Automatic liver segmentation and assessment of liver fibrosis using deep learning with MR T1-weighted images in rats.

OBJECTIVES: To validate the performance of nnU-Net in segmentation and CNN in classification for liver fibrosis using T1-weighted images. MATERIALS AND METHODS: In this prospective study, animal models of liver fibrosis were induced by injecting subcutaneously a mixture of Carbon tetrachloride and olive oil. A total of 99 male Wistar rats were successfully induced and underwent MR scanning with no contrast agent to get T1-weighted images. The regions of interest (ROIs) of the whole liver were delineated layer by layer along the liver edge by 3D Slicer. For segmentation task, all T1-weighted images were randomly divided into training and test cohorts in a ratio of 7:3. For classification, images containing the hepatic maximum diameter of every rat were selected and 80% images of no liver fibrosis (NLF), early liver fibrosis (ELF) and progressive liver fibrosis (PLF) stages were randomly selected for training, while the rest were used for testing. Liver segmentation was performed by the nnU-Net model. The convolutional neural network (CNN) was used for classification task of liver fibrosis stages. The Dice similarity coefficient was used to evaluate the segmentation performance of nnU-Net. Confusion matrix, ROC curve and accuracy were used to show the classification performance of CNN. RESULTS: A total of 2628 images were obtained from 99 Wistar rats by MR scanning. For liver segmentation by nnU-Net, the Dice similarity coefficient in the test set was 0.8477. The accuracies of CNN in staging NLF, ELF and PLF were 0.73, 0.89 and 0.84, respectively. The AUCs were 0.76, 0.88 and 0.79, respectively. CONCLUSION: The nnU-Net architecture is of high accuracy for liver segmentation and CNN for assessment of liver fibrosis with T1-weighted images.

Effect of Iron Deposition on Native T1 Mapping and Blood Oxygen Level Dependent for the Assessment of Liver Fibrosis in Rabbits With Carbon Tetrachloride Intoxication.

RATIONALE AND OBJECTIVES: This study aimed to explore the effect of iron deposition on native T1 mapping and blood oxygen level-dependent (BOLD) imaging in detecting liver fibrosis (LF) in a rabbit model. MATERIALS AND METHODS: An LF group (n = 100) was established by subcutaneously injecting 50% carbon tetrachloride (CCl4) oil solution, and 20 normal rabbits composed a control group. Native T1 mapping and BOLD were performed, and the T1native and R2* quantitative parameters were analyzed by receiver operating characteristic (ROC) and multiple logistic regression analyses, with histopathological results and liver iron content (LIC) serving as reference standards. RESULTS: In total, 18, 17, 16, 18, and 15 rabbits were histopathologically diagnosed with LF stages F0, F1, F2, F3, and F4, respectively. T1native (r = 0.47), R2* (r = 0.75) and LIC (r = 0.61) increased with LF stage progression (p < 0.001). Compared to T1native values, R2* performed better in diagnosing the LF stage, especially for distinguishing F1-F2 from F3-F4 (AUC = 0.66 vs. 0.91, p = 0.01). Combined with the LIC, both T1native and R2* showed improved diagnostic value in comparison to the individual imaging techniques, particularly for diagnosing F0 vs. F1-F2 and F0 vs. F1-F4, with AUC values of 0.90 vs. 0.70 (p = 0.01) and 0.93 vs. 0.77 (p = 0.01) for T1native + LIC vs. LIC, respectively. CONCLUSION: BOLD imaging performed better than native T1 mapping in predicting and diagnosing LF stage progression. The decrease in diagnostic accuracy caused by the deposition of liver iron is a potential pitfall in the assessment of LF with BOLD imaging and native T1 mapping.

Assessment of the Biological Activities of Egyptian Purslane (Portulaca oleracea) Extract after Incorporating Metal Nanoparticles, in Vitro and in Vivo Study.

OBJECTIVE: Egyptian Purslane (Portulaca oleracea) is rich in omega-3 fatty acids and a wide range of vitamins and phyto-constituents that were absorbed slowly due to their high molecular weights. Therefore, this study was designed to accelerate the absorption of these phyto-constituents and hence increase their bioavailability by incorporating silver (Ag-NPs) and zinc oxide nanoparticles (ZnO-NPs) due to their impressive properties. METHODS: The major phyto-constituents and different biological activities were quantified in aqueous extract before and after incorporating metal nanoparticles (M-NPs). The efficiency of ZnO-P. nano-extract was studied on cell cycle and apoptosis of human hepatocellular carcinoma (HEPG-2) cells. Then, both Ag- and ZnO-P. nano-extracts were studied against hepatic fibrosis induced by thioacetamide (TAA) in rats through undergoing different hematological and biochemical measurements in addition to the histopathological examination in hepatic tissues compared to the extract itself. RESULTS: The ZnO-P. nano-extract showed significantly (P<0.05) higher antioxidant and scavenging activity due to the existence of higher total polyphenolic content. Also, it exhibited a significantly (P<0.05) higher inhibitory effect on acetyl cholinesterase (AChE) activity and higher cytotoxic activity against HEPG-2 cells. Therefore, ZnO-P. nano-extract was studied against the cell cycle and apoptosis of HEPG-2 cells compared to the extract itself. It was found that ZnO-P. nano-extract was safer than Ag-P. nano-extract. Both Ag- and ZnO- P. nano-extracts were studied against the hepatic fibrosis induced by thioacetamide (TAA) compared to the native extract. It was noticed that ZnO-P. nano-extract exhibited an ameliorative effect against hepatic fibrosis by decreasing levels of inflammatory and fibrotic markers significantly (P<0.05) more than Ag-P. nano-extract. Furthermore, it improved the antioxidant status of the hepatic tissue in addition to restoring the histopathological architecture of liver tissue. CONCLUSION: ZnO-P. nano-extract showed higher in vitro and in vivo biological activities than Ag-P. nano-extract and native P. extract itself.

Usefulness of noninvasive diagnostic procedures for assessment of methotrexate hepatotoxicity in patients with rheumatoid arthritis.

Methotrexate (MTX) is recommended as a first-line treatment for rheumatoid arthritis (RA). There are no strict guidelines regarding monitoring for liver damage in RA patients. This study aimed to evaluate noninvasive diagnostic procedures in assessing liver fibrosis in RA patients. Ninety-six RA patients were recruited for this study. The procollagen III N-terminal peptide (PIIINP) serum level was measured in all patients. The Enhanced Liver Fibrosis score (ELF-1) was calculated for 82 patients. Transient elastography (TE) was performed in 91 patients, those examined were divided into two groups: a study and control group, comprising patients with and without risk factors for liver fibrosis, respectively. The TE result correlated only with the body mass index-BMI (p < 0.05); there was no correlation with the cumulative MTX dose (p = 0.33). The TE result was significantly higher in those with risk factors for liver fibrosis than in those without risk factors (TE result > = 7.1 kPa 28/42 vs 13/41, HR = 2.103, Mann-Whitney U test, approximately 0.02). There was a positive correlation between the PIIINP level and body weight (p = 0.028), cumulative MTX dose (p = 0.007), RA activity (p = 0.028) and diabetes mellitus (DM) (p = 0.001). There was a positive correlation between the ELF-1 score and age (p < 0.001), cumulative MTX dose (p = 0.007) and RA activity (p < 0.001). The PIIINP level and ELF-1 score are not organ specific, and readings may vary depending on RA activity. TE is organ specific and can be performed by a skilled ultrasonographer might be useful to assess actual liver condition.

Factors predicting staging and treatment initiation for patients with chronic hepatitis C infection: insurance a key predictor.

BACKGROUND: Chronic hepatitis C (HCV) infection affects over 2.4 million Americans and accounts for 18 000 deaths per year. Treatment initiation in this population continues to be low even after introduction of highly effective and shorter duration direct-acting antivirals. This study assesses factors that influence key milestones in the HCV care continuum. METHODS: Retrospective time-to-event analyses were performed to assess factors influencing liver fibrosis staging and treatment initiation among individuals confirmed with chronic HCV infection at University of Illinois Hospital and Health Sciences System between 1 August 2015 and 24 October 2016 and followed through 28 January 2018. Cox regression models were utilized for multivariable analyses. RESULTS: Individuals tested at the liver clinic (hazard ratio [HR] = 2.03; 95% confidence interval [CI]: 1.19-3.46) and at the federally qualified health center (HR = 3.51; 95% CI: 2.19-5.64) had higher instantaneous probability of being staged compared with individuals tested at the emergency department (ED) or inpatient setting. And probability of treatment initiation increased with advancing liver fibrosis especially for Medicaid beneficiaries (HR = 1.64; 95% CI: 1.35-1.99). CONCLUSIONS: The study demonstrates a need for improving access for patients with early stages of the disease in order to reduce HCV-related morbidity and mortality, especially those tested at nontraditional care locations such as the ED or the inpatient setting.

Standardization of diethylnitrosamine-induced hepatocellular carcinoma rat model with time based molecular assessment.

This study was intended (1) to develop a robust animal model for hepatocellular carcinoma (HCC) research, in which HCC tumors develop in a background of fibrosis or cirrhosis; and (2) to explore time-dependent regulatory changes in key molecular markers during disease advancement and HCC development. With the aim of establishing such HCC model, male Sprague-Dawley rats were injected with diethylnitrosamine (DEN) at a dose of 30 mg/kg twice a week for 10 weeks then once a week from 12th to 16th weeks. The rats were kept under observation until 18th week. At defined time intervals (2nd, 4th, 12th, and 18th week), serum biomarkers and microscopic components of tissue samples were used to investigate the chronic progression of liver disease, while gene and protein analysis was used to monitor expression patterns during HCC development. DEN-intoxicated rats manifested inflammation at week 4, fibrosis at week 12 and cirrhosis with early HCC tumors at week 18. Molecular analysis revealed that key markers of inflammation (Il-1ß, Il-6, and Tnf-α), fibrosis (Tgf-ß1, Col1α1, Col3α1, and Timp-1), and angiogenesis (Hif1-α and Vegf) were promptly (P ≤ 0.001) up-regulated at week 4, week 12 and week 18, respectively. Oxidative stress (iNos, Cyp2e1, and Sod1) and pro-apoptotic (Bax) markers showed significant upregulation from week 4 to week 12. However, Sod1 and Bax expressions dropped after week 12 and reached a minimum at 18th week. Strikingly, expressions of anti-apoptotic (Bcl-2) and cell proliferation (Pcna, Hgf, and Afp) markers were abruptly increased at week 18. Collectively, we describe an 18-week HCC model in DEN-intoxicated rats that exhibit chronic inflammation, oxidative imbalance, advance fibrosis/cirrhosis, halted apoptosis, and angiogenic sprouting, progressively.

Methotrexate does not increase the risk of liver fibrosis in patients with rheumatoid arthritis: assessment by ultrasound elastography (ARFI-MetRA study).

OBJECTIVES: Only limited data are available on the risk of liver fibrosis in patients with rheumatoid arthritis on long-term methotrexate treatment. To assess the risk of liver fibrosis in patients with rheumatoid arthritis treated with methotrexate, non-invasive, ultrasound-based elastography [acoustic radiation force impulse (ARFI) imaging] was applied. METHODS: In total, 119 patients were assessed using acoustic radiation force impulse (ARFI) imaging between July 2018 and April 2019. In a cross-sectional, single-centre study design, ARFI scores were compared between patient subgroups with (n = 65) and without (n = 54) methotrexate exposure. The main outcome variable was the mean fibrosis score as measured by the ARFI method. The mean shear wave velocity was calculated from 10 valid ARFI measurements for each patient. Inferential statistical analyses (between group) were performed using ANOVA for independent samples in the case of continuous outcome variables. RESULTS: Sixty-five patients with and fifty-four patients without MTX exposure were assessed using the ARFI elastography method. Participating patients on MTX medication (1.113 m/s) showed ARFI scores that were comparable to those of participants without MTX exposure (1.062 m/s); P = 0.228. The mean cumulative dose in the group of MTX-exposed patients was 3602 mg. CONCLUSION: The mean value of the repeated determination of liver density using ARFI imaging did not differ significantly between the MTX-exposed and MTX-naive patients with RA. No increased rate of liver fibrosis was found among RA patients treated with MTX.

Assessment of fibrotic pathways induced by environmental chemicals using 3D-human liver microtissue model.

Exposure to environmental chemicals, particularly those with persistent and bioaccumulative properties have been linked to liver diseases. Induction of fibrotic pathways is considered as a pre-requirement of chemical induced liver fibrosis. Here, we applied 3D in vitro human liver microtissues (MTs) composed of HepaRG, THP-1 and hTERT-HSC that express relevant hepatic pathways (bile acid, sterol, and xenobiotic metabolism) and can recapitulate key events of liver fibrosis (e.g. extracellular matrix-deposition). The liver MTs were exposed to a known profibrotic chemical, thioacetamide (TAA) and three representative environmental chemicals (TCDD, benzo [a] pyrene (BaP) and PCB126). Both TAA and BaP triggered fibrotic pathway related events such as hepatocellular damage (cytotoxicity and decreased albumin release), hepatic stellate cell activation (transcriptional upregulation of α-SMA and Col1α1) and extracellular matrix remodelling. TCDD or PCB126 at measured concentrations did not elicit these responses in the 3D liver MTs system, though they caused cytotoxicity in HepaRG monoculture at high concentrations. Reduced human transcriptome (RHT) analysis captured molecular responses involved in liver fibrosis when MTs were treated with TAA and BaP. The results suggest that 3D, multicellular, human liver microtissues represent an alternative, human-relevant, in vitro liver model for assessing fibrotic pathways induced by environmental chemicals.

Noninvasive assessment of liver fibrosis in Crohn's disease patients exposed to methotrexate.

BACKGROUND: Methotrexate is widely used to treat some inflammatory chronic disorders, though it is hampered by the risk of liver fibrosis. Many recommendations have been made to assess methotrexate-related hepatotoxicity, including liver biopsy. However, other noninvasive methods to assess liver fibrosis have been developed and could be implemented for patients treated with methotrexate. AIM: The aim of the study was to compare the prevalence of liver fibrosis by means of noninvasive methods [aspartate transaminase-to-platelet ratio index (APRI) Forns index, and transient elastography] in patients with Crohn's disease exposed or not to methotrexate, and to identify risk factors for liver fibrosis. METHODS: Prospective, cross-sectional study. All patients with Crohn's disease exposed to methotrexate were included and compared to an unselected cohort of outpatients with Crohn's disease never exposed to methotrexate. RESULTS: A total of 84 patients with Crohn's disease, 56 exposed to methotrexate, and 28 controls, were included. Significant liver fibrosis was found in 7% of methotrexate-exposed patients with Crohn's disease and 10% of controls as measured by transient elastography, and in 7% of controls as measured by the Forns index. No cases of liver fibrosis were detected by APRI. Only alcohol consumption, diabetes mellitus, and age were associated with significant liver fibrosis. CONCLUSIONS: Significant liver fibrosis is uncommon among patients with Crohn's disease, even among those exposed to methotrexate. The risk of liver fibrosis in Crohn's disease seems to depend on common risk factors for liver disease.

Severity assessment in mice subjected to carbon tetrachloride.

The Directive 2010/63 EU requires classifying burden and severity in all procedures using laboratory animals. This study evaluated the severity of liver fibrosis induction by intraperitoneal carbon tetrachloride (CCl4) injections in mice. 29 male C57BL/6N mice were treated three times per week for 4 weeks with an intraperitoneal injection (50 µl) of either 0.6 ml/kg body weight CCl4-vehicle solution, germ oil (vehicle-control) or handling only. Severity assessment was performed using serum analysis, behavioral tests (open field test, rotarod, burrowing and nesting behavior), fecal corticosterone metabolite (FCM) measurement, and survival. The most significant group differences were noticed in the second week of treatment when the highest AST (1463 ± 1404 vs. 123.8 ± 93 U/L, p < 0.0001) and nesting values were measured. In addition, respective animals showed lower moving distances (4622 ± 1577 vs. 6157 ± 2060 cm, p < 0.01) and velocity in the Open field, identified as main factors in principal component analysis (PCA). Overall, a 50% survival rate was observed within the treatment group, in which the open field performance was a good tracer parameter for survival. In summary, this study demonstrates the feasibility of assessing severity in mice using behavioral tests and highlight the open field test as a possible threshold parameter for risk assessment of mortality.

Dynamic contrast-enhanced MRI with Gd-EOB-DTPA for the quantitative assessment of early-stage liver fibrosis induced by carbon tetrachloride in rabbits.

PURPOSE: To explore quantitative parameters obtained by dynamic contrast-enhanced magnetic resonance imaging (DCE MRI) with Gd-EOB-DTPA in discriminating early-stage liver fibrosis (LF) in a rabbit model. MATERIALS AND METHODS: LF was established in 60 rabbits by the injection of 50% CCl4 oil solution, whereas 30 rabbits served as the control group. All rabbits underwent pathological examination to determine the LF stage using the METAVIR classification system. DCE MRI was performed, and quantitative parameters, including Ktrans, Kep, Ve, Vp and Re were measured and evaluated among the different LF stages using spearman correlation coefficients and receiver operating characteristic curve. RESULTS: In all, 24, 25, and 22 rabbits had stage F0, stage F1, and stage F2 LF, respectively. Ktrans (r = 0.803) increased, and Kep (r = -0.495) and Re (r = -0.701) decreased with LF stage progression (P < 0.001), while no significant correlation was found for Ve or Vp. Ktrans and Re were significantly different between all LF stage pairs compared (F0 vs. F1, F0 vs. F2, F1 vs. F2, F0 vs. F1-F2, P < 0.05). With the exception of F0 vs. F1, Kep differed significantly between stages (P < 0.05). The AUC of Ktrans was higher than that of other quantitative parameters, with an AUC of 0.92, 0.99, 0.94 and 0.92 for staging F0 vs. F1, F0 vs. F2, F1 vs. F2, and F0 vs. F1-F2, respectively. CONCLUSION: Among quantitative parameters of Gd-EOB-DTPA DCE MRI, Ktrans was the best predictor for quantitatively differentiating early-stage LF.

Assessment of Hepatoprotective Effect of Chokeberry Juice in Rats Treated Chronically with Carbon Tetrachloride.

The aim of this study was to compare the protective effects of chokeberry juice and silymarin against chemical-induced liver fibrosis in rats. Liver fibrosis was induced by CCl4 administered two days a week for six weeks. Two groups of rats were co-treated with chokeberry juice, 10 mL/kg/day. or silymarin as a positive control, 100 mg/kg/day for six weeks. Hepatic lipid peroxidation was suppressed by 50% and the activity of hepatic antioxidant enzymes was increased by 19%-173% in rats co-treated with CCl4 and substances tested as compared to rats administered CCl4 alone. Hepatic hydroxyproline was decreased by 24% only in rats treated with silymarin. The messenger RNA (mRNA) expression levels of fibrosis-related molecules, procollagen I, α-SMA, TIMP-1, TGFß, and TNFα, which were significantly increased in the liver of CCl4-treated rats, were not modulated by substances tested. Histological evaluation revealed a slight protective effect of silymarin against fibrosis. However, in CCl4 + chokeberry-treated rats, the density of vacuolated hepatocytes was significantly lower than that in silymarin administered animals. Chokeberry juice did not demonstrate an antifibrotic effect in the applied experimental model of fibrosis, and the effect of the known antifibrotic agent, silymarin, was very limited.

Endogenous Fluorescence Dissimilarity Assessment of Four Potential Biomarkers of Early Liver Fibrosis by Preservation Media Effect.

Build-up of extracellular matrix in liver fibrosis results in changes on endogenous molecules expression that may be studied through the fluorescence characterization of ex vivo liver samples. To the best of our knowledge, no investigations have provided in-depth evidence and discussion on the changes of the endogenous fluorescence in ex vivo tissue due to the effects of the preservation media. In this work, we contrast and analyze the endogenous fluorescence from tryptophan, vitamin A, hydroxyproline and elastin cross-links potential biomarkers of the liver fibrosis, in in vivo measurements and liver samples preserved on formaldehyde, and two standard preservation media. As it is known, chemical changes in tissue, caused by formaldehyde fixation, alter the endogenous fluorescence spectra. We propose the use of phosphate-buffered saline (PBS), and Iscove's Modified Dulbecco's Medium (IMDM) to elude the fluorescence changes. PBS and IMDM showed to maintain the endogenous fluorescence characteristics similar to in vivo conditions. The results of this work point the way for a more reliable assessment of endogenous fluorescence in ex vivo hepatic studies.

A pragmatic non-invasive assessment of liver fibrosis in patients with psoriasis, rheumatoid arthritis or Crohn's disease receiving methotrexate therapy.

BACKGROUND AND AIMS: The reported hepatotoxicity of methotrexate underlines the need for a repeated non-invasive and reliable evaluation of liver fibrosis. We estimated, using a non-invasive strategy, the prevalence of significant liver fibrosis in patients treated by methotrexate and the predictors of significant fibrosis (fibrosis≥F2). METHODS: Fibrosis was prospectively evaluated using 9 non-invasive tests in consecutive patients with psoriasis, rheumatoid arthritis, or Crohn's disease. Significant fibrosis was assessed without liver biopsy by defining a "specific method" (result given by the majority of the tests) and a "sensitive method" (at least one test indicating a stage≥F2). RESULTS: One hundred and thirty-one patients (66 Psoriasis, 40 rheumatoid arthritis, and 25 Crohn's disease) were enrolled, including 83 receiving methotrexate. Seven tests were performed on average per patient, with a complete concordance in 75% of cases. Fibroscan® was interpretable in only 61% of patients. The best performances (AUROC>0.9) for predicting significant fibrosis were obtained by tests dedicated to steatohepatitis (FibroMeter NAFLD, NFS and FPI). The prevalence of fibrosis≥F2 according to the "specific" or the "sensitive" assessment of fibrosis was 10% and 28%, respectively. Methotrexate exposure did not influence the fibrosis stage. Factors independently associated with significant fibrosis according our "sensitive method" were age, male gender, and metabolic syndrome. CONCLUSION: We provided a non-invasive approach for identifying liver fibrosis≥F2 by using 8 biochemical tests and Fibroscan®. In this population, the risk of significant fibrosis was related to age, male gender, and presence of metabolic syndrome, but was not influenced by methotrexate.

B-mode ultrasound for the assessment of hepatic fibrosis: a quantitative multiparametric analysis for a radiomics approach.

Hepatic fibrosis and cirrhosis are a growing global health problem with increasing mortality rates. Early diagnosis and staging of hepatic fibrosis represent a major challenge. Currently liver biopsy is the gold standard for fibrosis assessment; however, biopsy requires an invasive procedure and is prone to sampling error and reader variability. In the current study we investigate using quantitative analysis of computer-extracted features of B-mode ultrasound as a non-invasive tool to characterize hepatic fibrosis. Twenty-two rats were administered diethylnitrosamine (DEN) orally for 12 weeks to induce hepatic fibrosis. Four control rats did not receive DEN. B-mode ultrasound scans sampling throughout the liver were acquired at baseline, 10, and 13 weeks. Computer extracted quantitative parameters representing brightness (echointensity, hepatorenal index) and variance (heterogeneity, anisotropy) of the liver were studied. DEN rats showed an increase in echointensity from 37.1 ± SD 7.8 to 53.5 ± 5.7 (10 w) to 57.5 ± 6.1 (13 w), while the control group remained unchanged at an average of 34.5 ± 4.5. The three other features studied increased similarly over time in the DEN group. Histologic analysis showed METAVIR fibrosis grades of F2-F4 in DEN rats and F0-F1 in controls. Increasing imaging parameters correlated with increasing METAVIR grades, and anisotropy showed the strongest correlation (ρ = 0.58). Sonographic parameters combined using multiparametric logistic regression were able to differentiate between clinically significant and insignificant fibrosis. Quantitative B-mode ultrasound imaging can be implemented in clinical settings as an accurate non-invasive tool for fibrosis assessment.

Assessment of the possible roles of SB-269970 versus ketanserin on carbon tetrachloride-induced liver fibrosis in rats: Oxidative stress/TGF-ß1-induced HSCs activation pathway.

BACKGROUND: In liver fibrosis, a major morbid and mortal disease, oxidative stress motivation of hepatic stellate cells (HSCs)-into myofibroblasts terminated in collagen deposition remain the key pathophysiological deal. Serotonin (5-HT) through its HSCs-expressed receptors, especially 5-HT2A and 7, shows crucial events in fibrogenesis of chronic liver diseases. Molecular hepatic oxidative stress-fibrotic roles of 5-HT2A and 7 receptors antagonists (ketanserin and SB-269970 respectively) are still a challenging issue. METHODS: Seven groups of adult male Wistar rats (n=10) were used. A carbon tetrachloride (CCl4) solution was injected intraperitoneally twice weekly for 6 weeks. On the 7th week, rats developed liver fibrosis were treated either by ketanserin (1mg/kg/day, ip) or SB-269970 (2mg/kg/day, ip) for 14days. Survival rates, and serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels in addition to hepatic malondialdehyde (MDA) and reduced glutathione (GSH) levels, superoxide dismutase (SOD) and catalase (CAT) activities, and transforming growth factor-beta1 (TGF-ß1) and procollagen type I N-terminal propeptide (PINP) levels, beside the hepatic histopathological fibrotic changes, were evaluated. RESULTS: In CCl4-challenged rats, each therapeutic approach showed significant reductions in elevated serum ALT, and AST levels, hepatic MDA, TGF-ß1, and PINP levels, and histopathological hepatic fibrotic scores as well as significant elevations in survival rates, reduced hepatic GSH levels, and SOD, and CAT activities. Remarkably, significant ameliorative measurements were observed in SB-269970 treated group. CONCLUSION: Blockade of 5-HT2A and 7 receptors each alone could be a future reliable therapeutic approach in liver fibrosis through a reduction in oxidative stress/TGF-ß1-induced HSCs activation pathway.

Iron overload by Superparamagnetic Iron Oxide Nanoparticles is a High Risk Factor in Cirrhosis by a Systems Toxicology Assessment.

Superparamagnetic iron oxide nanoparticles (SPIONs) as a contrast agent have been widely used in magnetic resonance imaging for tumor diagnosis and theranostics. However, there has been safety concern of SPIONs with cirrhosis related to excess iron-induced oxidative stress. In this study, the impact of iron overload by SPIONs was assessed on a mouse cirrhosis model. A single dose of SPION injection at 0.5 or 5 mg Fe/kg in the cirrhosis group induced a septic shock response at 24 h with elevated serum levels of liver and kidney function markers and extended impacts over 14 days including high levels of serum cholesterols and persistent low serum iron level. In contrast, full restoration of liver functions was found in the normal group with the same dosages over time. Analysis with PCR array of the toxicity pathways revealed the high dose of SPIONs induced significant expression changes of a distinct subset of genes in the cirrhosis liver. All these results suggested that excess iron of the high dose of SPIONs might be a risk factor for cirrhosis because of the marked impacts of elevated lipid metabolism, disruption of iron homeostasis and possibly, aggravated loss of liver functions.

In vivo assessment of liver fibrosis using diffuse reflectance and fluorescence spectroscopy: a proof of concept.

A novel application of diffuse reflectance and fluorescence spectroscopy in the assessment of liver fibrosis is here reported. To induce different stages of liver fibrosis, a sufficient number of male Wistar rats were differentially exposed to chronic administration with carbon tetrachloride. Then, diffuse reflectance and fluorescence spectra were in vivo measured from the liver surface of each animal by a minimal invasive laparoscopic procedure. The liver fibrosis degree was conventionally determined by means of histological examination using the Mason's Trichrome stain, accompanied by hepatic expression of α-sma, and evaluation of the ALT/AST serum levels. The liver from rats exhibiting higher grades of fibrosis showed a significant increase in diffuse reflectance and fluorescence intensity when compared with control animals. At 365 nm, the diffuse reflectance spectrum exhibited an increase of 4 and 3-fold in mild and advanced fibrotic rats, respectively, when compared to the control group. Similarly, the fluorescence emission at 493 nm was 2-fold higher in fibrotic animals than in controls. By using fluorescence intensity, discrimination algorithms indicated 73% sensitivity and 94% specificity for recognition of hepatic fibrosis, while for diffuse reflectance, these values increased up to 85% and 100%, respectively. Taking into consideration there is a special need for developing new diagnostic approaches focused on detecting different stages of liver fibrosis with minimal invasiveness, these results suggest that diffuse reflectance and fluorescence spectroscopy could be worthy of further exploration in patients with liver disease.

Assessment of liver fibrosis with transient elastography and FibroTest in patients treated with methotrexate for chronic inflammatory diseases: a case-control study.

BACKGROUND & AIMS: Although methotrexate (MTX) is used in the effective treatment of inflammatory disorders, its use is hampered by the risk of liver fibrosis. Non-invasive methods for the diagnosis of liver fibrosis, such as transient elastography (FibroScan) and FibroTest could be useful for monitoring MTX-liver toxicity. The aim of this case-control study was to determine factors associated with liver fibrosis in a large cohort of patients requiring MTX. METHODS: Consecutive adults with various benign inflammatory diseases were prospectively assessed using FibroScan and FibroTest when they were treated with MTX (cases) or before beginning treatment (controls). RESULTS: Among 518 included patients, 44 patients (8.5%) had FibroScan and/or FibroTest results suggesting severe liver fibrosis. In a multivariate analysis, factors associated with abnormal markers of liver fibrosis were the body mass index >28 kg/m(2) and high alcohol consumption. Neither long MTX duration nor cumulative doses were associated with elevated FibroScan or FibroTest results. CONCLUSIONS: Severe liver fibrosis is a rare event in patients treated with MTX and is probably unrelated to the total dose. Patients with other risk factors for liver disease should be closely monitored with non-invasive methods before and during MTX treatment.

Ethyl carbamate in alcoholic beverages from Mexico (tequila, mezcal, bacanora, sotol) and Guatemala (cuxa): market survey and risk assessment.

Ethyl carbamate (EC) is a recognized genotoxic carcinogen, with widespread occurrence in fermented foods and beverages. No data on its occurrence in alcoholic beverages from Mexico or Central America is available. Samples of agave spirits including tequila, mezcal, bacanora and sotol (n=110), and of the sugarcane spirit cuxa (n=16) were purchased in Mexico and Guatemala, respectively, and analyzed for EC. The incidence of EC contamination was higher in Mexico than in Guatemala, however, concentrations were below international guideline levels (<0.15 mg/L). Risk assessment found the Margin of Exposure (MOE) in line with that of European spirits. It is therefore unlikely that EC plays a role in high rates of liver cirrhosis reported in Mexico.