Clinical features and gene mutation spectrum in children with sideroblastic anemia / 中国当代儿科杂志

OBJECTIVE@#To study the clinical features and gene mutation spectrum of children with sideroblastic anemia (SA) and the clinical value of targeted next-generation sequencing in the molecular diagnosis of children with SA.@*METHODS@#Clinical data were collected from 36 children with SA. Targeted next-generation sequencing was used to detect mutations in SA-related pathogenic genes and genes associated with heme synthesis and mitochondrial iron metabolism. The association between genotype and clinical phenotype was analyzed.@*RESULTS@#Of the 36 patients, 32 had congenital sideroblastic anemia (CSA) and 4 had myelodysplastic syndrome with ring sideroblasts (MDS-RS). Mutations in CSA-related genes were detected in 19 children (19/36, 53%), among whom 9 (47%) had ALAS2 mutation, 4 (21%) had SLC25A38 mutation, and 6 (32%) had mitochondrial fragment deletion. No pathogenic gene mutation was detected in 4 children with MDS-RS. Among the 19 mutations, 89% (17/19) were known mutations and 11% (2/19) were novel mutations. The novel mutation of the ALAS2 gene c.1153A>T(p.I385F) was rated as "possibly pathogenic" and the novel mutation of the SLC25A38 gene c.175C>T(p.Q59X) was rated as "pathogenic".@*CONCLUSIONS@#ALAS2 and SLC25A38 gene mutations are commonly seen in children with CSA, but mitochondrial gene fragment deletion also accounts for a relatively high proportion. For children with hypoplastic anemia occurring in infancy, mitochondrial disease should be considered.

A Novel Hemizygous I418S Mutation in the ALAS2 Gene in a Young Korean Man with X-Linked Sideroblastic Anemia

No abstract available.

Congenital sideroblastic anemia-a new family with identification of K156E mutation of ALAS2 gene and literature review / 中华血液学杂志

<p><b>OBJECTIVE</b>To raise awareness of molecular pathogenesis and treatment of congenital sideroblastic anemia (CSA).</p><p><b>METHODS</b>A complete blood count and iron metabolism were detected from the proband and other members of the family. Mutation analysis was performed on the complete coding regions of ALAS2 gene by common polymerase chain reaction (PCR) using genomic DNA as a template from members the family. ALAS2 mutations were detected by direct sequencing and mutation types were confirmed by sequencing followed by plasmid cloning.</p><p><b>RESULTS</b>The proband male presented with microcytic hypochromic anemia (hemoglobin 84 g/L, mean corpuscular volume 64 fL, mean corpuscular hemoglobin 16.5 pg), and iron overload (serum iron 44.7 μmol/L, serum ferritin 3 123 μg/L and transferrin saturation 0.84). A mutation 466 A>G predicting a Lys156Glu amino acid change was identified in the proband and 3 females from the family. The proband was hemizygous for this mutation and presented with microcytic anemia and iron overload, while all 3 heterozygous females showed marginally increased red cell distribution width without any other symptoms. The proband treated with 300 mg of pyridoxine per day and iron chelation therapy with deferoxamine for one year had durable hematopoietic patients improvements, including increase in hemoglobin to 98 g/L and decrease in serum ferritin to 1 580 μg/L.</p><p><b>CONCLUSION</b>This was a novel K156E substitution in ALAS2 gene identified in a 3-generation pedigree in China. Our findings emphasized the importance of gene based diagnosis of CSA, and CSA patient with ALAS2 mutation responded to pyridoxine treatment.</p>

Effect of treadmill exercise and nutrition supplement on activity and gene expression of rate-limiting enzyme of heme metabolism and globin / 中国应用生理学杂志

<p><b>AIM</b>To investigate the possible role of rate-limiting enzyme of heme metabolism and globin in the development of the low hemoglobin (Hb), red blood (cell) count (RBC) and hematocrit (Hct) after long-term exercise, and effect of nutrition supplement on sports anemia.</p><p><b>METHODS</b>Male Wistar rats were randomly assigned to three groups (n = 10): control (C), exercise (P) and exercise + nutrition (G). Animals in the P and G groups started treadmill running at 30 m/min, 0% grade, 1 min/time. Running time was gradually increased with 2 min/time during initial 5 weeks and final 4 weeks. In addition, running frequency was 2 times/day except initial 2 weeks. At the end of eleventh week, gene expression of 5-aminolevulinate synthase (ALAS), ferrochelatase, alpha-globin and beta-globin in bone marrow were measured with RT-PCR. Mean-while heme oxygenase 1 (HO-1) activity in liver was measured with immunohistochemical method.</p><p><b>RESULTS</b>Eleven weeks of exercise induced a significant increase in HO-1 and a significant increase in gene expression of beta-globin (P < 0.01, P < 0.05, respectively). Treatment with anti-sports anemia compound dosage led to no significant differences in rate-limiting enzyme of heme metabolism and globin in the exercised rats. The G group had a significantly higher HO-1 level in liver than the C group (P < 0.01). These finds showed that exercise was associated with no significant difference in heme synthetase and alpha-globin gene expression, and significant difference in heme catabolic enzyme and beta-globin gene expression.</p><p><b>CONCLUSION</b>The increase of HO-1 activity in liver might be one of the causes of the lower Hb, RBC and Hct status in exercised rats.</p>

Purification and production of the extracellular 5-aminolevulinate from recombiniant Escherichia coli expressing yeast ALAS / 生物工程学报

Aminolevulinic acid (ALA) is biosynthesized by the enzyme ALA synthase (ALAS). The ALA production has been studied using the overproducing ALAS from several bacteria in Escherchia coil, respectively. However, ALAS from eucaryote expressed in E. coli for producing ALA in the culture is not known. The extracellular ALA production and cell growth were investageted respectively using the recombinant E. coli overproducing Saccharomyces cerevisiae ALAS in shake-flask fermentations. The ALAS activity from the cell extract was assayed. The extracellular ALA was purified by the national-made large-dimension resins and confirmed by the capillary electrophoresis measurements. At 12h after induction at 37 degrees C, the extracellular ALA production was up to 162mg per liter LB culture at initial pH 6.5 with exogenous levulinate, succinate and and glycine at the concentration of 20 mmol/L respectively. The purity of ALA after purification is up to 90%.

Progress of study on sideroblastic anemia and its possible gene therapy--review / 中国实验血液学杂志

It was thought that delta-aminolevulinate synthase (ALAS) is the rate-limiting enzyme in the heme biosynthetic pathway. Actually there are two isozymes of ALAS and ALAS2 (erythroid delta-aminolevulinate synthase), they play the leading role in the hemoglobin biosynthetic pathway. Mutations in ALAS2 gene causes X-linked sideroblastic anemia (XLSA). About 25 different mutations in ALAS2 gene have been identified in XLSA patients and two of them were reported by our laboratory. It is possible to cure the patients with XLSA by gene therapy because it is a single gene disorder.

Evaluation of protective property of vitamin C supplementation against experimentaly induced lead toxicity in male albino rats

To investigate the protective property of vitamin C upon lead induced toxicity in male albino rats. Subjects: The biological effects of lead exposure at 500 ppm for 7 weeks in drinking water upon 40 male albino rats were investigated with and without 5% vitamin C supplementation in normal rat chow in four groups: group [I] is the negative control group, group [II] rats were supplemented with normal rat chow containing 5% vitamin C, group [III] rats were given 500 ppm lead acetate in drinking water and group [IV] rats were given 500 ppm lead acetate in drinking water and 5% vitamin C in normal rat chow. It was observed that lead content in the lead exposed group was significantly increased in kidneys, liver, brain, RBC's and serum by 18.6, 7.5, 7.3, 8.3 and 4.1-fold, respectively as compared with negative control group indicating that the kidney was the most deteriorated organ in lead toxicity with elevated serum creatinine level. With 5% vitamin C supplementation in the lead exposed group, the lead content in these organs and fluids significantly decreased by 45, 61, 31, 58 and 39%, respectively as compared with the lead exposed group. Also with vitamin C supplementation, AST, ALT and creatinine in serum significantly decreased by 22, 23 and 29%, respectively with a concomitant significant increase in delta ALAD activity, HB and HCT by 52, 35 and 53%, respectively as compared to the lead exposed group but with non-significant changes in serum iron as compared with both negative control and lead exposed groups. Current search amplify the beneficial protective effect of vitamin C supplementation against lead induced toxicity

Construction of recombinant vector expressing ALAS2 gene in X-linked sideroblastic anemia / 中国实验血液学杂志

X-linked sideroblastic anemia (XLSA) is caused by mutations of erythroid-specific 5-aminolevulinate synthetase (ALAS2) gene. In this study a eukaryotic expression vector of ALAS2 was constructed and transfected into eukaryotic cells to observe the expression of ALAS2 gene. The full length cDNA of ALAS2 gene was inserted into plasmid pDs-red2-N1, named pDs-red2-N1/ALAS2. Then, the vector was transfected into K562 cells via electroporation. At 48 hours after transfection, total RNA from K562 cells was extracted, expressions of ALAS2 gene and protein with red fluorescence in the K562 cells were detected by RT-PCR and flow cytometry, respectively. The vector was also transfected into COS 7 cells via liposome. Both mRNA and protein expression in COS7 cells were detected by RT-PCR and fluorescence microscopy. The result showed that after the pDs-red2-N1/ALAS2 eukaryotic expression vector was digested by KpnI and BamHI, two fragments of 4 700 bp and 1 764 bp were displayed by electrophoresis on agarose gel. Sequence method confirmed that the sequence was correct. RT-PCR amplified the total RNA extracted from the transfected K562 and COS7 cells, and could find mRNA of ALAS2 gene that can't be found in K562 and COS7 cells usually. The expressions of both fluorescein and ALAS2 were significantly increased. The percentage of positive cells reached about 19.2% and 10.7%, respectively. ALAS2 expression lasted for 10 days in COS7 cells and the peak was at the third day. It is concluded that the eukaryotic expression vector of ALAS2 gene is successfully constructed; K562 and COS7 cells transfected with the vector via electroporation and liposome can express ALAS2 protein. So, the vector has the potential in gene replacement and can be used for patients with XLSA in future.

The effect of lead bioaccumulation on haem biosynthetic enzymes in fish.

The bioaccumulations of lead in the liver and hepatic microsomes of fish after 1, 3, 7, 14, 28, and 45 days exposure were studied. In addition, the relationship between the bioaccumulated lead in both hepatic microsomes and the liver and their haem biosynthetic enzymes were studied. Lead toxicity was shown to result in a depression of the microsomal mixed function oxidase system, as assessed by a decrease in hepatic microsomal cytochrome P-450 and b5 content and by a decrease in the activity of the enzymes aniline hydroxylase and aminopyrine demethylase. Lead had a more marked effect on cytochrome P-450 than b5. The activity of the rate-limiting enzyme of haem biosynthesis, delta-aminolevulinic acid synthase, was inversely correlated with the microsomal cytochrome P-450 content. The activity of the heam biosynthetic enzymes delta-aminolevulinic acid dehydratase, coproporphyrinogen oxidase and ferrochelatase were decreased by increasing lead pretreatment. The activity of the haem catabolic enzyme, haem oxygenase, was increased by concentration and length of time to lead exposure.

Efecto de la tioctamina frente a la acción del hexaclorobenceno / Effect of the thioctamide in relation the hexachlorobenzene action

Se estudió el efecto de la intoxicación crónica con hexaclorobenceno en ratas, con y sin administración simultánea de tioctamida. En el grupo que recibió hexaclorobenceno solo, se produjo el esperado desarrollo de porfiria incrementándose la excreción urinaria y el contenido hepático de porfirinas y disminuyendo la actividad Uroporfirinógeno decarboxilasa. El contenido hepático de dienos conjugados no varió, en tanto que el de malondialdehido se incrementó en un grado estadísticamente no significativo. Estos resultados indicarían la existencia de un ligero proceso de peroxidación lipídica. La tioctamida (25 mg/Kg de peso) produjo efectos nocivos antes que protectores, detectados por un aumento de la actividad transaminasa glutámico pirúvica y una inhibición a nivel de la primera etapa de la Uroporfirinógeno decarboxilasa. Los resultados indicarían que: 1) altas dosis de tioctamida producen un decremento en la actividad Uroporfirinógeno decarboxilasa, enmascarando quizás su posible efecto protector frente a la acción del hexaclorobenceno por radicales libres; 2) la Uroporfirinógeno decarboxilasa es un parámetro más sensible que la medición de dienos conjugados o de melondialdehido para ensayar la producción de radicales libres por acción del hexaclorobenceno in vivo. De ser así, la tioctamida, ensayada a dosis menores y no tóxicas, a través de su habilidad como atrapante de radicales libres, quizás pueda proteger contra la acción del hexaclorobenceno.

Some hematological features of lead - intoxication

The goal of this study is to demonstrate the effect of exposure to lead on some hematological parameters. The study was carried out on twenty male rats exposed to different doses of lead acetate [one and two mg/kg body weight respectively]. Blood samples were taken after intraperitoneal injection for one week [acute effect] and eight weeks [chronic effect]. With low dose and after one week exposure statistically significant elevation were encountered with respect to blood lead level, Hb A[2]% and Hb F%. After eight weeks exposure significant reduction of erythrocyte count, hemoglobin concentration, erythrocyte delta amino-levulinic dehydratase and pyrimidine -5-nucleotidase were detected, with significant elevation of erythrocyte zinc protoporphyrin. Rats exposed to higher dose had obvious significant effect on the above mentioned parameters specially after prolonged exposure, with significant increase of Hb A[2]%

Synergistic effect of allyl isopropyl acetamide with cadmium on hepatic heme synthesis in chick embryo.

The effect of cadmium along with a porphyrogenic drug, allyl isopropyl acetamide, on the induction of 5-amino levulinic acid (ALA) synthetase, ALA dehydratase and heme level was studied. The interaction of cadmium with allyl isopropyl acetamide indicated that the decrease in hepatic heme level by cadmium or allyl isopropyl acetamide may occur in a synergistic manner, whereas the induction of ALA synthetase by cadmium or allyl isopropyl acetamide may not take place in the same manner. Further, neither allyl isopropyl acetamide treatment alone nor allyl isopropyl acetamide-cadmium treatment had any effect on ALA dehydratase activity.

Effect of triiodothyronine in the regulation of haem biosynthetic pathway

La administración de triyodotironina a animales tiroidectomizados, disminuyó en un 50% el contenido de citocromo P-450. La actividad de hemo oxigenasa no se modificó por el tratamiento con triyodotironina, ya sea solo o con una dosis subóptima de Cl2Co. Bajo las mismas condiciones la actividad de la amino levulínico sintelasa no fue afectada. La triyodotironina produjo un incremento del 100% en la actividad de triptófano pirrolasa. Tanto la holo como la enzima total fueron aumentadas en el mismo grado. La actividad de la porfobilinógeno deaminasa-uroporfirinógeno co-sintetasa, fue inducida en los animales tratados con triyodotironina, en un 67% por sobre los valores de los animales tiroidectomizadosm y sólo 32% con respecto a los animales con operación simulada. Nuestros resultados sugieren que bajo estimulación por triyodotironina, la disminución en el contenido de citocromo P-450 no es debida a un aumento en la velocidad de degradación del hemo, sino a una disociación de éste para incrementar el "pool" celular del hemo, y saturar en parte a la nueva apotriptófano pirrolasa sintetizada

Evaluacion de la capacidad de la desferrioxamina para revertir una severa porfiria inducida por hexaclorobenceno / Evaluation of desferrioxamine capacity to revert a severe porphyrid induced by hexachlorobenzene

El objeto del presente trabajo es investigar la capacidad de la desferrioxamina (DF) como quelante de hierro para disminuir o revertir una porfiria experimental severa inducida por hexaclorobenceno (HCB) en ratas. La DF se comienza a administrar después de 17 semanas de intoxicación con HCB y se continúa hasta la semana 27. Se cuantifican semanalmente excreciones en orina de ácido aminolevúlico (ALA), profobilinógeno y porfirinas. Al final del experimento se sacrifican los animales y se determinan porfirinas hepáticas y actividad de ALA sintetasa y porfirinógeno carboxi-liasa. Los resultados indican que el quelante adminsitrado no mejora el disturbio provocado por el HCB sobre el camino metabólico del hemo en las presentes condiciones. Se comparan estos resultados con los obtenidos cuando la DF se da conjuntamente con el HCB desde el comienzo de la administración del fungicida, situación en que sí el quelante retarda y atenúa el efecto porfirinogénico del HCB. Se discute el papel del hierro en la metabolización del HCB

interrelationship between Zinc; Copper and Iron in Lead intoxicated subjects

Inhibition of Hb biosynthesis in lead intoxicated subjects appears as a result of disorder in the interaction of Cu, Zn and Fe ions on delta-Am-inolevulinic acid dehydratase. [ALA-D]. Elevation of free serum copper and decrease of serum iron and zinc are the main cause of ALAD inhibition. Thus, it is safe to suggest that administration of Zn orally leads to replacement of cu from catalytic sites of this enzyme which is known as a zinc dependent enzyme

Progesterone, its A/B cis reduction and delta aminolevulinic acid synthetase in the developing hen

La 5 ß reductasa (Rasa) aumenta después de la eclosión en tejidos esteroidogénicos de la gallina en desarrollo, especialmente en el ovario derecho. La sintetasa del ácido delta aminolevulínico (ALAs) es más activa en estos tejidos que en hígado durante la mayoría de los estadios embrionarios. Pero después de la eclosión sólo aumenta en forma aguda ALAs hepática y adrenal; en ovario izquierdo dicha enzima crece moderadamente y en ovario derecho desciende a valores muy bajos. Cierta relación entre las curvas de ALAs y Rasa durante el desarrollo embrionario del ovario izquierdo y la adrenal sugieren que la 5 ß pregnanediona fuera un inductor natural de la ALAs en estas glándulas endocrinas funcionantes, por lo menos durante sus estadios embrionarios

Adaptación de Rhodopseudomonas palustris a cambios de condiciones atmosféricas / Adaptation of Rhodopseudomonas palustris to changes in atmospheric conditions

El burbujeo de O2 95%-N2 5% a un cultivo de Rhodopseudomonas palustris crecido en condiciones fotosintéticas provoca un cese inmediato de la síntesis de bacterioclorofila y una disminución de la actividad de la enzima delta-aminolevulinato sintetasa (ALA-S), sin alterar el crecimiento bacteriano. Al cesar el gaseado el nivel enzimático se recupera rápidamente pero el de bacterioclorofila lo hace más lentamente. El agregado de cloramfenicol al cesar el gaseado, no afecta la recuperación de la actividad enzimática. En cambio, si se lo agrega desde el inicio del gaseado, la actividad igual decae pero su recuperación es menor que en ausencia del antibiótico. La enzima de extractos de células gaseadas muestra activación espontánea a 40-C Se postula el efecto de distintos activadores sobre diversas formas de la enzima ALA-S para explicar estos hechos