Clearance Of Colloidal Carbon Research Articles

Hepatic irradiation augments engraftment of donor cells following hepatocyte transplantation

Engraftment of donor hepatocytes is a critical step that determines the success of hepatocyte transplantation. Rapid and efficient integration of donor cells would enable prompt liver repopulation of these cells in response to selective proliferative stimuli offered by a preparative regimen. We have earlier demonstrated that hepatic irradiation (HIR) in combination with a variety of hepatotrophic growth signals, such as partial hepatectomy and hepatocyte growth factor, can be used as a preparative regimen for liver repopulation of transplanted hepatocytes. In this study, we investigated the effects of HIR on engraftment of transplanted dipeptidyl peptidase IV (DPPIV)-positive hepatocytes in congeneic DPPIV-deficient rats. HIR-induced apoptosis of hepatic sinusoidal endothelial cells (SEC) within 6 hours of HIR resulted in dehiscence of the SEC lining in 24 hours. Although there was no change of the number of Kupffer cells after HIR, colloidal carbon clearance decreased 24 hours post HIR, indicating a suppression of phagocytic function. DPPIV+ donor cells were transplanted 24 hours after HIR (0-50 Gy). There was an HIR dose-dependent increase in the donor hepatocyte mass engrafted in the liver parenchyma. The number of viable transplanted hepatocytes present in hepatic sinusoids or integrated in the parenchyma was greater in the HIR-treated group at 3 and 7 days after transplantation compared with the sham controls. Finally, we validated these rodent studies in cynomolgus monkeys, demonstrating that a single 10-Gy dose of HIR was sufficient to enhance engraftment of donor porcine hepatocytes. These data indicate that transient disruption of the SEC barrier and inhibition of the phagocytic function of Kupffer cells by HIR enhances hepatocyte engraftment and the integrated donor cell mass. Thus, preparative HIR could be potentially useful to augment hepatocyte transplantation.

Effects of a somatostatin analogue (SMS 201-995) on hepatic and splenic reticulo-endothelial function in the rat.

The effects of a long acting somatostatin analogue, SMS 201-995, on reticulo-endothelial system (RES) activity were studied in rats. Administration of 2 micrograms SMS 201-995 subcutaneously twice a day for 7 days significantly increased the splenic and hepatic uptake of 99mTc-sulphur colloid and damaged 51mCr-red blood cells. Furthermore, SMS 201-995 administration significantly increased the plasma clearance of colloidal carbon as indicated by a lower area under the curve and an increased elimination constant. SMS 201-995 administration also significantly improved survival after intraperitoneal injection of Escherichia coli endotoxin. These results suggest that SMS 201-995 stimulates RES activity in rats. It is suggested that SMS 201-995 may be of value in stimulating RES activity in patients with cirrhosis and portal hypertension.

Muramyl dipeptide increases tolerance to shock and bacterial challenge in mice

We measured the phagocytic capacity of the reticulo-endothelial system by assay of the clearance of colloidal carbon from the blood stream in both normal mice and mice in which shock had been induced by intraperitoneal injection of hypertonic glucose. The group of shocked mice was further subdivided into those pretreated with the immunoadjuvant, muramyl dipeptide (MDP), and those given placebo. Shock reduced reticulo-endothelial phagocytosis (P less than 0.01), whereas pretreatment of shocked mice with MDP led to a hyperphagocytic state (P less than 0.01). In a second series of experiments we subjected shocked mice, pretreated with MDP or placebo, to a virulent Klebsiella pneumoniae infection. MDP significantly improved survival (P less than 0.01).

Immunomodulatory properties of Mycoplasma pulmonis: I. Characterization of the immunomodulatory activity

For many years, it has been recognized that Mycoplasma infection affects the host's immune system in different ways. In this work, experiments were performed to characterize the influence of Mycoplasma pulmonis infection on various immunological parameters and to follow the kinetics of their variations. A Balb/c mouse model was used to assess hematological evaluations, changes in spleen weight, antibody responses against sheep erythrocytes, neutrophil phagocytosis, colloidal carbon clearance, and anti- Mycoplasma antibody responses. At the hematological level, infected animals were found to have significantly increased total lymphocyte and polymorphonuclear leukocyte counts and an augmentation in spleen weight. Seven days after Mycoplasma infection, antibody responses against sheep erythrocytes were considerably diminished, and at days 7 and 14 after infection, phagocytic activity was also reduced. After 1 week of infection, the colloidal carbon clearance pattern was decreased, and during the whole infectious process, anti- Mycoplasma antibody titers were found to be low. Results from this part of research show a persistent infection that does not resolve in a short period, which is associated with a general dysfunction in the immune system and poor immune responses against several different antigens.

Xanthine oxidase released from reperfused hind limbs mediate kupffer cell activation, neutrophil sequestration, and hepatic oxidative stress in rats subjected to tourniquet shock.

We have shown previously that rats subjected to tourniquet shock develop an acute form of remote organ injury of the liver that is both Kupffer cell (KC) and polymorphonuclear (PMN) leukocyte dependent. Circulating plasma xanthine oxidase (XO) has been shown to be responsible for the development of endothelial dysfunction and for remote organ injury of the lung and intestine after ischemia-reperfusion protocols. We now hypothesize that XO is released from rat hind limbs upon reperfusion and that it is responsible for KC and PMN leukocyte activation in this shock model. Our results show that about 30% of rat gastrocnemius muscle xanthine dehydrogenase (XD) is converted to XO during the 5-h tourniquet period and that it is released into the femoral vein within 10 min of reperfusion. Total muscle xanthine oxidoreductase activity (XO + XD) decreases within 30 min of reperfusion and is paralleled by a corresponding increase in femoral vein lactic dehydrogenase. In addition, liver tissue XO increases significantly within 30 min of reperfusion without a corresponding conversion of endogenous XD. Conversion of hepatic XD becomes evident 60 min after reperfusion is initiated, as does XO, and alanine aminotransferase (ALT) release into the hepatic vein, presumably from damaged hepatocytes as a consequence of oxidative stress. Tissue myeloperoxidase activity also increases significantly after the 60-min reperfusion period. That XO mediates KC and PMN activation is supported by the following observations: a) the close relationships between plasma XO and the time courses of tumor necrosis factor-alpha TNFalpha release into the hepatic vein and colloidal carbon clearance by KCs; b) that colloidal carbon clearance, TNFalpha and ALT release, loss of tissue free thiols, lipid peroxidation (TBARS), and liver infiltration by PMN neutrophils can also be induced by the administration of exogenous XO to normal rats; and c) pretreatment of rats with allopurinol inhibits KC activation and liver leukocyte infiltration. These results suggest that XO, released from the ischemic limb on reperfusion, is taken up by the liver were it mediates KC and PMN neutrophil activation and thus contributes to the development of multiple system organ failure after hind limb reperfusion.

Activated Kupffer cells attenuate the liver response to the peroxisome proliferator perfluorooctanoic acid.

It has been suggested that peroxisome proliferators stimulate Kupffer cells, an effect which may be involved in their mechanism of action. To evaluate this hypothesis, this study was designed to investigate the effect of stimulating Kupffer cells on basal as well as induced peroxisomal enzyme activity. Twenty four hours following treatment of male Sprague-Dawley rats with the peroxisome proliferating agent perfluorooctanoic acid (PFOA), in corn oil or with corn oil alone, hepatic peroxisomal beta-oxidation was 4.6 +/- 0.2 and 1.8 +/- 0.1 U/g liver, respectively. As expected, PFOA did not influence the catalase activity. Stimulating Kupffer cells in vivo by zymosan A (25 mg/kg, i.v.) prior to treatment with corn oil or PFOA diminished basal as well as PFOA-induced peroxisomal beta-oxidation by 20-35%. Activation of Kupffer cells by zymosan A also diminished catalase activity by over 60%. Furthermore, PFOA reduced blood colloidal carbon clearance by 35% within 2 h of its administration. The data suggest that activation of Kupffer cells exerts a negative effect on basal as well as PFOA-induced peroxisomal enzyme activities. Data also suggest that PFOA inhibits Kupffer cells. Activated Kupffer cells may indeed produce factors which interfere with normal hepatic peroxisomal functions and responses.

The Pectic Polysaccharide from Bupleurum falcatum L. Enhances Clearance of Immune Complexes in Mice

The effect of pectic polysaccharide, bupleuran 2IIb, on clearance of immune complex from circulation in vivo was examined in mice. The immune complex clearance was enhanced by intraperitoneal treatment of bupleuran 2IIb in a dose dependent fashion, whereas clearance of colloidal carbon was reduced by bupleuran 2IIb-treatment. The body and organ weights were not significantly altered in bupleuran 2IIb-treated mice, and the corrected phagocytic index (α), indicating the phagocytic activity of mononuclear phagocytic cells was also not affected. The expression of Fc receptors for IgG (FcγRs) gene after administration of bupleuran 2IIb has been studied using reverse transcriptase-polymerase chain reaction (RT-PCR). Bupleuran 2IIb-treated mice showed significant enhanced expression of both FcγRI and FcγRII/FcγRIII mRNA in liver compared with control mice. These results suggest that bupleuran 2IIb activated the mononuclear phagocytic system on immune complex clearace in vivo through an increase of FcγRs expression, and resulted in the enhancement of immune complex clearance.

The Pectic Polysaccharide from Bupleurum falcatum L. Enhances Clearance of Immune Complexes in Mice

The effect of pectic polysaccharide, bupleuran 2IIb, on clearance of immune complex from circulation in vivo was examined in mice. The immune complex clearance was enhanced by intraperitoneal treatment of bupleuran 2IIb in a dose dependent fashion, whereas clearance of colloidal carbon was reduced by bupleuran 2IIb-treatment. The body and organ weights were not significantly altered in bupleuran 2IIb-treated mice, and the corrected phagocytic index (α), indicating the phagocytic activity of mononuclear phagocytic cells was also not affected. The expression of Fc receptors for IgG (FcγRs) gene after administration of bupleuran 2IIb has been studied using reverse transcriptase-polymerase chain reaction (RT-PCR). Bupleuran 2IIb-treated mice showed significant enhanced expression of both FcγRI and FcγRII/FcγRIII mRNA in liver compared with control mice. These results suggest that bupleuran 2IIb activated the mononuclear phagocytic system on immune complex clearace in vivo through an increase of FcγRs expression, and resulted in the enhancement of immune complex clearance.

Gadolinium chloride treatment attenuates hepatic platelet accumulation after lipopolysaccharide administration.

Intravenous administration of LPS to rats results in the accumulation of both neutrophils and platelets in the liver and the development of midzonal hepatocellular necrosis. The development of liver injury entails contributions from both cellular and soluble mediators, including neutrophils, platelets, Kupffer cells, tumor necrosis factor-alpha (TNF-alpha), and components of the coagulation system. Much remains unknown about the interactions among these mediators in the pathogenesis of liver injury in vivo. Accordingly, we conducted studies with gadolinium chloride (GdCl3), an agent that inhibits Kupffer cell phagocytosis, to evaluate the role of Kupffer cells in lipopolysaccharide (LPS)-mediated liver injury, elevation in plasma TNF-alpha activity, thrombocytopenia, hepatic platelet accumulation, and activation of the coagulation system. Female Sprague-Dawley rats were pretreated with GdCl3-6H2O (10 mg/kg, i.v.) or saline vehicle 24 h before the administration of LPS (4 mg/kg, i.v.) or saline vehicle. In a preliminary study, this GdCl3 treatment regimen decreased the clearance of colloidal carbon from blood, indicating inhibition of Kupffer cell phagocytosis. Pretreatment with GdCl3 attenuated LPS-induced liver injury, monitored as increased plasma alanine aminotransferase and isocitrate dehydrogenase activities and histologic analysis. Electron micrographs of livers from rats treated with LPS revealed platelets within the sinusoids as well as Kupffer cells with phagolysosomes containing material resembling platelets. Pretreatment with GdCl3 attenuated LPS-induced thrombocytopenia and hepatic platelet accumulation, as measured by radiolabeled platelets. Treatment with GdCl3 did not, however, alter the elevation in plasma TNF-alpha activity or the activation of the coagulation system, as evidenced by a decreased in plasma fibrinogen concentration. These results suggest that Kupffer cells contribute to LPS-induced hepatic platelet accumulation and raise the possibility that protection against LPS-induced hepatic injury by Kupffer cell inactivation may be due at least partly to decreased deposition of platelets within the liver.

Influence of deoxyspergualin on mononuclear phagocytic system in mice

The influence of deoxyspergualin (DSG) on mononuclear phagocytic system was examined in BALB/c mice. DSG did not influence on both the organ distribution of sheep red blood cells (SRBC) and blood clearance of colloidal carbon when the agent was administered intraperitoneally into mice at doses of 5.0 and 10.0 mg/kg for six consecutive days before antigen. Hemagglutination titers in experimental and control mice did not differ significantly when 10.0 mg/kg of DSG was given six times before SRBC injection. However, when administered during the course of the response (days 0 to 5) DSG (5.0 mg/kg/day) markedly suppressed antibody formation.

Does level of ligation influence results in a murine biliary obstruction model?

Despite advances in perioperative management, patients with extrahepatic biliary obstruction still experience a high rate of complications and death after surgery. The rat is commonly used as an experimental animal for research in obstructive jaundice. Ligation of the rat bile duct high in the liver hilum is assumed to produce a more severe model of biliary obstruction than low ligation. The differences are attributed to the ability of the rat bile duct to dilate. Differences in level of ligation may, thus, explain some discrepancies between studies. To test this hypothesis, female Lewis rats underwent high ligation (HL), low ligation (LL), and sham celiotomy. Colloidal carbon clearance, bilirubin, total serum bile acids, and hematocrit were measured 12 days later. Liver and spleen weight, presence or absence of ascites, infection, and adequacy of ligation were noted and the liver was processed for routine histology and electron microscopy. Although bilirubin levels were higher after HL than after LL, liver and spleen weight, total serum bile salts, and phagocytic constants K and alpha were not different between these two groups. Gross, histologic, and ultrastructural appearance did not differ between HL and LL groups. High ligation causes greater hyperbilirubinemia than low ligation, but does not alter other parameters including phagocytic constants. The present study does not confirm the hypothesis that HL creates a more severe model than LL; therefore, it is unlikely that differences in level of ligation explain variability in results between studies.

Depletion of neutrophils and modulation of Kupffer cell function in allyl alcohol-induced hepatotoxicity

The roles of neutrophils (PMNs) and Kupffer cells in hepatotoxicity caused by allyl alcohol in rats in vivo were examined. To test the involvement of PMNs in the response to allyl alcohol, the number of circulating PMNs was reduced to < 500/μl by treatment with immunoglobulin (Ig) isolated from serum of rabbits treated with rat PMNs (anti-PMN Ig). Rats received anti-PMN Ig or control Ig 6 h before and 6 h after administration of allyl alcohol (40 mg/kg, i.p.). Hepatotoxicity was assessed 18 h after allyl alcohol administration. In rats pretreated with control Ig, treatment with allyl alcohol resulted in hepatotoxicity as evidenced by an increase in the activity of alanine aminotransferase (ALT) in serum. Neutropenia did not attenuate hepatic injury caused by allyl alcohol. Leukopenia induced by pretreatment with cyclophosphamide also did not influence the hepatotoxic response to allyl alcohol. To inhibit the function of Kupffer cells, animals were treated with gadolinium chloride (GdCl 3; 10 mg/kg, i.v.) 24 h before administration of allyl alcohol. This dose of GdCl 3 decreased in situ clearance of colloidal carbon by 64%. Despite the inhibition of Kupffer cell function, ALT activity in serum was not different in allyl alcohol-treated rats pretreated with GdCl 3 and those pretreated with saline vehicle. Histopathologic evaluation of the livers confirmed a lack of protective effect of GdCl 3. These results suggest that neither neutrophils nor Kupffer cells play a major role in liver injury due to allyl alcohol.

Direct and Correlated Responses to Multitrait, Divergent Selection for Immunocompetence

Leghorn lines had been selected for an immunocompetence index based on four traits: antibody production to Mycoplasma gallisepticum (MG) and Pasteurella multocida (PM) vaccines, reticuloendothelial clearance of colloidal carbon (CCA), and cell-mediated, wing web response to phytohemagglutinin (PHA). The purpose of this study was to produce replicated lines of chickens with divergent levels of multitrait immunocompetence by index selection. The objectives of analyses of Generations 5 to 7 of this study was to characterize these lines with respect to immune-response traits, correlations among these traits, and correlated responses in other important production traits.Differences (P < .05) existed between the lines selected for high or low immune response and between the two replicates in mean breeding values and in individual immune-response traits. Averages of heritability estimates, weighted by number of offspring and pooled across three generations (two cycles of selection), estimated by using sire variance components and parent-offspring correlations were, respectively, .16 and .09 for the index, .31 and .08 for MG, .21 and −.02 for PM, .06 and .05 for CCA, and .08 and .12 for PHA. Realized heritabilities (response divided by effective selection differential) pooled across the two selection cycles, were .19 and .11 for the index, .06 and −.01 for MG, .44 and .32 for PM, 1.52 and −1.21 for CCA, and .48 and .15 for PHA, for Replicates 1 and 2, respectively. Phenotypic correlations among traits were generally small, and several estimates were negative. Estimates of genetic correlation varied widely. Juvenile and adult body weights, age of first egg, 32-wk egg weight, and rate of egg production were analyzed to evaluate effects of selection on these traits of direct economic importance. Very few differences were noted.

Macrophages, but not dendritic cells, accumulate colloidal carbon following administration in situ.

The dendritic cell system operates in situ to capture and present antigens in a form that is immunogenic to T cells. It is likely that dendritic cells require endocytic activity in order to process antigens. On the other hand, macrophages are considered to be the principal cells that internalize substrates in situ. We therefore investigated the phenotype of cells that scavenge the indigestible endocytic tracer, colloidal carbon, by phenotyping the endocytic cells with monoclonal antibodies that help distinguish macrophages from dendritic cells. Of some importance was the monoclonal N418, an antibody to the p150/90 leukocyte beta 2 integrin. FACS analyses on isolates from blood, spleen and peritoneal cavity showed that N418 reacts primarily with dendritic cells. N418 also stained dendritic profiles strongly in tissue sections of liver and spleen, but most of the cells that actively endocytosed carbon in both organs showed little or no N418 staining. Likewise, carbon could not be identified in cells that react with M342, which stains intracellular granules of dendritic cells. In contrast, the carbon-labeled cells in both liver and spleen were labeled with antibodies (SER-4, F4/80, FA11) that bind primarily to isolated macrophages. Therefore the clearance of colloidal carbon in situ reflects the scavenging activity of macrophages and not the endocytic activity that underlies the antigen presenting function of dendritic cells.

Vitamin A Potentiation of Carbon Tetrachloride Hepatotoxicity: Role of Liver Macrophages and Active Oxygen Species

Pretreatment of rats with large doses of vitamin A (VA) potentiates the hepatotoxicity of CCl 4. Because our previous studies indicate that VA treatment does not enhance CCl 4, metabolism but does enhance CCl 4-induced lipid peroxidation and activates liver Kupffer cells to release increased amounts of oxygen-centered free radicals, the current studies were designed to determine if VA treatment potentiates CCl 4-induced liver injury through increased release of reactive oxygen species. Plasma clearance of colloidal carbon, an index of Kupffer cell phagocytic activity, was enhanced two- to threefold in rats treated for 7 days with VA (retinol, 250,000 IU/kg per day). Accordingly, VA treatment alone caused Kupffer cell activation. To determine if these activated Kupffer cells could potentiate hepatic injury through release of reactive oxygen species upon Cd4 challenge, polyethylene glycol coupled-superoxide dismutase (PEG-SOD, 10,000 IU/kg) or -catalase (PEG-CAT, 40,000 IU/kg) was given iv 2 hr after CCl 4, (0.15 ml/kg, ip) to control or VA-pretreated rats to quench any released reactive oxygen. PEG-SOD and PEG-CAT effectively blocked VA potentiation of CCl 4, liver injury as assessed at 24 hr by change in plasma ALT. Methylpalmitate (MP, 2 g/kg), an inhibitor of Kupffer cell phagocytosis and related oxygen burst, also blocked the potentiation of liver injury when given iv 24 hr before CCI4 to VA-pretreated rats. At the doses used, PEG-SOD or PEG-CAT did not influence CCl 4 toxicity in control rats (at 0.15 or 2 ml CCl 4/kg). Importantly, SOD, CAT, and MP blocked the enhanced lipid peroxidation induced by CCl 4, in VA-pretreated rats. From these findings we conclude that the potentiation of CCl 4, liver injury by VA pretreatment is mediated, at least in part, by active oxygen species released from Kupffer cells and possibly other macrophages that are activated by VA. Supporting this conclusion is the failure of VA pretreatment to increase the release of LDH from suspension of hepatocytes incubated with CCl 4,.

The effect of Mycoplasma arthritidis infection on the kinetics of colloidal carbon clearance in mice

The activity of phagocytes from A/J mice was estimated by the carbon clearance test following injection of Mycoplasma arthritidis. Phagocytic activity was significantly depressed 12 h post-infection (P=0.001) and returned to normal values at 24 h. For animals examined 2 and 7 days post-infection, the overall phagocytic activity increased significantly (P<10−4). Phagocytic activity gradually decreased and returned to that of the control group by the end of the fourth week. The relative weights of liver and spleen were significantly increased from the 2nd day post infection (P=0.0028 and P=0.0014 respectively) and remained increased until the end of the experiment. The early depressive effect on phagocytic activity may be related to superantigen activity with the production of mediators such as macrophage deactivating factor. The later expansion of the macrophage population might bring about the stimulation of autoreactive clones of T and B cells and be responsible for the chronic arthritis that developed in the mycoplasma treated mice.

Ischemic injury in liver transplantation: difference in injury sites between warm and cold ischemia in rats.

Using liver allografts with warm or cold ischemia, we evaluated functional and morphological alterations in hepatocytes, sinusoidal endothelial cells and Kupffer cells in a rat transplantation model. All recipients of allografts with either 4 hr of cold or 30 min of warm ischemia lived more than 22 days and were judged viable. On the other hand, all recipients of grafts with 6 hr of cold or 60 min of warm ischemia died within 2 days and were therefore judged to be nonviable. With these viable and nonviable allograft models, hepatocyte function was evaluated by the bile output and serum glutamic-oxaloacetic transaminase, serum glutamic-pyruvic transaminase and serum lactate dehydrogenase levels; endothelial cell function was judged by the serum hyaluronic acid level, and Kupffer cell function was measured by an intravenous colloidal carbon clearance test. Hepatocyte injury was the prominent feature in warm ischemic grafts, especially in the nonviable ones. On the other hand, serum hyaluronic acid values were significantly higher in the nonviable cold ischemic group, compared with the viable counterpart, suggesting that the functional depression of endothelial cells was predominant in cold, nonviable livers. Histological examinations coincided with the above findings. The phagocytic activity of Kupffer cells was depressed by warm or cold ischemia, whereas the number of Kupffer cells was reduced in the warm ischemia group. We conclude that in liver allografts the main site of injury in warm ischemia is the hepatocytes and suggest that cold ischemia is associated with endothelial cell damage.

Effects of liposome encapsulated hemoglobin on the reticuloendothelial system.

The effects of different doses (4, 10, and 25%) of liposome encapsulated hemoglobin (LEH) were measured on the Reticuloendothelial System (RES) and Kupffer cells (KC) by i) colloidal carbon clearance in the rat in vivo and in the isolated perfused liver, ii) magnetometry, and iii) histological analysis. At the highest dose, in vivo carbon clearance rates (k) were half the rate as in controls at 2 and 12 hours post-treatment. By 24 hours post-treatment clearance rates were at control levels. Empty liposomes (LIP) caused a 2-fold decrease in k at 2 hours only. With both LEH and LIP, the effects were less severe when rats were given the lower doses. Magnetometric studies showed a decrease in KC phagosomal motion in the LEH-treated (25%) rats at 2 and 24 hours that returned to control levels at 2 weeks. Perfused livers from rats treated with a low dose of LEH cleared carbon at the same rate as LIP and Krebs Ringer Bicarbonate (KRB) controls. Histological examination showed minimal tissue damage in all test groups. Thus, LEH and LIP have some short-term deleterious effects on KC and the RES probably due to RES blockade rather than cellular damage.

Perfluorochemical emulsions decrease Kupffer cell phagocytosis

One drawback to using perfluorochemical emulsions as blood substitutes is that perfluorochemical particles are cleared from the blood by the reticuloendothelial system, primarily liver and spleen. We measured the impact of two perfluorochemical emulsions on clearance of colloidal carbon (less than 1 microns) and 51Cr-sheep red blood cells (about 8 microns) by the reticuloendothelial system in vivo and in the isolated perfused liver. Male rats were injected with 2 ml/100 gm body wt of Fluosol-DA or Oxypherol-ET for 4 consecutive days. Carbon (1 ml/100 gm body wt) or sheep red blood cells (0.05 ml of 5% vol/vol/100 gm body wt) were then injected intravenously (in vivo) or added to perfusate. Samples were taken at several time points for 1 hr. In the isolated perfused liver, carbon clearance was depressed by 25% 1 day after treatment. Rates returned to control levels by 12 days in Fluosol-DA-treated rats but remained depressed by 67% in Oxypherol-ET-treated rats. Sheep red blood cell (8 microns) clearance was two to five times slower than carbon clearance and depressed by 40% in livers from Fluosol-DA rats 1 day and 12 days after treatment. Added serum did not improve phagocytosis. In vivo carbon clearance remained normal in Fluosol-DA-treated rats but decreased by 74% in Oxypherol-ET-treated rats 1 day after treatment, returning to normal by 12 days. Clearance rates were similar in control rats in vivo and in the perfused liver. We conclude that the isolated perfused liver is a good model to measure liver clearance function. Although low doses of perfluorochemical emulsions may depress Kupffer cell phagocytosis, general reticuloendothelial system function is not significantly compromised.

Impaired immune responsiveness in Plasmodium berghei immune mice

Mice immunized against Plasmodium berghei parasites by drug-controlled infection exhibited decreased immunoresponsiveness against rabbit red blood cells (RRBC). Increasing RRBC antigen dose increased responsiveness, but agglutinating anti-RRBC antibodies of the IgG class remained undetectable. Clearance of colloidal carbon from the bloodstream of malaria-immunized mice was not different from controls. Removal of all the persistent parasites from immune mice did not restore responsiveness until 140 days after treatment, suggesting that the parasite per se did not influence responsiveness directly. Because of this, and because of the fact that priming of mice with RRBC before P. berghei immunization was not more effective than priming after immunization, it was concluded that antigen uptake and subsequent presentation were not impaired in P. berghei immune mice, in contrast to infected mice. Anti-RRBC antibodies were detected in serum of P. berghei immune mice, but regulation of responsiveness to RRBC by transfer of such immune mouse serum was not found. Immunoglobulin levels, especially of the IgG2 and IgG3 subclass were elevated in sera of P. berghei immune mice, which indicated an LPS-like polyclonal activation. The results also suggest that during drug-controlled infection, which leads to immunity against infection, a state of B-cell tolerance is induced.