Glycyrrhizin attenuates hepatic ischemia-reperfusion injury by suppressing HMGB1-dependent GSDMD-mediated kupffer cells pyroptosis.

Gasdermin D (GSDMD), a genetic substrate for inflammatory caspases, plays a central role in pyroptosis of macrophages and release of interleukin­1ß (IL-1ß), but was mainly referred to microbial infection. High mobility group box-1 (HMGB1), served as an alarm molecule during various pathological process, has been widely recognized to be involved in liver ischemia-reperfusion (I/R). Glycyrrhizin, a natural anti-inflammatory and antiviral triterpene in clinical use, was recently referred to have ability to prevent I/R induced liver injury by inhibiting HMGB1 expression and activity. However, the mechanisms responsible for damage amelioration subsequently to HMGB1 inhibition during liver I/R remain enigmatic. This study was designed to explore the functional role and molecular mechanism of glycyrrhizin in the regulation of I/R induced liver injury. We found that liver I/R promotes GSDMD-mediated pyroptotic cell death of Kupffer cells, which was inhibited by glycyrrhizin. Interestingly, endogenous HMGB1, not exogenous one, was involved in hypoxia-reoxygenation (H/R) induced pyroptosis. Moreover, GSDMD knockdown protects kupffer cells against H/R induced pyroptosis in vitro. Here, we report, for the first time, that glycyrrhizin attenuated tissue damage and kupffer cells pyroptosis during liver ischemia-reperfusion injury (LIRI) and identify a previously unrecognized HMGB1- dependent GSDMD- mediated signaling pathway in the mechanism of kupffer cells pyroptosis induced by H/R. Our findings provide the first demonstration of GSDMD-determined pyroptotic cell death responsible for I/R induced release of IL-1ß and this would provide a mandate to better understand the unconventional mechanisms of cytokine release in the sterile innate immune system.

Effect of Soothing Gan (Liver) and Invigorating Pi (Spleen) Recipes on TLR4-p38 MAPK Pathway in Kupffer Cells of Non-alcoholic Steatohepatitis Rats.

OBJECTIVE: To investigate the mechanism of inflflammatory-mediated toll-like receptor 4 (TLR4)-p38 mitogen-activated protein kinase (p38 MAPK) pathway in Kupffer cells (KCs) of non-alcoholic steatohepatitis (NASH) rats and the intervention effect of soothing Gan (Liver) and invigorating Pi (Spleen) recipes on this pathway. METHODS: After 1 week of acclimatization, 120 Sprague-Dawley male rats were randomly divided into 8 groups using a random number table (n=15 per group): normal group, model group, low-dose Chaihu Shugan Powder (, CHSG) group (3.2 g/kg), high-dose CHSG group (9.6 g/kg), low-dose Shenling Baizhu Powder (, SLBZ) group (10 g/kg), high-dose SLBZ (30 g/kg) group, and low- and highdose integrated recipe (L-IR, H-IR) groups. All rats in the model and treatment groups were fed with a high-fat diet (HFD). The treatments were administrated by gastrogavage once daily and lasted for 26 weeks. The liver tissues were detected with hematoxylin-eosin (HE) and oil red O staining. Levels of liver lipids, serum lipids and transaminases were measured. KCs were isolated from the livers of rats to evaluate the mRNA expressions of TLR4 and p38 MAPK by real-time flfluorescence quantitative polymerase chain reaction, and proteins expressions of TLR4, p-p38 MAPK and p38 MAPK by Western blot. Levels of inflammatory cytokines including tumor necrosis factor α (TNF-α), interleukin (IL)-1 and IL-6 in KCs were measured by enzyme-linked immunosorbent assay. RESULTS: After 26 weeks of HFD feeding, HE and oil red O staining showed that the NASH model rats successfully reproduced typical pathogenesis and histopathological features. Compared with the normal group, the model group exhibited significant increases in body weight, liver weight, liver index, serum levels of total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol, and aspartate aminotransferase as well as TC and TG levels in liver tissues, and significant decrease in serum level of high-density lipoprotein cholesterol (Plt;0.05 or Plt;0.01), while those indices were significantly ameliorated in the H-IR group (Plt;0.05 or Plt;0.01). Higher levels of TNF-α, IL-1 and IL-6 in KCs were observed in the model group compared with the normal group (Plt;0.01). Significant decreases in TNF-α, IL-1 and IL-6 were observed in the H-SLBZ, H-IR and L-IR groups compared with the model group (Plt;0.05 or Plt;0.01). The mRNA expressions of TLR4 and p38 MAPK and protein expressions of TLR4, p38 MAPK and p-p38 MAPK in KCs in the model group were significantly higher than the normal group (Plt;0.01), while those expression levels in the L-IR and H-IR groups were significantly lower than the model group (Plt;0.05 or Plt;0.01). CONCLUSION: Inflflammation in KCs might play an important role in the pathogenesis of NASH in rats. The data demonstrated the importance of TLR4-p38MAPK signaling pathway in KCs for the anti-inflflammatory effect of soothing Gan and invigorating Pi recipes.

Kupffer cell-monocyte communication is essential for initiating murine liver progenitor cell-mediated liver regeneration.

UNLABELLED: Liver progenitor cells (LPCs) are necessary for repair in chronic liver disease because the remaining hepatocytes cannot replicate. However, LPC numbers also correlate with disease severity and hepatocellular carcinoma risk. Thus, the progenitor cell response in diseased liver may be regulated to optimize liver regeneration and minimize the likelihood of tumorigenesis. How this is achieved is currently unknown. Human and mouse diseased liver contain two subpopulations of macrophages with different ontogenetic origins: prenatal yolk sac-derived Kupffer cells and peripheral blood monocyte-derived macrophages. We examined the individual role(s) of Kupffer cells and monocyte-derived macrophages in the induction of LPC proliferation using clodronate liposome deletion of Kupffer cells and adoptive transfer of monocytes, respectively, in the choline-deficient, ethionine-supplemented diet model of liver injury and regeneration. Clodronate liposome treatment reduced initial liver monocyte numbers together with the induction of injury and LPC proliferation. Adoptive transfer of monocytes increased the induction of liver injury, LPC proliferation, and tumor necrosis factor-α production. CONCLUSION: Kupffer cells control the initial accumulation of monocyte-derived macrophages. These infiltrating monocytes are in turn responsible for the induction of liver injury, the increase in tumor necrosis factor-α, and the subsequent proliferation of LPCs.

Amiloride reduces portal hypertension in rat liver cirrhosis.

OBJECTIVE: This study aimed to investigate the effect of amiloride on portal hypertension. Amiloride is known to inhibit Na(+)/H(+) exchangers on activated hepatic stellate cells. METHODS: Liver cirrhosis in rats was induced by bile duct ligation (BDL) or thioacetamide (TAA) administration. The effects of zymosan for Kupffer cell (KC) activation or a thromboxane (TX) analogue (U46619) were tested in isolated perfused livers of cirrhotic rats and in vivo. Downstream mechanisms were investigated using Rho kinase inhibitor (Y-27632) or amiloride. Acute and chronic effects of amiloride and canrenoate on portal pressure were compared in perfused livers and in vivo. TXB(2) efflux was measured by ELISA. The phosphorylation state of moesin (p-moesin) as an indicator of Rho kinase activity and expression of the thromboxane synthase were assessed by western blot analyses. The activity of hepatic stellate cells was analysed by western blot and staining for alpha-smooth muscle actin (alpha-SMA). RESULTS: In BDL rats, KC activation via zymosan increased portal pressure. This was attenuated by the Rho kinase inhibitor Y-27632. Increased thromboxane efflux following zymosan infusion remained unaltered by Y-27632. The infusion of amiloride attenuated zymosan- and U46619-induced increases in portal perfusion pressure. In vivo, direct administration of amiloride, but not of canrenoate, lowered portal pressure. In TAA and BDL rats, treatment with amiloride for 3 days reduced basal portal pressure and KC-induced increases in portal pressure whereas canrenoate had no effect. In livers of amiloride-treated animals, the phosphorylation state of moesin and the number of alpha-SMA positive cells were reduced. CONCLUSIONS: Amiloride lowers portal pressure in rat liver cirrhosis by inhibition of intrahepatic vasocontraction. Therefore, patients with cirrhosis and portal hypertension may benefit from amiloride therapy.

The role of p65 NF-kappaB/RelA in pancreatitis-induced Kupffer cell apoptosis.

Acute pancreatitis induces liver injury by upregulating Kupffer cell-derived Fas/FasL; on the other hand, acute pancreatitis induces apoptosis of Kupffer cells via NF-kappaB-dependent pathways. The balance between upregulation of Fas/FasL and Fas/FasL-induced apoptosis of its originator cell may determine the severity of pancreatitis-related liver injury. The aim of our study was to determine the role of p65 NF-kappaB/RelA in pancreatitis-induced Kupffer cell apoptosis. Acute pancreatitis was induced in NIH Swiss mice by a choline-deficient ethionine-supplement (CDE) diet. In vitro mouse Kupffer cell line was transfected with p65 siRNA and treated with pancreatic elastase to mimic pancreatitis. CDE pancreatitis upregulated nuclear translocation of p65 NF-kappaB/RelA, Fas/FasL, caspase-3, and DNA fragmentation in mice livers (all P < 0.001). In vitro, pancreatic elastase mimicked CDE-pancreatitis by upregulating nuclear translocation of p65 NF-kappaB/RelA, Fas/FasL, caspase-3, DNA fragmentation, and apoptosis in Kupffer cells (all P < 0.001). Transfection with p65 siRNA attenuated the elastase-induced nuclear translocation of p65 NF-kappaB/RelA, upregulation of Fas/FasL, caspase-3, DNA fragmentation, and apoptosis in Kupffer cells (all P < 0.001). Acute pancreatitis activates p65 NF-kappaB/RelA and induces apoptosis of Kupffer cells. Inhibition of p65NF-kappaB/RelA attenuates elastase-induced upregulation of proapoptotic pathways and apoptosis in Kupffer cells. The ability of Kupffer cells to autoregulate their stress response by inducing self-apoptosis warrants further investigation.

Swift increase in alcohol metabolism (SIAM): understanding the phenomenon of hypermetabolism in liver.

The swift increase in alcohol metabolism (SIAM) is a phenomenon defined as a rapid increase in hepatic respiration and alcohol metabolism after administration of a bolus dose of alcohol. Continuous exposure to alcohol is known to produce adaptive changes in liver alcohol and oxygen metabolism. A considerable burst of hepatic respiration can also occur after administration of a single large dose of alcohol and results in a near doubling of alcohol metabolism, a high demand for oxygen, and downstream or pericentral hypoxia. These dramatic changes in rates of alcohol metabolism and tissue concentrations of oxygen are not due to induced enzyme activity in liver. This phenomenon depends on activation of mitochondrial function, an increase in co-factor supply for nicotinamide adenine dinucleotide-dependent alcohol metabolism, depletion of glycogen reserves, liberation of fatty acids through activation of an adrenergic response to alcohol providing substrate for catalase, and activation of Kupffer cells, the hepatic resident macrophages responsible for production of cytokines and prostaglandins. An understanding of the mechanisms of hypermetabolism in liver can have vital ramifications for knowledge of both alcohol-related and alcohol-unrelated liver injury because hypoxia that is a result of hypermetabolism can compound effects of pharmaceuticals and environmental agents on the liver. Swift increase in alcohol metabolism is an excellent example of the complexity of cell-cell interactions in liver and extrahepatic regulation of biochemical and molecular events in this organ, and this important phenomenon shall be considered in studies of liver disease and biochemistry.

New method of delivering gene-altered Kupffer cells to rat liver: studies in an ischemia-reperfusion model.

BACKGROUND & AIMS: Kupffer cells play a major role in the pathogenesis of several diseases. They release physiologically active substances that often lead to localized tissue injury. Therefore, the aim of this study was to establish a model to protect the liver through supplementation of Kupffer cells that have been transduced by recombinant adenovirus. METHODS: Optimal conditions for intravenous injection in rats were established using carbon-labeled Kupffer cells. Adenoviral-transduced Kupffer cells encoding the Cu/Zn-SOD gene (Ad.SOD1) or beta-galactosidase reporter gene (Ad.LacZ) were transplanted into recipient rats. Twenty-four hours after transplantation, 70% hepatic ischemia-reperfusion was used to induce hepatic oxidative stress, and liver injury was determined 8 or 24 hours later. RESULTS: In initial experiments, 10%-20% of the injected carbon-labeled cells were localized in the host liver after 24 hours, representing approximately 1% of the total population of Kupffer cells. Pretreatment of the recipient with a single dose of cyclosporin A maximized Kupffer cell reseeding up to 4%-10% of the total Kupffer cell population, suggesting that efficiency is limited by host immune response. Moreover, reseeded Kupffer cells were retained in host livers for up to 14 days after transplant. In livers of animals injected with Kupffer cells transduced with Ad.LacZ, transgene expression was observed, indicating Kupffer cell functional integrity. Injection of Kupffer cells transduced with Ad.SOD1 significantly blunted the increase in serum transaminases and liver injury because of ischemia-reperfusion compared with controls. CONCLUSIONS: This novel approach allows delivery of transduced Kupffer cells in rats, which can be used as an investigative tool as well as a therapeutic strategy against inflammatory liver diseases.

Activation of human and mouse Kupffer cells by lipopolysaccharide is mediated by CD14.

Upregulation of CD14 in Kupffer cells has been implicated in the pathogenesis of several forms of liver injury, including alcoholic liver disease. However, it remains unclear whether CD14 mediates lipopolysaccharide (LPS) signaling in this specialized liver macrophage population. In this series of experiments, we determined the role of CD14 in LPS activation of Kupffer cells by using several complementary approaches. First, we isolated Kupffer cells from human livers and studied the effects of anti-CD14 antibodies on LPS activation of these cells. Kupffer cells were incubated with increasing concentrations of LPS in the presence and absence of recombinant human LPS binding protein (LBP). With increasing concentrations of LPS, human Kupffer cell tumor necrosis factor-alpha (TNF-alpha) production (a marker for Kupffer cell activation) increased in a dose-dependent manner in the presence and absence of LBP. In the presence of anti-human CD14 antibodies, the production of TNF-alpha was significantly diminished. Second, we compared LPS activation of Kupffer cells isolated from wild-type and CD14 knockout mice. Kupffer cells from CD14 knockout mice produced significantly less TNF-alpha in response to the same amount of LPS. Together, these data strongly support a critical role for CD14 in Kupffer cell responses to LPS.

n-3 Polyunsaturated fatty acid-enriched diet does not protect from liver injury but attenuates mortality rate in a rat model of systemic endotoxemia.

OBJECTIVE: We investigated the potential of dietary fish oil containing n-3 polyunsaturated fatty acids to attenuate hepatic injury and mortality rate of rats in response to systemic endotoxemia. DESIGN: Prospective, randomized, controlled animal study. SETTING: University laboratory. SUBJECTS: A total of 43 male Sprague Dawley rats. INTERVENTIONS: Rats were fed either fish oil supplement or regular standard lab chow. After 8 wks of feeding, each diet group was subjected to a single exposure of lipopolysaccharide (Escherichia coli, 10 mg/kg intravenously) or saline. Hepatic microvascular response and liver injury were assessed by in vivo analysis of Kupffer cell phagocytic activity, leukocyte-endothelial cell interaction, nutritive sinusoidal perfusion failure, and parenchymal cell apoptosis (intravital fluorescence epi-illumination technique) as well as bile flow, serum liver enzyme activities, and tissue histomorphology. MEASUREMENTS AND MAIN RESULTS: In animals fed a standard diet, livers at 16 hrs after lipopolysaccharide-exposure exhibited depressed Kupffer cell phagocytic activity, enhanced hepatic microvascular leukocyte activation, leukocytic tissue infiltration, sinusoidal perfusion failure, and parenchymal cell apoptosis. Hepatic microvascular injury was further accompanied by reduced bile flow and enhanced liver enzyme release. The fish oil enriched diet did not significantly change the multiple features of endotoxemia-associated liver injury; however, it maintained arterial blood pressure, systemic leukocyte count, and acid base balance and showed a tendency toward improved survival on lipopolysaccharide exposure with a 16 hr-survival rate of 80% (p =.06 vs. survival rate of 40% in animals fed a regular diet). Moreover, slightly increased serum concentrations of interleukin-10 coincided with enhanced concentrations of interleukin-6 in fish oil fed endotoxemic animals. Healthy, non-lipopolysaccharide-exposed, fish oil fed animals did not differ from those fed with the regular diet, except for dampened Kupffer cell phagocytic activity. CONCLUSIONS: Fish oil feeding does not protect from local endotoxemia-induced hepatic microvascular dysfunction. However, dietary modulation of inflammatory mediator response by macrophages, constituting an appropriate immune response, could add to the survival advantage seen in fish oil-fed animals on exposure to lipopolysaccharide.

Activation of in vivo Kupffer cell function by oral administration of Cordyceps sinensis in rats.

We investigated the effect of water extracts of Cordyceps sinensis (WECS) on Kupffer cell function in rats. Rats were received a single i.v. injection of a colloidal carbon solution and then the clearance rate from the blood were measured. The rats had been daily administered with WECS, p.o. at a dose of 200 mg/kg for 25 days until the day before the injection of colloidal carbon. The half-life of the colloidal carbon in the blood of rats administered WECS 200 mg/kg was significantly shorter than that of the control rats. This suggests that accelerated function of Kupffer cells is partially involved in the anti-metastatic action of WECS.

Mechanism of increased lung injury after acute pancreatitis in IL-10 knockout mice.

BACKGROUND: IL-10 is a potent anti-inflammatory cytokine which inhibits inflammatory cytokine release from all tissue sites. Hepatic cytokine release from Kupffer cells (KC) is one important source of inflammatory cytokines and may be the main one causing lung damage in acute pancreatitis (AP). Here we studied the KC contribution to lung injury in IL-10 knockout (KO) mice, in which tissue inflammatory cytokine release from all sites is unrestrained, and AP is more severe. METHODS: Three- to 4-week-old C57BL/6J mice and KO mice on a C57BL/6J background were used. Control mice received regular chow and gadolinium chloride (GD; 1 mg/100 g iv), to inhibit KC activity, or saline. Pancreatitis mice received a choline-deficient, ethionine-supplemented diet for 66 h to induce AP and saline or GD injections iv. After 66 h, lung tissue was assessed for edema, myeloperoxidase (MPO), and superoxide dismutase (SOD). Histology was scored in a blinded fashion. RESULTS: In pancreatitis KO mice, KC blockade had no effect on the degree of lung edema, lung neutrophil infiltration, and lung histology score. As expected, each of these parameters was more severe in the KO mice than in the normal mice: lung wet/dry ratio 5.3 +/- 0.2 versus 4.3 +/- 0.13; lung MPO (U/g) 1.9 +/- 0.2 versus 1.1 +/- 0.08; histology score 7.1 +/- 0.8 versus 5.3 +/- 0.5. CONCLUSION: Endogenous IL-10 is important in reducing the lung injury in this model of AP. KC-derived cytokines were of minor importance, compared to those derived from all other tissue sites.

Kupffer cells contain a glycine-gated chloride channel.

Here the effect of glycine on intracellular Ca2+ concentration ([Ca2+]i) in cultured Kupffer cells stimulated with lipopolysaccharide (LPS) was investigated to assess the possibility that they contain a glycine-gated chloride channel. LPS (10 micrograms/ml) increased [Ca2+]i rapidly, with peak values reaching 307 +/- 29 nM. Glycine (1 mM) prevented this increase nearly completely. Low concentrations of strychnine (1 microM), a glycine receptor antagonist, reversed the inhibitory effect of glycine completely; however, high concentrations of strychnine (1 mM) mimicked glycine. The effects of glycine and high-dose strychnine were prevented when cells were incubated in chloride-free buffer. Furthermore, potassium (25 mM) and LPS depolarized the Kupffer cell plasma membrane, whereas glycine caused hyperpolarization and prevented depolarization due to potassium and LPS. Moreover, tumor necrosis factor-alpha (TNF-alpha) production in cultured Kupffer cells due to LPS was decreased significantly by glycine. Therefore, it is concluded that Kupffer cells contain a glycine-gated chloride channel similar to that described previously in the central nervous system. Prevention of increases in [Ca2+]i due to LPS by activation of chloride influx reduced synthesis and release of toxic mediators by Kupffer cells.

Roles of oxidative stress in activation of Kupffer and Ito cells in liver fibrogenesis.

An increasing body of experimental evidence is emerging to incriminate oxidative stress as a pivotal signal for liver fibrogenesis. This paper reviews the results from our studies testing this hypothesis. In the rat model of alcoholic liver disease, the importance of oxidative stress was supported by marked accentuation of liver fibrosis by dietary supplementation of iron, a pro-oxidant, and the significant correlation of the liver malondialdehyde (MDA) and 4-hydroxynonenal (4HNE) levels with the hepatic collagen accumulation. Both MDA and 4HNE adduct epitopes were detected intensely and diffusely in close association with collagen deposition. The direct cause and effect relationship between MDA/4HNE and Ito cell stimulation was indicated by the demonstration of Ito cell collagen gene induction by these aldehydes in culture. In primary cultures of rat Kupffer cells (KC), addition of antioxidants such as alpha-tocopherol acetate and succinate suppressed mRNA expression and the release of interleukin (IL)-6 and tumour necrosis factor alpha (TNF alpha). In rats with biliary fibrosis, an increase in the liver MDA level was accompanied by enhanced mRNA expression of procollagen alpha 1(I) and transforming growth factor beta 1 in Ito cells; and that of TNF alpha and IL-6 in KC. Furthermore, the gel shift assay of KC nuclear extracts showed enhanced NF-kB DNA binding activity. These results support the proposal that enhanced oxidative stress constitutes an important signal for activation of Kupffer and Ito cells in experimental liver fibrogenesis.

Role of Kupffer cells in iodized oil embolization.

RATIONALE AND OBJECTIVES: Iodized oil is a common oily embolic agent used in chemoembolization for treating hepatic tumors. However, how the iodized oil is cleared from the liver has been an unsettled and controversial issue. In this study, the authors attempt to clarify whether Kupffer cells are involved in the clearance of iodized oil and to evaluate the effect of hepatic arterial injection of iodized oil on the functional status of Kupffer cells. METHODS: Iodized oil was injected into the proper hepatic artery in 42 Fischer 344 rats. In vivo microscopy was performed immediately after and 1, 3, 7, 15, 30, and 60 days after injection. Electron microscopy was performed after in vivo microscopy. RESULTS: Kupffer cells actively captured and phagocytosed iodized oil droplets in the hepatic circulation. The number and functional status of Kupffer cells in the liver were significantly increased after the injection of the iodized oil and returned to normal when the liver was cleared of the oil. CONCLUSIONS: Kupffer cells play an important role in clearing iodized oil from the liver. Iodized oil activates the immune defense system in the liver, which may have a synergistic effect in tumor treatment.

Modulation of Kupffer cell activity by Tinospora cordifolia in liver damage.

Kupffer cells are major determinants of outcome of liver injury. Their activity was therefore studied in a model of chronic liver disease. The effect of Tinospora cordifolia, an indigenous agent with proven hepatoprotective activity, was evaluated on Kupffer cell function, using carbon clearance test as a parameter. Rats were divided into two major groups. In Gp I which served as normal control t1/2 of carbon was 9.48 +/- 4.14 min. GpII received horse-serum in a dose of 0.5 ml/100 gm b.w. i.p. for a period of 12 weeks and was divided into three sub-groups. In Gp IIA at the end of 12 weeks half-life of carbon was found to be significantly increased to 19.86 +/- 7.95 min (p < 0.01). Indicating suppressed Kupffer cell function in chronic liver damage. In Gp IIB treated with vehicle for 4 more weeks there was significant prolongation of half-life to 38.32 +/- 10.61 min (p < 0.01), indicating perpetuation of damage in absence of damaging agent. Whereas in Gp IIc, treated with Tinospora cordifolia t 1/2 was decreased to 14.24 7.74 min (p < .01), as compared to vehicle control indicating a significant improvement in Kupffer cell function and a trend towards normalization.

Effects of combined therapies with protein-bound polysaccharide (PSK, Krestin) and fluorinated pyrimidine derivatives on experimental liver metastases and on the immunologic capacities of the hosts.

An experimental model is introduced for the study of liver metastases using intrasplenically injected EL-4 and Lewis lung tumor cells. Fluorinated pyrimidine derivatives, 1-(2-tetrahydrofuryl)-5-fluorouracil and 5-fluorouracil, showed inhibitory effects on the frequencies of liver metastases. Immunosuppressive effects of these drugs were compared at the doses capable of showing 50% inhibition of the development of metastatic nodules. These derivatives strongly suppressed the phagocytic activity and the number of Kupffer cells of the liver and then the humoral response against sheep red blood cells, the delayed hypersensitivity against picryl chloride. On the contrary, combined administration of protein-bound polysaccharide (PSK) and 1-(2-tetrahydrofuryl)-5-fluorouracil showed no inhibitory effect on these activities.

Vitamin A supplementation improves macrophage function and bacterial clearance during experimental salmonella infection.

The effects of additional but nontoxic amounts of vitamin A on susceptibility to salmonella infection was studied by comparing rates of bacterial clearance and phagocytosis. Forty-eight male Lewis rats were divided into a treatment group receiving a total of 6000 units of vitamin A palmitate weekly for 5 weeks and a control group was given an equal volume of saline. After completion of the treatment regimen, one-half from each group were infected intraperitoneally with 10(5) Salmonella typhimurium; the other half received intraperitoneal injection of saline. At this time no differences in weight gain were noted and all animals were sacrificed within 2 weeks. At 72 hr after bacterial challenge, all saline-treated control animals displayed bacteremia. Cultures of liver and splenic homogenates were positive in 89 and 100% of infected control animals vs 0 and 44% for treated animals during the first week of infection. Kupffer cell, peritoneal, and splenic macrophages of the vitamin A-treated group had greater phagocytic activity than controls as assessed by the percentage of cells ingesting yeast particles and by the number of particles ingested (phagocytic index). These results suggest that vitamin A in moderate amounts may benefit the host's response to infection by enhancing phagocytic cell function.

Endogeneous interferon alpha/beta produced by murine Kupffer cells augments liver-associated natural killing activity.

Nonparenchymal liver cells from untreated C3HeB/FeJ mice, when incubated in medium containing-10% fetal bovine serum or portal serum, produced significant amounts of interferon alpha/beta (IFN alpha/beta). In contrast, other cell populations (spleen, mononuclear blood cells and peritoneal cells) from C3HeB/FeJ mice or nonparenchymal liver cells from other strains of mice (C3H/HeJ, germ-free C3H/HeN and C57Bl/6J) produced little or no detectable IFN in fetal bovine serum under the same culture conditions. The cells in the nonparenchymal liver cell population responsible for IFN alpha/beta production were adherent, phagocytic, silica-sensitive, carbonyl-iron-sensitive, and Thy1.2-, presumably Kupffer cells or resident liver macrophages. IFN alpha/beta production by cultured Kupffer cells was not observed if medium containing fetal bovine serum or portal serum was treated with polymyxin B or if Kupffer cells were cultured in serum-free medium. This suggested that small amounts of endotoxin in fetal bovine or portal serum stimulated Kupffer cells to produce IFN alpha/beta. Possibly, Kupffer cells are in a different state of activation/maturation than peritoneal and splenic macrophages since the sensitivity of resident Kupffer cells from C3HeB/FeJ mice to the stimulatory effects of endotoxin. The endogenous production of IFN alpha/beta by Kupffer cells from C3HeB/FeJ mice can augment liver-associated natural killer (NK) activity against YAC-1 cells (4h) and induce liver-associated cytotoxic activity, not restricted by the major histocompatibility complex, against NK resistant P815 mastocytoma cells (18 h).

Fatty acid intake and Kupffer cell function: fish oil alters eicosanoid and monokine production to endotoxin stimulation.

Diets high in n-3 fatty acids appear to have an anti-inflammatory effect, which is thought to be due to decreased macrophage prostaglandin (PG) and thromboxane (Tx) production after incorporation of these fatty acids into cell membrane phospholipids. The effect of n-3 fatty acids incorporation on macrophage monokine release in response to septic stimuli is not well established. Kupffer cells, the fixed macrophages of the liver, were obtained from rats fed diets with fat sources derived from corn oil (CO, control), fish oil (FO, high in n-3 fatty acids), or safflower oil (SO, high in n-6 fatty acids) for 2 or 6 weeks. After exposure to bacterial lipopolysaccharide, Kupffer cells from rats fed FO for 2 or 6 weeks produced less PG and Tx than Kupffer cells from rats fed CO or SO. After 2 weeks of defined diets, interleukin-1 (IL-1) and tumor necrosis factor release were not affected by dietary fat source. In contrast, after 6 weeks of feeding, Kupffer cells from both the FO and the SO groups released less IL-1 and tumor necrosis factor when triggered by lipopolysaccharide than Kupffer's cells from animals fed the control diet that contained CO. These data suggest that altered monokine release from macrophages may contribute to the anti-inflammatory effect of diets high in n-3 fatty acids. Also shown in our results is that prolonged changes in membrane phospholipid content induced by dietary fat source can influence not only PG and Tx production but monokine release as well.

Decreased phagocytic and bactericidal activity of the hepatic reticuloendothelial system during chronic ethanol treatment and its restoration by levamisole.

The effect of chronic ethanol feeding (3 weeks) on the phagocytic and the bactericidal activity of hepatic RES versus viable Escherichia coli was studied using the isolated rat liver perfused with a serum-containing medium. Controls or ethanol-fed animals were used as liver or serum donors. The bactericidal activity of serum from ethanol-fed rats was similar to controls and accounted for the disappearance of nearly one-tenth of the bacterial inoculum from the system. When control livers were perfused with a medium containing serum from ethanol-fed animals, phagocytosis was comparable to controls while intracellular killing was greatly reduced. When livers were isolated from ethanol-fed rats, phagocytosis was significantly depressed and no killing occurred, irrespective of the source of serum. Levamisole was able to restore the macrophage activity depressed by ethanol. Our data indicate that the direct effect of ethanol on hepatic macrophages plays a major role in reducing the bactericidal activity of hepatic RES, although alterations of serum factors may contribute to an ineffective preparation of bacteria for intracellular killing.