no evidence that specific polyvalent antivenoms, used in doses of up to 80 ml, prevented any of the effects of N. nigricollis venom. Clinical laboratory diagnosis is discussed. In the past many bites were wrongly classified as viper bites on the basis of clinical findings. Immunodiagnosis is a promising method for assessing the true importance of N. nigricollis bite in West Africa.

Mediation systems in bacterial lipopolysaccharide-induced hypotension and disseminated intravascular coagulation. I. The role of complement.

We have studied the role of complement in lipopolysaccharide (LPS)-induced hypotension and disseminated intravascular coagulation (DIC) by comparing the effects of injection of three preparations of LPS from E. Coli 0111:B4, S. minnesota Re595, and S. marcescens. Injections of nonlethal doses of these LPS preparations into normal rabbits produced decreases in mean arterial blood pressure during a 5-h period. When rabbits treated with cobra venom factor (CoF) to deplete C3 were injected with the various LPS preparations, mean arterial pressures fell at a rate and extent essentially identical to that observed in normal rabbits. Rabbits genetically deficient in C6 also demonstrated LPS-induced hypotensive changes. Only minimal, or no changes in plasma C3 levels or serum CH50 values were detected in normal rabbits after LPS injection. Hypotensive changes were also induced in rabbits when complement was rapidly activated by intravenous injection of CoF. In contrast to the hypotension induced by LPS, the fall in arterial pressure associated with the consumption of complement was short lived and required the rapid consumption of considerable amounts of C3. The occurrence of DIC noted in normal rabbits injected with each preparation of LPS was not inhibited in either rabbits treated with cobra factor or in C6-deficient rabbits. The DIC was most pronounced after injection of Re595 and S. marcescens LPS. Injection of the various LPS preparations produced a rapid disappearance of circulating neutrophils and mononuclear cells, which occurred with the same kinetics and to the same extent in normal, CoF-treated, and C6-deficient rabbits. Injection of either Re595 LPS or S. marcescens LPS produced a biphasic disappearance of circulating 51Cr-platelets. In contrast, injection of 0111:B4 LPS affected only slightly the rate of disappearance of 51Cr-platelets. Depletion of C3 by cobra factor treatment had no effect on the disappearance of platelets in animals injected with 0111:B4. In marked contrast cobra factor treatment greatly reduced the initial rapid disappearance of platelets in rabbits injected with either Re595 or S. marcescens LPS, but had no effect in the secondary disappearance phase.

Relationship of serum complement levels to events of the malarial paroxysm.

Malarial paroxysms due to Plasmodium vivax were studied for alterations in whole serum complement (C') and certain C' components. The objective was to relate C' values with events of the parasite cycle during schizogony and with the febrile pattern. Substantial decreases in C' were found in 9 of 18 paroxysms studied during relapse. In contrast, only one of 22 paroxysms occuring during the primary attack was associated with a striking depression in C', and this case exhibited certain characteristics of a relapse paroxysm. The mean change in C' levels during paroxysms in relapse (-23%) was significantly different from paroxysms of the primary attack (-2%). Depletion of C' was associated directly with degree of parasitemia and presence of complement-fixing (CF) antibody. Lowest levels of C' were found within a few hours after completion of schizont repture and peak fever. C4 levels reflected changes in whole serum C' and appeared to be a more sensitive indicator of C' alterations during malaria. While the alterations in C4 as well as C1 and C2 indicated that the classical C' pathway was involved, some preliminary results showed little or no depletion of late components, C3 and C6. Overall results are compatible with C' activation and depletion during or soon after schizont repture if parasite density is sufficiently high and if CF antibody is present.

Role of C3b in the breakdown of C3 in hypocomplementaemic mesangiocapillary glomerulonephritis.

Abstract Breakdown of C3 by hypocomplementaemic sera from patients with mesangiocapillary nephritis (M.C.G.N.) was shown to be mediated by the alternative pathway of complement activation. Evidence indicated that a fragment of C3 (C3b) is required for activation of the alternative pathway in nephritic serum. It is concluded that depletion of C3 in patients with M.C.G.N. is responsible for the known failure of their sera to break down pure C3 and for the normal survival of radio-labelled C3 preparations observed in such patients. These observations clearly demonstrate the activation of the complement system by the alternative pathway in human disease.

The Relationship of Complement Consumption to Immune Fever

Abstract The administration of cobra venom factor or soluble human serum albumin (HSA)-anti-HSA complexes in antigen excess to rabbits resulted in a lowering of total hemolytic complement and C3 titers which persisted more than 24 hr. When such animals were challenged with complexes made up in 10 times antigen excess, the consumption of complement, febrile and thrombocytopenic responses were significantly diminished from those seen in untreated controls, and the duration of post-challenge leukopenia was shortened. These observations suggest a role for complement consumption, particularly of components C3-9, in the manifestation of these responses. Pyrogenic tolerance to complexes could not be explained by their more rapid removal from the circulation or a failure of the thermoregulatory centers to respond to preformed leukocytic pyrogen, suggesting that impairment in complement consumption either by depletion of C3 or interference with the activation of C1 might be responsible for this state. In contrast to these findings using complexes in 10 times antigen excess, similar prior treatment of immunized rabbits given HSA or of normal rabbits that received a mixture of soluble and insoluble complexes in 3 times antibody excess failed to suppress febrile or hematologic responses. If the formation of antigen-antibody complexes is essential for the production of these responses in immunized animals challenged with specific antigen, it appears that the large, rapidly-cleared complexes formed near equivalence are most likely involved in producing these biologic activities. Whereas some of the properties of these complexes make them more able to fix complement in vitro, it seems unlikely that complement-mediated mechanisms are significantly involved in the pathogenesis of the in vivo responses measured in these investigations.

Acute Viral Infection: Tissue Injury Mediated by Anti-Viral Antibody through a Complement Effector System

Abstract Anti-viral antibody was able to injure virus-infected cells in the presence of a complement effector system. In lymphocytic choriomeningitis (LCM) and mumps infected culture cells, viral antigen was present on the surface of cells and accessible to interact with anti-viral antibody. Once complement had been activated, cellular injury ensued with the amount of damage directly related to the proportion of cells containing viral antigen. Mice treated with cobra factor were protected from the lethal effects of LCM challenge while showing no deficiency in either infectious viral replication or presence and activity of sensitized lymphoid cells. Such mice showed significant depletion of C3 and total complement levels. Once cobra factor treatment was ended, protection disappeared rapidly, and C3 and total complement levels returned to normal.

Effect of C4 depletion on the utilization of the terminal components of guinea-pig complement by endotoxin.

ENDOTOXIN and antigen-antibody complexes (Ag-Ab) are potent inflammatory agents which induce marked pathophysiological effects in a number of animal species. Both these agents lead to increased vascular permeability, local accumulations of polymorphonuclear leucocytes and alterations in the coagulability of blood1. There is mounting evidence that these similarities derive from endotoxins and Ag-Ab being activators of the complement (C) system, an important mediator of inflammation1'2. When endotoxins or preformed Ag-Ab are reacted with normal guinea-pig (GP) serum so as to induce equivalent consumption of whole haemolytic C, there are, however, striking differences in the consumption profiles of the individual C components3. Although interaction of Ag-Ab with GP serum results in the consumption of CI, C4, and C2 as well as C3 to C9, the interaction of endotoxin with GP serum leads to a marked depletion of C3 to C9 with little or no detectable loss of C1,C4 and C2 (ref. 3).

Acute immune complex disease in rabbits. The role of complement and of a leukocyte-dependent release of vasoactive amines from platelets.

By depletion of C3 from rabbits undergoing acute experimental immune complex disease with an anticomplementary factor in cobra venom, it has been possible to demonstrate that deposition of the complexes in arteries and glomeruli does not require the complement components reacting after C2. Immunological reactions, in which platelets release their vasoactive amines, have been examined in rabbits undergoing immune complex disease. A correlation was obtained between the presence of a complement-independent reaction which required blood leukocytes, antigen and platelets, the deposition of immune complexes, and the induction of glomerulonephritis. C3 depletion did, however, have a marked alleviating effect on the severity of the arterial lesions. Neutrophil accumulation and the subsequent necrotizing arteritis were prevented. In contrast, the character and severity of the glomerulonephritis was not altered by depletion of later-acting complement components.