[MRI-Based Assessment of Hepatic Iron Overload and Steatosis in Patients Undergoing Hemodialysis:Current Status,Challenges,and Perspectives].

Long-term treatment of anemia involving frequent blood transfusions and intravenous iron administration increases the risks of hepatic iron overload and steatosis in the patients undergoing hemodialysis.Pathological accumulation of iron damages hepatocytes,not only elevating the risks of progressive hepatic fibrosis and cirrhosis but also potentially accelerating the process of hepatic steatosis.Iron overload and steatosis may interact with each other,exacerbating liver damage and ultimately leading to further deterioration of hepatic fibrosis and cirrhosis.MRI characterized by non-invasiveness and high repeatability,enables the simultaneous quantitative assessment of hepatic iron and fat content,providing crucial information for early diagnosis and intervention of liver diseases.In recent years,researchers have achieved significant advances in the application of MRI in the diagnosis and treatment of liver diseases.MRI can accurately reflect the extent of hepatic iron overload and steatosis in patients and predict the risk of liver diseases.This article reviews the latest advances,challenges,and perspectives in the application of MRI in assessing hepatic iron overload and steatosis in the patients undergoing hemodialysis,aiming to offer valuable references for clinical practice.

Pharmacological Prevention of Ectopic Erythrophagocytosis by Cilostazol Mitigates Ferroptosis in NASH.

Hepatic iron overload (HIO) is a hallmark of nonalcoholic fatty liver disease (NAFLD) with a poor prognosis. Recently, the role of hepatic erythrophagocytosis in NAFLD is emerging as a cause of HIO. We undertook various assays using human NAFLD patient pathology samples and an in vivo nonalcoholic steatohepatitis (NASH) mouse model named STAMTM. To make the in vitro conditions comparable to those of the in vivo NASH model, red blood cells (RBCs) and platelets were suspended and subjected to metabolic and inflammatory stresses. An insert-coculture system, in which activated THP-1 cells and RBCs are separated from HepG2 cells by a porous membrane, was also employed. Through various analyses in this study, the effect of cilostazol was examined. The NAFLD activity score, including steatosis, ballooning degeneration, inflammation, and fibrosis, was increased in STAMTM mice. Importantly, hemolysis occurred in the serum of STAMTM mice. Although cilostazol did not improve lipid or glucose profiles, it ameliorated hepatic steatosis and inflammation in STAMTM mice. Platelets (PLTs) played an important role in increasing erythrophagocytosis in the NASH liver. Upregulated erythrophagocytosis drives cells into ferroptosis, resulting in liver cell death. Cilostazol inhibited the augmentation of PLT and RBC accumulation. Cilostazol prevented the PLT-induced increase in ectopic erythrophagocytosis in in vivo and in vitro NASH models. Cilostazol attenuated ferroptosis of hepatocytes and phagocytosis of RBCs by THP-1 cells. Augmentation of hepatic erythrophagocytosis by activated platelets in NASH exacerbates HIO. Cilostazol prevents ectopic erythrophagocytosis, mitigating HIO-mediated ferroptosis in NASH models.

Association between hepatic iron overload assessed by magnetic resonance imaging and glucose intolerance states in the general population.

BACKGROUND AND AIM: While there is evidence that iron overload disorders are associated with type 2 diabetes, the relationship between hepatic iron overload and prediabetes remains unclear. We aimed to investigate the association between hepatic iron overload, as assessed by magnetic resonance imaging (MRI), and different glucose intolerance states in the population-based Study. METHODS AND RESULTS: We included data from 1622 individuals with MRI data, who did not have known type 2 diabetes (T2DM). Using an oral glucose tolerance testing, participants were classified as having isolated impaired fasting glucose (i-IFG), isolated impaired glucose tolerance (i-IGT), combined IFG and IGT (IFG + IGT) or previously unknown T2DM. Hepatic iron and fat contents were assessed through quantitative MRI. We undertook linear and multinomial logistic regression models adjusted for potential confounders and MRI-assessed hepatic fat content to examine the association of hepatic iron overload with different glucose intolerance states or continuous markers of glucose metabolism. MRI-assessed hepatic iron overload was positively associated only with both 2-h plasma glucose (ß = 0.32; 95%CI 0.04-0.60) and the combined IFG + IGT category (relative risk ratio = 1.87; 95%CI 1.15-3.06). No significant associations were found between hepatic iron overload and other glucose intolerance states or biomarkers of glucose metabolism, independently of potential confounders. CONCLUSIONS: MRI-assessed hepatic iron overload was associated with higher 2-h glucose concentrations and the combined IFG + IGT category, but not with other glucose intolerance states. Our findings suggest a weak adverse impact of hepatic iron overload on glucose metabolism, but further studies are needed to confirm these findings.

Hepatic STAMP2 alleviates polychlorinated biphenyl-induced steatosis and hepatic iron overload in NAFLD models.

Polychlorinated biphenyls (PCBs) have been associated with neurotoxicity, hepatoxicity, oncogenicity, and endocrine-disrupting effects. Although the recent studies have demonstrated that PCB exposure leads to nonalcoholic fatty liver disease (NAFLD), the underlying mechanism has remained unsolved. In this study, we examined the hepatic effects of a PCB mixture, Aroclor 1260, whose composition mimics human bioaccumulation patterns, and PCB 126 in C57BL/6 mice. Male C57Bl/6 mice were fed a standard diet or a 60% high-fat diet and exposed to Aroclor 1260 (10 mg/kg or 20 mg/kg) or PCB 126 (1 mg/kg or 5 mg/kg) by intraperitoneal injection for a total of four injections (2, 3, 4, and 5 weeks) for 6 weeks. In mice, both Aroclor 1260 and PCB 126-induced liver damage, hepatic steatosis and inflammation. We also observed that PCB exposure-induced hepatic iron overload (HIO). We previously demonstrated that hepatic six transmembrane protein of prostate 2 (STAMP2) may represent a suitable therapeutic target for NAFLD patients. Thus, we further examined whether hepatic STAMP2 is involved in PCB-induced NAFLD. We observed that hepatic STAMP2 was significantly decreased in PCB-induced NAFLD models in vivo and in vitro. Furthermore, overexpression of hepatic STAMP2 using an adenoviral delivery system resulted in improvement of PCB-induced steatosis and HIO in vivo and in vitro. Our findings indicate that enhancing hepatic STAMP2 expression represents a potential therapeutic avenue for the treatment of PCB exposure-induced NAFLD.

Recombinant FGF21 Attenuates Polychlorinated Biphenyl-Induced NAFLD/NASH by Modulating Hepatic Lipocalin-2 Expression.

Although recent studies have demonstrated that polychlorinated biphenyls (PCB) exposure leads to toxicant-associated steatohepatitis, the underlying mechanism of this condition remains unsolved. Male C57Bl/6 mice fed a standard diet (SD) or 60% high fat diet (HFD) were exposed to the nondioxin-like PCB mixture Aroclor1260 or dioxin-like PCB congener PCB126 by intraperitoneal injection for a total of four times for six weeks. We observed hepatic injury, steatosis, inflammation, and fibrosis in not only the Aroclor1260-treated mice fed a HFD but the PCB126-treated mice fed either a SD or a HFD. We also observed that both types of PCB exposure induced hepatic iron overload (HIO). Noticeably, the expression of hepatic lipocalin-2 (LCN2) was significantly increased in the PCB-induced nonalcoholic fatty liver disease (NAFLD)/nonalcoholic steatohepatitis (NASH) models. The knockdown of LCN2 resulted in improvement of PCB-induced lipid and iron accumulation in vitro, suggesting that LCN2 plays a pivotal role in PCB-induced NAFLD/NASH. We observed that recombinant FGF21 improved hepatic steatosis and HIO in the PCB-induced NAFLD/NASH models. Importantly, recombinant FGF21 reduced the PCB-induced overexpression of hepatic LCN2 in vivo and in vitro. Our findings indicate that recombinant FGF21 attenuates PCB-induced NAFLD/NASH by modulating hepatic lipocalin-2 expression. Our data suggest that hepatic LCN2 might represent a suitable therapeutic target for improving PCB-induced NAFLD/NASH accompanying HIO.

Free-breathing R2∗ mapping of hepatic iron overload in children using 3D multi-echo UTE cones MRI.

PURPOSE: To enable motion-robust, ungated, free-breathing R2∗ mapping of hepatic iron overload in children with 3D multi-echo UTE cones MRI. METHODS: A golden-ratio re-ordered 3D multi-echo UTE cones acquisition was developed with chemical-shift encoding (CSE). Multi-echo complex-valued source images were reconstructed via gridding and coil combination, followed by confounder-corrected R2∗ (=1/ T2∗ ) mapping. A phantom containing 15 different concentrations of gadolinium solution (0-300 mM) was imaged at 3T. 3D multi-echo UTE cones with an initial TE of 0.036 ms and Cartesian CSE-MRI (IDEAL-IQ) sequences were performed. With institutional review board approval, 85 subjects (81 pediatric patients with iron overload + 4 healthy volunteers) were imaged at 3T using 3D multi-echo UTE cones with free breathing (FB cones), IDEAL-IQ with breath holding (BH Cartesian), and free breathing (FB Cartesian). Overall image quality of R2∗ maps was scored by 2 blinded experts and compared by a Wilcoxon rank-sum test. For each pediatric subject, the paired R2∗ maps were assessed to determine if a corresponding artifact-free 15 mm region-of-interest (ROI) could be identified at a mid-liver level on both images. Agreement between resulting R2∗ quantification from FB cones and BH/FB Cartesian was assessed with Bland-Altman and linear correlation analyses. RESULTS: ROI-based regression analysis showed a linear relationship between gadolinium concentration and R2∗ in IDEAL-IQ (y = 8.83x - 52.10, R2 = 0.995) as well as in cones (y = 9.19x - 64.16, R2 = 0.992). ROI-based Bland-Altman analysis showed that the mean difference (MD) was 0.15% and the SD was 5.78%. However, IDEAL-IQ R2∗ measurements beyond 200 mM substantially deviated from a linear relationship for IDEAL-IQ (y = 5.85x + 127.61, R2 = 0.827), as opposed to cones (y = 10.87x - 166.96, R2 = 0.984). In vivo, FB cones R2∗ had similar image quality with BH and FB Cartesian in 15 and 42 cases, respectively. FB cones R2∗ had better image quality scores than BH and FB Cartesian in 3 and 21 cases, respectively, where BH/FB Cartesian exhibited severe ghosting artifacts. ROI-based Bland-Altman analyses were 2.23% (MD) and 6.59% (SD) between FB cones and BH Cartesian and were 0.21% (MD) and 7.02% (SD) between FB cones and FB Cartesian, suggesting a good agreement between FB cones and BH (FB) Cartesian R2∗ . Strong linear relationships were observed between BH Cartesian and FB cones (y = 1.00x + 1.07, R2 = 0.996) and FB Cartesian and FB cones (y = 0.98x + 1.68, R2 = 0.999). CONCLUSION: Golden-ratio re-ordered 3D multi-echo UTE Cones MRI enabled motion-robust, ungated, and free-breathing R2∗ mapping of hepatic iron overload, with comparable R2∗ measurements and image quality to BH Cartesian, and better image quality than FB Cartesian.

Orphan Nuclear Receptor ERRγ Is a Transcriptional Regulator of CB1 Receptor-Mediated TFR2 Gene Expression in Hepatocytes.

Orphan nuclear receptor estrogen-related receptor γ (ERRγ) is an important transcription factor modulating gene transcription involved in endocrine control of liver metabolism. Transferrin receptor 2 (TFR2), a carrier protein for transferrin, is involved in hepatic iron overload in alcoholic liver disease (ALD). However, TFR2 gene transcriptional regulation in hepatocytes remains largely unknown. In this study, we described a detailed molecular mechanism of hepatic TFR2 gene expression involving ERRγ in response to an endocannabinoid 2-arachidonoylglycerol (2-AG). Treatment with 2-AG and arachidonyl-2'-chloroethylamide, a selective cannabinoid receptor type 1 (CB1) receptor agonist, increased ERRγ and TFR2 expression in hepatocytes. Overexpression of ERRγ was sufficient to induce TFR2 expression in both human and mouse hepatocytes. In addition, ERRγ knockdown significantly decreased 2-AG or alcohol-mediated TFR2 gene expression in cultured hepatocytes and mouse livers. Finally, deletion and mutation analysis of the TFR2 gene promoter demonstrated that ERRγ directly modulated TFR2 gene transcription via binding to an ERR-response element. This was further confirmed by chromatin immunoprecipitation assay. Taken together, these results reveal a previously unrecognized role of ERRγ in the transcriptional regulation of TFR2 gene expression in response to alcohol.

Ultrashort echo time imaging for quantification of hepatic iron overload: Comparison of acquisition and fitting methods via simulations, phantoms, and in vivo data.

BACKGROUND: Current R2*-MRI techniques for measuring hepatic iron content (HIC) use various acquisition types and fitting models. PURPOSE: To evaluate the accuracy and precision of R2*-HIC acquisition and fitting methods. STUDY TYPE: Signal simulations, phantom study, and prospective in vivo cohort. POPULATION: In all, 132 patients (58/74 male/female, mean age 17.7 years). FIELD STRENGTH/SEQUENCE: 2D-multiecho gradient-echo (GRE) and ultrashort echo time (UTE) acquisitions at 1.5T. ASSESSMENT: Synthetic MR signals were created to mimic published GRE and UTE methods, using different R2* values (25-2000 s-1 ) and signal-to-noise ratios (SNR). Phantoms with varying iron concentrations were scanned at 1.5T. In vivo data were analyzed from 132 patients acquired at 1.5T. R2* was estimated by fitting using three signal models. Accuracy and precision of R2* measurements for UTE acquisition parameters (SNR, echo spacing [ΔTE], maximum echo time [TEmax ]) and fitting methods were compared for simulated, phantom, and in vivo datasets. STATISTICAL TESTS: R2* accuracy was determined from the relative error and by linear regression analysis. Precision was evaluated using coefficient of variation (CoV) analysis. RESULTS: In simulations, all models had high R2* accuracy (error <5%) and precision (CoV <10%) for all SNRs, shorter ΔTE (≤0.5 msec), and longer TEmax (≥10.1 msec); except the constant offset model overestimated R2* at the lowest SNR. In phantoms and in vivo, all models produced similar R2* values for different SNRs and shorter ΔTEs (slopes: 0.99-1.06, R2 > 0.99, P < 0.001). In all experiments, R2* results degraded for high R2* values with longer ΔTE (≥1 msec). In vivo, shorter and longer TEmax gave similar R2* results (slopes: 1.02-1.06, R2 > 0.99, P < 0.001) for the noise subtraction model for 25≤R2*≤2000 s-1 . However, both quadratic and constant offset models, using shorter TEmax (≤4.7 msec) overestimated R2* and yielded high CoVs up to ∼170% for low R2* (<250 s-1 ). DATA CONCLUSION: UTE with TEmax ≥ 10.1 msec and ΔTE ≤ 0.5 msec yields accurate R2* estimates over the entire clinical HIC range. Monoexponential fitting with noise subtraction is the most robust signal model to changes in UTE parameters and achieves the highest R2* accuracy and precision. LEVEL OF EVIDENCE: 2 Technical Efficacy: Stage 2 J. Magn. Reson. Imaging 2019;49:1475-1488.

Autoregressive moving average modeling for hepatic iron quantification in the presence of fat.

BACKGROUND: Measuring hepatic R2* by fitting a monoexponential model to the signal decay of a multigradient-echo (mGRE) sequence noninvasively determines hepatic iron content (HIC). Concurrent hepatic steatosis introduces signal oscillations and confounds R2* quantification with standard monoexponential models. PURPOSE: To evaluate an autoregressive moving average (ARMA) model for accurate quantification of HIC in the presence of fat using biopsy as the reference. STUDY TYPE: Phantom study and in vivo cohort. POPULATION: Twenty iron-fat phantoms covering clinically relevant R2* (30-800 s-1 ) and fat fraction (FF) ranges (0-40%), and 10 patients (four male, six female, mean age 18.8 years). FIELD STRENGTH/SEQUENCE: 2D mGRE acquisitions at 1.5 T and 3 T. ASSESSMENT: Phantoms were scanned at both field strengths. In vivo data were analyzed using the ARMA model to determine R2* and FF values, and compared with biopsy results. STATISTICAL TESTS: Linear regression analysis was used to compare ARMA R2* and FF results with those obtained using a conventional monoexponential model, complex-domain nonlinear least squares (NLSQ) fat-water model, and biopsy. RESULTS: In phantoms and in vivo, all models produced R2* and FF values consistent with expected values in low iron and low/high fat conditions. For high iron and no fat phantoms, monoexponential and ARMA models performed excellently (slopes: 0.89-1.07), but NLSQ overestimated R2* (slopes: 1.14-1.36) and produced false FFs (12-17%) at 1.5 T; in high iron and fat phantoms, NLSQ (slopes: 1.02-1.16) outperformed monoexponential and ARMA models (slopes: 1.23-1.88). The results with NLSQ and ARMA improved in phantoms at 3 T (slopes: 0.96-1.04). In patients, mean R2*-HIC estimates for monoexponential and ARMA models were close to biopsy-HIC values (slopes: 0.90-0.95), whereas NLSQ substantially overestimated HIC (slope 1.4) and produced false FF values (4-28%) with very high SDs (15-222%) in patients with high iron overload and no steatosis. DATA CONCLUSION: ARMA is superior in quantifying R2* and FF under high iron and no fat conditions, whereas NLSQ is superior for high iron and concurrent fat at 1.5 T. Both models give improved R2* and FF results at 3 T. LEVEL OF EVIDENCE: 2 Technical Efficacy Stage: 2 J. Magn. Reson. Imaging 2019;50:1620-1632.

Radial Ultrashort TE Imaging Removes the Need for Breath-Holding in Hepatic Iron Overload Quantification by R2* MRI.

OBJECTIVE: The objective of this study is to evaluate radial free-breathing (FB) multiecho ultrashort TE (UTE) imaging as an alternative to Cartesian FB multiecho gradient-recalled echo (GRE) imaging for quantitative assessment of hepatic iron content (HIC) in sedated patients and subjects unable to perform breath-hold (BH) maneuvers. MATERIALS AND METHODS: FB multiecho GRE imaging and FB multiecho UTE imaging were conducted for 46 test group patients with iron overload who could not complete BH maneuvers (38 patients were sedated, and eight were not sedated) and 16 control patients who could complete BH maneuvers. Control patients also underwent standard BH multiecho GRE imaging. Quantitative R2* maps were calculated, and mean liver R2* values and coefficients of variation (CVs) for different acquisitions and patient groups were compared using statistical analysis. RESULTS: FB multiecho GRE images displayed motion artifacts and significantly lower R2* values, compared with standard BH multiecho GRE images and FB multiecho UTE images in the control cohort and FB multiecho UTE images in the test cohort. In contrast, FB multiecho UTE images produced artifact-free R2* maps, and mean R2* values were not significantly different from those measured by BH multiecho GRE imaging. Motion artifacts on FB multiecho GRE images resulted in an R2* CV that was approximately twofold higher than the R2* CV from BH multiecho GRE imaging and FB multiecho UTE imaging. The R2* CV was relatively constant over the range of R2* values for FB multiecho UTE, but it increased with increases in R2* for FB multiecho GRE imaging, reflecting that motion artifacts had a stronger impact on R2* estimation with increasing iron burden. CONCLUSION: FB multiecho UTE imaging was less motion sensitive because of radial sampling, produced excellent image quality, and yielded accurate R2* estimates within the same acquisition time used for multiaveraged FB multiecho GRE imaging. Thus, FB multiecho UTE imaging is a viable alternative for accurate HIC assessment in sedated children and patients who cannot complete BH maneuvers.

Does fat suppression via chemically selective saturation affect R2*-MRI for transfusional iron overload assessment? A clinical evaluation at 1.5T and 3T.

PURPOSE: Fat suppression (FS) via chemically selective saturation (CHESS) eliminates fat-water oscillations in multiecho gradient echo (mGRE) R2*-MRI. However, for increasing R2* values as seen with increasing liver iron content (LIC), the water signal spectrally overlaps with the CHESS band, which may alter R2*. We investigated the effect of CHESS on R2* and developed a heuristic correction for the observed CHESS-induced R2* changes. METHODS: Eighty patients [female, n = 49; male, n = 31; mean age (± standard deviation), 18.3 ± 11.7 y] with iron overload were scanned with a non-FS and a CHESS-FS mGRE sequence at 1.5T and 3T. Mean liver R2* values were evaluated using three published fitting approaches. Measured and model-corrected R2* values were compared and statistically analyzed. RESULTS: At 1.5T, CHESS led to a systematic R2* reduction (P < 0.001 for all fitting algorithms) especially toward higher R2*. Our model described the observed changes well and reduced the CHESS-induced R2* bias after correction (linear regression slopes: 1.032/0.927/0.981). No CHESS-induced R2* reductions were found at 3T. CONCLUSION: The CHESS-induced R2* bias at 1.5T needs to be considered when applying R2*-LIC biopsy calibrations for clinical LIC assessment, which were established without FS at 1.5T. The proposed model corrects the R2* bias and could therefore improve clinical iron overload assessment based on linear R2*-LIC calibrations. Magn Reson Med 76:591-601, 2016. © 2015 Wiley Periodicals, Inc.

Identification of transferrin receptor 1 as a hepatitis C virus entry factor.

Hepatitis C virus (HCV) is a liver tropic pathogen that affects ∼170 million people worldwide and causes liver pathologies including fibrosis, cirrhosis, steatosis, iron overload, and hepatocellular carcinoma. As part of a project initially directed at understanding how HCV may disrupt cellular iron homeostasis, we found that HCV alters expression of the iron uptake receptor transferrin receptor 1 (TfR1). After further investigation, we found that TfR1 mediates HCV entry. Specifically, functional studies showed that TfR1 knockdown and antibody blocking inhibit HCV cell culture (HCVcc) infection. Blocking cell surface TfR1 also inhibited HCV pseudoparticle (HCVpp) infection, demonstrating that TfR1 acts at the level of HCV glycoprotein-dependent entry. Likewise, a TfR1 small-molecule inhibitor that causes internalization of surface TfR1 resulted in a decrease in HCVcc and HCVpp infection. In kinetic studies, TfR1 antibody blocking lost its inhibitory activity after anti-CD81 blocking, suggesting that TfR1 acts during HCV entry at a postbinding step after CD81. In contrast, viral spread assays indicated that HCV cell-to-cell spread is less dependent on TfR1. Interestingly, silencing of the TfR1 trafficking protein, a TfR-1 specific adaptor protein required for TfR1 internalization, also inhibited HCVcc infection. On the basis of these results, we conclude that TfR1 plays a role in HCV infection at the level of glycoprotein-mediated entry, acts after CD81, and possibly is involved in HCV particle internalization.

Spike-in SILAC proteomic approach reveals the vitronectin as an early molecular signature of liver fibrosis in hepatitis C infections with hepatic iron overload.

Hepatitis C virus (HCV)-induced iron overload has been shown to promote liver fibrosis, steatosis, and hepatocellular carcinoma. The zonal-restricted histological distribution of pathological iron deposits has hampered the attempt to perform large-scale in vivo molecular investigations on the comorbidity between iron and HCV. Diagnostic and prognostic markers are not yet available to assess iron overload-induced liver fibrogenesis and progression in HCV infections. Here, by means of Spike-in SILAC proteomic approach, we first unveiled a specific membrane protein expression signature of HCV cell cultures in the presence of iron overload. Computational analysis of proteomic dataset highlighted the hepatocytic vitronectin expression as the most promising specific biomarker for iron-associated fibrogenesis in HCV infections. Next, the robustness of our in vitro findings was challenged in human liver biopsies by immunohistochemistry and yielded two major results: (i) hepatocytic vitronectin expression is associated to liver fibrogenesis in HCV-infected patients with iron overload; (ii) hepatic vitronectin expression was found to discriminate also the transition between mild to moderate fibrosis in HCV-infected patients without iron overload.

Iron-Induced Hepatocarcinogenesis-Preventive Effects of Nutrients.

The liver is a primary organ that stores body iron, and plays a central role in the regulation of iron homeostasis. Hepatic iron overload (HIO) is a prevalent feature among patients with chronic liver diseases (CLDs), including alcoholic/nonalcoholic liver diseases and hepatitis C. HIO is suggested to promote the progression toward hepatocellular carcinoma because of the pro-oxidant nature of iron. Iron metabolism is tightly regulated by various factors, such as hepcidin and ferroportin, in healthy individuals to protect the liver from such deteriorative effects. However, their intrinsic expressions or functions are frequently compromised in patients with HIO. Thus, various nutrients have been reported to regulate hepatic iron metabolism and protect the liver from iron-induced damage. These nutrients are beneficial in HIO-associated CLD treatment and eventually prevent iron-mediated hepatocarcinogenesis. This mini-review aimed to discuss the mechanisms and hepatocarcinogenic risk of HIO in patients with CLDs. Moreover, nutrients that hold the potential to prevent iron-induced hepatocarcinogenesis are summarized.

Active transport nanochelators for the reduction of liver iron burden in iron overload.

Liver dysfunction and failure account for a major portion of premature deaths in patients suffering from various iron associated pathogeneses, particularly primary and secondary iron overload disorders, despite intensive treatment. The liver is a central player in iron homeostasis and a major iron storage organ, and currently, there are no active approaches for the excretion of excess liver iron. Herein, we report a new method for the rapid reduction of iron burden in iron overload diseases by developing a new class of liver targeted nanochelators with favorable pharmacokinetics and biodistribution. The new nanochelators bypass the reticuloendothelial system and specifically target hepatocytes without non-specific accumulation in other organs. The targeted nanochelators bound and neutralized excess iron in the liver and from the vasculature and, eventually leading to rapid hepatobiliary excretion of labile iron. Further, these rapidly excreted nanochelators did not induce toxicity in the liver, were highly cytocompatible in both iron overload and non-loaded conditions, and were promising in mitigating iron triggered free radical oxidative damage. These studies provide key insights into the development of organ targeted nanochelating systems and the rapid reduction of iron burden in vivo. This methodology allows for further development of nanotherapeutics for specific iron overload diseases.

Dual gradient echo in-phase and out of phase sequences in assessment of hepatic iron overload in patients with beta-thalassemia, would be better?

PURPOSE: To evaluate the diagnostic accuracy of the dual gradient-echo (GRE) in- and out-of-phase sequences as a quantitative tool for hepatic iron overload in comparison with MRI R2* relaxometry in paediatric patients with beta-thalassemia. METHOD: Sixty-three patients with beta-thalassemia major (transfusion-dependent) or beta-thalassemia intermedia (transfusion- and non-transfusion-dependent) were referred from the paediatric department (haematology unit) to the radiology department at a university hospital. The paediatrician conducted a clinical examination for the studied group, assessed their laboratory data, conducted R2* relaxometry and dual gradient echo sequences to calculate R2* and relative signal intensity index at the axial mid-section of the liver, and studied their correlation. A 1.5 Tesla MR scanner was used (Achieva; Philips Medical Systems, the Netherlands). Data were fed to the computer and analysed using the IBM SPSS software package version 20.0 (Armonk, NY: IBM Corp). The Kolmogorov-Smirnov test was used to verify the normality of distribution. The significance of the results was determined at the 5% level. The Chi-square, Fisher's exact correction, Pearson coefficient, and Bland-Altman tests were used. RESULTS: Dual gradient-echo in- and out-of-phase sequences using visual assessment accurately assessed 93.65% of our patient group with hepatic iron overload. A significant correlation was found between the relative signal intensity index and hepatic MRI R2* relaxometry (p < 0.001, r = 0.861). CONCLUSIONS: Dual gradient-echo in and out-of-phase sequences are good imaging tools for hepatic iron detection and quantification. These sequences showed good correlation with R2* relaxometry (r = 0.861, p < 0.001).

Iron oxide nanoparticles aggravate hepatic steatosis and liver injury in nonalcoholic fatty liver disease through BMP-SMAD-mediated hepatic iron overload.

Nonalcoholic fatty liver disease (NAFLD) is the leading hepatic manifestation of metabolic syndrome worldwide, and is clinically accompanied by iron overload. As the increasing application of iron oxide nanoparticles (IONPs) on the imaging and diagnosis in NAFLD, the potential hepatic effect and mechanism of IONPs on NAFLD should be well studied. Here, we demonstrate that carboxyl-modified (COOH-IONPs) and amino-coated IONPs (NH2-IONPs) exhibit no significant hepatic toxicity in normal mice at the clinical injection dose, but aggravate SREBP-1c-mediated de novo lipogenesis (DNL) in the livers of mice with NAFLD induced by high-fat diet (HFD) and in HepG2 cells incubated with oleic acid (OA), especially in those treated by the positive NH2-IONPs. In the present study, mice receiving IONPs for 7 day show mild iron overload in the liver and exhibit enhanced hepatic inflammation in NAFLD. The BMP-SMAD pathway is initiated by hepatic iron overload and is aggravated in NAFLD. In conclusion, BMP-SMAD-mediated hepatic iron overload aggravated lipid accumulation in the liver and hepatic inflammatory responses, implying that effective measures in addition to hepatic iron overload are needed for individuals at the risk of IONPs in NAFLD.

Determination of Non-Invasive Biomarkers for the Assessment of Fibrosis, Steatosis and Hepatic Iron Overload by MR Image Analysis. A Pilot Study.

The reference diagnostic test of fibrosis, steatosis, and hepatic iron overload is liver biopsy, a clear invasive procedure. The main objective of this work was to propose HSA, or human serum albumin, as a biomarker for the assessment of fibrosis and to study non-invasive biomarkers for the assessment of steatosis and hepatic iron overload by means of an MR image acquisition protocol. It was performed on a set of eight subjects to determine fibrosis, steatosis, and hepatic iron overload with four different MRI sequences. We calibrated longitudinal relaxation times (T1 [ms]) with seven human serum albumin (HSA [%]) phantoms, and we studied the relationship between them as this protein is synthesized by the liver, and its concentration decreases in advanced fibrosis. Steatosis was calculated by means of the fat fraction (FF [%]) between fat and water liver signals in "fat-only images" (the subtraction of in-phase [IP] images and out-of-phase [OOP] images) and in "water-only images" (the addition of IP and OOP images). Liver iron concentration (LIC [µmol/g]) was obtained by the transverse relaxation time (T2* [ms]) using Gandon's method with multiple echo times (TE) in T2-weighted IP and OOP images. The preliminary results showed that there is an inverse relationship (r = -0.9662) between the T1 relaxation times (ms) and HSA concentrations (%). Steatosis was determined with FF > 6.4% and when the liver signal was greater than the paravertebral muscles signal, and thus, the liver appeared hyperintense in fat-only images. Hepatic iron overload was detected with LIC > 36 µmol/g, and in these cases, the liver signal was smaller than the paravertebral muscles signal, and thus, the liver behaved as hypointense in IP images.

Dietary Iron Overload Differentially Modulates Chemically-Induced Liver Injury in Rats.

Hepatic iron overload is well known as an important risk factor for progression of liver diseases; however, it is unknown whether it can alter the susceptibility to drug-induced hepatotoxicity. Here we investigate the pathological roles of iron overload in two single-dose models of chemically-induced liver injury. Rats were fed a high-iron (Fe) or standard diet (Cont) for four weeks and were then administered with allyl alcohol (AA) or carbon tetrachloride (CCl4). Twenty-four hours after administration mild mononuclear cell infiltration was seen in the periportal/portal area (Zone 1) in Cont-AA group, whereas extensive hepatocellular necrosis was seen in Fe-AA group. Centrilobular (Zone 3) hepatocellular necrosis was prominent in Cont-CCl4 group, which was attenuated in Fe-CCl4 group. Hepatic lipid peroxidation and hepatocellular DNA damage increased in Fe-AA group compared with Cont-AA group. Hepatic caspase-3 cleavage increased in Cont-CCl4 group, which was suppressed in Fe-CCl4 group. Our results showed that dietary iron overload exacerbates AA-induced Zone-1 liver injury via enhanced oxidative stress while it attenuates CCl4-induced Zone-3 liver injury, partly via the suppression of apoptosis pathway. This study suggested that susceptibility to drugs or chemical compounds can be differentially altered in iron-overloaded livers.

Reduction of Liver Iron Load in Adult Patients with ß-Thalassemia Major Treated with Modern Chelation Modalities.

BACKGROUND: Management of beta-thalassemia major (TM) requires life-long hemotransfusions leading to iron overload. Iron elimination is enhanced by the use of modern chelators. AIM: To assess the effect of modern chelation therapy by dynamics of serum ferritin concentration and liver MRI T2*. PATIENTS AND METHODS: Forty-six patients with TM (male to female ratio =1:1, mean age 33.2±10.9 years) were prospectively studied between 2011 and 2014. Twenty-one patients (45.7%) were treated with deferasirox, 17 (37%) - with deferiprone, and 8 (17.3%) - with deferiprone in combination with deferoxamine. Ferritin was measured by ELISA. MRI T2* was assessed by Siemens Magnetom Avanto 1.5T. The patients were allocated into 3 groups based on their initial ferritin level and liver MRI T2*. Statistical analysis was performed using SPSS v. 18 for Windows. Data were analysed by descriptive analysis, analysis of variance and correlative analysis, means were compared using t-test and one-way ANOVA. RESULTS: In 2011, 9 (19.5%) patients had normal liver MRI T2*; in 2014 they were 17 (37%). The patients with mild grade liver siderosis were 12 (26%) in 2011, and in 2014 they were 14 (30.4%). In 2011, the patients with moderate liver siderosis were 14 (30.4%), and in 2014 - 12 (26.0%). Eleven patients (23.9%) had severe liver siderosis in 2011 and only two patients (4.0%) were diagnosed with the condition in 2014. CONCLUSION: A reduction of iron overload was found in all studied groups. This positive effect is attributed to the use of modern chelators and the ease of access to accurate monitoring.