Maresin 1 mitigates liver steatosis in ob/ob and diet-induced obese mice.

BACKGROUND/OBJECTIVES: The aim of this study was to characterize the effects of Maresin 1 (MaR1) in obesity-related liver steatosis and the mechanisms involved. METHODS: MaR1 effects on fatty liver disease were tested in ob/ob (2-10 µg kg-1 i.p., 20 days) and in diet-induced obese (DIO) mice (2 µg kg-1, i.p., or 50 µg kg-1, oral gavage for 10 days), as well as in cultured hepatocytes. RESULTS: In ob/ob mice, MaR1 reduced liver triglycerides (TG) content, fatty acid synthase (FAS) and stearoyl-CoA desaturase-1 protein expression, while increased acetyl-CoA carboxylase (ACC) phosphorylation and LC3II protein expression, in parallel with a drop in p62 levels. Similar effects on hepatic TG, ACC phosphorylation, p62 and LC3II were observed in DIO mice after MaR1 i.p. injection. Interestingly, oral gavage of MaR1 also decreased serum transaminases, reduced liver weight and TG content. MaR1-treated mice exhibited reduced hepatic lipogenic enzymes content (FAS) or activation (by phosphorylation of ACC), accompanied by upregulation of carnitine palmitoyltransferase (Cpt1a), acyl-coenzyme A oxidase (Acox1) and autophagy-related proteins 5 and 7 (Atg5-7) gene expression, along with increased number of autophagic vacuoles and reduced p62 protein levels. MaR1 also induced AMP-activated protein kinase (AMPK) phosphorylation in DIO mice and in primary hepatocytes, and AMPK inhibition completely blocked MaR1 effects on Cpt1a, Acox1, Atg5 and Atg7 expression. CONCLUSIONS: MaR1 ameliorates liver steatosis by decreasing lipogenic enzymes, while inducing fatty acid oxidation genes and autophagy, which could be related to AMPK activation. Thus, MaR1 may be a new therapeutic candidate for reducing fatty liver in obesity.

Insulin-like growth factor I improves intestinal barrier function in cirrhotic rats.

BACKGROUND AND AIMS: In liver cirrhosis, disruption of the intestinal barrier facilitates bacterial translocation and spontaneous bacterial peritonitis. Insulin-like growth factor I (IGF-I) is an anabolic hormone synthesised by hepatocytes that displays hepatoprotective activities and trophic effects on the intestine. The aim of this study was to investigate the effect of IGF-I on intestinal barrier function in cirrhotic rats. METHODS: In rats with carbon tetrachloride induced cirrhosis, we investigated the effect of IGF-I therapy on: (a) portal pressure; (b) intestinal histology and permeability to endotoxin and bacteria; (c) intestinal expression of cyclooxygenase 2 (COX-2) and tumour necrosis factor alpha (TNF-alpha), two factors that influence in a positive and negative manner, respectively, the integrity of the intestinal barrier; (d) intestinal permeability to 3H-mannitol in rats with bile duct ligation (BDL); and (e) transepithelial electrical resistance (TER) of polarised monolayers of rat small intestine epithelial cells. RESULTS: IGF-I therapy reduced liver collagen expression and portal pressure in cirrhotic rats, induced improvement in intestinal histology, and caused a reduction in bacterial translocation and endotoxaemia. These changes were associated with diminished TNF-alpha expression and elevated COX-2 levels in the intestine. IGF-I reduced intestinal permeability in BDL rats and enhanced barrier function of the monolayers of epithelial intestinal cells where lipopolysaccharide (LPS) caused a decrease in TER that was reversed by IGF-I. This effect of IGF-I was associated with upregulation of COX-2 in LPS treated enterocytes. CONCLUSIONS: IGF-I enhances intestinal barrier function and reduces endotoxaemia and bacterial translocation in cirrhotic rats. IGF-I therapy might be useful in the prevention of spontaneous bacterial peritonitis in liver cirrhosis.

Expression of insulin-like growth factor I by activated hepatic stellate cells reduces fibrogenesis and enhances regeneration after liver injury.

BACKGROUND/AIM: Hepatic stellate cells (HSCs) express alpha-smooth muscle actin (alphaSMA) and acquire a profibrogenic phenotype upon activation by noxious stimuli. Insulin-like growth I (IGF-I) has been shown to stimulate HSCs proliferation in vitro, but it has been reported to reduce liver damage and fibrogenesis when given to cirrhotic rats. METHODS: The authors used transgenic mice (SMP8-IGF-I) expressing IGF-I under control of alphaSMA promoter to study the influence of IGF-I synthesised by activated HSCs on the recovery from liver injury. RESULTS: The transgene was expressed by HSCs from SMP8-IGF-I mice upon activation in culture and in the livers of these animals after CCl4 challenge. Twenty four hours after administration of CCl4 both transgenic and wild type mice showed similar extensive necrosis and increased levels of serum transaminases. However at 72 hours SMP8-IGF-I mice exhibited lower serum transaminases, reduced hepatic expression of alphaSMA, and improved liver morphology compared with wild type littermates. Remarkably, at this time all eight CCl4 treated wild type mice manifested histological signs of liver necrosis that was severe in six of them, while six out of eight transgenic animals had virtually no necrosis. In SMP8-IGF-I mice robust DNA synthesis occurred earlier than in wild type animals and this was associated with enhanced production of HGF and lower TGFbeta1 mRNA expression in the SMP8-IGF-I group. Moreover, Colalpha1(I) mRNA abundance at 72 hours was reduced in SMP8-IGF-I mice compared with wild type controls. CONCLUSIONS: Targeted overexpression of IGF-I by activated HSCs restricts their activation, attenuates fibrogenesis, and accelerates liver regeneration. These effects appear to be mediated in part by upregulation of HGF and downregulation of TGFbeta1. The data indicate that IGF-I can modulate the cytokine response to liver injury facilitating regeneration and reducing fibrosis.

Effect of ursodeoxycholic acid on methionine adenosyltransferase activity and hepatic glutathione metabolism in rats.

BACKGROUND AND AIMS: Both bile salts and glutathione participate in the generation of canalicular bile flow. In this work, we have investigated the effect of different bile salts on hepatic glutathione metabolism. METHODS: Using the isolated and perfused rat liver, we studied hepatic glutathione content, and metabolism and catabolism of this compound in livers perfused with taurocholate, ursodeoxycholate, or deoxycholate. RESULTS: We found that in livers perfused with ursodeoxycholate, levels of glutathione and the activity of methionine adenosyltransferase (an enzyme involved in glutathione biosynthesis) were significantly higher than in livers perfused with other bile salts. In ursodeoxycholate perfused livers, methionine adenosyltransferase showed a predominant tetrameric conformation which is the isoform with highest activity at physiological concentrations of substrate. In contrast, the dimeric form prevailed in livers perfused with taurocholate or deoxycholate. The hepatic activities of gamma-glutamylcysteine synthetase and gamma-glutamyltranspeptidase, enzymes involved, respectively, in biosynthetic and catabolic pathways of glutathione, were not modified by bile salts. CONCLUSIONS: Ursodeoxycholate specifically enhanced methionine adenosyltransferase activity and hepatic glutathione levels. As glutathione is a defensive substance against oxidative cell damage, our observations provide an additional explanation for the known hepatoprotective effects of ursodeoxycholate.

S-adenosyl-L-methionine protects the liver against the cholestatic, cytotoxic, and vasoactive effects of leukotriene D4: a study with isolated and perfused rat liver.

Cysteinyl-leukotrienes can cause cholestasis and liver damage when administered at nanomolar concentrations. Using the isolated and perfused rat liver we analyzed whether S-adenosyl-L-methionine (SAMe) may protect this organ against the noxious effects of leukotriene-D4 (LTD4). We observed that a 2 nmol bolus of this compound decreased bile flow (-12.6% +/- 1.6%, P < .02), and bile salt excretion (-23.5% +/- 2.2%, P < .02; both compared with baseline values), caused the release of glutamic-oxaloacetic transaminase (GOT) and lactic dehydrogenase (LDH) to the hepatic effluent, and increased significantly the perfusion pressure as compared with a control group not receiving LTD4 (6.0 +/- 1.1 vs. 0.2 +/- 0.02 mm hg, respectively; P < .001). The cholestatic effect of LTD4 was attenuated by infusion of SAMe which, at rates of 67 and 100 microg/min, totally prevented the decrease in bile salt excretion. Likewise, in SAMe infused livers, the release to the effluent of GOT and LDH was lower than in the group receiving LTD4 only, and was even lower than in the control group. We also found that the increase in perfusion pressure induced by LTD4 was prevented by SAMe in a dose-dependent manner. Of interest, SAMe increased the biliary excretion of the eicosanoid in a dose-related fashion. We conclude that SAMe reverts the cholestatic, cytotoxic, and hemodynamic effects of LTD4 on the liver, and that these protective effects might be partly because of a stimulation of the biliary excretion of the leukotriene.

Enhanced urinary excretion of cGMP in liver cirrhosis. Relationship to hemodynamic changes, neurohormonal activation, and urinary sodium excretion.

Cyclic guanosine monophosphate (cGMP) has been proposed to mediate peripheral arterial vasodilation in liver cirrhosis. Nitric oxide and natriuretic peptides are the main signals for cGMP generation. Variation in urinary cGMP excretion parallels changes in plasma cGMP levels. Our aim was to determine urinary excretion of cGMP (UcGMPV) and to investigate its relationship to systemic hemodynamics, neurohumoral activity and renal sodium excretion in cirrhosis. Urinary excretion of cGMP was measured in 19 healthy subjects and 20 patients with alcoholic cirrhosis. Systemic hemodynamic parameters, blood volume (BV), plasma atrial natriuretic factor (ANF), and the endothelium-dependent vasodilator substance P (SP) were determined in all patients and in five healthy subjects. Urinary cGMPV was higher in the group of patients (736 pg/min; 50-3229 pg/min) than in controls (126 pg/min; 0-1657 pg/min) (P < 0.01). In addition, UcGMPV inversely correlated with the systemic vascular resistance and directly with cardiac output, blood volume, SP, ANF, and Pugh's score. By Cox regression analysis, only systemic vascular resistance remained inversely associated with UcGMPV. In conclusion, urinary cGMP excretion is increased in cirrhosis. It is suggested that increased cGMP generation may be related to the hyperkinetic circulation in human cirrhosis.

Taurocholate-stimulated leukotriene C4 biosynthesis and leukotriene C4-stimulated choleresis in isolated rat liver.

BACKGROUND/AIMS: Cysteinyl-containing leukotrienes seem to exert a cholestatic effect. However, leukotriene inhibitors were found to reduce bile salt efflux in isolated rat hepatocytes, suggesting a role for leukotrienes in bile flow formation. METHODS: In the isolated rat liver, the effects of two different concentrations of leukotriene C4 on bile flow and bile salt excretion are analyzed, as well as the possible effect of taurocholate on the hepatic production of cysteinyl-containing leukotrienes. RESULTS: Leukotriene C4 (0.25 fmol) increased bile salt excretion (+22.2%; P < 0.05), whereas a much higher dose (0.25 x 10(6) fmol) showed the known cholestatic effect, reducing bile salt excretion (-25.9%; P < 0.01). These dose-dependent biphasic effects were specific because they could be prevented by the simultaneous administration of cysteinyl-containing leukotriene antagonists. On the other hand, taurocholate administration induced a dose-dependent increase in biliary excretion of cysteinyl-containing leukotrienes. Furthermore, taurocholate increased messenger RNA levels of 5-lipoxygenase, a key enzyme in leukotriene biosynthesis. Taurocholate increase of hepatocyte intracellular calcium was not significant, suggesting that taurocholate effects are not mediated by stimulation of calcium metabolism. CONCLUSIONS: These results constitute evidence for the existence of a positive feedback mechanism by which bile salts stimulate the synthesis of leukotrienes that, in turn, stimulate bile salt excretion.

Role of leukotrienes B4 and C4 in liver allograft rejection.

Previous studies have shown that eicosanoids may act in vitro as immunoregulatory substances. In this study, the concentrations of leukotriene B4 (LTB4) and leukotriene C4 (LTC4) were measured in a model of allograft rejection. Six orthotopic allotransplants of the liver were performed in dogs without the administration of immunosuppressives. LTB4 levels showed an increase coinciding with the start of rejection, significant differences being present between the basal levels and those measured 24 h post-revascularization (P < 0.05), and every day from the 3rd postoperative day (P < 0.01). LTB4 rose before the parameters generally used in evaluating rejection. LTC4 levels increased significantly (P < 0.001) in the first 24 h, and experienced no further variations. LTB4 may play an important role in the mechanisms which bring about the response to the allograft. This substance could be a specific and early marker for rejection.

Inhibitors of the lipoxygenase arachidonic acid pathway impair glycocholate efflux in isolated rat hepatocytes.

Lipoxygenase arachidonic acid metabolites mediate secretory processes in several tissues, but their possible involvement in liver transport functions is still unknown. This study evaluated the influence of the lipoxygenase inhibitor nordihydroguayaretic acid (NDGA), the cyclooxygenase inhibitor indomethacin (INDO), and the dual cyclo and lipoxygenase inhibitors 3-amino-1-[m-(trifluoromethyl)-phenyl]-2-pyrazoline (BW 755c) and eicosatetraynoic acid (ETYA) on the handling of glycocholic acid (GC) by isolated rat hepatocytes. No drug modified cell viability or oxygen consumption in hepatocytes. In 30-min incubations with 50 microM GC the initial rate of GC uptake (V0) in control hepatocytes was 1.15 +/- 0.09 nmol.mg protein-1.min-1. The cellular GC content remained constant from 10 to 30 min (steady-state phase), the 30-min value being 6.63 +/- 0.35 nmol.mg protein-1. NDGA (10-50 microM), BW 755c (25-200 microM) and ETYA (5-100 microM), prevented the steady-state phase occurring, thus determining a progressive accumulation of GC in cells with time. As compared to controls, 50 microM NDGA (+37%, p less than 0.01), 200 microM BW 755c (+39%, p less than 0.01) and 5 microM ETYA (+19%, p less than 0.05) induced the highest increases in the amount of GC in cells at 30 min, in all cases V0 being unchanged. Concentrations of BW 755c and ETYA above those indicated also decreased V0. Both V0 and the amount of cellular GC in the steady-state phase were proportionally decreased by high INDO concentrations (25-100 microM) which did not modify the morphology of the uptake curve. Since experiments with dual and lipoxygenase inhibitors suggested an impairment of GC efflux, the initial rate of GC efflux (V0ef) was measured in hepatocytes preloaded with 50 microM GC and transferred to a GC-free medium. In controls, V0ef was 1.12 +/- 0.12 nmol.mg protein-1.min-1. BW 755c (200 microM) and NDGA (50 microM) reduced V0ef by 45 and 38%, respectively. The kinetic analysis of the effect of 200 microM BW 755c on the efflux process using hepatocytes preloaded with GC from 5 to 200 microM disclosed a non-competitive inhibition. Vmax was reduced from 1.37 +/- 0.15 to 0.89 +/- 0.10 (p less than 0.01), whereas Km was unchanged (3.79 +/- 0.33 vs. 4.25 +/- 0.54, N.S.). In summary, inhibitors of the lipoxygenase arachidonic acid pathway impaired the efflux of GC from isolated rat hepatocytes. The hypothesis is raised that oxidized metabolites of arachidonic acid may participate on the secretion of bile salts in these cells.

Renal prostacyclin influences renal function in non-azotemic cirrhotic patients treated with furosemide.

The influence of prostaglandins on renal function changes induced by furosemide was analyzed in 21 non-azotemic cirrhotic patients with ascites. Patients were studied in two periods of 120 min immediately before and after furosemide infusion (20 mg, ev). Furosemide caused an increase in creatinine clearance in 15 patients (group A: 99 +/- 7 vs. 129 +/- 5 ml/min; mean +/- S.E.) and a reduction in the remaining six (group B: 102 +/- 13 vs. 71 +/- 9 ml/min). Parallel changes were observed in the urinary excretion of 6-Keto-prostaglandin-F1 alpha (metabolite of renal prostacyclin) which augmented after furosemide in 14 of the 15 patients from group A (478 +/- 107 vs. 1034 +/- 159 pg/min, p less than 0.001) and decreased in all patients from group B (1032 +/- 240 vs. 548 +/- 136 pg/min, p less than 0.05). In contrast, the urinary excretion of prostaglandin E2 was stimulated by furosemide in all patients (group A, 92 +/- 19 vs. 448 +/- 60 pg/min, p less than 0.001; and group B, 209 +/- 63 vs. 361 +/- 25 pg/min, p less than 0.05). In all of the patients furosemide-induced changes (post- minus pre-furosemide values) in creatinine clearance were closely correlated in a direct and linear fashion with those in 6-Keto-prostaglandin-F1 alpha (r = 0.74; p less than 0.001). These changes were associated with a higher furosemide-induced natriuresis in group A than in group B (641 +/- 68 vs. 302 +/- 46 mumol/min, p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

[Effect of energy administration on the amino acid level in the postoperative phase]. / Auswirkungen von Energiezufuhr auf die Aminosäuren-Spiegel während der postoperativen Phase.

A total of 34 cancer patients, all of them subjected to radical surgery of the stomach or large bowel were studied. Group I (n = 5) received during the first postoperative days total parenteral nutrition with a caloric support of 35-45 kcal/kg/day and 12,5 gr N in a 8,5% L-aminoacid solution (Freamine II). Group II (n = 9) received an isotonic solution of 3% L-aminoacid without caloric support. Serum amino acids (AA) were determined daily (Perkin-Elmer KLA-1 Analyzer), as well as nitrogen balance (NB) and serum albumin (Alb) on the preoperative, 1st, and 6th postoperative day: Both groups experienced a progressive increase of serum AA during the period of study. Group II showed levels of branched-AA significantly higher than group I, as well as the total of essential-AA. MET, GLY and PHE were considerably elevated in both groups. ALA did not change in group I showing subnormal values in group II. NB was significantly higher in group I, but none of the groups studied has recovered the initial values of Alb after six days of treatment.