Renal Iron Research Articles

MTOR Inhibition improves anaemia and reduces organ damage in a murine model of sickle cell disease.

Mechanistic target of rapamycin (mTOR) has been shown to play an important role in red blood cell physiology, with inhibition of mTOR signalling leading to alterations in erythropoiesis. To determine if mTOR inhibition would improve anaemia in sickle cell disease (SCD), mice with SCD were treated with the dual mTORC1/2 inhibitor, INK128. One week after daily oral drug treatment, erythrocyte count, haemoglobin, and haematocrit were all significantly increased while reticulocyte counts were reduced. These parameters remained stable during 3weeks of treatment. Similar effects were observed following oral treatment with the mTORC1 inhibitor, sirolimus. Sirolimus treatment prolonged the lifespan of sickle cell erythrocytes in circulation, reduced spleen size, and reduced renal and hepatic iron accumulation in SCD mice. Following middle cerebral artery occlusion, stroke size was reduced in SCD mice treated with sirolimus. In conclusion, mTOR inhibition is protective against anaemia and organ damage in a murine model of SCD.

Effects of Bone Marrow Infiltration By Multiple Myeloma on Erythropoiesis

Diseases that infiltrate the bone marrow disrupt erythropoiesis leading to anemia. In multiple myeloma (MM), anemia severity can be correlated with degree of marrow infiltration by myeloma cells. Infiltrating MM may impair the function and structure of erythroblastic islands (EBIs), the marrow erythropoietic niches. An EBI consists of a central macrophage surrounded by colony-forming units-erythroid/proerythroblasts (CFU-E/pro-EBs) and their progeny, the differentiating erythroblasts. Cytokines produced by MM cells, such as Fas ligand (FL), tumor necrosis factor (TNF), and TNF-related apoptosis-inducing ligand (TRAIL), can induce erythroid cell apoptosis. Physical displacement of the erythroid cells away from central macrophages by MM can destroy the EBIs. Non-erythrotoxic therapies that kill MM cells while sparing erythropoietic cells allow quantification of erythropoiesis and marrow MM infiltration before and after treatment of newly diagnosed MM patients. Marrow biopsies from 15 newly diagnosed MM patients were obtained before and after 4 courses of non-erythrotoxic induction therapy with bortezomib, dexamethasone, and lenalidomide (Richardson et al, Blood 2010). CBCs and serum MM paraprotein quantifications were obtained with the marrow biopsies and before each course of therapy. No patient had renal insufficiency, iron or cobalamin deficiency, erythropoietin (EPO) therapy, or RBC transfusion.At diagnosis, percentages of marrow space occupied by MM and erythroid cells were negatively correlated. Percentages of marrow space infiltrated by MM (range = 2.3 - 72.3%) were also negatively correlated with hemoglobin (Hb), hematocrit (Hct) and RBCs. One patient had a partial response: marrow myeloma decreasing from 27.5% to 5.3%. All other patients had reductions in marrow myeloma to < 2.2%. The 8 patients with < 30% MM infiltration at diagnosis had no change (-1.4% to 1.8%) in marrow space occupied by erythroid cells following therapy, whereas 7 patients with > 35% MM infiltration at diagnosis increased marrow space occupied by erythroid cells following therapy (3.4 to 19.2%). Hb, Hct, and RBCs did not change during therapy in patients with < 30% MM infiltration, but those with > 35% myeloma infiltration at diagnosis had progressive increases in Hb, Hct, and RBCs during therapy.These clinical data were used to study the relationship between marrow infiltration by MM and erythropoiesis. Mathematical models of MM infiltration effects on marrow EBI structure/function were developed and tested in simulations. A previously developed hybrid discrete-continuous model of erythropoiesis based on EBI (Eymard et al, J Math Biol 2015) was extended to a larger area of marrow containing multiple EBIs. In the model, CFU-E/proEBs have 3 fates-- self-renewal, differentiation, and apoptosis--that depend upon factors produced systemically, such as glucocorticoids and EPO, and locally, such as stem cell factor and bone morphogenetic protein 4 by central macrophages and FL by mature erythroblasts. Intracellular regulatory networks were modeled with ordinary differential equations and extracellular concentrations by partial differential equations. Under normal conditions, EBIs achieve a steady-state that produces new RBCs at rates which maintain normal Hb, Hct and RBCs. At early times after the section of bone marrow is infiltrated by small foci of proliferating MM cells, EBI function is not affected. With further proliferation, infiltrating MM cells occupy more marrow space, inducing erythroid cell apoptosis by producing FL, TNF or TRAIL and by displacing erythroid cells from central macrophages, thereby destroying EBIs. However, central macrophages of destroyed islands persist or are replaced by differentiation of monocyte-macrophage precursors. After MM cells are killed by therapy, the residual macrophages can interact with burst-forming units-erythroid (BFU-E), thereby reestablishing EBIs. If the MM infiltrate is not sufficiently reduced after a course of therapy, it can physically interfere with the macrophage-BFU-E interaction, preventing the reestablishment of an EBI and full recovery of RBC production until a subsequent therapy reduces the infiltrate sufficiently that the EBI is reestablished. The model is consistent with the clinical data and may apply to other marrow infiltrative diseases including myelofibrosis, systemic infections, or other malignancies. DisclosuresNo relevant conflicts of interest to declare.

A Mechanistic Investigation of the TTP-like State Associated with Intravenous Abuse of Opana® ER

Background: Since 2012, a growing list of case reports has emerged describing the occurrence of a TTP-like state following intravenous (i.v.) abuse of extended release oxymorphone (Opana® ER). The pathophysiology resulting from intravenous exposure to Opana® ER has demonstrated a spectrum of severity with unclear etiology, and approaches to treatment/management have ranged from early plasma exchange to supportive care alone. As a result these cases have spurred a collaborative investigation to better understand mechanisms of toxicity. Given the temporal relation between the reformulation of the drug product to achieve extended release and the onset of reported cases, we investigated the inert ingredient mixture as a possible causal factor.Methods: Guinea pigs were used as an animal model to understand the hematopathologic and nephrotoxic potential of the inert ingredient mixture of Opana® ER, which was provided by Endo Pharmaceuticals under a research collaborative agreement. Animal studies were approved by Institutional Animal Care and Use Committees and conducted in adherence with NIH guidelines. Animals were studied over various i.v. dosing schemes (single injections of 0.1, 0.3, or repeated dosing of 0.3mg/kg (5 doses at 1.5hr intervals)) with either the entire inert ingredient mixture ("PEO+") or high molecular weight polyethylene oxide alone ("PEO")-the major constituent of the inert ingredient mixture. The dose range was based on estimates of the inert ingredient mass introduced during the adulteration and injection of the drug product. Additional in vitro and cell-based assessments of coagulation (thromboelastography, thrombin generation) and ADAMTS13 function were carried out to better understand in vivo observations.Results: Intravenous administration of the solubilized inert ingredient mixture in guinea pigs resulted in dose-dependent pathologic changes in blood parameters and kidney function. Hematological changes included a dose dependent increase in free plasma hemoglobin (Hb), decreased hematocrit and platelet count. Acute kidney injury was characterized by dose-dependent renal cortical iron deposition, plasma creatinine increase, distal convoluted tubular injury (NGAL mRNA induction), and glomerular damage (hemoglobinuria) (Table 1). At high doses, red blood cell aggregation and renal hypoxia (HIF-1 accumulation) was observed. These observations were similar following the administration of PEO alone, and occurred independent of changes to ADAMTS13 and its cleavage of Von Willebrand Factor. PEO, when spiked into blood in vitro, does not directly modulate coagulability as indicated by global assays of coagulation.Conclusions: The present animal study suggests that i.v. exposure to the inert ingredients included in Opana® ER can trigger hematologic responses resembling elements of TTP, namely intravascular hemolysis and acute renal injury/failure. However, the renal injury observed in the guinea pig model is distinct from classic TTP, which involves spontaneous platelet clumping and subsequent microvascular thrombosis; instead, the nephrotoxicity likely arises from a multifactorial process encompassing renal tubular injury and glomerular damage, arising secondary to hemolysis, extracellular Hb/heme/iron mediated oxidative tissue damage and compromised tissue oxygenation.Table 1Guinea pig dose group(mg of PEO+/kg)Cmax of plasma free Hb [mg/ml]AUC0-24h of plasma free Hb [mg/ml x h]hematocrit at 24h [%]platelet count at 24h [K/µl]NGAL induction** (renal cortex)plasma creatinine at 24h [µmol/L]hemoglobinuria at end of study [mg/ml]iron deposition (renal cortex)[µg/g tissue]Control0.8 ±0.1 (n=4)16.2 ±4.8 (n=4)36 ±1.7 (n=5)205 ±11.6 (n=5)1 ±0.3 (n=3)87.0 ±9.3 (n=4)0.1 (n=1)33 ±3.5 (n=3)0.1 single dose1.4 ±0.6 (n=11)20.5 ±5.75 (n=11)35 ±3.1 (n=5)157 ±20.5 (n=5)1 ±0.5 (n=3)104 ±41.3 (n=4)0.6 (n=2)49 ±7.3 (n=3)0.3 single dose2 ±0.3 (n=13)36.5 ±5.1 (n=13)31 ±1.1 (n=6)135 ±11.2 (n=6)1.6 ±0.4 (n=3)145 ±36.7 (n=4)1.2 (n=2)153 ±2.5 (n=3)0.3 multi dose (5 injections at 1.5h intervals)*5.1 ±1.3 (n=5)87.8 ±15.8 (n=4)28 ±0.8 (n=4)62 ±12.4 (n=4)24 ±0.8 (n=4)421.1 ±33.8 (n=4)9.8 ±5 (n=3)139 ±9.5 (n=4)data represented as mean ±SEM, n = number of animals*endpoint 24h, all other groups 48h**Fold induction over baseline DisclosuresWong:Haemonetics Corporation: Research Funding.

Renal iron overload in rats with diabetic nephropathy.

Diabetic nephropathy (DN) remains incurable and is the main cause of end-stage renal disease. We approached the pathophysiology of DN with systems biology, and a comprehensive profile of renal transcripts was obtained with RNA-Seq in ZS (F1 hybrids of Zucker and spontaneously hypertensive heart failure) rats, a model of diabetic nephropathy. We included sham-operated lean control rats (LS), sham-operated diabetic (DS), and diabetic rats with induced renal ischemia (DI). Diabetic nephropathy in DI was accelerated by the single episode of renal ischemia. This progressive renal decline was associated with renal iron accumulation, although serum and urinary iron levels were far lower in DI than in LS. Furthermore, obese/diabetic ZS rats have severe dyslipidemia, a condition that has been linked to hepatic iron overload. Hence, we tested and found that the fatty acids oleic acid and palmitate stimulated iron accumulation in renal tubular cells invitro. Renal mRNAs encoding several key proteins that promote iron accumulation were increased in DI. Moreover, renal mRNAs encoding the antioxidant proteins superoxide dismutase, catalase, and most of the glutathione synthetic system were suppressed, which would magnify the prooxidant effects of renal iron loads. Substantial renal iron loads occur in obese/diabetic rats. We propose that in diabetes, specific renal gene activation is partly responsible for iron accumulation. This state might be further aggravated by lipid-stimulated iron uptake. We suggest that progressive renal iron overload may further advance renal injury in obese/diabetic ZS rats.

Organ iron accumulation in chronically transfused children with sickle cell anaemia: baseline results from the TWiTCH trial.

Transcranial Doppler (TCD) With Transfusions Changing to Hydroxyurea (TWiTCH) trial is a randomized, open-label comparison of hydroxycarbamide (also termed hydroxyurea) versus continued chronic transfusion therapy for primary stroke prevention in patients with sickle cell anaemia (SCA) and abnormal TCD. Severity and location of iron overload is an important secondary outcome measure. We report the baseline findings of abdominal organ iron burden in 121 participants. At enrollment, patients were young (9·8±2·9years), predominantly female (60:40), and previously treated with transfusions (4·1±2·4years) and iron chelation (3·1±2·1years). Liver iron concentration (LIC; 9·0±6·6mg/g dry weight) and serum ferritin were moderately elevated (2696±1678μg/l), but transferrin was incompletely saturated (47·2±23·6%). Spleen R2* was 509±399Hz (splenic iron ~13·9mg/g) and correlated with LIC (r(2) =0·14, P=0·0008). Pancreas R2* was increased in 38·3% of patients but not to levels associated with endocrine toxicity. Kidney R2* was increased in 80·7% of patients; renal iron correlated with markers of intravascular haemolysis and was elevated in patients with increased urine albumin-creatinine ratios. Extra-hepatic iron deposition is common among children with SCA who receive chronic transfusions, and could potentiate oxidative stress caused by reperfusion injury and decellularized haemoglobin.

Atypical haemolytic uraemic syndrome treated with the complement inhibitor eculizumab: the experience of the Australian compassionate access cohort.

This study aimed to report the clinical characteristics and outcomes of Australian patients treated with eculizumab for atypical haemolytic uraemic syndrome (aHUS). A retrospective cohort study was undertaken of all patients in Australia treated with eculizumab provided in a compassionate access programme for a clinical diagnosis of aHUS using prospectively collected clinical data. A total of 10 patients with a median age of 23.5 years (interquartile range (IQR) 24.83 years) received compassionate access eculizumab for aHUS in Australia. Eight patients were female, and three had a family history of aHUS. Three received eculizumab for an initial acute aHUS presentation, three for relapsing and refractory acute aHUS, two for de novo aHUS post-renal transplantation, and one each for aHUS recurrence post-transplantation and facilitation of transplantation with a history of aHUS. The median duration of eculizumab therapy has been 911.5 days (IQR 569 days) with a cumulative exposure of 9184 days. At baseline all patients had renal and extra-renal aHUS involvement, with up to three non-renal organs affected. All but one patient, who died from uncontrollable gastrointestinal aHUS manifestations, have continued. The nine continuing patients achieved remission of aHUS. Two of the four patients requiring renal replacement therapy (RRT) at eculizumab commencement subsequently ceased RRT. Clinical events occurring in this cohort while on eculizumab treatment included neutropenia (two), posterior reversible encephalopathy syndrome (one), cardiomyopathy (one), pulmonary embolus (one), antibody-mediated rejection resulting in renal graft failure (one), iron deficiency (one), gastrointestinal haemorrhage (one) and death (one). Eculizumab has been an effective therapy for aHUS in this cohort, including when other therapies have failed.

Aging in Sickle Cell Disease: Co-morbidities and New Issues in Management

Availability of hydroxyurea (HU) coupled with early therapeutic interventions has increased the life expectancy of patients with sickle cell disease. Hence, the sickle cell community needs to be aware of common diseases of aging that survivors are predisposed to. We chose to investigate the sickle cell disease-related complications as well as non sickle cell disease-related medical problems of aging in 45 sickle cell patients over the age of 40 years. The most frequent chronic complications of sickle cell disease were elevated tricuspid regurgitant jet velocity on echocardiogram, chronic renal disease, iron overload and leg ulcers. Medical co-morbidities in this patient group included hypertension, diabetes mellitus (DM), hypercholesterolemia and symptomatic coronary artery disease (CAD). In our cohort, only 38.0% had a primary care doctor. Only 11.0% over age 50 had a screening colonoscopy, and of the women, 42.0% had a screening mammography. Medical co-morbidities and lack of health maintenance in older sickle cell patients are likely to impact overall health and mortality. Aging patients with sickle cell disease may benefit from a primary medical home for age appropriate comprehensive health care.

Chelation of dietary iron prevents iron accumulation and macrophage infiltration in the type I diabetic kidney

We previously reported that the functional deletion of p21, a cyclin-dependent kinase inhibitor, in mice attenuated renal cell senescence in streptozotocin (STZ)-induced type 1 diabetic mice. In the present study, we investigated the effect of iron chelation on renal cell senescence and inflammation in the type 1 diabetic kidney. STZ-treated mice showed increase in iron accumulation, tubular cell senescence and macrophage infiltration at week 28 in the kidney. Administering deferasirox, which removes only dietary iron, significantly attenuated iron accumulation in proximal tubules and the number of infiltrating F4/80-positive cells without effecting blood glucose, hematocrit or hemoglobin levels. In contrast however, deferasirox did not influence renal cell senescence. The lack of p21 decreased the renal tubular iron accumulation and did not change tubular cell senescence. Interestingly, the STZ-treated animals showed an increase in p16, another cyclin-dependent kinase inhibitor. The results suggest that type 1 diabetes increases renal tubular iron accumulation and macrophage infiltration through a p21-dependent mechanism, and that the chelation of dietary iron attenuates these responses.

Quantitative T2* magnetic resonance imaging for renal iron overload assessment: normal values by age and sex.

Few studies of renal iron content have been performed with multiecho gradient-echo (ME-GRE) T2* magnetic resonance imaging (MRI). We assessed the feasibility and reproducibility of ME-GRE T2* MRI for measuring regional and global renal T2* values, and established the lower limits of normal in healthy subjects, also correlating the measured values with age and sex. Twenty consecutive healthy subjects (13 men and 7 women, mean age 29.1 ± 7.2 years, range 19-42 years) underwent MRI examinations using a 1.5 T magnet and an ME-GRE T2* sequence. For each kidney, T2* was measured in anterior, posterolateral, and posteromedial renal parenchymal regions. The mean T2* value was calculated as the average of the two kidneys T2* values. For the mean kidney T2* value, the coefficients of variation for intra- and inter-operator reproducibility were 1.76% and 6.23%, respectively. The lower limit of normal for the mean kidney T2* value was 31 ms (median 51.39 ± 10.09). There was no significant difference between left and right kidney T2* values (p = 0.578). No significant correlation was found between T2* values and subjects' age or sex. Renal ME-GRE T2* appears to be a feasible and reproducible technique. The renal T2* values showed no dependence on sex or age.

Association between renal iron accumulation and renal interstitial fibrosis in a rat model of chronic kidney disease.

Iron accumulation is associated with the pathophysiology of chronic kidney disease (CKD). Renal fibrosis is a final common feature that contributes to the progression of CKD; however, little is known about the association between renal iron accumulation and renal interstitial fibrosis in CKD. Here we investigate the effects of iron chelation on renal interstitial fibrosis in a rat model of CKD. CKD was induced by 5/6 nephrectomy in Sprague-Dawley rats. At 8 weeks after operation, 5/6 nephrectomized rats were administered an oral iron chelator, deferasirox (DFX), in chow for 8 weeks. Other CKD rats were given a normal diet. Sham-operative rats given a normal diet served as a control. CKD rats exhibited hypertension, glomerulosclerosis and renal interstitial fibrosis. Iron chelation with DFX did not change hypertension and glomerulosclerosis; however, renal interstitial fibrosis was attenuated in CKD rats. Consistent with these findings, renal gene expression of collagen type III and transforming growth factor-β was increased in CKD rats compared with the controls, while iron chelation suppressed these increments. In addition, a decrease in vimentin along an increase in E-cadherin in renal gene expression was observed in CKD rats with iron chelation. CKD rats also showed increased CD68-positive cells in the kidney, whereas its increase was attenuated by iron deprivation. Similarly, increased renal gene expression of CD68, tumor necrosis factor-α and monocyte chemoattractant protein-1 was suppressed in CKD rats with iron chelation. Renal iron accumulation seems to be associated with renal interstitial fibrosis in a rat model of CKD.

Renal anaemia and iron deficiency: introducing the PIVOTAL trial

Iron deficiency and anaemia are often associated with chronic kidney disease and can lead to significant fatigue, altered cognitive function and a reduced quality of life. In November 2013, Kidney Research UK began coordinating the PIVOTAL trial—the largest renal clinical trial ever undertaken in the UK. Angela Lumsdon explains the rationale behind the study.

Higher Endogenous Erythropoietin Levels in Hemodialysis Patients with Hepatitis C Virus Infection and Effect on Anemia

Study for influence of chronic Hepatitis C (HCV) on endogenous erythropoietin production and on anemia in dialysis patients remains inconclusive. We hypothesize that chronic hemodialysis patients with co-existing Hepatitis C infection will have higher hemoglobin levels than chronic hemodialysis patients without hepatitis C infection. Secondly, we hypothesize that the higher hemoglobin levels will be associated with higher erythropoietin levels. Therefore we conducted a cross-sectional study of chronic hemodialysis patients with and without hepatitis C infection and evaluated associations with hemoglobin and erythropoietin levels. Our primary outcome was level of hemoglobin. Secondary outcome included association of hemoglobin and erythropoietin levels. 57 chronic hemodialysis patients (33 male, 24 female, mean age 46.05 ± 12.7 years) were included. The mean time spent on hemodialysis was 7.16 ± 6.2 years. None of the patients received any recombinant EPO therapy. Biochemical analyses include ALT, AST, Albumin, C-Reactive Protein, cholesterol levels and complete blood counts. Iron status of patients (transferrin saturation and serum ferritin levels) and parathyroid hormone were measured. Endogenous EPO serum levels were measured by a standardized enzyme-linked immunoassay. 23 of the hemodialysed patients (38.5%) were HCV (+). There was no difference in age, sex, distribution of primary renal diseases, iron status, albumin, C-Reactive-Protein and parathyroid hormone levels between HCV (+) and (-) patients. Mean duration time on dialysis was higher in HCV (+) than HCV (-) patients. Hemoglobin levels were similar between study groups. However serum endogenous erythropoietin levels were significantly higher in HCV (+) patients than HCV (-) patients (19.6 ± 10 mUI/ml vs 7.8 ± 7.7 mUI/ml, p = 0.03). No correlation has been found between the severity of anemia and HCV infection. However, HCV (+) hemodialysed patients had higher serum endogenous erythropoietin levels as compared to HCV (-) patients. Further studies are needed to clarify why high endogenous erythropoietin level does not improve anemia in HCV infected hemodialysis patients.

Fibroblast growth factor 23 and acute kidney injury.

Fibroblast growth factor 23 (FGF23), which is produced in bone, participates in the maintenance of phosphate metabolism and can serve as a biomarker for adverse cardiovascular outcomes in patients with chronic kidney disease and end-stage renal disease. Circulating FGF23 rapidly increases after acute kidney injury (AKI), preceding other known markers such as neutrophil gelatinase-associated lipocalin and serum creatinine. The increase in FGF23 in AKI appears to be independent of parathyroid hormone, vitamin D signaling pathways, and dietary phosphate. The potential mechanisms include: (1) increased production of FGF23 in the bone by yet-to-be-identified factors; (2) ectopic production of FGF23 by injured renal tubules; and (3) decreased renal clearance of circulating FGF23. Circulating FGF23 determined by intact FGF23 enzyme-linked immunosorbent assay (ELISA) is a more reliable biomarker of AKI than FGF23 C-terminal ELISA (a mixed readout of C-terminal fragment and intact FGF23). Given that FGF23 can be ectopically expressed in differentiated renal tubules and iron modulates FGF23 metabolism, an effect of iron on FGF23 expression in renal tubules is conceivable but remains to be confirmed. More clinical and experimental studies are required to validate the use of circulating FGF23 as a biomarker for the early identification of AKI and prediction of short- and long-term adverse outcomes post-AKI. More importantly, the biologic effect of increased FGF23 in AKI needs to be defined.

Marked Hyperferritinemia Does Not Predict for Hemophagocytic Lymphohistiocytosis (HLH) in the Adult Population

IntroductionMarkedly elevated serum ferritin levels are often thought to be exclusively seen in rheumatologic and inflammatory disorders including adult-onset Still’s disease (AOSD), macrophage activation syndrome (MAS), and hemophagocytic lymphohistocytosis (HLH). In the pediatric population, ferritin >10,000 µg/L is highly sensitive and specific for HLH (Allen CE, et al. Pediatr Blood Cancer. 2008). In order to determine what conditions are associated with profoundly elevated ferritin levels in the adult population, we performed a retrospective analysis of patients with serum ferritin >50,000 µg/L in a large academic healthcare system.MethodsUsing a centralized clinical data registry, we searched for all patients at Brigham and Women’s Hospital (BWH), Massachusetts General Hospital (MGH), and Faulkner Hospital (FH) who had serum ferritin levels >50,000 µg/L between 8/1/1988-1/29/2014 for BWH and MGH, and 8/9/1995-1/29/2014 for FH. For patients who had multiple ferritin levels >50,000 µg/L, we used the highest value. Patients <18 years old were excluded and the medical charts of the remaining patients were manually reviewed. We collected demographic data including age, race, and gender. Each patient was categorized as having HLH, MAS, rheumatologic/inflammatory disorder, hematologic malignancy, solid tumor, renal failure, sepsis/infection, liver failure, iron overload, hemolytic anemia, or none of these conditions. Causes were not considered mutually exclusive. Univariate and multivariable linear regressions were used to evaluate the association of those factors with ferritin levels modeled using the logarithmic transformation of the actual ferritin levels. The Wilcoxon rank sum test was also performed along with the univariate analysis. Those covariates statistically significant at an alpha level of 0.2 were included in the multivariable stepwise linear regression model. In the multivariable analysis, a p-value <0.05 was considered statistically significant.ResultsWe identified 113 total patients with ferritin levels above 50,000 µg/L at BWH, MGH, and FH during the study period. The patients ranged from 20 years old to 88 years old (mean 56 years old). The majority of patients were male (58%). The racial distribution included White (75%), Black (16%), Hispanic (4%), Asian (2%), and unknown (3%). Ferritin levels ranged from 50,129 to 439,500 µg/L with a median of 82,930 µg/L (notably, two patients had ferritin levels more than the assay of >100,000 µg/L). The characteristics of the patients with ferritin levels >50,000 µg/L included renal disease (65%), liver disease (54%), sepsis/infection (46%), hematologic malignancy (33%), rheumatologic/inflammatory conditions (18%), HLH (17%), iron overload (11%), hemolytic anemia (4%), solid malignancy (4%), and MAS (3%) (table 1). The majority of patients with HLH were thought to have secondary HLH triggered by a malignancy (9/19) or infection (6/19), with the minority having no clear precipitating cause (4/19). All 9 patients with malignancy-associated HLH had a hematologic malignancy (4/9 had T-cell lymphoma, 3/9 had DLBCL, 1/9 had transformed AML, and 1/9 had erythroleukemia). The stepwise linear regression showed that presence of hemolytic anemia was the only variable associated with a differential ferritin level (β=1.51, p=0.03).ConclusionsExtremely elevated ferritin levels are associated with many conditions and most often seen in patients with hematologic malignancies, liver failure, and renal failure. Although less common in our cohort, those patients with hemolytic anemia had higher ferritin levels compared to patients without hemolytic anemia. In contrast to what has been described in the pediatric population, there does not appear to be any value above which ferritin is specific for HLH in the adult population. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Identifying Renal Microstructural and Blood Flow Changes in Sickle Cell Disease Using Quantitative MRI Techniques

BACKGROUND: In patients with sickle cell disease (SCD), renal disease is strikingly high, with up to one third of adults developing chronic kidney disease. Renal disease is associated with considerable morbidity and mortality although it has not been as well studied as other SCD-related complications. Studies which could provide insight into clinically silent renal injury are limited. To date, no large studies of functional or structural renal changes have been published, particularly in the context of sickle cell crises. Magnetic Resonance Imaging (MRI) is safe and capable of providing quantitative assessments of disease without ionizing radiation. Clinically available MRI techniques have already been used to assess both acute and chronic kidney disease. This is the first study to evaluate these MRI techniques in SCD renal disease.OBJECTIVE: The objective of this pilot study was to evaluate the capability of quantitative MRI techniques to assess both structural changes and renal blood flow in patients with SCD.METHODS: Pediatric and adult patients were recruited to an IRB approved study from the Sickle Cell Anemia Centers at UH Rainbow Babies & Children’s Hospital and University Hospitals Case Medical Center. Six pediatric patients (age 12-18 years, 1 male, 5 female, eGFR 110-185 mL/min/1.73m2) have been recruited to date. Healthy controls were also recruited. Diffusion (DTI) and Arterial Spin Labeling (ASL) scans were performed to assess medullary microstructure and cortical perfusion (non-contrast), respectively. Apparent Diffusion Coefficient (ADC) and Fractional Anisotropy (FA) maps were calculated using established methods. Mean renal T2* was measured to assess iron burden. A Student’s t-test was performed to compare the mean renal DTI and ASL results between subjects and healthy controls.RESULTS: In the six pediatric subjects, medullary FA values were decreased, suggesting degradation of medullary microstructure. Both cortical ADC and medullary FA were significantly reduced, indicative of microstructural changes in SCD subjects compared to controls. ASL perfusion maps, likewise, showed reduced cortical perfusion for SCD subjects in comparison to controls. Preliminary differences in blood flow and microstructure were seen between SCD subjects on hydroxyurea (HU) and those not on HU. Analysis of the T2* data showed increased renal iron deposition that did not appear to correlate with serum ferritin.CONCLUSIONS: Our results suggest that quantitative diffusion and ASL MRI are sensitive to medullary microstructural and cortical blood flow changes in pediatric patients with SCD. Preliminarily, iron deposition in the kidney appears to relate to chronic hemolysis rather than transfusional iron. These sensitive techniques may be useful in assessing the impact of therapeutic interventions, such as HU, on renal function in children. There may also be utility in assessing the impact of acute hemolysis/sickling on renal microstructure and perfusion. DisclosuresPiccone:Novartis Pharmaceuticals Cooperation: Speakers Bureau.

EFFICACY AND SAFETY OF IRON (III)–HYDROXIDE SUCROSE COMPLEX IN CORRECTION OF ANEMIA STAGE 5D CHRONIC KIDNEYDISEASE HEMODIALYSIS PATIENTS NOT TREATED BY ERYTHROPOIESIS–STIMULATING AGENTS (PROSPECTIVEANALYSIS)

The aim of study was to evaluat the efficacy and safety ofSUFER® (iron (III) sucrose complex) in correction of anemia stage 5D chronic kidney disease hemodialysis patients.&#x0D; Methods. This study included thirty patients undergoing regular hemodialysis (mean age 48,81±3,24 years, mean duration of dialysis 30,43±9,25 months) with renal anemia and iron deficiency. All patients were treated with SUFER® intravenously 200 mg three times a week. Correction dose was determined according to the manufacturer’s recommendations.&#x0D; Results. Mean level offerritin was significantly increased from 125,15 ± 21,46 ng / ml to 375,56 ± 64,12 ng / ml (p &lt;0,001), transferrin saturation – from 17,48 ± 2,71% to 38,21 ± 4,90 ng / ml (p &lt;0,001). 23 (76.67%) patients achieved target levels offerritin and transferrin, 12 (40% ) adverse events.&#x0D; Conclusions. SUFER® is effective and safety drug in dialysis patients.

Angiotensin II alters the expression of duodenal iron transporters, hepatic hepcidin, and body iron distribution in mice.

Angiotensin II (ANG II) has been shown to affect iron metabolism through alteration of iron transporters, leading to increased cellular and tissue iron contents. Serum ferritin, a marker of body iron storage, is elevated in various cardiovascular diseases, including hypertension. However, the associated changes in iron absorption and the mechanism underlying increased iron content in a hypertensive state remain unclear. The C57BL6/J mice were treated with ANG II to generate a model of hypertension. Mice were divided into three groups: (1) control, (2) ANG II-treated, and (3) ANG II-treated and ANG II receptor blocker (ARB)-administered (ANG II-ARB) groups. Mice treated with ANG II showed increased serum ferritin levels compared to vehicle-treated control mice. In ANG II-treated mice, duodenal divalent metal transporter-1 and ferroportin (FPN) expression levels were increased and hepatic hepcidin mRNA expression and serum hepcidin concentration were reduced. The mRNA expression of bone morphogenetic protein 6 and CCAAT/enhancer-binding protein alpha, which are regulators of hepcidin, was also down-regulated in the livers of ANG II-treated mice. In terms of tissue iron content, macrophage iron content and renal iron content were increased by ANG II treatment, and these increases were associated with reduced expression of transferrin receptor 1 and FPN and increased expression of ferritin. These changes induced by ANG II treatment were ameliorated by the administration of an ARB. Angiotensin II (ANG II) altered the expression of duodenal iron transporters and reduced hepcidin levels, contributing to the alteration of body iron distribution.

Pathophysiology of the Belgrade rat.

The Belgrade rat is an animal model of divalent metal transporter 1 (DMT1) deficiency. This strain originates from an X-irradiation experiment first reported in 1966. Since then, the Belgrade rat’s pathophysiology has helped to reveal the importance of iron balance and the role of DMT1. This review discusses our current understanding of iron transport homeostasis and summarizes molecular details of DMT1 function. We describe how studies of the Belgrade rat have revealed key roles for DMT1 in iron distribution to red blood cells as well as duodenal iron absorption. The Belgrade rat’s pathology has extended our knowledge of hepatic iron handling, pulmonary and olfactory iron transport as well as brain iron uptake and renal iron handling. For example, relationships between iron and manganese metabolism have been discerned since both are essential metals transported by DMT1. Pathophysiologic features of the Belgrade rat provide us with a unique and interesting animal model to understand iron homeostasis.

P528 RESTLESS LEG SYNDROME (RLS) IS ASSOCIATED WITH HEPATIC ENCEPHALOPATHY (HE) IN DECOMPENSATED CIRRHOSIS. A CLINICAL PILOT STUDY

Objective: The prevalence of RLS is close to 10% in the general population; which adversely affects the quality of life (QOL). Exact etiology of RLS is still unknown. Iron deficiency, small intestinal bacterial overgrowth (SIBO) and inflammatory bowel disease (IBD) have clear associations with RLS. Decompensated cirrhosis with portal hypertension has multiorgan involvement causing minimal and overt encephalopathy, sleep disturbances ( dysnomia, parasomnia) and stupor; all of which has a clear association with sub acute bacterial peritonitis ( SBP ) which has a precipitating clinical state along with SIBO. This clinical study evaluates the association of RLS in HE amongst decompensated cirrhotics. Methods: One hundred and eight (n=108) patients were recruited and subdivided into three subgroups. Group A (n=36) decompensated cirrhotic (mean MELD of 16, OHE 20/36 (55%), MHE 16/36 (44%), esophageal varices grade II 24/36 (67%). Group B (n=36) chronic liver disease without cirrhosis with mean MELD 6: Alcohol related 9/36 (25%), NASH 12/ 36 (33%), HCV 12/36 (33%), HBV 1/36 (3%) and AIH 2/36 (6%). Group C (n=36) healthy controls. Initially all received Xifaxan 550mg orally twice daily for 10 days to eradicate coexisting SIBO. All underwent Methane breath test for SIBO. Baseline labs: Serum levels for renal function, ferritin, iron studies, hemoglobin and hematocrit, ammonia, Celiac and IBD serology, stool lactoferrin & calprotectin and urine for toxicology screening were collected. Groups A and B underwent neuro-psychometric and flicker testing for MHE and OHE and sleep testing for RLS (with Mayo RLS questionnaire). Exclusion: Chronic iron deficiency, Celiac disease, IBD, major depression, IBS, benzodiazepines, narcotics, alcohol, anti-psychotics and use of illicit drugs. Results: Group A 24/36 (67%) had RLS: [ OHE 16/20 (80%), MHE 8/16 (50%), esophageal varices 8/10 (80%), alcoholic cirrhotic 10/14 (71%), CHC 3/ 6 (50%), NASH 3/6 (50%) and SIBO 14/36 (39%) ]. Group B 1/36 (3%) RLS and SIBO 7/ 36 (19%). Group C : 2/36 (6%) RLS and SIBO 3/36 (8%). All individuals were confirmed by sleep study and RLS questionnaire. Serum ammonia has no impact on RLS. Conclusion: This clinical trial postulates that decompensated cirrhotics with Hepatic encephalopathy have high incidence of RLS with portal hypertension. Larger trials will validate this finding.

Resistance exercise decreases iron absorption without deteriorating the iron status in rats (907.2)

To investigate the effects of resistance exercise on the dietary iron balance and the iron status, rats were divided into a sedentary (S) or an exercise (E) group and given different diets in terms of energy and iron sufficiency for three weeks. The energy‐deficient groups consumed 70% of the energy that was consumed by the energy‐sufficient groups. The iron‐deficient groups consumed a diet containing half of the iron (20 mg Fe/kg diet) of a standard diet. The E groups showed significantly higher fecal iron excretion compared to the S groups, thus resulting in a significantly lower apparent absorption, regardless of the energy and iron sufficiency of the diet. The iron status in the blood was not significantly lower in the E groups than the S groups. The iron content in the liver, heart and spleen did not differ between the E and S groups. The renal iron content of the rats that consumed the diet with sufficient energy but deficient iron was significantly higher in the E group than in the S group, while no such differences were observed in the kidneys in the groups that consumed other diets. These results suggest that, although resistance exercise decreases the dietary iron absorption, it does not appear to worsen the iron status in rats.