Iron Grading Research Articles

Diagnostic value of T2 relaxation time for hepatic iron grading in rat model of fatty and fibrotic liver.

The objective of this study was to assess the quantitative diagnostic value of T2 relaxation time for determining liver iron grades in the presence of fat and fibrosis. Sixty Sprague-Dawley (SD) male rats were randomly divided into control (10 rats) and model (50 rats) groups. The model group of coexisting iron, steatosis, and liver fibrosis was induced by intraperitoneal injection of carbon tetrachloride (CCl4) dissolved in edible vegetable oil (40% v/v). The control group received an intraperitoneal injection of 0.9% saline. All rats underwent multi-echo gradient and spin echo (M-GRASE) magnetic resonance imaging, and the T2 relaxation time of the liver was measured. The rats were killed immediately after imaging, and liver specimens were extracted for histological evaluation of steatosis, iron, and fibrosis. The relationship and differences between T2 relaxation time and liver fibrosis stage, as well as the pathological grade of hepatic steatosis, were assessed by Spearman's rank correlation coefficient, non-parametric Mann-Whitney test, and the Kruskal-Wallis test. The area under the receiver operating characteristic curve and interaction analysis were used to quantify the diagnostic performance of T2 relaxation time for detecting different degrees of liver iron grades. Six normal control rats and 34 model rats were included in this study. Fibrosis stages were F0 (n = 6), F1 (n = 6), F2 (n = 8), F3 (n = 10), and F4 (n = 10). Steatosis grades were S0 (n = 5), S1 (n = 8), S2 (n = 12), and S3 (n = 15). Hepatocyte or Kupffer cell iron grades were 0 (n = 7), 1 (n = 9), 2 (n = 12), 3 (n = 10), and 4 (n = 2). The liver fibrosis stages were positively correlated with the iron grades (P < 0.01), and the iron grades and fibrosis stages were negatively correlated with the T2 relaxation time (P < 0.01). The T2 relaxation times exhibited strongly significant differences among rats with different histologically determined iron grades (P < 0.01). Pairwise comparisons between each grade of liver iron indicated significant differences between all iron grades, except between grades 0 and 1, and between grades 1 and 2 (P > 0.05). The T2 relaxation time of the liver had an area under the receiving operating characteristic curve (AUC) of 0.965 (95% CI 0.908-0.100, P < 0.001) for distinguishing rats with a pathological grade of hepatic iron (grade ≥ 1) from those without, an AUC of 0.871 (95% CI 0.757-0.985, P < 0.001) for distinguishing rats with no iron overload (grade ≤ 1) from rats with moderate or severe iron overload (grade ≥ 2), and an AUC of 0.939 (95% CI 0.865-1.000, P < 0.001) for distinguishing rats with no to moderate iron overload (grade ≤ 2) from rats with severe iron overload (grade 3). The interaction of different pathological grades of iron, steatosis, and fibrosis has a negligible influence on the T2 relaxation time (P > 0.05). In conclusion, T2 relaxation time can assess histologically determined liver iron grades, regardless of coexisting liver steatosis or fibrosis; therefore, it is suitable for distinguishing between the presence and absence of iron deposition and it is more accurate for higher iron grading.

A study of Iron store in Bone marrow and its correlation in various anemia

incidence of anaemia is a very common but prevalence of anaemia is disproportionately high in developing countries. With proper clinical history, haematological and bone marrow examination findings proper evaluation and management of patients with anaemia can be made. Microscopic examination of bone marrow aspirate is the gold standard for assessing marrow iron store. However, conventional Gale’s method assesses iron in marrow fragments alone which provides little valuable information about functional iron deficiency seen in many chronic diseases. Thisstudy is conducted to measure iron grading using perl’s Prussian blue staining in various hematological conditions. A descriptive study of Perl’s Prussian blue stained bone marrow aspirate smears of 130 patients with anaemia. A CBC, Reticulocyte count and Bone marrow iron was assessed by the Gale’s method in this study.After excluding 9 cases of dry tap, 121 cases were evaluated in our study. 78.57% of cases (61 cases) diagnosedas microcytic anaemia had iron store very low (grade 0 and 1). In patients with megaloblastic anaemia (15 cases), 46.7% of cases (7 cases) had iron store grade was normal, but in 8 cases iron grade was low. In patients with dimorphic anaemia (23 cases), 78.3% of cases (18 cases) had iron store grade was normal.Bone marrow aspiration with application of Prussian blue stain is useful to investigate patients with suspected iron deficiency anaemia, anaemia of chronic disease, megaloblastic anaemia and acute leukemia.

Automated Whole-Liver MRI Segmentation to Assess Steatosis and Iron Quantification in Chronic Liver Disease.

Background Standardized manual region of interest (ROI) sampling strategies for hepatic MRI steatosis and iron quantification are time consuming, with variable results. Purpose To evaluate the performance of automatic MRI whole-liver segmentation (WLS) for proton density fat fraction (PDFF) and iron estimation (transverse relaxometry [R2*]) versus manual ROI, with liver biopsy as the reference standard. Materials and Methods This prospective, cross-sectional, multicenter study recruited participants with chronic liver disease who underwent liver biopsy and chemical shift-encoded 3.0-T MRI between January 2017 and January 2021. Biopsy evaluation included histologic grading and digital pathology. MRI liver sampling strategies included manual ROI (two observers) and automatic whole-liver (deep learning algorithm) segmentation for PDFF- and R2*-derived measurements. Agreements between segmentation methods were measured using intraclass correlation coefficients (ICCs), and biases were evaluated using Bland-Altman analyses. Linear regression analyses were performed to determine the correlation between measurements and digital pathology. Results A total of 165 participants were included (mean age ± standard deviation, 55 years ± 12; 96 women; 101 of 165 participants [61%] with nonalcoholic fatty liver disease). Agreements between mean measurements were excellent, with ICCs of 0.98 for both PDFF and R2*. The median bias was 0.5% (interquartile range, -0.4% to 1.2%) for PDFF and 2.7 sec-1 (interquartile range, 0.2-5.3 sec-1) for R2* (P < .001 for both). Margins of error were lower for WLS than ROI-derived parameters (-0.03% for PDFF and -0.3 sec-1 for R2*). ROI and WLS showed similar performance for steatosis (ROI AUC, 0.96; WLS AUC, 0.97; P = .53) and iron overload (ROI AUC, 0.85; WLS AUC, 0.83; P = .09). Correlations with digital pathology were high (P < .001) between the fat ratio and PDFF (ROI r = 0.89; WLS r = 0.90) and moderate (P < .001) between the iron ratio and R2* (ROI r = 0.65; WLS r = 0.64). Conclusion Proton density fat fraction and transverse relaxometry measurements derived from MRI automatic whole-liver segmentation (WLS) were accurate for steatosis and iron grading in chronic liver disease and correlated with digital pathology. Automated WLS estimations were higher, with a lower margin of error than manual region of interest estimations. © RSNA, 2021 Online supplemental material is available for this article. See also the editorial by Moura Cunha and Fowler in this issue.

Digital pathology: accurate technique for quantitative assessment of histological features in metabolic-associated fatty liver disease.

Histological evaluation of metabolic-associated fatty liver disease (MAFLD) biopsies is subjective, descriptive and with interobserver variability. To examine the relationship between different histological features (fibrosis, steatosis, inflammation and iron) measured with automated whole-slide quantitative digital pathology and corresponding semiquantitative scoring systems, and the distribution of digital pathology measurements across Fatty Liver Inhibition of Progression (FLIP) algorithm and Steatosis, Activity and Fibrosis (SAF) scoring system METHODS: We prospectively included 136 consecutive patients who underwent liver biopsy for MAFLD at three Spanish centres (January 2017-January 2020). Biopsies were scored by two blinded pathologists according to the Non-alcoholic Steatohepatitis (NASH) Clinical Research Network system for fibrosis staging, the FLIP/SAF classification for steatosis and inflammation grading and Deugnier score for iron grading. Proportionate areas of collagen, fat, inflammatory cells and iron deposits were measured with computer-assisted digital image analysis. A test-retest experiment was performed for precision repeatability evaluation. Digital pathology showed strong correlation with fibrosis (r=0.79; P<0.001), steatosis (r=0.85; P<0.001) and iron (r=0.70; P<0.001). Performance was lower when assessing the degree of inflammation (r=0.35; P<0.001). NASH cases had a higher proportion of collagen and fat compared to non-NASH cases (P<0.005), whereas inflammation and iron quantification did not show significant differences between categories. Repeatability evaluation showed that all the coefficients of variation were ≤1.1% and all intraclass correlation coefficient values were ≥0.99, except those of collagen. Digital pathology allows an automated, precise, objective and quantitative assessment of MAFLD histological features. Digital analysis measurements show good concordance with pathologists´ scores.

Quantification of liver iron overload disease with laser ablation inductively coupled plasma mass spectrometry

BackgroundHereditary hemochromatosis is the most frequent, identified, genetic disorder in Caucasians affecting about 1 in 1000 people of Northern European ancestry, where the associated genetic defect (homozygosity for the p.Cys282Tyr polymorphism in the HFE gene) has a prevalence of approximately 1:200. The disorder is characterized by excess iron stores in the body. Due to the incomplete disease penetrance of disease-associated genotype, genetic testing and accurate quantification of hepatic iron content by histological grading of stainable iron, quantitative chemical determination of iron, or imaging procedures are important in the evaluation and staging of hereditary hemochromatosis.MethodsWe here established novel laser ablation inductively coupled plasma mass spectrometry protocols for hepatic metal bio-imaging for diagnosis of iron overload.ResultsWe demonstrate that these protocols are a significant asset in the diagnosis of iron overload allowing iron measurements and simultaneous determination of various other metals and metalloids with high sensitivity, spatial resolution, and quantification ability.ConclusionsThe simultaneous measurement of various metals and metalloids offers unique opportunities for deeper understanding of metal imbalances. Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) is a highly powerful and sensitive technique for the analysis of a variety of solid samples with high spatial resolution. We conclude that this method is an important add-on to routine diagnosis of iron overload and associated hepatic metal dysbalances resulting thereof.

Tissue iron quantification in chronic liver diseases using MRI shows a relationship between iron accumulation in liver, spleen, and bone marrow

To investigate iron loading within the liver, pancreas, spleen, and bone marrow using magnetic resonance imaging (MRI) transverse relaxation rate (R2*), in patients with diffuse liver diseases; to evaluate the relationships between iron accumulation in these tissue compartments; and to assess the association between tissue iron overload and the pattern of hepatic cellular iron distribution (hepatocytes versus Kupffer cells). Fifty-six patients with diffuse liver diseases had MRI-derived R2* values, using a multi-echo chemical-shift encoded MRI sequence, of the liver, pancreas, spleen, and vertebral bone marrow. All patients had liver biopsy samples scored for hepatic iron grading (0-4) and iron cellular distribution (within hepatocytes only or within both hepatocytes and Kupffer cells). Liver R2* increased with histological iron grade (RS=0.58, p<0.001) and correlated with spleen (RS=0.71, p<0.001) and bone marrow R2* (RS=0.66, p<0.001), but not with pancreatic R2* (RS=0.22, p=0.096). Splenic and bone marrow R2* values were also correlated (RS=0.72, p<0.001). Patients with iron inside Kupffer cells had the highest R2* in liver, spleen and bone marrow. Patients with chronic diffuse liver diseases have concomitant hepatic, splenic, and bone marrow iron loading. The highest hepatic iron scores and iron inside Kupffer cells were associated with the highest splenic and bone marrow deposits, suggesting systemic iron accumulation in the mononuclear phagocytic system.

Virtual iron concentration imaging based on dual-energy CT for noninvasive quantification and grading of liver iron content: An iron overload rabbit model study.

To assess the accuracy of liver iron content (LIC) quantification and grading ability associated with clinical LIC stratification using virtual iron concentration (VIC) imaging on dual-energy CT (DECT) in an iron overload rabbit model. Fifty-one rabbits were prepared as iron-loaded models by intravenous injection of iron dextran. DECT was performed at 80 and 140 kVp. VIC images were derived from an iron-specific algorithm. Postmortem LIC assessments were conducted on an inductively coupled plasma (ICP) spectrometer. Correlation between VIC and LIC was analyzed. VIC were stratified according to the corresponding clinical LIC thresholds of 1.8, 3.2, 7.0, and 15.0 mg Fe/g. Diagnostic performance of stratification was evaluated by receiver operating characteristic analysis. VIC linearly correlated with LIC (r = 0.977, P < 0.01). No significant difference was observed between VIC-derived LICs and ICP (P > 0.05). For the four clinical LIC thresholds, the corresponding cutoff values of VIC were 19.6, 25.3, 36.9, and 61.5 HU, respectively. The highest sensitivity (100%) and specificity (100 %) were achieved at the threshold of 15.0 mg Fe/g. Virtual iron concentration imaging on DECT showed potential ability to accurately quantify and stratify hepatic iron accumulation in the iron overload rabbit model. • Virtual iron concentration (VIC) linearly correlates with liver iron content (LIC). • VIC accurately quantifies LIC. • VIC accurately grades LIC based on clinical LIC stratification.

Hepatic iron deposition in patients with sickle cell disease: Role of breath-hold multiecho T2∗-weighted MRI sequence

IntroductionThe purpose of this study is to detect the role of breath-hold multiecho T2∗-weighted MRI, in quantification of hepatic iron deposition in patients with sickle cell disease. MethodsThirty-seven patients underwent 1.5-T MRI of the liver that included a multiecho T2∗-weighted sequence. Hepatic T2∗ iron grading was done for each patient by placing regions of interest in the hepatic parenchyma. Hepatic T2∗ values were correlated with histopathological iron grade. Liver biopsy was done for all patients. Written consent was obtained from all patients prior to MRI studies. ResultsThirty-two patients (86.5 %) had evidence of hepatic iron deposition on histopathological examination, including eight (25%) with grade 3, eleven (34%) with grade 2 and thirteen patients (41%) with grade 1.Patients with negative iron deposition histologically, had T2∗ values ranging from 28–32ms. For the patients with positive hepatic iron deposition, hepatic T2∗ decreased with increasing iron grade.Statistical analysis showed that for differentiation of hepatic iron deposition grade 3 from grades 1 and 2, hepatic T2∗ less than 13ms had a sensitivity and specificity of 100 % and 98%, respectively. ConclusionBreath-hold multiecho T2∗-weighted MRI sequence offers an accurate estimation of hepatic iron deposition.

English

BACKGROUND: Iron deficiency anemia is one of the most prevalent forms of malnutrition. The list of diagnostic tools is quite long starting from the basic hemogram, p.s for comment, reticulocyte count to iron studies including serum ferritin, serum iron, total iron binding capacity. Evaluation of the bone marrow iron stores is the gold standard. AIM: To perform a correlation study between Perl's Prussian blue stained bone marrow aspirate smears and serum iron and serum ferritin. MATERIALS AND METHODS: A descriptive study of Perl's Prussian blue stained bone marrow aspirate smears of 40 adult patients with moderate to severe anemia was performed at the Department of Pathology, Gandhi Medical College, Bhopal. Bone marrow was assessed by Gale's method and correlated with serum ferritin and serum iron. RESULTS: Gale's grading method revealed decreased iron stores in 30%; normal iron stores in 55% cases, and 15% cases had increased iron stores. The co-relational coefficient between bone marrow iron grade and serum ferritin came out to be 0.59 with an F- test value of 55.10.The correlation coefficient between bone marrow grade and serum iron is 0.76 and the F-test value -117.7. CONCLUSION: Although bone marrow iron grading is still gold standard but there exists a positive correlation between bone marrow iron with serum ferritin and serum iron thus iron status can be assessed by serum ferritin and serum iron, bone marrow being an invasive procedure can be avoided, also it will be helpful

Diagnostic accuracy of dual-echo (in- and opposed-phase) T1-weighted gradient recalled echo for detection and grading of hepatic iron using quantitative and visual assessment.

Detection and quantification of hepatic iron with dual-echo gradient recalled echo (GRE) has been proposed as a rapid alternative to other magnetic resonance imaging (MRI) techniques. Co-existing steatosis and T1 weighting are limitations. This study assesses the accuracy of routine dual-echo GRE. Between 2010 and 2013, 109 consecutive patients underwent multi-echo (ME) MRI and dual-echo GRE for quantification of hepatic iron. Liver iron concentration (LIC) was calculated from ME-MRI. Relative signal intensity (RSI) and fat signal fraction (FSF) were calculated from dual-echo GRE. Four radiologists subjectively evaluated dual-echo GRE (±subtraction). Diagnostic accuracy was compared between techniques and correlated with biopsy using Fisher's exact test, Spearman correlation and regression. The sensitivity of visual detection of iron ranged from 48 to 55%. Subtraction did not increase sensitivity (p < 0.001). Inter-observer variability was substantial (κ = 0.72). The specificity of visual detection of iron approached 100% with false-positive diagnoses observed using subtraction. LIC showed a higher correlation with histopathological iron grade (r = 0.94, p < 0.001) compared with RSI (r = 0.65, p = 0.02). Univariate regression showed an association between RSI and LIC (B = 0.98, p < 0.001, CI 0.73-1.23); however, the association was not significant with multi-variate regression including FSF (p = 0.28). Dual-echo GRE has low sensitivity for hepatic iron. Subtraction imaging can result in false-positive diagnoses. • Routine liver MRI studies cannot effectively screen patients for iron overload. • Concomitant hepatic steatosis and iron limits diagnostic accuracy of routine liver MRI. • Dual-echo GRE subtraction imaging causes false-positive diagnoses of iron overload. • Dedicated MRI techniques should be used to diagnose and quantify iron overload.

Intensive method of assessment and classification of the bone marrow iron status: A study of 80 patients

The increasing prevalence of multiple co-morbidities among anemic patients with chronic diseases have made the use of serum ferritin (which is also an acute phase reactant) and transferrin saturation more challenging in diagnosing iron deficiency. Microscopic examination of bone marrow aspirate is the gold standard" for assessing marrow iron store. However, conventional Gale's method assesses iron in marrow fragments alone which provides little valuable information about functional iron deficiency seen in many chronic diseases. To perform an intensive bone marrow iron grading by assessing iron in fragments, in macrophages around fragments and in erythroblasts and to correlate the marrow iron store results with serum ferritin. A descriptive study of Perl's Prussian blue stained bone marrow aspirate smears of 80 adult patients with moderate to severe anemia. Bone marrow iron was assessed by both the Gale's method and the intensive method and correlated with serum ferritin. The intensive grading system revealed normal iron stores in 37.5% cases, depleted iron stores in 16.25% patients while 23.75% and 22.5% patients had functional iron deficiency and combined deficiency, respectively. Mean log ferritin concentration was significantly lower in patients with depleted iron stores (0.91 μg/l) in comparison to those with normal iron stores (2.13 μg/l; P = 0.001), functional iron deficiency (2.65 μg/l; P = 0.000), or combined deficiency (2.04 μg/l; P = 0.002). Intense marrow iron examination provides a useful iron status classification which is of particular importance in cases of chronic diseases and inflammation.

The importance of spleen, spleen iron, and splenectomy for determining total body iron load, ferrikinetics, and iron toxicity in thalassemia major patients

The importance of spleen, spleen iron and splenectomy has been investigated in 28 male and 19 female β-thalassemia major (β-ΤΜ), adult patients. In one study, an increase from about five (615 g; 19.5 × 11.0 × 6.0 cm) to twenty (2030 g; 25.0 × 17.5 × 12.0 cm) times higher than the normal size and weight of spleen has been observed in twenty patients following splenectomy. In a second study, the mean size for the liver (19.4 cm, range 13.5–26.0 cm) and spleen (15.6 cm, range 7.0–21.0 cm) measured by magnetic resonance imaging (MRI) and by ultrasound imaging for spleen (15.1 cm, range 9.0–21.0 cm) of 16 patients indicated that on average the spleen is about 80% of the size of the liver. In the third study, comparison of the iron load using MRI T2* and iron grading of stained biopsies indicated that substantial but variable amounts of excess iron are stored in the spleen (0–40%) in addition to that in the liver. Following splenectomy, total body iron storage capacity is reduced, whereas serum ferritin (p = 0.0085) and iron concentration in other organs appears to increase despite the reduction in the rate of transfusions (p = 0.0001) and maintenance of hemoglobin levels (p = 0.1748). Spleen iron seems to be cleared faster than liver iron using effective chelation protocols. Spleen iron is a major constituent of the total body iron load in β-ΤΜ patients and should be regularly monitored and targeted for chelation. Normalization of the body iron stores at an early age could maintain the spleen in near normal capacity and secondary effects such as cardiac and other complications could be avoided.

Lack of association of primary iron overload and common HFE gene mutations with liver cirrhosis in adult Indian population

To find out the association of common HFE mutations (viz., C282Y and H63D) with primary iron overload (PIL) in liver cirrhosis (CLD) patients of Indian origin. Polymerase chain reaction-restriction fragment length polymorphism method was used for screening C282Y and H63D mutation in 496 CLD patients (hepatitis B virus associated cirrhosis (HBVc) = 74, hepatitis C virus associated cirrhosis (HCV) = 50, alcoholic cirrhosis with hepatitis (ALcW) = 38, alcoholic cirrhosis without hepatitis (ALc) = 92, cryptogenic cirrhosis (CC) = 242) and 502 healthy controls. Transferrin saturation of >45 or serum ferritin of >300ng/mL (males)/>200ng/mL (females) with normal total exogenous iron intake was suggestive of PIL. Histological liver iron grading was done by Perl's Prussian blue stain. Of 496 patients, 13 (2.6; 9 CC, 2 ALc, 1 HBVc, 1 AlcW) had PIL. However, only two (15.3) of 13 patients (1 CC and 1 HBVc) were positive for H63D heterozygous mutation. All the subjects were found to be C282Y wild type, except a single case of double heterozygous (C282Y/H63D) who however, did not have PIL. Overall frequency of H63D allele in patients and controls was not significantly different (5.95 and 4.58 respectively, p = 0.17). A highly significant H63D allele frequency (p < 0.005) was observed in HBVc (10.82) and ALcW (11.84) groups but they were not associated with PIL. The frequency of PIL, and the HFE gene mutaion (C282Y) are both rare in Indian patients and explain why hemochromatosis is a rare cause of liver cirrhosis in India. A highly significant H63D allele frequency in HBV and alcohol-related cirrhosis suggest a possible predisposing role for liver fibrosis of this allele.

Iron overload: accuracy of in-phase and out-of-phase MRI as a quick method to evaluate liver iron load in haematological malignancies and chronic liver disease

The purpose of this prospective study was to evaluate the accuracy of in-phase and out-of-phase imaging to assess hepatic iron concentration in patients with haematological malignancies and chronic liver disease. MRI-based hepatic iron concentration (M-HIC, μmol g(-1)) was used as a reference standard. 42 patients suspected of having iron overload and 12 control subjects underwent 1.5 T in- and out-of-phase and M-HIC liver imaging. Two methods, semi-quantitative visual grading made by two independent readers and quantitative relative signal intensity (rSI) grading from the signal intensity differences of in-phase and out-of-phase images, were used. Statistical analyses were performed using the Spearman and Kruskal-Wallis tests, receiver operator curves and κ coefficients. The correlations between M-HIC and visual gradings of Reader 1 (r = 0.9534, p < 0.0001) and Reader 2 (r = 0.9456, p < 0.0001) were higher than the correlations of the rSI method (r = 0.7719, p < 0.0001). There was excellent agreement between the readers (weighted κ = 0.9619). Both visual grading and rSI were similar in detecting liver iron overload: rSI had 84.85% sensitivity and 100% specificity; visual grading had 85% sensitivity and 100% specificity. The differences between the grades of visual grading were significant (p < 0.0001) and the method was able to distinguish different degrees of iron overload at the threshold of 151 μmol g(-1) with 100% positive predictive value and negative predictive value. Detection and grading of liver iron can be performed reliably with in-phase and out-of-phase imaging. Liver fat is a potential pitfall, which limits the use of rSI.

The Significance of Hepatic Iron Overload in Allogenic Stem Cell Transplant Recipients.

Abstract 4313 IntroductionIron overload has increasingly been recognized as a cause of morbidity and mortality among allogenic hematopoietic stem cell transplant (HSCT) recipients. Estimation of liver iron is considered the gold standard method that reflects the overall body iron burden. AimWe attempted to correlate the severity of liver iron overload to various clinical parameters in allogenic HSCT recipients. MethodsWe studied 49 adult allogenic HSCT recipients who had a minimum of one liver biopsy performed after transplant. All patients had been diagnosed, transplanted and followed up in our institution since 1998. Biopsies were performed due to abnormal liver function tests under the initial clinical suspicion of GVHD with a mean of 185 days (range: 23-844 days) post-transplant. Sections from each biopsy were stained with hematoxylin and eosin, Masson trichome and Perl's iron stains. We used the histological semiquantitative grading of liver iron (grade 0-4), evidence of GVHD (score 0-3) and fibrosis (stage 0-4). Iron overload was defined as a score of 2 and above (correlating with hepatic iron concentration >1.4 mg/g dry weight). Various clinical parameters were also assessed including age, sex, underlying malignancy, number of transfusions, source and type of stem cell transplant, liver function tests, clinical grading of GVHD, significant bacteremia, other infections, survival and cause of death. ResultsIn this cohort, 28 of 49 patients had iron overload. The overall prevalence of iron overload was 57% (95% confidence interval, 42%-71%). Iron overload in 7 of the 49 patients (14%) was the only histological explanation of liver dysfunction. The degree of iron overload was directly correlated to the number of transfusions (r=0.69, p<0.001), serum ferritin (r=0.55, p=0.004) and the development of significant bacteremia (r=0.33, p=0.019). The correlation to the histologic liver-GVHD grade was close to statistical significance (p=0.062). 37 of the 49 patients (76%) died after HSCT at a mean of 677.8 days (range: 52-3821 days) post-transplant. The overall mortality did not correlate statistically to the degree of iron overload. The mortality was higher in patients with high serum bilirubin (direct or total) and increasing clinical degree of acute GVHD. Sixteen patients died secondary to infection (bacterial=15, fungal=1). However, there was no significant evidence of increased risk of infection-related mortality with iron overload (Hazard Ratio = 1.16, 95% CI, 0.76-1.77, p = 0.496). ConclusionIron overload is a common adverse effect of allogenic HSCT (57% prevalence). Iron overload occurred in some of our patients after as few as 10 red blood cell units. In this cohort, iron overload statistically correlated to the number of transfusions, development of significant bacteremia and clinical acute GVHD. We did not find a statistically significant association with mortality, likely due to our small sample size and complex clinical picture. Iron overload was the only histological explanation of abnormal liver functions in 14% of our patients. More studies are needed to determine the impact of iron overload in long-term morbidity and mortality in allogenic transplant patients. Disclosures:No relevant conflicts of interest to declare.

Improved method for assessing iron stores in the bone marrow

Background:Bone marrow iron microscopy has been the “gold standard” method of assessing iron deficiency. However, the commonly used method of grading marrow iron remains highly subjective.Aim:To improve the bone marrow...

Hepatic Iron Quantitation and Its Relationship with Disease Measures and Histologically Assessed Iron Content

The study was undertaken to determine the relationships between the histological finding of hepatic injury, necrosis, fibrosis and iron staining with biochemical measures of hepatic injury and hepatic iron content. One hundred fourteen consecutive subjects undergoing liver biopsy were studied. Data pertaining to biopsy reports, histology activity index, hepatic iron concentration, hepatic iron index, and histological grading of iron were recorded and analyzed using regression analysis. Statistical relationships were found between the hepatic iron index, histological grading of iron and hepatic iron concentration, whereas histology activity index was related to hepatic grading of iron, but not to hepatic iron concentration. The pathologist's assessment of the histological grading of iron relates directly to the histology activity index and hepatic iron concentration. Serum measures of iron, total iron-binding capacity and ferritin can be used to monitor hepatic iron content more economically and simply than with hepatic iron concentration measurement on liver biopsy.

Evidence for a sequential transfer of iron amongst ferritin, transferrin and transferrin receptor during duodenal absorption of iron in rat and human

To elucidate the sequential transfer of iron amongst ferritin, transferrin and transferrin receptor under various iron status conditions. Incorporation of 59Fe into mucosal and luminal proteins was carried out in control WKY rats. The sequential transfer of iron amongst ferritin, transferrin and transferrin receptor was carried out in iron deficient, control and iron overloaded rats. The duodenal proteins were subjected to immunoprecipitation and quantitation by specific ELISA and in situ localization by microautoradiography and immunohistochemistry in tandem duodenal sections. Human duodenal biopsy (n = 36) collected from subjects with differing iron status were also stained for these proteins. Ferritin was identified as the major protein that incorporated iron in a time-dependent manner in the duodenal mucosa. The concentration of mucosal ferritin was significantly higher in the iron excess group compared to control, iron deficient groups (731.5 +/- 191.96 vs 308.3 +/- 123.36, 731.5 +/- 191.96 vs 256.0 +/- 1.19, P < 0.005), while that of luminal transferrin which was significantly higher than the mucosal did not differ among the groups (10.9 +/- 7.6 vs 0.87 +/- 0.79, 11.1 +/- 10.3 vs 0.80 +/- 1.20, 6.8 +/- 4.7 vs 0.61 +/- 0.63, P < 0.001). In situ grading of proteins and iron, and their superimposition, suggested the occurrence of a sequential transfer of iron. This was demonstrated to occur through the initial binding of iron to luminal transferrin then to absorptive cell surface transferrin receptors. The staining intensity of these proteins varied according to the iron nutrition in humans, with intense staining of transferrin receptor observed in iron deficient subjects. It is concluded that the intestine takes up iron through a sequential transfer involving interaction of luminal transferrin, transferrin-transferrin receptor and ferritin.

Gender and liver fibrosis in chronic hepatitis: the role of iron status

The role of gender in the progression of fibrosis in chronic hepatitis C is still under investigation. To investigate whether gender affects the progression of liver disease and/or hides other risk factors. A prospective series of 121 consecutive patients with chronic hepatitis C underwent liver biopsy. Grading and staging for chronic hepatitis were each evaluated according to Ishak's classification. In univariate and multivariate analysis on the whole group of patients, male gender was not associated either with significant liver fibrosis (Ishak's score > 2) or with cirrhosis (Ishak's score > 4). On the contrary, in univariate analysis on patients aged < or = 50 years, male gender was nearly significantly (P = 0.06) predictive of liver fibrosis, whereas it was not in patients > 50 years. Hepatic iron grading, along with age, was an independent factor associated with fibrosis. Moreover, the values of all the variables which describe iron status were significantly higher in males aged < or = 50 years in comparison with females of the same age. In chronic hepatitis C, male gender may be predictive of liver fibrosis only in patients aged < or = 50 years. Among fibrogenetic factors hidden by gender, iron status could play a major role.

Hepatic iron concentration does not predict response to standard and pegylated-IFN/ribavirin therapy in patients with chronic hepatitis C

Background/Aims Iron overload is common among patients with chronic hepatitis C (CHC). In this study the role of hepatic iron concentration (HIC) and serum iron parameters was assessed to determine response to standard and pegylated interferon (IFN)/ribavirin combination therapy in patients with CHC. Methods Liver biopsies were obtained from 169 IFN-naı̈ve patients ( m=115, f=54, age: 40.8±10.7) with CHC. 140 patients were treated with standard IFN/ribavirin, 29 patients with pegylated-IFN/ribavirin. Biopsy specimens were evaluated according to the DiBisceglie scoring system and iron grading. HIC was determined by atomic absorption spectroscopy. Ferritin and transferrin saturation and presence of HFE-C282Y and H63D gene mutations were determined at baseline. Results Nonresponders to combination therapy had higher serum ferritin levels at baseline ( p<0.01). There was no difference of HIC, transferrin saturation levels, and the HFE-mutation status between responders and nonresponders. Logistic regression analysis revealed serum ferritin as an independent predictor of response. HIC correlated with the DiBisceglie score ( r=0.352, p<0.001), iron grading ( r=0.352, p<0.001) and serum ferritin ( r=0.335, P<0.001). Conclusions Pretreatment liver iron concentration does not predict response to combination therapy in patients with CHC. In contrast, high baseline serum ferritin levels are predictors of poor response to antiviral therapy.