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Hepatic insulin resistance is recognized as a driver of type 2 diabetes and fatty liver disease but specific therapies are lacking. Here we explore the potential of human induced pluripotent stem cells (iPSCs) for modeling hepatic insulin resistance in vitro, with a focus on resolving the controversy about the impact of inflammation in the absence of steatosis. For this, we establish the complex insulin signaling cascade and the multiple inter-dependent functions constituting hepatic glucose metabolism in iPSC-derived hepatocytes (iPSC-Heps). Co-culture of these insulin-sensitive iPSC-Heps with isogenic iPSC-derived pro-inflammatory macrophages induces glucose output by preventing insulin from inhibiting gluconeogenesis and glycogenolysis and activating glycolysis. Screening identifies TNFα and IL1β as the mediators of insulin resistance in iPSC-Heps. Neutralizing these cytokines together restores insulin sensitivity in iPSC-Heps more effectively than individual inhibition, reflecting specific effects on insulin signaling and glucose metabolism mediated by NF-κB or JNK. These results show that inflammation is sufficient to induce hepatic insulin resistance and establish a human iPSC-based in vitro model to mechanistically dissect and therapeutically target this metabolic disease driver.

Background and aimsAccumulation of visceral adipose tissue is associated with hepatic inflammation and fibrosis, suggestive of its metabolic and inflammatory properties. We aimed to examine the histologic findings of visceral and subcutaneous adipose tissue and to associate these findings with clinical and radiologic characteristics in patients with cirrhosis.MethodsIncluded were 55 adults with cirrhosis who underwent liver transplantation from 3/2017-12/2018 and had an abdominal computed tomography (CT) scan within 6 months prior to transplant. Visceral-to-subcutaneous adipose tissue ratio (VSR) was calculated using visceral (VATI) and subcutaneous adipose tissue index (SATI) quantified by CT at the L3-vertebral level and normalized for height (cm2/m2). VAT (greater omentum), SAT (abdominal wall), and skeletal muscle (rectus abdominis) biopsies were collected at transplant.ResultsMajority of patients had VAT inflammation (71%); only one patient (2%) had SAT inflammation. Patients with VAT inflammation had similar median VATI (42 vs 41 cm2/m2), lower median SATI (64 vs 97 cm2/m2), and higher median VSR (0.63 vs 0.37, p = 0.002) than patients without inflammation. In univariable logistic regression, VSR was associated with VAT inflammation (OR 1.47, 95%CI 1.11-1.96); this association remained significant even after adjusting for age, sex, BMI, HCC, or MELD-Na on bivariable analyses.ConclusionIn patients with cirrhosis undergoing liver transplantation, histologic VAT inflammation was common, but SAT inflammation was not. Increased VSR was independently associated with VAT inflammation. Given the emerging data demonstrating the prognostic value of VSR, our findings support the value of CT-quantified VSR as a prognostic marker for adverse outcomes in the liver transplant setting.

STING, Tmem173, is involved in liver injury caused by both infectious and sterile inflammatory models. Its role in toxic liver injury and non-alcoholic fatty liver disease (NAFLD), however, is less clear. While a few groups have investigated its role in NAFLD pathogenesis, results have been conflicting. The objective of this study was to clarify the exact role of STING in toxic liver injury and NAFLD models. Goldenticket mice (Tmem173gt), which lack STING protein, were subjected to either a toxic liver injury with tunicamycin (TM) or one of two dietary models of non-alcoholic fatty liver disease: high fructose feeding or Fructose-Palmitate-Cholesterol (FPC) feeding. Three days after TM injection, Tmem173gt mice demonstrated less liver injury (average ALT of 54 ± 5 IU/L) than control mice (average ALT 108 ± 24 IU/L). In contrast, no significant differences in liver injury were seen between WT and Tmem173gt mice fed either high fructose or FPC. Tmem173gt mice only distinguished themselves from WT mice in their increased insulin resistance. In conclusion, while STING appears to play a role in toxic liver injury mediated by TM, it plays little to no role in two dietary models of NAFLD. The exact role of STING appears to be stimulus-dependent.

Fibroblast growth factor 23 (FGF23) is a bone marrow cell produced hormone that functions in the intestine and kidney to regulate phosphate homeostasis. Increased serum FGF23 is a well-established predictor of mortality in renal disease, but recent findings linking increased levels to hepatic and cardiac diseases have suggested that other organs are sources of FGF23 or targets of its effects. The potential ability of the liver to produce FGF23 in response to hepatocellular injury was therefore examined. Very low levels of Fgf23 mRNA and FGF23 protein were detected in normal mouse liver, but the amounts increased markedly during acute liver injury from the hepatotoxin carbon tetrachloride. Serum levels of intact FGF23 were elevated during liver injury from carbon tetrachloride. Chronic liver injury induced by a high fat diet or elevated bile acids also increased hepatic FGF23 levels. Stimulation of toll-like receptor (TLR) 4-driven inflammation by gut-derived lipopolysaccharide (LPS) underlies many forms of liver injury, and LPS induced Fgf23 in the liver as well as in other organs. The LPS-inducible cytokines IL-1β and TNF increased hepatic Fgf23 expression as did a TLR2 agonist Pam2CSK3. Analysis of Fgf23 expression and FGF23 secretion in different hepatic cell types involved in liver injury identified the resident liver macrophage or Kupffer cell as a source of hepatic FGF23. LPS and cytokines selectively induced the hormone in these cells but not in hepatocytes or hepatic stellate cells. FGF23 failed to exert any autocrine effect on the inflammatory state of Kupffer cells but did trigger proinflammatory activation of hepatocytes. During liver injury inflammatory factors induce Kupffer cell production of FGF23 that may have a paracrine proinflammatory effect on hepatocytes. Liver-produced FGF23 may have systemic hormonal effects as well that influence diseases in in other organs.

Background and aimsInduced pluripotent stem cells (iPSCs) provide an important tool for the generation of patient-derived cells, including hepatocyte-like cells, by developmental cues through an endoderm intermediate. However, most iPSC lines fail to differentiate into endoderm, with induction resulting in apoptosis.Approach and resultsTo address this issue, we built upon published methods to develop an improved protocol. We discovered that doxycycline dramatically enhances the efficiency of iPSCs to endoderm differentiation by inhibiting apoptosis and promoting proliferation through the protein kinase B pathway. We tested this protocol in >70 iPSC lines, 90% of which consistently formed complete sheets of endoderm. Endoderm generated by our method achieves similar transcriptomic profiles, expression of endoderm protein markers, and the ability to be further differentiated to downstream lineages.ConclusionsFurthermore, this method achieves a 4-fold increase in endoderm cell number and will accelerate studies of human diseases in vitro and facilitate the expansion of iPSC-derived cells for transplantation studies.

X-box binding protein-1 (XBP1) is a transcription factor that plays a central role in controlling cellular responses to endoplasmic reticulum (ER) stress. Under stress conditions, the transcriptionally active form of XBP1 is generated via splicing of Xbp1 mRNA by the ER-resident protein inositol-requiring enzyme-1 (IRE1α). Genetic deletion of XBP1 has multiple consequences: some resulting from the loss of the transcription factor per se, and others related to compensatory activation of IRE1α. The objective of the current study was to investigate the effects of XBP1 deletion in adult mouse liver and determine to what extent they are direct or indirect. XBP1 was deleted from hepatocytes in adult Xbp1fl/fl mice using AAV8-Transthyretin-Cre (Xbp1Δhep). Xbp1Δhep mice exhibited no liver disease at baseline, but developed acute biochemical and histologic liver injury in response to a dietary challenge with fructose for 4 weeks. Fructose-mediated liver injury in Xbp1Δhep mice coincided with heightened IRE1α activity, as demonstrated by Xbp1 mRNA splicing, JNK activation, and regulated IRE1α-dependent RNA decay (RIDD). Activation of eIF2α was also evident, with associated up-regulation of the pro-apoptotic molecules CHOP, BIM, and PUMA. To determine whether the adverse consequences of liver-specific XBP1 deletion were due to XBP1 loss or heightened IRE1α activity, we repeated a fructose challenge in mice with liver-specific deletion of both XBP1 and IRE1α (Xbp1Δhep;Ire1aΔhep). Xbp1Δhep;Ire1aΔhep mice were protected from fructose-mediated liver injury and failed to exhibit any of the signs of ER stress seen in mice lacking XBP1 alone. The protective effect of IRE1α deletion persisted even with long-term exposure to fructose. Xbp1Δhep mice developed liver fibrosis at 16 weeks, but Xbp1Δhep;Ire1aΔhep mice did not. Overall, the results indicate that the deleterious effects of hepatocyte-specific XBP1 deletion are due primarily to hyperactivation of IRE1α. They support further exploration of IRE1α as a contributor to acute and chronic liver diseases.

Perinatal hepatic inflammation can have devastating consequences. Monocytes play an important role in the initiation and resolution of inflammation, and their diverse functions can be attributed to specific cellular subsets: pro-inflammatory or classical monocytes (Ly6cHi) and pro-reparative or non-classical monocytes (Ly6cLo). We hypothesized that inherent differences in Ly6cHi classical monocytes and Ly6cLo non-classical monocytes determine susceptibility to perinatal hepatic inflammation in late gestation fetuses and neonates. We found an anti-inflammatory transcriptional profile expressed by Ly6cLo non-classical monocytes, and a physiologic abundance of these cells in the late gestation fetal liver. Unlike neonatal pups, late gestation fetuses proved to be resistant to rhesus rotavirus (RRV) mediated liver inflammation. Furthermore, neonatal pups were rendered resistant to RRV-mediated liver injury when Ly6cLo non-classical monocytes were expanded. Pharmacologic inhibition of Ly6cLo non-classical monocytes in this setting restored susceptibility to RRV-mediated disease. These data demonstrate that Ly6cLo monocytes promote resolution of perinatal liver inflammation in the late gestation fetus, where there is a physiologic expansion of non-classical monocytes, and in the neonatal liver upon experimental expansion of these cells. Therapeutic strategies directed towards enhancing Ly6cLo non-classical monocyte function may mitigate the detrimental effects of perinatal liver inflammation.

Macronutrient metabolism is a highly orchestrated process, with adipose tissue and liver each playing central roles in nutrient uptake, processing, transport, and storage. These 2 tissues form an important metabolic circuit, particularly as it relates to lipids as the primary storage form of excess energy. The function of the circuit is influenced by many factors, including the quantity and type of nutrients consumed and their impact on the overall health of the tissues. In this review we begin with a brief summary of the homeostatic disposition of lipids between adipose tissue and liver and how these processes can become dysregulated in obesity. We then explore how specific dietary nutrients and nutrient combinations can exert unique influences on the liver-adipose tissue axis.

Although autoimmune hepatitis (AIH) is more common in women and affects people of all races/ethnicities, there is currently limited information regarding the relationship between race/ethnicity and AIH, especially in the context of underserved populations. We aim to evaluate the relationship between race/ethnicity and AIH and better characterize its clinical features among different racial groups. We conducted a 15-year retrospective analysis, from January 2002 to June 2017, of patients seen at Zuckerberg San Francisco General Hospital (ZSFG). Sixty-three AIH patients and 2049 non-AIH controls were eligible for the study. The main predictor of interest was race/ethnicity, and the main outcome of interest was AIH diagnosis; other secondary measures recorded include clinical features such as ALT, bilirubin, and biopsy fibrosis at presentation. In a multivariable model adjusting for age and sex, we found that black (OR 9.6, 95% CI 1.8-178), Latino (OR 25.0, 95% CI 5.3-448), and Asian/Pacific Islander (API) (OR 10.8, 95% CI 2.2-196) race/ethnicity were associated with increased odds of an AIH diagnosis compared to the white reference group. Among people of colour with AIH, there were no significant differences in baseline ALT (p = .45), total bilirubin at presentation (p = .06), fibrosis at presentation (p = .74), and hospitalization (p = .27). Race/ethnicity is an independent risk factor for AIH. The clinical features of AIH did not differ significantly among black, Latino, and API patients.

Background & aimsThe factors that distinguish metabolically healthy obesity from metabolically unhealthy obesity are not well understood. Diet has been implicated as a determinant of the unhealthy obesity phenotype, but which aspects of the diet induce dysmetabolism are unknown. The goal of this study was to investigate whether specific macronutrients or macronutrient combinations provoke dysmetabolism in the context of isocaloric, high-energy diets.MethodsMice were fed 4 high-energy diets identical in calorie and nutrient content but different in nutrient composition for 3 weeks to 6 months. The test diets contained 42% carbohydrate (sucrose or starch) and 42% fat (oleate or palmitate). Weight and glucose tolerance were monitored; blood and tissues were collected for histology, gene expression, and immunophenotyping.ResultsMice gained weight on all 4 test diets but differed significantly in other metabolic outcomes. Animals fed the starch-oleate diet developed more severe hepatic steatosis than those on other formulas. Stable isotope incorporation showed that the excess hepatic steatosis in starch-oleate-fed mice derived from exaggerated adipose tissue lipolysis. In these mice, adipose tissue lipolysis coincided with adipocyte necrosis and inflammation. Notably, the liver and adipose tissue abnormalities provoked by starch-oleate feeding were reproduced when mice were fed a mixed-nutrient Western diet with 42% carbohydrate and 42% fat.ConclusionsThe macronutrient composition of the diet exerts a significant influence on metabolic outcome, independent of calories and nutrient proportions. Starch-oleate appears to cause hepatic steatosis by inducing progressive adipose tissue injury. Starch-oleate phenocopies the effect of a Western diet; consequently, it may provide clues to the mechanism whereby specific nutrients cause metabolically unhealthy obesity.

Diets containing excess carbohydrate and fat promote hepatic steatosis and steatohepatitis in mice. Little is known, however, about the impact of specific carbohydrate/fat combinations on liver outcome. This study was designed to determine whether high-energy diets with identical caloric density but different carbohydrate and fat composition have unique effects on the liver. Four experimental diets were formulated with 60%kcal carbohydrate and 20%kcal fat, each in nearly pure form from a single source: starch-oleate, starch-palmitate, sucrose-oleate and sucrose-palmitate. The diets were fed to mice for 3 or 12 weeks for analysis of lipid metabolism and liver injury. All mice developed hepatic steatosis over 12 weeks, but mice fed the sucrose-palmitate diet accumulated more hepatic lipid than those in the other three experimental groups. The exaggerated lipid accumulation in sucrose-palmitate-fed mice was attributable to a disproportionate rise in hepatic de novo lipogenesis. These mice accrued more hepatic palmitate and exhibited more evidence of liver injury than any of the other experimental groups. Interestingly, lipogenic gene expression in mice fed the custom diets did not correlate with actual de novo lipogenesis. In addition, de novo lipogenesis rose in all mice between 3 and 12 weeks, without feedback inhibition from hepatic steatosis. The pairing of simple sugar (sucrose) and saturated fat (palmitate) in a high-carbohydrate/moderate-fat diet induces more de novo lipogenesis and liver injury than other carbohydrate/fat combinations. Diet-induced liver injury correlates positively with hepatic de novo lipogenesis and is not predictable by isolated analysis of lipogenic gene expression.

Nonalcoholic fatty liver disease (NAFLD) is a disorder characterized by excess accumulation of fat in hepatocytes (nonalcoholic fatty liver (NAFL)); in up to 40% of individuals, there are additional findings of portal and lobular inflammation and hepatocyte injury (which characterize nonalcoholic steatohepatitis (NASH)). A subset of patients will develop progressive fibrosis, which can progress to cirrhosis. Hepatocellular carcinoma and cardiovascular complications are life-threatening co-morbidities of both NAFL and NASH. NAFLD is closely associated with insulin resistance; obesity and metabolic syndrome are common underlying factors. As a consequence, the prevalence of NAFLD is estimated to be 10-40% in adults worldwide, and it is the most common liver disease in children and adolescents in developed countries. Mechanistic insights into fat accumulation, subsequent hepatocyte injury, the role of the immune system and fibrosis as well as the role of the gut microbiota are unfolding. Furthermore, genetic and epigenetic factors might explain the considerable interindividual variation in disease phenotype, severity and progression. To date, no effective medical interventions exist that completely reverse the disease other than lifestyle changes, dietary alterations and, possibly, bariatric surgery. However, several strategies that target pathophysiological processes such as an oversupply of fatty acids to the liver, cell injury and inflammation are currently under investigation. Diagnosis of NAFLD can be established by imaging, but detection of the lesions of NASH still depend on the gold-standard but invasive liver biopsy. Several non-invasive strategies are being evaluated to replace or complement biopsies, especially for follow-up monitoring.

UnlabelledIdentification of microRNAs (miRNAs) that regulate lipid metabolism is important to advance the understanding and treatment of some of the most common human diseases. In the liver, a few key miRNAs have been reported that regulate lipid metabolism, but since many genes contribute to hepatic lipid metabolism, we hypothesized that other such miRNAs exist. To identify genes repressed by miRNAs in mature hepatocytes in vivo, we injected adult mice carrying floxed Dicer1 alleles with an adenoassociated viral vector expressing Cre recombinase specifically in hepatocytes. By inactivating Dicer in adult quiescent hepatocytes we avoided the hepatocyte injury and regeneration observed in previous mouse models of global miRNA deficiency in hepatocytes. Next, we combined gene and miRNA expression profiling to identify candidate gene/miRNA interactions involved in hepatic lipid metabolism and validated their function in vivo using antisense oligonucleotides. A candidate gene that emerged from our screen was lipoprotein lipase (Lpl), which encodes an enzyme that facilitates cellular uptake of lipids from the circulation. Unlike in energy-dependent cells like myocytes, LPL is normally repressed in adult hepatocytes. We identified miR-29a as the miRNA responsible for repressing LPL in hepatocytes, and found that decreasing hepatic miR-29a levels causes lipids to accumulate in mouse livers.ConclusionOur screen suggests several new miRNAs are regulators of hepatic lipid metabolism. We show that one of these, miR-29a, contributes to physiological lipid distribution away from the liver and protects hepatocytes from steatosis. Our results, together with miR-29a's known antifibrotic effect, suggest miR-29a is a therapeutic target in fatty liver disease.

Non-alcoholic steatohepatitis (NASH) is a disorder marked by alterations in hepatic lipid homeostasis as well as liver injury in the form of cell death, inflammation and fibrosis. Research into the pathophysiology of NASH is dynamic. New concepts from the fields of cell biology, microbiology, immunology and genetics are being tested for their applicability to NASH; discoveries in each of these areas are enriching our understanding of this complex disease. This review summarizes how recent developments from the bench and bedside are merging with more traditional concepts to reshape our view of NASH pathogenesis. Highlights include human studies that emphasize the role of de novo lipogenesis in NASH and experimental work uncovering a role for the inflammasome in NASH. Genetic predispositions to NASH are being clarified, and intestinal microbiome is emerging as a determinant of fatty liver. These unique ideas are now taking their place within an integrated picture of NASH pathogenesis.

Myofibroblasts are the major source of extracellular matrix components that accumulate during tissue fibrosis, and hepatic stellate cells (HSCs) are believed to be the major source of myofibroblasts in the liver. To date, robust systems to genetically manipulate these cells have not been developed. We report that Cre under control of the promoter of Pdgfrb (Pdgfrb-Cre) inactivates loxP-flanked genes in mouse HSCs with high efficiency. We used this system to delete the gene encoding α(v) integrin subunit because various α(v)-containing integrins have been suggested as central mediators of fibrosis in multiple organs. Such depletion protected mice from carbon tetrachloride-induced hepatic fibrosis, whereas global loss of β₃, β₅ or β₆ integrins or conditional loss of β₈ integrins in HSCs did not. We also found that Pdgfrb-Cre effectively targeted myofibroblasts in multiple organs, and depletion of the α(v) integrin subunit using this system was protective in other models of organ fibrosis, including pulmonary and renal fibrosis. Pharmacological blockade of α(v)-containing integrins by a small molecule (CWHM 12) attenuated both liver and lung fibrosis, including in a therapeutic manner. These data identify a core pathway that regulates fibrosis and suggest that pharmacological targeting of all α(v) integrins may have clinical utility in the treatment of patients with a broad range of fibrotic diseases.

Background and aimsStudies investigating insulin resistance (IR) in chronic hepatitis C virus (HCV) infection have used surrogate measures of IR that have limited reliability. We aimed to describe the distribution and risk factors associated with IR and its change over time in HCV using direct measurement.MethodsOne hundred two non-cirrhotic, non-diabetic, HCV-infected subjects underwent clinical, histologic, and metabolic evaluation, and 27 completed repeat evaluation at 6 months. Insulin-mediated glucose uptake was measured by steady-state plasma glucose (SSPG) concentration during the insulin suppression test.ResultsThree subjects with diabetes were excluded and 95 completed all testing. SSPG ranged from 39 to 328 mg/dL (mean 135 mg/dL) and was stable over time (mean SSPG change -0.3 mg/dL). SSPG was associated with Latino ethnicity (Coef 67, 95 % CI 37-96), BMI (Coef 19 per 5 kg/m(2), 95 % CI 5-32), ferritin (Coef 1.4 per 10 ng/ml, 95 % CI 0.2-2.5), male gender (Coef -48, 95 % CI -80 to -16), and HDL (Coef -16, 95 % CI -28 to -5 mg/dL). Current tobacco use (Coef 55, 95 % CI 19-90), steatosis (Coef -44, 95 % CI -86 to -3), and increases in BMI (Coef 30 per 5 kg/m(2), 95 % CI 6-53) and triglyceride (Coef 3.5 per 10 mg/dL, 95 % CI 0.3-6.7) predicted change in SSPG.ConclusionsThere was a wide spectrum of insulin resistance in our HCV population. Host factors, rather than viral factors, appeared to more greatly influence insulin action and its change in HCV.

PurposeTo evaluate the feasibility of using contrast material-enhanced computed tomographic (CT) measurements of hepatic fractional extracellular space (fECS) and macromolecular contrast material (MMCM) uptake to measure severity of liver fibrosis.Materials and methodsAll procedures were approved by and executed in accordance with University of California, San Francisco, institutional animal care and use committee regulations. Twenty-one rats that received intragastric CCl(4) for 0-12 weeks were imaged with respiratory-gated micro-CT by using both a conventional contrast material and a novel iodinated MMCM. Histopathologic hepatic fibrosis was graded qualitatively by using the Ishak fibrosis score and quantitatively by using morphometry of the fibrosis area. Hepatic fECS and MMCM uptake were calculated for each examination and correlated with histopathologic findings by using uni- and multivariate linear regressions.ResultsIshak fibrosis scores ranged from a baseline of 0 in untreated animals to a maximum of 5. Histopathologic liver fibrosis area increased from 0.46% to 3.5% over the same interval. Strong correlations were seen between conventional contrast-enhanced CT measurements of fECS and both the Ishak fibrosis scores (R(2) = 0.751, P < .001) and the fibrosis area (R(2) = 0.801, P < .001). Strong negative correlations were observed between uptake of MMCM in the liver and Ishak fibrosis scores (R(2) = 0.827, P < .001), as well as between uptake of MMCM in the liver and fibrosis area (R(2) = 0.643, P = .001). Multivariate linear regression analysis showed a trend toward independence for fECS and MMCM uptake in the prediction of Ishak fibrosis scores, with an R(2) value of 0.86 (P = .081 and P = .033, respectively).ConclusionContrast-enhanced CT measurements of fECS and MMCM uptake are individually capable of being used to estimate the degree of early hepatic fibrosis in a rat model.Supplemental materialhttp://radiology.rsna.org/lookup/suppl/doi:10.1148/radiol.12112452/-/DC1.

UnlabelledHepatic stellate cells (HSCs) are liver-specific mesenchymal cells that play vital roles in liver development and injury. Our knowledge of HSC biology is limited by the paucity of in vivo data. HSCs and sinusoidal endothelial cells (SECs) reside in close proximity, and interactions between these two cell types are potentially critical for their development and function. Here, we introduce a transgenic zebrafish line, Tg(hand2:EGFP), that labels HSCs. We find that zebrafish HSCs share many similarities with their mammalian counterparts, including morphology, location, lipid storage, gene-expression profile, and increased proliferation and matrix production, in response to an acute hepatic insult. Using the Tg(hand2:EGFP) line, we conducted time-course analyses during development to reveal that HSCs invade the liver after SECs do. However, HSCs still enter the liver in mutants that lack most endothelial cells, including SECs, indicating that SECs are not required for HSC differentiation or their entry into the liver. In the absence of SECs, HSCs become abnormally associated with hepatic biliary cells, suggesting that SECs influence HSC localization during liver development. We analyzed factors that regulate HSC development and show that inhibition of vascular endothelial growth factor signaling significantly reduces the number of HSCs that enter the liver. We also performed a pilot chemical screen and identified two compounds that affect HSC numbers during development.ConclusionOur work provides the first comprehensive description of HSC development in zebrafish and reveals the requirement of SECs in HSC localization. The Tg(hand2:EGFP) line represents a unique tool for in vivo analysis and molecular dissection of HSC behavior.

UnlabelledBile acids are known to play important roles as detergents in the absorption of hydrophobic nutrients and as signaling molecules in the regulation of metabolism. We tested the novel hypothesis that naturally occurring bile acids interfere with protein-mediated hepatic long chain free fatty acid (LCFA) uptake. To this end, stable cell lines expressing fatty acid transporters as well as primary hepatocytes from mouse and human livers were incubated with primary and secondary bile acids to determine their effects on LCFA uptake rates. We identified ursodeoxycholic acid (UDCA) and deoxycholic acid (DCA) as the two most potent inhibitors of the liver-specific fatty acid transport protein 5 (FATP5). Both UDCA and DCA were able to inhibit LCFA uptake by primary hepatocytes in a FATP5-dependent manner. Subsequently, mice were treated with these secondary bile acids in vivo to assess their ability to inhibit diet-induced hepatic triglyceride accumulation. Administration of DCA in vivo via injection or as part of a high-fat diet significantly inhibited hepatic fatty acid uptake and reduced liver triglycerides by more than 50%.ConclusionThe data demonstrate a novel role for specific bile acids, and the secondary bile acid DCA in particular, in the regulation of hepatic LCFA uptake. The results illuminate a previously unappreciated means by which specific bile acids, such as UDCA and DCA, can impact hepatic triglyceride metabolism and may lead to novel approaches to combat obesity-associated fatty liver disease.

Purpose: Nonalcoholic fatty liver disease is an important cause of chronic liver disease. There are limited methods for monitoring metabolic changes during progression to steatohepatitis. Hyperpolarized 13C MRSI (HP 13C MRSI) was used to measure metabolic changes in a rodent model of fatty liver disease. Methods: Fifteen Wistar rats were placed on a methionine‐ and choline‐deficient (MCD) diet for 1–18 weeks. HP 13C MRSI, T2‐weighted imaging, and fat‐fraction measurements were obtained at 3 T. Serum aspartate aminotransaminase, alanine aminotransaminase, and triglycerides were measured. Animals were sacrificed for histology and measurement of tissue lactate dehydrogenase (LDH) activity. Results: Animals lost significant weight (13.6% ± 2.34%), an expected characteristic of the MCD diet. Steatosis, inflammation, and mild fibrosis were observed. Liver fat fraction was 31.7% ± 4.5% after 4 weeks and 22.2% ± 4.3% after 9 weeks. Lactate‐to‐pyruvate and alanine‐to‐pyruvate ratios decreased significantly over the study course; were negatively correlated with aspartate aminotransaminase and alanine aminotransaminase (r = −[0.39–0.61]); and were positively correlated with triglycerides (r = 0.59–0.60). Despite observed decreases in hyperpolarized lactate signal, LDH activity increased by a factor of 3 in MCD diet–fed animals. Observed decreases in lactate and alanine hyperpolarized signals on the MCD diet stand in contrast to other studies of liver injury, where lactate and alanine increased. Observed hyperpolarized metabolite changes were not explained by alterations in LDH activity, suggesting that changes may reflect co‐factor depletion known to occur as a result of oxidative stress in the MCD diet. Conclusion: HP 13C MRSI can noninvasively measure metabolic changes in the MCD model of chronic liver disease. [ABSTRACT FROM AUTHOR]

Insulin initiates diverse hepatic metabolic responses, including gluconeogenic suppression and induction of glycogen synthesis and lipogenesis (1), (2). The liver possesses a rich sinusoidal capillary network with a higher degree [...]

The analgesic acetaminophen is widely known for its potential to cause severe and sometimes lethal liver injury. When ingested in large amounts, acetaminophen overwhelms the normal metabolic pathways of glucuronidation [...], The clinical syndrome of acetaminophen-induced liver injury represents the combined result of drug toxicity and a potent innate immune response that follows drug-induced cell death. In this issue of the JCI, Imaeda and colleagues report that DNA released from dying hepatocytes is a key stimulus of innate immune activation in the acetaminophen-treated mouse liver (see the related article beginning on page 305). They present evidence indicating that hepatocyte DNA promotes immune activation by acting as a danger-associated molecular pattern (DAMP) that stimulates cytokine production in neighboring sinusoidal endothelial cells via Tlr9 and the Nalp3 inflammasome.

[alpha]-Naphthylisothiocyanate (ANIT) is a hepatotoxicant that causes acute cholestatic hepatitis with infiltration of neutrophils around bile ducts and necrotic hepatocytes. The objective of this study was to determine whether the [[beta].sub.2]-integrin CD18, which plays an important role in leukocyte invasion and cytotoxicity, contributes to ANIT-induced hepatic inflammation and liver injury. Mice with varying levels of leukocyte CD18 expression were treated with ANIT and monitored for hepatic neutrophil influx and liver injury over 48 h. Mice that were partially deficient in CD18 (30% of normal levels) developed periportal inflammation and widespread hepatic necrosis after ANIT treatment in a pattern identical to that in wild-type (WT) mice. In contrast, mice that completely lack CD18 (CD18 null) were resistant to ANIT toxicity. Forty-eight hours after ANIT, CD18-null mice displayed 60% lower serum alanine aminotransferase (ALT) levels and 75% less hepatic necrosis, as shown by morphometry, than WT mice. This was true despite evidence that ANIT still provoked hepatic neutrophil influx in CD18-null mice. WT mice could also be protected from ANIT-induced hepatocellular necrosis, by depleting the animals of neutrophils. Notably, neither CD 18-null mice nor neutrophil-depleted WT mice exhibited any attenuation of bile duct injury or cholestasis due to ANIT. We conclude from these experiments that neutrophils invade ANIT-treated livers in a CD 18-independent fashion but utilize CD18 to induce hepatocellular cytotoxicity. The results emphasize that neutrophil-mediated amplification of ANIT-induced liver injury is directed toward hepatocytes rather than cholangiocytes. In fact, the data indicate that the majority of ANIT toxicity toward hepatocytes in vivo is neutrophil driven. bile duct; cholangiocyte; hepatocyte; hepatotoxicity; inflammation; liver doi:10.1152/ajpgi.00458.2005

Author(s): Betty Y Y Tam [1, 10, 11]; Kevin Wei [1, 11]; John S Rudge [2]; Jana Hoffman [1]; Joceyln Holash [2]; Sang-ki Park [3, 10]; Jenny Yuan [1]; Colleen [...]

[alpha]-Naphthylisothiocyanate (ANIT) is a hepatotoxin that causes severe neutrophilic inflammation around portal tracts and bile ducts. The chemotactic signals that provoke this inflammatory response are unknown. In this study, we addressed the possibility that ANIT upregulates CXC chemokines in the liver and that these compounds mediate hepatic inflammation and tissue injury after ANIT treatment. Mice treated with a single dose of ANIT (50 mg/kg) exhibited rapid hepatic induction of the CXC chemokine macrophage inflammatory protein-2 (MIP-2). MIP-2 derived primarily from hepatocytes, with no apparent contribution by biliary cells. In ANIT-treated mice, the induction of MIP-2 coincided with an influx of neutrophils to portal zones; this hepatic neutrophil recruitment was suppressed by 50% in mice that lack the receptor for MIP-2 (CXCR[2.sup.-/-]). Interestingly, despite their markedly reduced degree of hepatic inflammation, CXCR[2.sup.-/-] mice displayed just as much hepatocellular injury and cholestasis after ANIT treatment as wild-type mice. Moreover, after long-term exposure, ANIT CXCR[2.sup.-/-] mice developed liver fibrosis that was indistinguishable from that in wild-type mice. In summary, our data show that CXC chemokines are responsible for some of the hepatic inflammation that occurs in response to ANIT but that these compounds are not essential to the pathogenesis of either acute or chronic ANIT hepatotoxicity. hepatotoxicity; cholangiocyte; neutrophil; fibrosis; macrophage inflammatory protein-2