In vivo self-aggregation and efficient preparation of recombinant lichenase based on ferritin / 生物工程学报

Enzyme separation, purification, immobilization, and catalytic performance improvement have been the research hotspots and frontiers as well as the challenges in the field of biocatalysis. Thus, the development of novel methods for enzyme purification, immobilization, and improvement of their catalytic performance and storage are of great significance. Herein, ferritin was fused with the lichenase gene to achieve the purpose. The results showed that the fused gene was highly expressed in the cells of host strains, and that the resulted fusion proteins could self-aggregate into carrier-free active immobilized enzymes in vivo. Through low-speed centrifugation, the purity of the enzymes was up to > 90%, and the activity recovery was 61.1%. The activity of the enzymes after storage for 608 h was higher than the initial activity. After being used for 10 cycles, it still maintained 50.0% of the original activity. The insoluble active lichenase aggregates could spontaneously dissolve back into the buffer and formed the soluble polymeric lichenases with the diameter of about 12 nm. The specific activity of them was 12.09 times that of the free lichenase, while the catalytic efficiency was 7.11 times and the half-life at 50 ℃ was improved 11.09 folds. The results prove that the ferritin can be a versatile tag to trigger target enzyme self-aggregation and oligomerization in vivo, which can simplify the preparation of the target enzymes, improve their catalysis performance, and facilitate their storage.

Cardiac dysfunction and ferritin as early markers of severity in pediatric sepsis / Disfunção cardíaca e a ferritina como marcadores precoces de gravidade na sepse pediátrica

Abstract Objective: The aim of this study was to verify the association of echocardiogram, ferritin, C-reactive protein, and leukocyte count with unfavorable outcomes in pediatric sepsis. Methods: A prospective cohort study was carried out from March to December 2014, with pediatric critical care patients aged between 28 days and 18 years. Inclusion criteria were diagnosis of sepsis, need for mechanical ventilation for more than 48 h, and vasoactive drugs. Serum levels of C-reactive protein, ferritin, and leukocyte count were collected on the first day (D0), 24 h (D1), and 72 h (D3) after recruitment. Patients underwent transthoracic echocardiography to determine the ejection fraction of the left ventricle on D1 and D3. The outcomes measured were length of hospital stay and in the pediatric intensive care unit, mechanical ventilation duration, free hours of VM, duration of use of inotropic agents, maximum inotropic score, and mortality. Results: Twenty patients completed the study. Patients with elevated ferritin levels on D0 had also fewer ventilator-free hours (p = 0.046) and higher maximum inotropic score (p = 0.009). Patients with cardiac dysfunction by echocardiogram on D1 had longer hospital stay (p = 0.047), pediatric intensive care unit stay (p = 0.020), duration of mechanical ventilation (p = 0.011), maximum inotropic score (p = 0.001), and fewer ventilator-free hours (p = 0.020). Conclusion: Cardiac dysfunction by echocardiography and serum ferritin value was significantly associated with unfavorable outcomes in pediatric patients with sepsis.

Resumo Objetivo: Verificar a associação do ecocardiograma, da ferritina, da proteína C reativa (PCR) e da contagem de leucócitos com desfechos desfavoráveis na sepse pediátrica. Métodos: Estudo de coorte prospectivo, de março a dezembro de 2014, com pacientes críticos pediátricos entre 28 dias e 18 anos. Critérios de inclusão foram diagnóstico de sepse, necessidade de ventilação mecânica (VM) por mais de 48 horas e uso de drogas vasoativas. Avaliaram‐se os níveis séricos PCR, ferritina, contagem de leucócitos, no recrutamento (D0), 24 horas (D1) e 72 horas (D3) após o recrutamento. No D1 e no D3 todos os pacientes foram submetidos a ecocardiograma transtorácico para determinação da Fração de Ejeção (FE) do ventrículo esquerdo. Os desfechos avaliados foram tempo de internação hospitalar e na Unidade de Terapia Intensiva Pediátrica (UTIP); duração da VM; horas livres de VM; duração do uso de inotrópicos; escore de inotrópicos máximo e mortalidade. Resultados: Vinte pacientes completaram o estudo. Ferritina elevada no D0 associou‐se com menor tempo livre de ventilação (p = 0,046) e maior escore de inotrópicos máximo (p = 0,009). A disfunção cardíaca pelo ecocardiograma no D1 relacionou‐se com maior tempo de internação hospitalar (p = 0,047), de UTIP (p = 0,020), VM total (p = 0,011), escore de inotrópicos máximo (p = 0,001) e menor tempo livre de VM (p = 0,020). Conclusão: A disfunção cardíaca pelo ecocardiograma e o valor de ferritina sérica associaram‐se significativamente com desfechos desfavoráveis nos pacientes pediátricos com sepse.

Body Iron Stores and Cardiovascular Health.

A substantial body of epidemiological and experimental data suggests the significance of serum iron as an important cardiovascular risk factor. There are many conflicting reports on the hypothesis that body iron stores are associated with risk of coronary heart disease. The likely mechanism by which iron may play a pathogenic role in cardiovascular disease has been postulated. The recent discovery of the HFE C282Y mutation commonly seen in hereditary hemochromatosis and its reported association with increased risk of coronary heart disease has increased the need to evaluate its role in the development of cardiovascular disease. Overall, understanding the role of iron in relation to cardiovascular health may be an important tool to help stratify risk for cardiovascular disease.

Genetic characterization of Toxoplasma gondii isolates from eared doves (Zenaida auriculata) in Brazil / Caracterização genética de isolados de Toxoplasma gondii de pombos (Zenaida auriculata) no Brasil

Eared doves (Zenaida auriculata), which are common in urban, rural and wild areas in many regions of Brazil, are frequently prey for domestic cats. Therefore Toxoplasma gondii isolates obtained from doves may reflect greater environmental diversity than those from other hosts. The aim of the present study was to evaluate T. gondii seroprevalence, isolate and genotype strains from Z. auriculata. Serum and tissue samples were collected from 206 doves for use in the modified agglutination test (MAT) and mouse bioassay. The prevalence of T. gondii antibodies in the doves was 22.3% (46/206), with titers ranging from 16 to 4096, and T. gondii strains were isolated from 12 of these doves. Five genotypes were detected by means of PCR-RFLP, including ToxoDB genotypes #1, #6, #17 and #65, and one genotype that had not previously been described (ToxoDB#182). This was the first report on isolation of T. gondii from Z. auriculata. This study confirmed the genetic diversity of T. gondii isolates and the existence of clonal type II (ToxoDB genotype #1) in Brazil.

Pombos silvestres (Zenaida auriculata), comuns em áreas urbanas, rurais e selvagens em muitas regiões do Brasil, são frequentemente predados por gatos domésticos. Sendo assim, os isolados de T. gondii obtidos de pombos podem refletir uma maior diversidade ambiental do que os outros hospedeiros. O objetivo do presente estudo foi avaliar a soroprevalência, isolar e genotipar T. gondii de Z. auriculata. Amostras de soro e tecido foram coletadas de 206 pombos para o teste de aglutinação modificado (MAT) e o bioensaio em camundongos. A prevalência de anticorpos contra T. gondii em pombos foi 22,3% (46/206), com títulos variando de 16 a 4096, e T. gondii foi isolado de 12 pombos. Cinco genótipos foram detectados por PCR-RFLP, incluindo os genótipos ToxoDB #1, #6, #17, #65 e um genótipo não descrito anteriormente (ToxoDB#182). Esse é o primeiro relato de isolamento de T. gondii de Z. auriculata. Este estudo também confirmou a diversidade dos isolados de T. gondii e a presença de tipo clonal II (ToxoDB #1) no Brasil.

Glial cell activity is maintained during prolonged inflammatory challenge in rats

We evaluated the expression of glial fibrillary acidic protein (GFAP), glutamine synthetase (GS), ionized calcium binding adaptor protein-1 (Iba-1), and ferritin in rats after single or repeated lipopolysaccharide (LPS) treatment, which is known to induce endotoxin tolerance and glial activation. Male Wistar rats (200-250 g) received ip injections of LPS (100 µg/kg) or saline for 6 days: 6 saline (N = 5), 5 saline + 1 LPS (N = 6) and 6 LPS (N = 6). After the sixth injection, the rats were perfused and the brains were collected for immunohistochemistry. After a single LPS dose, the number of GFAP-positive cells increased in the hypothalamic arcuate nucleus (ARC; 1 LPS: 35.6 ± 1.4 vs control: 23.1 ± 2.5) and hippocampus (1 LPS: 165.0 ± 3.0 vs control: 137.5 ± 2.5), and interestingly, 6 LPS injections further increased GFAP expression in these regions (ARC = 52.5 ± 4.3; hippocampus = 182.2 ± 4.1). We found a higher GS expression only in the hippocampus of the 6 LPS injections group (56.6 ± 0.8 vs 46.7 ± 1.9). Ferritin-positive cells increased similarly in the hippocampus of rats treated with a single (49.2 ± 1.7 vs 28.1 ± 1.9) or repeated (47.6 ± 1.1 vs 28.1 ± 1.9) LPS dose. Single LPS enhanced Iba-1 in the paraventricular nucleus (PVN: 92.8 ± 4.1 vs 65.2 ± 2.2) and hippocampus (99.4 ± 4.4 vs 73.8 ± 2.1), but had no effect in the retrochiasmatic nucleus (RCA) and ARC. Interestingly, 6 LPS increased the Iba-1 expression in these hypothalamic and hippocampal regions (RCA: 57.8 ± 4.6 vs 36.6 ± 2.2; ARC: 62.4 ± 6.0 vs 37.0 ± 2.2; PVN: 100.7 ± 4.4 vs 65.2 ± 2.2; hippocampus: 123.0 ± 3.8 vs 73.8 ± 2.1). The results suggest that repeated LPS treatment stimulates the expression of glial activation markers, protecting neuronal activity during prolonged inflammatory challenges.

Iron and ferritin changes in multiple organs failure after trauma / 法医学杂志

OBJECTIVE@#To investigate the changes of iron content in serum and liver, ferritin content in serum, percentage of myeloperoxidase (MPO) positive granulocyte in rabbits after different serious trauma and to explore the relationship between these changes and multiple organ failure (MOF).@*METHODS@#Rabbit trauma models were established. Iron content in serum and liver, ferritin content in serum and the percentage of MPO positive granulocyte were measured at different time after trauma.@*RESULTS@#After trauma, iron content in serum decreased sharply in early period (12-36h) and increased gradually to normal level in mild traumatic group after 60 h. Iron content in serum remained lower level in severe traumatic and death group 60 h after trauma. Iron content in liver obviously increased in death group. The changes of ferritin content in serum in mild traumatic were not obvious. Ferritin contents in serum in severe injury group and death group were slightly higher in early period and decreased in later period. The percentage of MPO positive granulocyte increased in early period after trauma. The percentage began to decrease 6 d after trauma and returned to normal level in mild traumatic group. The percentage obviously was significantly lower than normal levels in severe traumatic group and death group 6 d after trauma. Some rabbits died 60 h-6 d after severe trauma, and the pathological changes in the other organs were consistent with MOF.@*CONCLUSION@#Trauma can cause the serum iron, ferritin levels and percentage of MPO positive granulocyte changes. Severe trauma can cause uncompensated changes of these indicators, which could be the main mechanisms of MOF and death.

Purification and characterization of mycoferritin from Aspergillusflavus MTCC 873.

The fungus Aspergillus flavus MTCC 873, a non-toxigenic isolate demonstrated its capability to synthesize mycoferritin (MF) upon induction with iron in yeast extract sucrose (YES) medium. The molecular mass, yield, iron and carbohydrate contents of the MF were 440 kDa, 0.015 mg/g of wet mycelia, 0.8 and 30.4%, respectively. Native gel-electrophoresis revealed a band corresponding to dimeric form of equine spleen ferritin (ESF). Subunit analysis by SDS-PAGE revealed a single protein band with an apparent molecular mass of 24 kDa, suggesting similar sized subunits in the structure of apoferritin shell. Immunological cross-reactivity was observed with the anti-fish liver ferritin. Transmission electron microscopy (TEM) revealed an apparent particle size of 100 Å. N-terminal amino acid sequence of MF revealed a sequence of SLPLQDYA, which showed identities with other eukaryotic ferritin sequences. The spectral characteristics (UV/VIS, fluorescence and circular dichroic spectra) were similar to ESF. The fungus, unlike A. parasiticus 255 (non-toxigenic) was incapable of producing aflatoxins, when grown in YES media.

Serum biochemical markers in rheumatoid arthritis.

Rheumatoid arthritis (RA) characterized by local and systemic effects of inflammation has a wide range of biochemical markers implicated directly or indirectly to its pathogenesis. In the present study, homocysteine, cortisol, adenosine deaminase (ADA), ferritin, malondialdehyde (MDA) and -tocopherol in serum of RA patients and healthy individuals were estimated to assess if they contribute to the disease process. The markers of disease activity such as erythrocyte sedimentation rate (ESR) and rheumatoid factor (RF) were also measured. The study group included a total of 45 subjects, including 30 RA patients and the rest being healthy individuals. RA group showed a significant increase in the levels of homocysteine, ADA and MDA, and a significant decrease in α-tocopherol compared to the healthy individuals. However, cortisol and ferritin levels did not show any significant change. Also, there was no significant correlation between the studied serum markers and markers of disease activity. Our results indicate that these biochemical markers contribute independently to the pathogenesis of RA.

Low dose 'Sprinkles'-- an innovative approach to treat iron deficiency anemia in infants and young children.

Iron supplementation programs using pediatric tablets or drops have not been successful in the control of anemia amongst infants and children in India. Sprinkles is an innovative multi-micronutrient home fortification strategy to control iron deficiency and anemia. OBJECTIVE: We aimed to determine the hematologic response to different doses and forms of iron in Sprinkles and iron drops. SETTING: Twenty two villages of Vadu Rural Health Program, KEM Hospital, Pune. DESIGN: Double blind clustered randomized community-based trial. SUBJECTS: Children (n=432) aged 6 to 18 mo age with Hb between 70 to 100 g/L were enrolled. METHODS: Selected villages were randomized into 5 groups: Sprinkles 12.5, 20 or 30 mg ferrous fumarate, Sprinkles 20 mg micronized ferric pyrophosphate or drops 20 mg ferrous glycine sulphate (DROPS) for 8 weeks. Household socio-demographic information was collected at baseline. Side effects and compliance were monitored through weekly visits. Hemoglobin was estimated at baseline, 3 and 8 weeks. Ferritin was assessed at baseline and 8 weeks. RESULTS: Baseline characteristics were similar across all groups. Hemoglobin increased significantly (P<0.0001) in all groups at 8 weeks with no difference between groups. Ferritin increased (P<0.0001) significantly in all groups with no difference across the groups. Compliance (overall range: 42 to 62 %) was lowest for DROPS. Side effects were significantly higher among DROPS compared to Sprinkles (p>0.05). CONCLUSIONS: Sprinkles 12.5 mg FF dose is as efficacious as higher doses of iron in Sprinkles or DROPS in increasing hemoglobin. Sprinkles FF 12.5 mg is recommended as it has fewer reported side effects and better compliance compared to DROPS.

Antioxidative response mechanisms in halophytes: their role in stress defence.

Normal growth and development of plants is greatly dependent on the capacity to overcome environmental stresses. Environmental stress conditions like high salinity, drought, high incident light and low or high temperature cause major crop losses worldwide. A common denominator in all these adverse conditions is the production of reactive oxygen species (ROS) within different cellular compartments of the plant cell. Plants have developed robust mechanisms including enzymatic or nonenzymatic scavenging pathways to counter the deleterious effects of ROS production. There are a number of general reviews on oxidative stress in plants and few on the role of ROS scavengers during stress conditions. Here we review the regulation of antioxidant enzymes during salt stress in halophytes, especially mangroves. We conclude that (i) antioxidant enzymes protect halophytes from deleterious ROS production during salt stress, and (ii) genetic information from mangroves and other halophytes would be helpful in defining the roles of individual isoforms. This information would be critical in using the appropriate genes for oxidative stress defence for genetic engineering of enhanced stress tolerance in crop systems.

Iron homeostasis in the lung

Iron is essential for many aspects of cellular function. However, it also can generate oxygen-based free radicals that result in injury to biological molecules. For this reason, iron acquisition and distribution are tightly regulated. Constant exposure to the atmosphere results in significant exposure of the lungs to catalytically active iron. The lungs have a mechanism for detoxification to prevent associated generation of oxidative stress. Those same proteins that participate in iron uptake in the gut are also employed in the lung to transport iron intracellularly and sequester it in an inactive form within ferritin. The release of metal is expedited (as transferrin and ferritin) from lung tissue to the respiratory lining fluid for clearance by the mucocilliary pathway or to the reticuloendothelial system for long-term storage. This pathway is likely to be the major method for the control of oxidative stress presented to the respiratory tract.

Effects of Chronic Aluminum Administration on Blood and Liver Iron-Related Parameters in Mice

In this study, the effects of chronically administered aluminum on iron metabolism-related parameters of liver and blood of mice were investigated. An additional purpose to determine how chronic aluminum administration together with vitamin E as an antioxidant to mice changed the parameters related to iron metabolism. For these purposes, we used 21 adult female Balb-c mice in this study. The animals were divided into three groups: one group with aluminum administered chronically, another group with aluminum plus vitamin E administered chronically, and the control group. Serum levels of hemoglobin, ferritin, iron, transferrin, hematocrit, total iron binding capacity (TIBC), as well as percentage of transferrin saturation were determined in all groups. In addition, the liver tissue levels of ferritin and iron were analyzed. Hemoglobin and hematocrit levels of the aluminum group and aluminum plus vitamin E group were significantly decreased compared to the control. In conclusion, no changes occurred in the serum iron related parameters although Al induced anemia in mice when Al administered chronically. There was an increase in the levels of liver iron and ferritin with Al, but Vit E had no effect on the changes of all blood and liver parameters caused by Al.

Ferritina sérica: uso como marcador tumoral para el adenocarcinoma renal: informe preliminar / Serum ferritin: its use as tumor marker for renal adenocarcinoma: preliminary report

Se estudiaron 20 pacientes portadores de adenocarcinoma renal en diversas etapas clínicas, a los cuales se efectuaron determinaciones de ferritina sérica antes del tratamiento efectuado, después del mismo y bimestralmente, durante su seguimiento para investigar su empleo como potencial marcador tumoral. La ferritina sérica se halló elevada en los pacientes con cáncer renal en comparación con el grupo testigo; existió un incremento de la misma al aumentar la etapa clínica así como una disminución en los pacientes con regresión y estabilización de la neoplasia. En los pacientes con progresión aparecieron pruebas de persistencia de la elevación de la ferritina sérica. Según los resultados obtenidos, se justifica considerar a la ferritina sérica como un potencial marcador tumoral para el adenocarcinoma renal

Idiopathic hemochromatosis: a study in a large Puerto Rican family

We studied 13 first-degree relatives in a large family with an index case of idiopathic hemochromatosis to detect the relatives with evidence of iron overloading. Serum iron, total iron binding capacity (TIBC), and serum ferritin levels were measured in all family members. We also performed HLA typing to identify the relatives who are homozygous with the proband and genetically predisposed to develop the disease. The family was composed of the parents and 12 siblings including the index case. The mean age of the siblings was 25 years. None presented with evidence of iron overload by the iron biochemical tests. HLA typing demonstrated six homozygous siblings with the proband. In separate analysis these siblings did not present abnormalities in any of the iron biochemical tests. These homozygous relatives were followed for one year after initial evaluation and none presented abnormalities in the iron studies during this period. These results are contradictory to other previous studies done in families with idiopathic hemochromatosis. The most feasible explanations for these findings are the young age of these siblings and the predominance of females among them. We consider that these homozygous relatives must be followed for their life-times with iron studies to detect a possible increase in iron stores as expected in later ages

Frecuencia de las alteraciones del metabolismo del hierro / Frequency of the iron metabolism disorders

MARCO TEORICO: Los desórdenes del metabolismo del hierro constituyen el problema nutricional de mayor prevalencia en el mundo, sobre todo en países en desarrollo como el nuestro. OBJETIVO: Conocer la frecuencia de las alteraciones del metabolismo del hierro de la población que acude al Laboratorio del Hospital A.B.C. a través de la determinación de Hb en g/dL y el porcentaje de Saturación de transferrina TIPO DE ESTUDIO: Retrospectivo RESULTADOS: Existe deficienia de hierro en promedio en 1 de cada 4 individuos estudiados (p<0.05). CONCLUSION: Es necesario establecer un sistema de detección para el tratamiento específico y oportuno de este problema.

Evalución diagnóstica de la hemoglobina capilar y de los indicadores de nutrición en hierro en la respuesta al tratamiento, con hierro oral en la niñez / Diagnostic evaluation of capillary hemoglobin and iron nutritional parameters in the response to treatment with oral iron in childhood

La anemia por deficiencia de hierro se puede estimar mediante el incremento significativo de las cifras de hemoglobina (Hb) después de la suplementación con hierro. Cincuenta y cinco niños de 6 meses a 3 años de edad seleccionados en base a Hb capilar <= 11,5 g/d1 fueron tratados con hierro oral (3 mg/kg/día) durante 3 meses. En estos niños se analizaron los cambios pre y postratamiento en: Hb, índices eritrocíticos, índice de saturación de la transferrina, protoporfirina-zinc eritrocítica y ferritina sérica. Además se evaluó la capacidad diagnóstica de la Hb capilar, así como de los parámetros mencionados, en la respuesta al tratamiento. Se consideró respuesta ascenso en Hb venosa >= 1 g/dl. La magnitud del ascenso en Hb venosa postratamiento (0 a 4.2 g/dl) mostró correlación inversa con el nivel de Hb basal (r = 0.726). Los cambios promedio post-pretratamiento mostraron diferencias significativas (t pareada) en todas las mediciones de laboratorio estudiadas. En la distribución de frecuencias de los parámetros pretratamiento analizados, la Hb mostró menor traslape de valores entre los niños con y sin respuesta. Empleando a la Hb capilar (<= 11,5 g/dl) como prueba diagnóstica de deficiencia de hierro, se obtuvo respuesta en el 65,5% o sea en 1 de cada 1.5 niños tratados. Este porcentaje de respuestas disminuyó cuando se utilizó como criterio de deficiencia la anormalidad en 2 o más de las mediciones bioquímicas, pero también se redujeron los tratamientos innecesarios. Podemos concluir que la Hb capilar sola permitió identificar el mayor número de niños con respuesta al tratamiento con hierro