Evaluation of the antitumoral effect sunitinib eluting beads in VX2 liver tumour in a rabbit model.

Background: We demonstrated that DC Bead (Biocompatibles UK, Ltd) could be loaded with sunitinib and injected intra-arterially in the rabbit without unexpected toxicity. The purpose of this study is to evaluate the antitumoral effect of sunitinib eluting beads in the VX2 tumor model of liver cancer. Methods: VX2 tumors were implanted in the left liver lobe of New-Zealand white rabbits. Animals were assigned to 3 groups: Group 1 (n=6) received 1.5mg of sunitinib loaded in 0.05ml of 100-300um DC Bead, group 2 (n=5) received 0.05ml of 100-300um DC Bead, group 3 (n=5) received 0.05ml NaCl 0.9% in the left hepatic artery. One animal in each group was sacrificed at 24 hours and the others were followed for survival until day 15. Liver enzymes were measured daily. In group 1, plasmatic sunitinib concentration were measured daily by LC MS/MS tandem mass spectroscopy. At day 15 all living animals were sacrificed. After sacrifice, the livers were harvested for determination of the VEGF receptor tyrosine kinase activity by western blot and histopathological examination. Results: In group 1, no animals died during follow-up. In group 2, 2 animals died during follow-up on day x. In control group 3, 3 animals died during follow up on day x. In group 1 plasmatic sunitinib levels remained under therapeutic concentration throughout the experiment. Very high concentrations of sunitinib were measured in the liver tissue 24 and 15 days after embolization. Inhibition of the phosphorylation of the RTK was demonstrated at 24h and 15 days in groups 1. Sunitinib eluting beads seemed to penetrate in the tumor more effectively and there was more necrosis around the beads than their bland counterparts. Conclusions: Administration of sunitinib eluting beads in VX2 carrying rabbits resulted invery high drug concentrations at the site of embolization with minimal systemic passage. Despite the very high tissular sunitinib concentration we did not observe any additional toxicity with loaded beads. Sunitinib eluting beads inhibit the activation of RTK's triggered by ischemia and seem to prolong survival of the treated animals. Therefore we consider that local treatment with sunitinib may provide a promising approach for the treatment of liver cancer.

CD1d-restricted NK T cells are dispensable for specific antibody responses and protective immunity against liver stage malaria infection in mice.

Immunization with a single dose of irradiated sporozoites is sufficient to induce protection against malaria in wild-type mice. Although this protection is classically attributed to conventional CD4+ and CD8+ T cells, several recent reports have suggested an important role for CD1-restricted NK T cells in immunity to malaria. In this study, we directly compared the ability of C57BL/6 wild-type and CD1-deficient mice to mount a protective immune response against Plasmodium berghei sporozoites. Our data indicate that CD1-restricted NK T cells are not required for protection in this model system. Moreover, specific IgG antibody responses to the P. berghei circumsporozoite repeat sequence were also unaffected by CD1 deficiency. Collectively, our data demonstrate that CD1-restricted NK T cells are dispensable for protective immunity to liver stage P. berghei infection.

Effective size of two feral domestic cat populations (Felis catus L): effect of the mating system.

A variety of behavioural traits have substantial effects on the gene dynamics and genetic structure of local populations. The mating system is a plastic trait that varies with environmental conditions in the domestic cat (Felis catus) allowing an intraspecific comparison of the impact of this feature on genetic characteristics of the population. To assess the potential effect of the heterogenity of males' contribution to the next generation on variance effective size, we applied the ecological approach of Nunney & Elam (1994) based upon a demographic and behavioural study, and the genetic 'temporal methods' of Waples (1989) and Berthier et al. (2002) using microsatellite markers. The two cat populations studied were nearly closed, similar in size and survival parameters, but differed in their mating system. Immigration appeared extremely restricted in both cases due to environmental and social constraints. As expected, the ratio of effective size to census number (Ne/N) was higher in the promiscuous cat population (harmonic mean = 42%) than in the polygynous one (33%), when Ne was calculated from the ecological method. Only the genetic results based on Waples' estimator were consistent with the ecological results, but failed to evidence an effect of the mating system. Results based on the estimation of Berthier et al. (2002) were extremely variable, with Ne sometimes exceeding census size. Such low reliability in the genetic results should retain attention for conservation purposes.

Normal weight obesity: relationship with lipids, glycaemic status, liver enzymes and inflammation

BACKGROUND AND AIMS: Normal weight obesity (NWO) is defined as an excessive body fat associated with a normal body mass index (BMI) and has been associated with early inflammation, but its relationship with cardiovascular risk factors await investigation. METHODS AND RESULTS: Cross-sectional study including 3213 women and 2912 men aged 35-75 years to assess the clinical characteristics of NWO in Lausanne, Switzerland. Body fat was assessed by bioimpedance. NWO was defined as a BMI<25 kg/m(2) and a % body fat ≥66(th) gender-specific percentiles. The prevalence of NWO was 5.4% in women and less than 3% in men, so the analysis was restricted to women. NWO women had a higher % of body fat than overweight women. After adjusting for age, smoking, educational level, physical activity and alcohol consumption, NWO women had higher blood pressure and lipid levels and a higher prevalence of dyslipidaemia (odds-ratio=1.90 [1.34-2.68]) and fasting hyperglycaemia (odds-ratio=1.63 [1.10-2.42]) than lean women, whereas no differences were found between NWO and overweight women. Conversely, no differences were found between NWO and lean women regarding levels of CRP, adiponectin and liver markers (alanine aminotransferase, aspartate aminotransferase and gamma glutamyl transferase). Using other definitions of NWO led to similar conclusions, albeit some differences were no longer significant. CONCLUSION: NWO is almost nonexistent in men. Women with NWO present with higher cardiovascular risk factors than lean women, while no differences were found for liver or inflammatory markers. Specific screening of NWO might be necessary in order to implement cardiovascular prevention.

Can near infrared spectroscopy of the liver monitor tissue oxygenation?

Measurement of the hepatic oxygenation index by near infrared spectroscopy is a suitable method to estimate the oxygenation and can be a non-invasive means to continuously monitor tissue perfusion and to detect early haemodynamic disturbances in critically ill children.

Genome-wide association study identifies variants associated with progression of liver fibrosis from HCV infection.

Polymorphisms in IL28B were shown to affect clearance of hepatitis C virus (HCV) infection in genome-wide association (GWA) studies. Only a fraction of patients with chronic HCV infection develop liver fibrosis, a process that might also be affected by genetic factors. We performed a 2-stage GWA study of liver fibrosis progression related to HCV infection. We studied well-characterized HCV-infected patients of European descent who underwent liver biopsies before treatment. We defined various liver fibrosis phenotypes on the basis of METAVIR scores, with and without taking the duration of HCV infection into account. Our GWA analyses were conducted on a filtered primary cohort of 1161 patients using 780,650 single nucleotide polymorphisms (SNPs). We genotyped 96 SNPs with P values <5 × 10(-5) from an independent replication cohort of 962 patients. We then assessed the most interesting replicated SNPs using DNA samples collected from 219 patients who participated in separate GWA studies of HCV clearance. In the combined cohort of 2342 HCV-infected patients, the SNPs rs16851720 (in the total sample) and rs4374383 (in patients who received blood transfusions) were associated with fibrosis progression (P(combined) = 8.9 × 10(-9) and 1.1 × 10(-9), respectively). The SNP rs16851720 is located within RNF7, which encodes an antioxidant that protects against apoptosis. The SNP rs4374383, together with another replicated SNP, rs9380516 (P(combined) = 5.4 × 10(-7)), were linked to the functionally related genes MERTK and TULP1, which encode factors involved in phagocytosis of apoptotic cells by macrophages. Our GWA study identified several susceptibility loci for HCV-induced liver fibrosis; these were linked to genes that regulate apoptosis. Apoptotic control might therefore be involved in liver fibrosis.

Peroxisome proliferator-activated receptor-alpha-null mice have increased white adipose tissue glucose utilization, GLUT4, and fat mass: Role in liver and brain.

Activation of the peroxisome proliferator-activated receptor (PPAR)-alpha increases lipid catabolism and lowers the concentration of circulating lipid, but its role in the control of glucose metabolism is not as clearly established. Here we compared PPARalpha knockout mice with wild type and confirmed that the former developed hypoglycemia during fasting. This was associated with only a slight increase in insulin sensitivity but a dramatic increase in whole-body and adipose tissue glucose use rates in the fasting state. The white sc and visceral fat depots were larger due to an increase in the size and number of adipocytes, and their level of GLUT4 expression was higher and no longer regulated by the fed-to-fast transition. To evaluate whether these adipocyte deregulations were secondary to the absence of PPARalpha from liver, we reexpresssed this transcription factor in the liver of knockout mice using recombinant adenoviruses. Whereas more than 90% of the hepatocytes were infected and PPARalpha expression was restored to normal levels, the whole-body glucose use rate remained elevated. Next, to evaluate whether brain PPARalpha could affect glucose homeostasis, we activated brain PPARalpha in wild-type mice by infusing WY14643 into the lateral ventricle and showed that whole-body glucose use was reduced. Hence, our data show that PPARalpha is involved in the regulation of glucose homeostasis, insulin sensitivity, fat accumulation, and adipose tissue glucose use by a mechanism that does not require PPARalpha expression in the liver. By contrast, activation of PPARalpha in the brain stimulates peripheral glucose use. This suggests that the alteration in adipocyte glucose metabolism in the knockout mice may result from the absence of PPARalpha in the brain.

TRAIL receptor-mediated JNK activation and Bim phosphorylation critically regulate Fas-mediated liver damage and lethality.

TNF-related apoptosis-inducing ligand (TRAIL) is a member of the TNF family with potent apoptosis-inducing properties in tumor cells. In particular, TRAIL strongly synergizes with conventional chemotherapeutic drugs to induce tumor cell death. Thus, TRAIL has been proposed as a promising future cancer therapy. Little, however, is known regarding what the role of TRAIL is in normal untransformed cells and whether therapeutic administration of TRAIL, alone or in combination with other apoptotic triggers, may cause tissue damage. In this study, we investigated the role of TRAIL in Fas-induced (CD95/Apo-1-induced) hepatocyte apoptosis and liver damage. While TRAIL alone failed to induce apoptosis in isolated murine hepatocytes, it strongly amplified Fas-induced cell death. Importantly, endogenous TRAIL was found to critically regulate anti-Fas antibody-induced hepatocyte apoptosis, liver damage, and associated lethality in vivo. TRAIL enhanced anti-Fas-induced hepatocyte apoptosis through the activation of JNK and its downstream substrate, the proapoptotic Bcl-2 homolog Bim. Consistently, TRAIL- and Bim-deficient mice and wild-type mice treated with a JNK inhibitor were protected against anti-Fas-induced liver damage. We conclude that TRAIL and Bim are important response modifiers of hepatocyte apoptosis and identify liver damage and lethality as a possible risk of TRAIL-based tumor therapy.

Differential distribution of tau proteins in developing cat cerebral cortex and corpus callosum.

During the postnatal development of cat visual cortex and corpus callosum the molecular composition of tau proteins varied with age. In both structures, they changed between postnatal days 19 and 39 from a set of two juvenile forms to a set of at least two adult variants with higher molecular weights. During the first postnatal week, tau proteins were detectable with TAU-1 antibody in axons of corpus callosum and visual cortex, and in some perikarya and dendrites in the visual cortex. At later ages, tau proteins were located exclusively within axons in all cortical layers and in the corpus callosum. Dephosphorylation of postnatal day 11 cortical tissue by alkaline phosphatase strongly increased tau protein immunoreactivity on Western blots and in numerous perikarya and dendrites in all cortical layers, in sections, suggesting that some tau forms had been unmasked. During postnatal development the intensity of this phosphate-dependent somatodendritic staining decreased, but remained in a few neurons in cortical layers II and III. On blots, the immunoreactivity of adult tau to TAU-1 was only marginally increased by dephosphorylation. Other tau antibodies (TAU-2, B19 and BR133) recognized two juvenile and two adult cat tau proteins on blots, and localized tau in axons or perikarya and dendrites in tissue untreated with alkaline phosphatase. Tau proteins in mature tissue were soluble and not associated with detergent-resistant structures. Furthermore, dephosphorylation by alkaline phosphatase resulted in the appearance of more tau proteins in soluble fractions. Therefore tau proteins seem to alter their degree of phosphorylation during development. This could affect microtubule stability as well as influence axonal and dendritic differentiation.

Assessment of liver tumor response by high-field (3T) MRI after radiofrequency ablation: Short- and mid-term evolution of diffusion parameters within the ablation zone.

PURPOSE: To compare the apparent diffusion coefficient (ADC) values of malignant liver lesions on diffusion-weighted MRI (DWI) before and after successful radiofrequency ablation (RF ablation). MATERIALS AND METHODS: Thirty-two patients with 43 malignant liver lesions (23/20: metastases/hepatocellular carcinomas (HCC)) underwent liver MRI (3.0T) before (<1month) and after RF ablation (at 1, 3 and 6months) using T2-, gadolinium-enhanced T1- and DWI-weighted MR sequences. Jointly, two radiologists prospectively measured ADCs for each lesion by means of two different regions of interest (ROIs), first including the whole lesion and secondly the area with the visibly most restricted diffusion (MRDA) on ADC map. Changes of ADCs were evaluated with ANOVA and Dunnett tests. RESULTS: Thirty-one patients were successfully treated, while one patient was excluded due to focal recurrence. In metastases (n=22), the ADC in the whole lesion and in MRDA showed an up-and-down evolution. In HCC (n=20), the evolution of ADC was more complex, but with significantly higher values (p=0.013) at 1 and 6months after RF ablation. CONCLUSION: The ADC values of malignant liver lesions successfully treated by RF ablation show a predictable evolution and may help radiologists to monitor tumor response after treatment.

Complex organization of CTF/NF-I, C/EBP, and HNF3 binding sites within the promoter of the liver-specific vitellogenin gene.

Vitellogenin genes are expressed specifically in the liver of female oviparous vertebrates under the strict control of estrogen. To explain this tissue-specific expression, we performed a detailed analysis of the Xenopus laevis vitellogenin gene B1 promoter by DNase I footprinting and gel mobility-shift assays. We characterized five binding sites for the ubiquitous factor CTF/NF-I. Two of these sites are close to the TATA-box, whereas the others are located on both sides of the estrogen responsive unit formed by two imperfect estrogen response elements. Moreover two liver-enriched factors, C/EBP and HNF3, were found to interact with multiple closely spaced proximal promoter elements in the first 100 base pairs upstream of the TATA-box. To confirm the physiological significance of this in vitro analysis, in vivo DNase I footprinting experiments were carried out using the ligation-mediated polymerase chain reaction technique. The various cis-elements characterized in vitro as binding sites for known transcription factors and more particularly for liver-enriched transcription factors are efficiently recognized in vivo as well, suggesting that they play an important role in the control of the liver-specific vitellogenin gene B1 expression.

Injection of partially purified estrogen receptor protein from Xenopus liver nuclei into oocytes activates the silent vitellogenin locus.

Injection of extracts from Xenopus liver nuclei that are enriched 2000 times in estradiol receptor into Xenopus oocytes induces transcription of the silent vitellogenin locus, which is activated in liver by estradiol, but not of the albumin locus, which is active in liver but suppressed by high levels of estradiol. Transcription initiates within the 5'-end region of the gene we have studied and probably continues into the 3' third. The activation seems to be very efficient, but most of the primary transcripts are probably rapidly and inaccurately processed. New proteins are also made and secreted by the oocytes.

MicroRNA-122 modulates the rhythmic expression profile of the circadian deadenylase Nocturnin in mouse liver.

Nocturnin is a circadian clock-regulated deadenylase thought to control mRNA expression post-transcriptionally through poly(A) tail removal. The expression of Nocturnin is robustly rhythmic in liver at both the mRNA and protein levels, and mice lacking Nocturnin are resistant to diet-induced obesity and hepatic steatosis. Here we report that Nocturnin expression is regulated by microRNA-122 (miR-122), a liver specific miRNA. We found that the 3'-untranslated region (3'-UTR) of Nocturnin mRNA harbors one putative recognition site for miR-122, and this site is conserved among mammals. Using a luciferase reporter construct with wild-type or mutant Nocturnin 3'-UTR sequence, we demonstrated that overexpression of miR-122 can down-regulate luciferase activity levels and that this effect is dependent on the presence of the putative miR-122 recognition site. Additionally, the use of an antisense oligonucleotide to knock down miR-122 in vivo resulted in significant up-regulation of both Nocturnin mRNA and protein expression in mouse liver during the night, resulting in Nocturnin rhythms with increased amplitude. Together, these data demonstrate that the normal rhythmic profile of Nocturnin expression in liver is shaped in part by miR-122. Previous studies have implicated Nocturnin and miR-122 as important post-transcriptional regulators of both lipid metabolism and circadian clock controlled gene expression in the liver. Therefore, the demonstration that miR-122 plays a role in regulating Nocturnin expression suggests that this may be an important intersection between hepatic metabolic and circadian control.

Traitement des tumeurs malignes du foie par radiologie interventionnelle: techniques ablatives [Interventional radiology procedures for malignancies of the liver: ablation procedures].

Percutaneous ablative procedures allow curative treatment of stage BCLC 0 or BCLC A hepatocellular carcinoma, as well as liver metastases of colorectal cancer. Several methods exist including radiofrequency ablation, the most commonly used. These techniques can be used in combination with surgical excision or alone if surgery is contraindicated. They are associated with significantly reduced mortality as compared to surgery.