Kupffer cells are a critical component of the mononuclear phagocytic system and are central to both the hepatic and systemic response to pathogens. Kupffer cells are reemerging as critical mediators of both liver injury and repair. Kupffer cells exhibit a tremendous plasticity; depending on the local metabolic and immune environment, then can express a range of polarized phenotypes, from the proinflammatory M1 phenotype to the alternative/M2 phenotype. Multiple M2 phenotypes can be distinguished, each involved in the resolution of inflammation and wound healing. Here, we have provided an update on recent research that has contributed to the developing delineation of the contribution of Kupffer cells to different types of liver injury, with an emphasis on alcoholic and nonalcoholic liver diseases. These recent advances in our understanding of Kupffer cell function and regulation will likely provide new insights into the potential for therapeutic manipulation of Kupffer cells to promote the resolution of inflammation and enhance wound healing in liver disease.
Because the histological and biochemical progression of liver disease is similar in alcoholic steatohepatitis (ASH) and nonalcoholic steatohepatitis (NASH), we hypothesized that the genetic susceptibility to these liver diseases would be similar. To identify potential candidate genes that regulate the development of liver fibrosis, we studied a chromosome substitution strain (CSS-17) that contains chromosome 17 from the A/J inbred strain substituted for the corresponding chromosome on the C57BL/6J (B6) genetic background. Previously, we identified quantitative trait loci (QTLs) in CSS-17, namely obesity-resistant QTL 13 and QTL 15 (Obrq13 and Obrq15, respectively), that were associated with protection from diet-induced obesity and hepatic steatosis on a high-fat diet.
The effect of moderate alcohol consumption on liver fibrosis is not well understood, but evidence suggests that adenosine may play a role in mediating the effects of moderate ethanol on tissue injury. Ethanol increases the concentration of adenosine in the liver. Adenosine 2A receptor (A2AR) activation is known to enhance hepatic stellate cell (HSC) activation and A2AR deficient mice are protected from fibrosis in mice. Making use of a novel mouse model of moderate ethanol consumption in which female C57BL/6J mice were allowed continued access to 2% (vol/vol) ethanol (11% calories) or pair-fed control diets for 2 days, 2 weeks or 5 weeks and superimposed with exposure to CCl4, we tested the hypothesis that moderate ethanol consumption increases fibrosis in response to carbon tetrachloride (CCl4) and that treatment of mice with an A2AR antagonist prevents and/or reverses this ethanol-induced increase in liver fibrosis. Neither the expression or activity of CYP2E1, required for bio-activation of CCl4, nor AST and ALT activity in the plasma were affected by ethanol, indicating that moderate ethanol did not increase the direct hepatotoxicity of CCl4. However, ethanol feeding enhanced HSC activation and exacerbated liver fibrosis upon exposure to CCl4. This was associated with an increased sinusoidal angiogenic response in the liver. Treatment with A2AR antagonist both prevented and reversed the ability of ethanol to exacerbate liver fibrosis.
Cell-cycle induction in hepatocytes protects from prolonged tissue damage after toxic liver injury. Early growth response (Egr)-1(-/-) mice exhibit increased liver injury after carbon tetrachloride (CCl(4)) exposure and reduced TNF-? production. Because TNF-? is required for prompt cell-cycle induction after liver injury, here, we tested the hypothesis that Egr-1 is required for timely hepatocyte entry into the cell cycle after CCl(4)-induced liver injury. Acute liver injury was induced by a single injection of CCl(4). Assays were employed to assess indices of the cell cycle in liver after CCl(4) exposure. Bromodeoxyuridine incorporation peaked in wild-type mice at 48 h after CCl(4) but was reduced by 80% in Egr-1(-/-) mice. Proliferating-cell nuclear-antigen immunohistochemistry revealed blocks in cell-cycle entry and progression to DNA synthesis in Egr-1-deficient mice 48 h after CCl(4). Cyclin D, important for G0/G1 progression, was reduced at baseline and 36 h after CCl(4). Cyclin E1, required for G1/S-phase transition, was reduced in Egr-1(-/-) mice 24 and 48 h after CCl(4) exposure and was associated with reduced phosphorylation of the retinoblastoma protein. Proliferation in Egr-1(-/-) mice was delayed, rather than blocked, because indices of cell-cycle progression were restored 72 h after CCl(4) exposure. We concluded that Egr-1 was required for prompt cell-cycle entry (G0- to G1-phase) and G1/S-phase transition after toxic liver injury. These data support the hypothesis that Egr-1 provides hepatoprotection in the CCl(4)-injured liver, attributable, in part, to timely cell-cycle induction and progression.
Obesity is a global epidemic with more than 1 billion overweight adults and at least 300 million obese patients worldwide. Diabetes is characterized by a defect in insulin secretion or a decrease in sensitivity to insulin, which results in elevated fasting blood glucose. Both obesity and elevated fasting glucose are risk factors for nonalcoholic fatty liver disease, a disease spectrum that includes hepatic steatosis (nonalcoholic fatty liver), nonalcoholic steatohepatitis (NASH), fibrosis, and cirrhosis. Increased adiposity and insulin resistance contribute to the progression from NASH to fibrosis through the development of a profibrotic mileau in the liver, including increased hepatocellular death, increased reactive oxygen species generation, and an altered adipokine/cytokine balance. This review will summarize recent advances in our understanding of the pathological interactions among excessive fat accumulation, insulin resistance, and hepatic fibrogenesis and discuss specific molecular pathways that may be of interest in the development of therapeutic interventions to prevent and/or reverse hepatic fibrosis.
Improvements in treatment and management for pediatric central nervous system (CNS) tumors have increased survival rates, allowing clinicians to focus on long-term sequelae, including sleep disorders. The objective of this study was to describe a series of CNS tumor survivors who had sleep evaluations that included polysomnography (PSG) with attention to sleep disorder in relation to the tumor site.
In a previous study, we conducted telephone interviews with parents 6 to 10 months after their childs death from cancer, using open-ended questions to identify the type and frequency of cancer-related symptoms that most concerned them during the last week of their childs life. Because the parents identified many clinically striking symptoms (n=109) that were not of most concern to them, we conducted a secondary analysis of these interviews (48 mothers and four fathers of 52 patients) to identify descriptive factors associated with the parents level of concern. Six descriptive factors were associated with symptoms of most concern and 10 factors with symptoms not of most concern. Ten of these 16 factors occurred in both categories, indicating that clinicians should directly query parents to identify the symptoms that concern parents the most. Six factors differed between the two categories, and only one (the continuous distress caused by a symptom that is unrelieved) was unique to the category of symptoms of most concern. Five factors (symptom present for at least one week, symptom not seen as remarkable by the parent or causing no distress to the child, symptom well managed, symptom improved, and symptoms for which the parent felt adequately prepared) were unique to the category of symptoms not of most concern. By inquiring about symptoms of most concern and factors that influence parental concern, clinicians may be better able to direct care efforts to reduce patients and parents distress and support parents during the difficult end-of-life period.
Inflammatory gene expression plays a pathological role in acute and chronic hepatic inflammation, yet, inflammation also promotes liver repair by inducing protective mechanisms to limit collateral tissue damage by priming hepatocytes for proliferation. Early growth response (Egr)-1, a transcription factor that regulates inflammatory gene expression, plays a pathological role in many animal models of acute and chronic inflammatory disease. Here, we tested the hypothesis that Egr-1 is beneficial after toxic liver injury.
The transcription factor early growth response (Egr)-1 regulates the expression of genes required for execution of the wound healing response. Multiple cycles of injury, coupled to incomplete wound healing, lead to fibrosis. Therefore, we hypothesized that Egr-1 is required for the development of hepatic fibrosis. To test this hypothesis, we exposed wild-type and egr-1(-/-) mice to acute or chronic carbon tetrachloride (CCl(4)). Acute CCl(4) exposure established a profibrotic milieu in the liver, including activation of hepatic stellate cells as well as expression of type 1 collagen genes and tissue inhibitor of matrix metalloproteinase 1 in both wild-type and egr-1(-/-) mice. This response was exacerbated in egr-1(-/-) mice. After chronic CCl(4) exposure, hepatic fibrosis was established in both genotypes; however, the fibrotic response was profoundly worsened in Egr-1-deficient mice. Importantly, enhanced fibrosis in egr-1(-/-) mice was accompanied by a robust activation of the oval cell response, suggesting more severe liver injury and/or reduced hepatocyte proliferation when compared with wild-type mice. Hepatic expression of genes indicative of oval cell activation, as well as the number of cells expressing A6, a mouse oval cell marker, was greater in egr-1(-/-) mice. Taken together, these data reveal novel roles for Egr-1 as a negative regulator of both CCl(4)-induced hepatic fibrosis and the oval cell response.
The development of alcoholic liver disease (ALD) is a complex process involving both the parenchymal and non-parenchymal cells in the liver. Enhanced inflammation in the liver during ethanol exposure is an important contributor to injury. Kupffer cells, the resident macrophages in liver, are particularly critical to the onset of ethanol-induced liver injury. Chronic ethanol exposure sensitizes Kupffer cells to activation by lipopolysaccharide via Toll-like receptor 4. This sensitization enhances production of inflammatory mediators, such as tumor necrosis factor-alpha and reactive oxygen species, that contribute to hepatocyte dysfunction, necrosis, apoptosis, and fibrosis. Impaired resolution of the inflammatory process probably also contributes to ALD. The resolution of inflammation is an active, highly coordinated response that can potentially be manipulated via therapeutic interventions to treat chronic inflammatory diseases. Recent studies have identified an adiponectin/interleukin-10/heme oxygenase-1 (HO-1) pathway that is profoundly effective in dampening the enhanced activation of innate immune responses in primary cultures of Kupffer cells, as well as in an in vivo mouse model of chronic ethanol feeding. Importantly, induction of HO-1 also reduces ethanol-induced hepatocellular apoptosis in this in vivo model. Based on these data, we hypothesize that the development of therapeutic agents to regulate HO-1 and its downstream targets could be useful in enhancing the resolution of inflammation during ALD and preventing progression of early stages of liver injury.
Plasminogen activator inhibitor-1 (PAI-1) is an acute phase protein that has been shown to play a role in experimental fibrosis caused by bile duct ligation (BDL) in mice. However, its role in more severe models of hepatic fibrosis (e.g., carbon tetrachloride; CCl(4)) has not been determined and is important for extrapolation to human disease. Wild-type or PAI-1 knockout mice were administered CCl(4) (1 ml/kg body wt ip) 2x/wk for 4 wk. Plasma (e.g., transaminase activity) and histological (e.g., Sirius red staining) indexes of liver damage and fibrosis were evaluated. Proliferation and apoptosis were assessed by PCNA and TdT-mediated dUTP nick-end labeling (TUNEL) staining, respectively, as well as by indexes of cell cycle (e.g., p53, cyclin D1). In contrast to previous studies with BDL, hepatic fibrosis was enhanced in PAI-1(-/-) mice after chronic CCl(4) administration. Indeed, all indexes of liver damage were elevated in PAI-1(-/-) mice compared with wild-type mice. This enhanced liver damage correlated with impaired hepatocyte proliferation. A similar effect on proliferation was observed after one bolus dose of CCl(4), without concomitant increases in liver damage. Under these conditions, a decrease in phospho-p38, coupled with elevated p53 protein, was observed; these results suggest impaired proliferation and a potential G(1)/S cell cycle arrest in PAI-1(-/-) mice. These data suggest that PAI-1 may play multiple roles in chronic liver diseases, both protective and damaging, the latter mediated by its influence on inflammation and fibrosis and the former via helping maintain hepatocyte division after an injury.
Ethanol metabolism promotes the formation of a variety of reactive aldehydes in the liver. These aldehydes can rapidly form covalent protein adducts. Accumulating evidence indicates that these protein adducts may contribute to ethanol-mediated liver injury. Overproduction of gamma-ketoaldehydes, levuglandins (LGs) and isolevuglandins, is implicated in the pathogenesis of several chronic inflammatory diseases. gamma-Ketoaldehydes can form protein adducts orders of magnitude more quickly than 4-hydroxynonenal (4-HNE) or malondialdehyde. We hypothesized that ethanol-induced oxidative stress in vivo results in overproduction of LGE(2)- and isoLGE(2)-protein adducts in mouse liver. Female C57BL/6 mice were allowed free access to an ethanol-containing diet for up to 39 days or pair-fed control diets. Pathological markers of ethanol-induced hepatic injury including serum alanine aminotransferase, hepatic triglyceride, and CYP2E1 were elevated in response to ethanol feeding. Ethanol-induced formation of isoLGE(2)-, LGE(2)-, and 4-HNE-protein adducts in mouse liver was dependent on both dose and duration of ethanol feeding. Deficiency of cyclooxygenase 1 or 2 did not prevent ethanol-induced isoLGE(2) or LGE(2) adducts in the liver, but adduct formation was reduced in both TNFR1- and CYP2E1-deficient mice. In summary, ethanol feeding enhanced gamma-ketoaldehyde-protein adduct production via a TNFR1/CYP2E1-dependent, but cyclooxygenase-independent, mechanism in mouse liver.
Parents of terminally ill children with cancer frequently ask clinicians when their child will die. Such information helps parents prepare for the childs death. To identify how parents perceived when their childs cancer-related death would occur, we conducted a secondary analysis of telephone interviews with 49 bereaved parents 6-10 months after their childs death to extract their descriptions of this occurrence. The parents knew in advance that their child was going to die, but they described when their childs death would occur in three different ways: anticipated (parents observed changes that alerted them that death was imminent; n=22, 52.4%), surprising (parents were surprised that their child died on that particular day; n=13, 31.0%), and overdue (parents had been waiting for the end of their childs apparent suffering; n=7, 16.7%). These categories did not differ by patients diagnosis, sex, or location of death but differed slightly by symptom patterns. Parents who reported the occurrence of their childs death as surprising reported fewer symptom changes on the last day of their childs life, compared with the last week of life, than did the parents in the other two categories. These findings indicate that parents of children with terminal cancer can perceive when their childs death would occur very differently: Some are surprised, whereas others feel they have waited too long for their childs release from suffering. Clinicians can use these descriptions and the associated symptom patterns to help families prepare for their childs last week and last day.
The innate immune system has been implicated in the pathogenesis of alcoholic liver disease. Although innate immunity is usually considered an early response to injury, previous work implicating innate immunity in ethanol-induced liver injury focuses primarily on long-term ethanol exposure. We investigated the early period of ethanol exposure to determine whether there were temporal associations between activation of innate immune responses and known correlates of liver injury. Female C57BL/6 mice were allowed free access to an ethanol-containing Lieber-DeCarli diet or were pair-fed a control diet. Within 4 days of ethanol exposure, we observed a striking spike in expression of hepatic proinflammatory cytokines-including tumor necrosis factor alpha (TNF-alpha), interleukin-6, and interferon-gamma-prior to hepatic triglyceride accumulation or increased plasma alanine aminotransferase activities, as well as before the induction of cytochrome P450 2E1 or oxidative stress. This early spike in inflammatory cytokines coincided with deposition of C3b-iC3b/C3c (C3b) in the liver. This deposition, resulting from the cleavage of the third component of the complement system (C3), is evidence for activation of complement in response to ethanol. C3(-/-) mice were protected from the early, ethanol-induced increase in hepatic TNF-alpha expression. Ethanol increased C3b deposition in mice deficient in C3a receptor or C5a receptor, as well as in wild-type mice depleted of hepatic macrophages; however, there was no increase in hepatic TNF-alpha in the absence of C3a receptor, C5a receptor, or hepatic macrophages. In contrast, the absence of Toll-like receptor 4 (TLR-4) had no effect on the early, ethanol-induced increase in either C3b or TNF-alpha.
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