Unexpected Cause of Persistent Microcytosis and Neurological Symptoms in a Child: Niemann-Pick Disease Type C.

Atypical microcytic anemias are rare diseases of iron/heme metabolism that can be diagnostically challenging. We report the case of a 2-year-old twin boy with neurodevelopmental delay and persistent microcytosis in whom atypical microcytic anemias was initially suspected. He had low blood iron and transferrin saturation with normal/high ferritin despite iron therapy. Hemoglobinopathies were excluded by conventional/DNA studies. Hepcidin was high but iron-refractory-iron-deficiency anemia was ruled out by a genetic panel. Bone marrow aspiration revealed foamy cells and iron depletion. A genetic study confirmed the diagnosis of Niemann-Pick disease type C which was finally considered the origin of microcytosis through anemia of chronic disease.

Microcytic to hypochromic ratio as a discriminant index of thalassaemia trait in subjects with hypochromic anaemia.

INTRODUCTION: Differentiating between thalassaemia and iron deficiency anaemia (IDA) in hypochromic anaemia is a challenge to pathologists as it influences the choice of subsequent specialized confirmatory tests. In this study, we aimed to evaluate the performance of microcytic to hypochromic ratio (MicroR/ Hypo-He, M/H ratio) as a discriminant index in hypochromic anaemia. MATERIALS AND METHODS: A retrospective study was carried out on 318 subjects with hypochromic anaemia, which comprised 162 IDA and 156 thalassaemia trait subjects with α-thalassemia, ß-thalassemia and HbE trait. Optimal cut-off value, sensitivity and specificity of M/H ratio for thalassaemia trait discrimination was determined using Receiver Operating Characteristic (ROC) analysis. RESULTS: Subjects with thalassaemia trait showed higher MicroR compared to IDA ( p< 0.001) while subjects with IDA demonstrated higher Hypo-He than thalassaemia trait (p < 0.001). M/H ratio was significantly higher in thalassaemia trait compared to IDA, with medians of 3.77 (interquartile range: 2.57 - 6.52) and 1.73 (interquartile range: 1.27 - 2.38), respectively (p < 0.001). M/H ratio ≥ 2.25 was the optimal cut-off value for discriminating thalassaemia trait from IDA in hypochromic anaemia, with the area under ROC curve (AUC) of 0.83, sensitivity of 80.8% and specificity of 71.6%. CONCLUSIONS: M/H ratio is a useful discriminant index to distinguish thalassaemia trait from IDA in hypochromic anaemia prior to diagnostic analysis for thalassaemia confirmation. High M/H ratio is suggestive of thalassaemia trait than of IDA. However, more studies are required to establish the role of M/H ratio as a screening tool for thalassaemia discrimination in hypochromic anaemia.

α-Globin Genotypes Associated with Hb H Disease: A Report from Oman and a Review of the Literature from the Eastern Mediterranean Region.

α-Thalassemia (α-thal) is the most common autosomal recessive hemoglobinopathy. There is a vast diversity and geographical variability in underlying genotypes in Hb H (ß4) patients. Herein, we describe the genotypes found in the largest report of Omani Hb H patients. Moreover, we reviewed and summarized the literature published from the Eastern Mediterranean region. A retrospective review of all genetically confirmed Hb H disease patients diagnosed between 2007 and 2017 at Sultan Qaboos University Hospital, Muscat, Oman, was performed. Hematological parameters and clinical presentations were assessed. Both α-globin genes were screened for deletional and nondeletional mutations using a stepwise diagnostic strategy as described before. A total of 52 patients (27 females and 25 males) with a mean age of 20.6 years (range 0.23-80.0) were molecularly confirmed to carry Hb H disease. The patients had a hemoglobin (Hb) level of 9.3 g/dL (range 5.7-13.0) and mean corpuscular volume (MCV) of 58.4 fL (range 48.2-82.1). A total of eight genotype combinations were identified, with α2 polyadenylation signal mutation (polyA1) (AATAAA>AATAAG (αPA1α/αPA1α), often cited as αT-Saudiα/αT-Saudiα, being the most common (53.8%) followed by -α3.7/- -MED I (28.8%). Our cohort also included patients with combinations of αPA1 with other Hb variants: αPA1α/αPA1α with Hb S (HBB: c.20A>T) trait (n = 2), -α3.7/αPA1α (n = 2) and αcodon 19α (HBA2: c.56delG)/αPA1α (n = 1). Nondeletional Hb H disease due to the αPA1 mutation is the most common in Omanis. Molecular diagnosis is necessary for accurate confirmation of the diagnosis of α-thal, determination of underlying genotypes, follow-up and counseling.

Nondeletional α-Thalassemia: Two New Mutations on the α2 Gene.

About 10.0% of α-thalassemia (α-thal) cases are due to point mutations, small deletions, or insertions of one or more bases on the α genes that can alter mRNA processing at the transcription, translation, or post-translation level; these cases are called nondeletional α-thalassemias (α-thal). Most occur within the domain of the α2 gene without changes in the expression of the α1 gene. We present two new frameshift mutations on the HBA2 gene, associated with a nondeletional α-thal phenotype. The probands were referred to our clinic because of persistent microcytosis and hypochromia. The molecular characterization was performed by automatic sequencing of the α-globin genes. Two new mutations were detected on the HBA2 gene; HBA2: c.85delG, p.(Ala29fs*21), and HBA2: c.268_280delCACAAGCTTCGGG, p.(His90Trpfs*9). These new mutations cause a change of the reading frame, the first on codon 28 and the second from codons 89 to 93. In the first mutation, the result is an altered amino acid sequence and a premature termination codon at position 87, while the elimination of 13 bp generates a protein of 95 residues and in this case, the premature termination codon is at position 96. These types of mutation are among the most damaging changes to the coding of a protein. Not only do they lead to changes in the length of the polypeptide, but they also vary the chemical composition, which would result in a nonfunctional protein. The importance of identifying these new mutations lies in their possible association with α0-thal, which could lead to a severe thalassemia.

Association between Different Polymorphic Markers and ß-Thalassemia Intermedia in Central Iran.

ß-Thalassemia intermedia (ß-TI) is a clinical condition characterized by moderate, non transfusional anemia and hepatosplenomegaly. The main objective of this study was to determine the molecular basis of the clinical phenotype of ß-TI in Iran. To elucidate the mild phenotype of many patients with ß-TI, we screened for three prevalent ß-globin gene mutations [IVS-II-1 (G>A) HBB: c.315+1G>A, IVS-I-110 (G>A) HBB: c.93-21G>A and IVS-I-5 (G>C) [HBB: c.92+5G>C], deletions on the α-globin genes, XmnI polymorphisms and restriction fragment length polymorphism (RFLP) haplotypes on the ß-globin gene cluster in 50 ß-TI patients. Fifty-eight percent of the patients (29 cases) were associated with the mentioned mutations. We showed that the HBB: c.315+1G>A mutation is linked to haplotype [+ - + +] (57.69%). This haplotype is in linkage disequilibrium with the XmnI polymorphism (NG_000007.3: g.42677C>T) and has been associated with increased expression of Hb F in ß-TI patients. The XmnI polymorphism is defined in association with this prevalent mutation. Two patients had a single α-globin gene deletion [-α3.7 (rightward) deletion]. The main genetic factor in mild phenotype ß-TI patients is the linkage of an XmnI polymorphism (NG_000007.3: g.42677C>T) with the HBB: c.315+1G>A (80.76%), which is associated with increased production of Hb F and coinheritance of haplotype [+ - + +] with ß-TI, especially with the homozygous HBB: c.315+1G>A mutation. Molecular basis of ß-TI could be explained by the involvement of different factors that tend to develop the disease phenotype.

La patologización del celibato en la medicina española (1820-1920) / The pathologization of celibacy in spanish medicine (1820-1920)

En la España de la Restauración, la novela naturalista y la prensa anticlerical insistían en que la causa principal de la depravación sexual de los sacerdotes era el voto de castidad. ¿En qué medida las tecnologías de saber y de poder médicos de la época permitían defender esa tesis? Este es el asunto abordado en el artículo. En primer lugar se examinan los antecedentes ilustrados de la ofensiva higienista contra el celibato sexual. En segundo lugar se analiza la controversia suscitada por Monlau con su defensa higiénica de la castidad sexual. Los argumentos de Monlau tienen lugar en un contexto de propaganda a favor del celibato suscitada por la Iglesia Católica. En tercer lugar se exploran los argumentos médicos que conectaban causalmente la continencia absoluta con las desviaciones sexuales, en particular la pederastia. Por último se indican las circunstancias que, a comienzos del siglo XX, llevaron a reactivar la defensa médica de la abstinencia sexual entre los jóvenes, anunciando un nuevo prototipo de masculinidad

In the Spain of the Restoration, the naturalist novel and the anticlerical press insisted that the main cause of the sexual depravity of priests was the vow of chastity. To what extent the technologies of knowledge and medical power of the time allowed to defend that thesis? This is the issue addressed in the article. First, we examine the enlightened background of the hygienist offensive against sexual celibacy. Secondly, the controversy raised by Monlau with his hygienic defense of sexual chastity is analyzed. The arguments of Monlau take place in a context of propaganda in favor of celibacy raised by the Spanish Catholic Church. Third, the medical arguments that causally connected absolute continence with sexual deviations, particularly "pederasty", are explored. Finally, we indicate the circumstances that, at the beginning of the 20th century, led to reactivate the medical defense of sexual abstinence among young people, announcing a new pattern of masculinity

The AMP-activated protein kinase beta 1 subunit modulates erythrocyte integrity.

Failure to maintain a normal in vivo erythrocyte half-life results in the development of hemolytic anemia. Half-life is affected by numerous factors, including energy balance, electrolyte gradients, reactive oxygen species, and membrane plasticity. The heterotrimeric AMP-activated protein kinase (AMPK) is an evolutionarily conserved serine/threonine kinase that acts as a critical regulator of cellular energy balance. Previous roles for the alpha 1 and gamma 1 subunits in the control of erythrocyte survival have been reported. In the work described here, we studied the role of the beta 1 subunit in erythrocytes and observed microcytic anemia with compensatory extramedullary hematopoiesis together with splenomegaly and increased osmotic resistance.

NCOA4 Deficiency Impairs Systemic Iron Homeostasis.

The cargo receptor NCOA4 mediates autophagic ferritin degradation. Here we show that NCOA4 deficiency in a knockout mouse model causes iron accumulation in the liver and spleen, increased levels of transferrin saturation, serum ferritin, and liver hepcidin, and decreased levels of duodenal ferroportin. Despite signs of iron overload, NCOA4-null mice had mild microcytic hypochromic anemia. Under an iron-deprived diet (2-3 mg/kg), mice failed to release iron from ferritin storage and developed severe microcytic hypochromic anemia and ineffective erythropoiesis associated with increased erythropoietin levels. When fed an iron-enriched diet (2 g/kg), mice died prematurely and showed signs of liver damage. Ferritin accumulated in primary embryonic fibroblasts from NCOA4-null mice consequent to impaired autophagic targeting. Adoptive expression of the NCOA4 COOH terminus (aa 239-614) restored this function. In conclusion, NCOA4 prevents iron accumulation and ensures efficient erythropoiesis, playing a central role in balancing iron levels in vivo.

Intestinal DMT1 is critical for iron absorption in the mouse but is not required for the absorption of copper or manganese.

Divalent metal-ion transporter-1 (DMT1) is a widely expressed iron-preferring membrane-transport protein that serves a critical role in erythroid iron utilization. We have investigated its role in intestinal metal absorption by studying a mouse model lacking intestinal DMT1 (i.e., DMT1(int/int)). DMT1(int/int) mice exhibited a profound hypochromic-microcytic anemia, splenomegaly, and cardiomegaly. That the anemia was due to iron deficiency was demonstrated by the following observations in DMT1(int/int) mice: 1) blood iron and tissue nonheme-iron stores were depleted; 2) mRNA expression of liver hepcidin (Hamp1) was depressed; and 3) intraperitoneal iron injection corrected the anemia, and reversed the changes in blood iron, nonheme-iron stores, and hepcidin expression levels. We observed decreased total iron content in multiple tissues from DMT1(int/int) mice compared with DMT1(+/+) mice but no meaningful change in copper, manganese, or zinc. DMT1(int/int) mice absorbed (64)Cu and (54)Mn from an intragastric dose to the same extent as did DMT1(+/+) mice but the absorption of (59)Fe was virtually abolished in DMT1(int/int) mice. This study reveals a critical function for DMT1 in intestinal nonheme-iron absorption for normal growth and development. Further, this work demonstrates that intestinal DMT1 is not required for the intestinal transport of copper, manganese, or zinc.

Abnormal erythroid maturation leads to microcytic anemia in the TSAP6/Steap3 null mouse model.

Genetic ablation of the ferrireductase STEAP3, also known as TSAP6, leads to severe microcytic and hypochromic red cells with moderate anemia in the mouse. However, the mechanism leading to anemia is poorly understood. Previous results indicate that TSAP6/Steap3 is a regulator of exosome secretion. Using TSAP6/Steap3 knockout mice, we first undertook a comprehensive hematologic characterization of the red cell compartment, and confirmed a dramatic decrease in the volume and hemoglobin content of these erythrocytes. We observed marked anisocytosis as well as the presence of fragmenting erythrocytes. Consistent with these observations, we found by ektacytometry decreased membrane mechanical stability of knockout red cells. However, we were unable to document significant changes in the expression levels of the major skeletal and transmembrane proteins to account for this decrease in the membrane stability. Furthermore, there were no differences in red cell survival between wild type and knockout animals. However, when we monitored erythropoiesis, we found a decreased number of proerythroblasts in the bone marrow of TSAP6/Steap3(-/-) animals. In addition, progression from the proerythroblastic to the orthochromatic stage was affected, with accumulation of cells at the polychromatic stage. Altogether, our findings demonstrate that abnormal erythroid maturation is the main cause of anemia in these mice.

DMT1-mutant erythrocytes have shortened life span, accelerated glycolysis and increased oxidative stress.

BACKGROUND/AIMS: Deficiency of the divalent metal transporter 1 (DMT1) leads to hypochromic microcytic anemia. We have previously shown that DMT1 deficiency impairs erythroid differentiation and induces apoptosis of erythroid cells. Here we analyzed metabolic processes and survival of mature erythrocytes in order to address potential involvement of erythrocyte defect in the pathophysiology of the disease. METHODS: FACS analysis was used to determine the half-life of erythrocytes (CFSE fluorescence), phosphatidylserine exposure (Annexin V binding), cytosolic Ca(2+) (Fluo3/AM fluorescence) and reactive oxygen species (ROS; DCF fluorescence). Enzyme activities were determined by standard biochemical methods. The concentration of ATP and ADP was measured on HPLC-MS/MS. RESULTS: We observed an accelerated clearance of CFSE-labeled DMT1-mutant erythrocytes from circulating blood when compared to wild-type erythrocytes. In vitro, DMT1-mutant erythrocytes showed significantly increased Annexin V binding after exposure to hyperosmotic shock and glucose depletion. Despite exaggerated anti-oxidative defense, higher ROS levels were present in DMT1-mutant erythrocytes. Accelerated anaerobic glycolysis and reduced ATP/ADP ratio detected in DMT1-mutant erythrocytes indicate enhanced demand for ATP. CONCLUSIONS: We propose that DMT1 deficiency negatively affects metabolism and life span of mature erythrocytes; two other aspects of defective erythropoiesis which contribute to the pathophysiology of the disease.

Microcytic hypochromic anemia: should high performance liquid chromatography be used routinely for screening anemic and antenatal patients?

BACKGROUND: Hemoglobinopathies are the most common inherited red cell disorders worldwide. Identification of these disorders is immensely important epidemiologically and for improved management protocols. AIM AND OBJECTIVES: Our aim was to determine the prevalence of hemoglobinopathies in patients with microcytic hypochromic anemia and to assess the suitability of using high performance liquid chromatography (HPLC) routinely for screening antenatal cases and patients with anemia. MATERIALS AND METHODS: A total of 4335 cases received from Mar 2007 to Nov 2011 were studied for various hemoglobinopathies and variants on BIO RAD 'VARIANT' analyzer. RESULTS: Of the 4335 cases studied, 2119 were antenatal cases, 1710 patients with other disorders and 506 family studies. Of these, 688 cases displayed abnormal hemoglobin fractions on HPLC of which 140 were antenatal women. There were 455 cases of ß thalassemia trait, 24 ß thalassemia major, 20 thalassemia inter-media, 54 sickle cell trait, fivesickle cell disease, 21 double heterozygous ß thalassemia-sickle cell trait, nineand 4 Hb D- Punjab heterozygous and homozygous respectively, three Hb D ß Thalassemia trait, 20 and 37 Hb E homozygous and heterozygous respectively, three Hb E ß Thalassemia trait and four cases of Hb Q India. Twenty nine adults had isolated HbF elevation. CONCLUSION: Our study found a high prevalence (15.8%) of hemoglobinopathies amongst microcytic hypochromic anemia and antenatal cases. An accurate diagnosis helps in preventing unnecessary iron loading. Screening all antenatal cases with anemia helps in timely antenatal counseling, thus preventing the psychological trauma of bearing a transfusion dependent child for life.

Biomarkers of hypochromia: the contemporary assessment of iron status and erythropoiesis.

Iron status is the result of the balance between the rate of erythropoiesis and the amount of the iron stores. Direct consequence of an imbalance between the erythroid marrow iron requirements and the actual supply is a reduction of red cell hemoglobin content, which causes hypochromic mature red cells and reticulocytes. The diagnosis of iron deficiency is particularly challenging in patients with acute or chronic inflammatory conditions because most of the biochemical markers for iron metabolism (serum ferritin and transferrin ) are affected by acute phase reaction. For these reasons, interest has been generated in the use of erythrocyte and reticulocyte parameters, available on the modern hematology analyzers. Reported during blood analysis routinely performed on the instrument, these parameters can assist in early detection of clinical conditions (iron deficiency, absolute, or functional; ineffective erythropoiesis, including iron restricted or thalassemia), without additional cost. Technological progress has meant that in recent years modern analyzers report new parameters that provide further information from the traditional count. Nevertheless these new parameters are exclusive of each manufacturer, and they are patented. This is an update of these new laboratory test biomarkers of hypochromia reported by different manufactures, their meaning, and clinical utility on daily practice.

Zebrafish erythropoiesis and the utility of fish as models of anemia.

Erythrocytes contain oxygen-carrying hemoglobin to all body cells. Impairments in the generation of erythrocytes, a process known as erythropoiesis, or in hemoglobin synthesis alter cell function because of decreased oxygen supply and lead to anemic diseases. Thus, understanding how erythropoiesis is regulated during embryogenesis and adulthood is important to develop novel therapies for anemia. The zebrafish, Danio rerio, provides a powerful model for such study. Their small size and the ability to generate a large number of embryos enable large-scale analysis, and their transparency facilitates the visualization of erythroid cell migration. Importantly, the high conservation of hematopoietic genes among vertebrates and the ability to successfully transplant hematopoietic cells into fish have enabled the establishment of models of human anemic diseases in fish. In this review, we summarize the current progress in our understanding of erythropoiesis on the basis of zebrafish studies and highlight fish models of human anemias. These analyses could enable the discovery of novel drugs as future therapies.

Disseminated abscessation complicated with bone marrow abscess caused by Arcanobacterium pyogenes in a goat.

An 8-month-old, Nubian wether with a history of systemic illness was euthanatized for a pathological examination. At necropsy, the presence of disseminated abscessation and cellulitis in the limbs was noted. Other postmortem findings associated with the visceral disease in this animal included multiple abscess lesions, mainly in the lungs, kidneys, phalanxes and vertebrae. Histopathologically, lesions of arteriolitis were found as evidenced by bacterial embolisms in pulmonary and renal arteriola, indicating a bacteremia in the patient. Arcanobacterium pyogenes was consistently isolated from 8 lesions of abscessations, including the lesions of subcutaneous abscesses as well as bone marrow abscess in phalanxes and thoracic vertebrae. This is the first published report of disseminated arcanobacterial infection with bone marrow abscess of both the phalanxes and vertebrae in goat.

Chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature (CANDLE) syndrome.

Several syndromes manifest as recurrent daily fevers, skin lesions, and multisystem inflammation. We describe 4 patients with early-onset recurrent fevers, annular violaceous plaques, persistent violaceous eyelid swelling, low weight and height, lipodystrophy, hepatomegaly, and a range of visceral inflammatory manifestations. Laboratory abnormalities included chronic anemia, elevated acute-phase reactants, and raised liver enzymes. Histopathologic examination of lesional skin showed atypical mononuclear infiltrates of myeloid lineage and mature neutrophils. Our patients have a distinctive early-onset, chronic inflammatory condition with atypical or immature myeloid infiltrates in the skin. We propose the acronym CANDLE (chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature) syndrome for this newly described disorder, which is probably genetic in origin.