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T R Billiar

Publications and source records attributed to T R Billiar.

At least 253 records · Page 14Linked to original sources
PubMed1990

Hepatocyte modulation of Kupffer cell prostaglandin E2 production in vitro.

It is likely that dynamic interactions between hepatocytes and Kupffer cells contribute to the responses of these cell types both under normal conditions and during sepsis. In this study, we examined the influences of hepatocytes on the concentration of the inflammatory mediator PGE2 in Kupffer cell cultures. Evidence to suggest that cultured rat hepatocytes both metabolize PGE2 and produce a substance that promotes LPS-stimulated Kupffer cell PGE2 biosynthesis include the following: 1) PGE2 levels in Kupffer cell: hepatocyte coculture were lower than the levels in Kupffer cell cultures early after LPS stimulation; 2) 36 h after LPS, coculture PGE2 levels exceeded the levels in Kupffer cell cultures despite the demonstrated capacity for hepatocytes to metabolize PGE2; 3) a transferable, non-dialyzable, and heat-unstable factor in hepatocyte supernatant promoted PGE2 production when added to Kupffer cells with LPS or after LPS; 4) there was no increased PGE2 synthesis when the hepatocyte supernatant was added without LPS or if hepatocyte supernatant was preincubated with the Kupffer cells for 6 or 18 h before LPS administration; 5) there was an inability of the hepatocyte factor to promote PGE2 production in response to other macrophage-activating agents, including calcium ionophore A23187 or phorphol myristate acetate; and 6) there was no increased cell replication or protein synthesis in the Kupffer cell cultures following hepatocyte supernatant incubation. The increased Kupffer cell PGE2 production by the hepatocyte supernatant was not due to contamination of the hepatocyte supernatant by endotoxin or PGE2. These in vitro results raise the possibility that hepatocytes have the capacity to modulate local PGE2 levels by two distinct mechanisms. Hepatocytes can metabolize PGE2 as well as release factor(s) which promote LPS-induced PGE2 production by Kupffer cells.

Animals↗
PubMed1989

Hepatocytes produce nitrogen oxides from L-arginine in response to inflammatory products of Kupffer cells.

A metabolic pathway by which L-arginine (L-arg) is converted to the biologically active compound NO. has recently been described in macrophages (M phi) and endothelial cells. This report demonstrates that transferable products from activated Kupffer cells (KC) induce the conversion of large quantities of L-arg to nitrogen oxides within hepatocytes (HC). In M phi and endothelial cells, citrulline and NO2-/NO3- are the stable endproducts of this metabolic pathway. In contrast, HC L-arg metabolism resulted in significantly greater production of NO2-/NO3- than citrulline. The generation of NO. within HC was associated with a concurrent decrease in total protein synthesis.

Animals↗
PubMed1989

An L-arginine-dependent mechanism mediates Kupffer cell inhibition of hepatocyte protein synthesis in vitro.

The hepatic failure associated with severe sepsis is characterized by specific, progressive, and often irreversible defects in hepatocellular metabolism (1). Although the etiologic microbe can often be identified, the direct causes and mechanisms of the hepatocellular dysfunction are poorly understood. We have hypothesized that Kupffer cells (KC), which interact with ambient septic stimuli, respond by providing signals to adjacent hepatocytes (HC) in sepsis . Furthermore, we have provided evidence (2, 3) that KC activated by LPS from Gram-negative bacteria can induce profound changes in the function of neighboring HC in coculture. In our model, coculture of either KC (2) or peritoneal macrophages (Mphi)(3) with HC normally promotes HC protein synthesis ([(3)H]leucine incorporation). The addition of LPS or killed Escherichia colt' to such cocultures induces a profound decrease in HC protein synthesis, as well as qualitative changes ([(35)S]methionine, SDS-gel electrophoresis) in protein synthesis without inducing HC death (2, 3) . In this report we show that the inhibition in protein synthesis is mediated via an L-arginine-dependent mechanism. The metabolism of L-arginine by activated Mphi to substances with cytostatic and even lethal effects on target cells is a relatively recent discovery. After the description by Stuehr and Marletta (4, 5) that LPS- triggered Mphi produced nitrite/nitrate (NO(2)(-)/NO(3)(-)), Hibbs et al. (6, 7) and Iyengar et al. (8) demonstrated that L-arginine was the substrate for the formation of both these nitrogen end products and citrulline. A role for the arginine-dependent mechanism in Mphi tumor cytotoxicity (6, 7) and microbiostatic activity (9) has been suggested. However, the in vivo functions of this novel Mphi mechanism have not yet been defined, but it is possible that there are both physiologic as well as pathologic roles. Our in vitro results raise the possibility that some metabolic responses to microbial invasion maybe partially mediated by the L-arginine-dependent mechanism. What other metabolic responses are affected and the possible pathologic consequences remain to be studied.

Animals↗
PubMed1989

Kupffer cell cytotoxicity to hepatocytes in coculture requires L-arginine.

Activated macrophages convert L-arginine to citrulline and unstable nitrogen oxides that have cytotoxic properties. We recently have shown that the inhibition of protein synthesis in Kupffer cell (KC):hepatocyte (HC) coculture, following exposure to gram-negative bacterial endotoxin (lipopolysaccharide), is due to the metabolism of L-arginine by this cytotoxic pathway. Although this finding supports a role for activated KCs and the L-arginine-dependent mechanism in the HC dysfunction seen in sepsis, it and previous studies have failed to demonstrate direct damage to HCs by adjacent KCs. The current study was undertaken to determine if KCs exposed to lipopolysaccharide could directly damage HCs and, if so, whether the damage was dependent on the metabolism of L-arginine. By using the release of aspartate aminotransferase as a marker of HC damage, it was found that a significant aspartate aminotransferase release by KC:HC cocultures in response to lipopolysaccharide occurred only if L-arginine was present. In addition, requirements for significant aspartate aminotransferase release included KC:HC ratios of 7.5:1 or greater and L-arginine concentrations of 1 mmol or more. Although the KC-induced damage was mild, these results show that in vitro HC damage in KC:HC coculture does require the metabolism of L-arginine and supports a hypothesis that toxic L-arginine metabolites may contribute to liver cell damage in patients with sepsis.

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PubMed1989

Evidence that rat Kupffer cells stimulate and inhibit hepatocyte protein synthesis in vitro by different mechanisms.

Kupffer cell control of hepatocyte protein synthesis may be an important mechanism involved in the regulation of normal liver function and may be one mechanism responsible for the alterations in liver function seen during sepsis. The present series of in vitro experiments compare the response to various inflammatory stimuli of hepatocytes cocultured with Kupffer cells with that of hepatocytes cultured alone. In the absence of inflammatory stimuli, Kupffer cells stimulated hepatocyte protein synthesis. Lipopolysaccharide or gentamicin-killed Escherichia coli triggered Kupffer cell-mediated inhibition of cocultured hepatocyte protein synthesis but had no effect on protein synthesis of hepatocytes cultured alone. Phorbol myristate acetate, muramyl dipeptide, and calcium ionophore had no effect on hepatocytes cultured alone but resulted in a loss of Kupffer cell-mediated stimulation of cocultured hepatocyte protein synthesis without inhibition. Addition of dexamethasone to cocultures prevented the Kupffer cell-mediated inhibition of hepatocyte protein synthesis triggered by lipopolysaccharide, but did not block Kupffer cell-mediated stimulation in the absence of lipopolysaccharide. The data suggest that Kupffer cells can stimulate and inhibit hepatocyte protein synthesis by independent mechanisms. Kupffer cells may be important regulators of hepatocellular function in health and disease.

Acetylmuramyl-Alanyl-Isoglutamine↗
PubMed1989

Modulation of Kupffer cell membrane phospholipid function by n-3 polyunsaturated fatty acids.

Dietary n-3 polyunsaturated fatty acids (PUFAs) have been reported to improve clinical outcome in a number of inflammatory diseases including burns and sepsis. One mechanism contributing to the anti-inflammatory effect is the incorporation of n-3 PUFAs into membrane phospholipids which decreases macrophage eicosanoid production. We hypothesize that an additional mechanism for their effects is an alteration of membrane signal transduction that decreases macrophage responsiveness to inflammatory stimuli. Kupffer cells, the fixed macrophages of the liver, were obtained from rats pair fed diets for 6 weeks with 15% of calories supplied as menhaden (high n-3), corn (control), or safflower (high n-6) oils. The effects of the dietary oils on Kupffer cell membrane signal transduction and eicosanoid production were assessed by measuring inositol phospholipid (PI) metabolism, intracellular calcium responses, and prostaglandin E2 (PGE2) production to the inflammatory signals endotoxin (LPS) and platelet activating factor (PAF). The menhaden oil diet resulted in significant incorporation of n-3 PUFAs into total cellular PUFAs compared to corn and safflower oil. (total n-3 PUFAs, 28.1% menhaden vs 2.1% corn vs 1.2% safflower, P less than 0.03). This incorporation altered signal transduction of PAF as both PI turnover (65% +/- 10% of corn oil) and calcium response (0.6-fold vs 5.0-fold for corn oil) were significantly reduced in the menhaden oil group. (P less than 0.05) The menhaden oil diet also reduced significantly PGE2 production in response to PAF and LPS (corn, 348 +/- 23 pg/ml; menhaden, 48 +/- 6 pg/ml, P less than 0.01). We conclude that, in addition to modulating eicosanoid production, n-3 PUFAs can also alter macrophage membrane signal transduction.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Hepatocytes enhance Kupffer cell-mediated tumor cell cytostasis in vitro.

Kupffer cells (KC) are believed to play a major role in protecting the liver from metastases. In vitro, activated KC mediate both tumor cell cytostasis and cytolysis. Because hepatocytes (HC) occupy a position adjacent to KC in vivo, we investigated the influence of HC on KC tumoricidal activity. Using an in vitro assay of KC-mediated tumor cell cytostasis against murine P815 mastocytoma cells, we found that the presence of HC in the culture profoundly increased KC tumoricidal activity. HC enhanced KC inhibition of P815 proliferation and lowered the concentration of lipopolysaccharide and interferon-gamma necessary to activate the KC to a tumoricidal state. This stimulatory HC effect was dependent on the number of HC present and was transferable in cell-free supernatants, indicating that it was mediated by a soluble secreted product of HC. Furthermore, unlike other macrophage-priming or -potentiating factors, the transferable HC factor(s) was effective only if added simultaneously with lipopolysaccharide or interferon-gamma and not effective if added before these activating agents. These data show that HC produce a soluble mediator that enhances KC tumoricidal activity, suggesting that HC and KC interactions may be critical to the antitumor defense mechanisms of the liver.

Animals↗
PubMed1989

Role of monokines in altering hepatic metabolism in sepsis.

Multiple organ failure continues to be the primary cause of death after trauma and sepsis. This clinical syndrome represents the transition from a hypermetabolic response to injury to a syndrome of progressive organ failures and death. Risk factors include: perfusion deficits, persistent foci of dead or injured tissue, an uncontrolled focus of infection, the presence of the respiratory distress syndrome, persistent hypermetabolism, and preexisting fibrotic liver disease. Once initiated, most treatment modalities for the organ failure syndrome become progressively ineffective including: ventilation, antibiotics, nutrition, and surgery. The best treatment remains prevention and rapid control of risk factors including restoration of oxygen transport and aggressive nutrition support. There seems to be no treatment "magic bullet" either experimentally or clinically once the syndrome has occurred. The metabolic response to injury involves alterations in physiology and in the metabolism of carbohydrate, fat and amino acids. These changes seem to reflect the modulation of the end-organs by the mediator systems activated in response to the stress stimuli. The transition from hypermetabolism to organ failure appears to reflect the clinical appearance of liver failure. It is hypothesized that this liver failure may represent a state of regulatory dysfunction induced in large part by the activated hepatic macrophage, the Kupffer cell. The activation of these macrophages is hypothesized to represent the final stage of a series of stimulating events, eg. hypoxia, endotoxin, bacteria, and gut translocated toxins. The precise monokine(s) responsible are not yet completely characterized, although Interleukin-1 (IL-1) and tumor necrosis factor (TNF) appear to be involved as do prostaglandins (Pg) such as PgE2.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Hepatic dysfunction in multiple systems organ failure as a manifestation of altered cell-cell interaction.

Multiple organ failure continues to be the primary cause of death after trauma and sepsis. This clinical syndrome follows shock and resusitation and the transition from a hypermetabolic response to a syndrome of progressive organ failures and death. Risk factors include: perfusion deficits, persistent foci of dead or injured tissue, an uncontrolled focus of infection, the presence of the respiratory distress syndrome, and preexisting fibrotic liver disease. The imitation of the syndrome represents the clinical manifestation of hepatic failure. It is hypothesized that this hepatic failure results from paracrine amplification with Kupffer cell induced hepatocyte cytotoxicity. The best treatment remains prevention and rapid control of risk factors including restoration of oxygen transport and aggressive nutrition support. There seems to be no treatment "magic bullet" either experimentally or clinically once the syndrome has occurred.

Animals↗
PubMed1989

Evidence that activation of Kupffer cells results in production of L-arginine metabolites that release cell-associated iron and inhibit hepatocyte protein synthesis.

Activated macrophages are known to metabolize L-arginine to unstable intermediates that induce cytotoxic activity through the release of mitochondrial iron in target cells. We have recently shown that rat Kupffer cells (KC) use L-arginine to inhibit cocultured hepatocyte (HC) protein synthesis. Based on these two facts, a hypothesis is proposed that endotoxin-triggered KC release intermediates of L-arginine metabolism that inhibit HC protein synthesis by oxidizing iron at critical mitochondrial enzymes, thus interfering with mitochondrial respiration and energy production. Results of experiments testing this hypothesis showed that the metabolism of L-arginine was required for inhibition of protein synthesis and iron release and that the end products of L-arginine metabolism did not possess cytostatic or cytotoxic activity toward HC. The possibility that this cell culture phenomenon might provide insights into the mechanism of hepatic insufficiency in sepsis is raised.

Animals↗
PubMed1988

Splenectomy alters Kupffer cell response to endotoxin.

The spleen is a potential source for lymphokines, substances known to prime or activate macrophages. Liver macrophages or Kupffer cells are directly exposed to these products via the portal circulation. To determine whether a loss of splenic factors would alter Kupffer cell responses, we studied the effect of splenectomy or sham operation on Kupffer cell responses to endotoxin lipopolysaccharide (LPS). We determined Kupffer cell activation using an in vitro rat hepatocyte-Kupffer cell coculture system in which Kupffer cells normally mediate a significant decrease in hepatocyte protein synthesis if triggered by LPS. We found that Kupffer cells from splenectomized rats were significantly less responsive to LPS three to 60 days after splenectomy. Kupffer cells from sham-operated animals responded normally to LPS. This was contrasted to an increased sensitivity to LPS in the first two days following splenectomy. These data suggest that both splenectomy and a loss of splenic factors alter Kupffer cell responses to septic stimuli. We propose that the decreased sensitivity to LPS after splenectomy may be due to a loss of the priming effects of splenic lymphokines. This decreased responsiveness might contribute to the increased incidence of overwhelming infection after splenectomy.

Animals↗
PubMed1988

Effect of interleukin 2 on Kupffer cell activation. Interleukin 2 primes and activates Kupffer cells to suppress hepatocyte protein synthesis in vitro.

Interleukin 2 (IL-2) is an essential mediator of the immune response and has also been shown to be protective in experimental models of sepsis. Macrophages have IL-2 receptors but their function is unknown. We investigated the effect of IL-2 on Kupffer cells, the fixed macrophages of the liver, using an in vitro rat hepatocyte-Kupffer cell coculture system. In this model, endotoxin (lipopolysaccharide) triggers Kupffer cells to induce suppression of hepatocyte protein synthesis. We found that pretreatment with 10 U/mL or more of IL-2 primed Kupffer cells, significantly reducing the concentration of lipopolysaccharide necessary to trigger Kupffer cell-mediated suppression of hepatocyte protein synthesis. Higher concentrations of IL-2 (greater than or equal to 1 x 10(4) U/mL) alone were capable of priming and triggering Kupffer cells to suppress hepatocyte protein synthesis. These data show that IL-2 increases Kupffer cell sensitivity to endotoxin, suggesting that IL-2 may play an important role in regulating macrophage responses to septic stimuli.

Animals↗
PubMed1988

Endotoxin modulation of hepatocyte secretory and cellular protein synthesis is mediated by Kupffer cells.

We hypothesize that alterations of hepatocyte function in sepsis are modulated by endotoxin (lipopolysaccharide)-triggered Kupffer cells. In the present experiments the effect of lipopolysaccharide on secreted and cellular proteins synthesized by hepatocytes cultured alone or cocultured with Kupffer cells was investigated using polyacrylamide gel electrophoresis. Lipopolysaccharide had no direct effect on the types or amounts of secreted proteins synthesized by hepatocytes alone. Kupffer cells coculturing resulted in increased synthesis of some hepatocyte proteins (68k and 23k) whose production was not altered by lipopolysaccharide. In contrast, addition of lipopolysaccharide to the hepatocyte-Kupffer cell coculture substantially decreased synthesis of several proteins (73k, 66k, and 35k). Despite an overall decrease in protein synthesis of hepatocytes cocultured with Kupffer cells after the addition of lipopolysaccharide, there was increased synthesis of several individual proteins (58k and 44k). Similar effects were seen in synthesis of cellular protein. The addition of recombinant interleukin 1 to hepatocytes alone or in coculture had no effect on the amount or type of protein synthesized. We conclude that Kupffer cells regulate the types and amounts of individual proteins synthesized by hepatocytes via mediators other than interleukin 1.

Animals↗
PubMed1988

The role of intestinal flora on the interactions between nonparenchymal cells and hepatocytes in coculture.

Kupffer cells are exposed directly to a number of factors in the portal circulation that can modify or regulate their responses to septic stimuli. The gut represents a potential source of a number of these factors including endotoxin, lymphokines, and prostaglandins. We examined Kupffer cells from germfree rats and germfree rats exposed to endotoxin or bacteria via their GI tracts to determine the importance of the intestinal flora in maintaining or modulating Kupffer cell responses. Kupffer cells from germfree animals were reduced in numbers and failed to respond to LPS in Kupffer cell: hepatocyte coculture. When germfree rats were exposed to bacterial endotoxin or bacteria via the gastrointestinal tract their Kupffer cells increased in numbers to normal and the cells responded to LPS in culture. Intestinal overgrowth with Escherichia coli for 2 days activated the Kupffer cells and significantly increased Kupffer cell sensitivity to LPS. These data suggest that the environment of the gastrointestinal tract is important for normal Kupffer cell responses and that intestinal bacterial overgrowth can modify Kupffer cell responses to septic stimuli.

Animals↗
PubMed1988

Intestinal gram-negative bacterial overgrowth in vivo augments the in vitro response of Kupffer cells to endotoxin.

A number of disease states and therapeutic maneuvers common to surgical patients can result in changes in the intestinal flora, permitting bacterial overgrowth and translocation of bacteria to gut lymphoid tissue. It is possible that these changes in gut flora increase portal levels of several factors that are capable of altering macrophage activation state, including endotoxin, lymphokines, and eicosanoids. Since Kupffer cells are directly exposed to gut factors via the portal circulation, changes in intestinal flora may influence Kupffer cell responses. Using germfree rats, it has previously been shown that the presence of gut bacterial flora is important in inducing Kupffer cells to respond to endotoxin, and that an overgrowth of gram-negative bacteria can further augment Kupffer cell responses, supporting the above-mentioned hypothesis. The current set of experiments examines how intestinal gram-negative bacterial overgrowth in normal adult rats effects the response of Kupffer cells to septic stimuli. Kupffer cells were obtained from conventional rats with induced intestinal overgrowth with Escherichia coli C25 for 2 or 7 days. After 2 days of overgrowth, Kupffer cells were only slightly less responsive to lipopolysaccharide (LPS) than control Kupffer cells. However, after 7 days of overgrowth, when placed in coculture with normal hepatocytes, Kupffer cells were significantly more responsive to LPS (p less than 0.001), inducing a greater degree of suppression in hepatocyte protein synthesis at lower LPS concentrations. When cultured alone, Kupffer cells from these animals also produced more interleukin-1 (p less than 0.002) and prostaglandin E2 (PGE2) (p less than 0.009) in response to LPS. These results show that intestinal gram-negative bacterial overgrowth in conventional rats can have direct influences on the response of hepatic macrophages to septic stimuli, and provides further support to the hypothesis that imbalances in the intestinal flora can effect the responses of immune cells in other sites of the body.

Animals↗
PubMed1988

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.

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