SA2RAGE: a new sequence for fast B1+ -mapping.

At high magnetic field strengths (≥ 3T), the radiofrequency wavelength used in MRI is of the same order of magnitude of (or smaller than) the typical sample size, making transmit magnetic field (B1+) inhomogeneities more prominent. Methods such as radiofrequency-shimming and transmit SENSE have been proposed to mitigate these undesirable effects. A prerequisite for such approaches is an accurate and rapid characterization of the B1+ field in the organ of interest. In this work, a new phase-sensitive three-dimensional B1+-mapping technique is introduced that allows the acquisition of a 64 × 64 × 8 B1+-map in ≈ 20 s, yielding an accurate mapping of the relative B1+ with a 10-fold dynamic range (0.2-2 times the nominal B1+). Moreover, the predominant use of low flip angle excitations in the presented sequence minimizes specific absorption rate, which is an important asset for in vivo B1+-shimming procedures at high magnetic fields. The proposed methodology was validated in phantom experiments and demonstrated good results in phantom and human B1+-shimming using an 8-channel transmit-receive array.

Determinants of vitellogenin B1 promoter architecture. HNF3 and estrogen responsive transcription within chromatin.

The liver-specific vitellogenin B1 promoter is efficiently activated by estrogen within a nucleosomal environment after microinjection into Xenopus laevis oocytes, consistent with the hypothesis that significant nucleosome remodeling over this promoter is not a prerequisite for the activation by the estrogen receptor (ERalpha). This observation lead us to investigate determinants other than ERalpha of chromatin structure and transcriptional activation of the vitellogenin B1 promoter in this system and in vitro. We find that the liver-enriched transcription factor HNF3 has an important organizational role for chromatin structure as demonstrated by DNase I-hypersensitive site mapping. Both HNF3 and the estrogen receptor activate transcription synergistically and are able to interact with chromatin reconstituted in vitro with three positioned nucleosomes. We propose that HNF3 is the cellular determinant which establishes a promoter environment favorable to a rapid transcriptional activation by the estrogen receptor.

Transcriptional potentiation of the vitellogenin B1 promoter by a combination of both nucleosome assembly and transcription factors: an in vitro dissection.

The Xenopus laevis vitellogenin B1 promoter was assembled into nucleosomes in an oocyte extract. Subsequent RNA polymerase II-dependent transcription from these DNA templates fully reconstituted in chromatin in a HeLa nuclear extract was increased 50-fold compared with naked DNA. Remarkably, under specific conditions, production of a high level of transcripts occurred at very low DNA (1 ng/microliter) and HeLa nuclear protein (1.6 micrograms/microliters) concentrations. When partially reconstituted templates were used, transcription efficiency was intermediate between that of fully reconstituted and naked DNA. These results implicate chromatin in the process of the transcriptional activation observed. Depletion from the oocyte assembly extract of an NF-I-like factor which binds in the promoter region upstream of the TATA box (-114 to -101) or deletion from the promoter of the region interacting with this factor reduced the transcriptional efficiency of the assembled templates by a factor of 5, but transcription of these templates was still 10 times higher than that of naked DNA. Together, these results indicate that the NF-I-like factor participates in the very efficient transcriptional potentiation of the vitellogenin B1 promoter which occurs during nucleosome assembly.

Spectrally selective B1-insensitive T2 magnetization preparation sequence.

A T(2) magnetization-preparation (T(2) Prep) sequence is proposed that is insensitive to B(1) field variations and simultaneously provides fat suppression without any further increase in specific absorption rate (SAR). Increased B(1) inhomogeneity at higher magnetic field strength (B(0) > or = 3T) necessitates a preparation sequence that is less sensitive to B(1) variations. For the proposed technique, T(2) weighting in the image is achieved using a segmented B(1)-insensitive rotation (BIR-4) adiabatic pulse by inserting two equally long delays, one after the initial reverse adiabatic half passage (AHP), and the other before the final AHP segment of a BIR-4 pulse. This sequence yields T(2) weighting with both B(1) and B(0) insensitivity. To simultaneously suppress fat signal (at the cost of B(0) insensitivity), the second delay is prolonged so that fat accumulates additional phase due to its chemical shift. Numerical simulations as well as phantom and in vivo image acquisitions were performed to show the efficacy of the proposed technique.

Contaminação endógena por Aspergillus spp. em milho pós-colheita no Estado do Paraná

Sessenta amostras de milho pós-colheita foram avaliadas quanto à contaminação fúngica endógena e o potencial toxígeno de espécies do gênero Aspergillus e seus teleomorfos. Quarenta grãos aparentemente sadios de cada amostra foram desinfestados em NaClO e incubados em câmara úmida a 25±1ºC para exteriorização dos fungos, que posteriormente foram isolados em ágar Czapek-Dox. Foram identificadas as espécies Aspergillus flavus, A. parasiticus, Eurotium amstelodami e E. chevalieri. O potencial toxígeno dos fungos A. flavus e A. parasiticus foi avaliado quanto à síntese de aflatoxinas em meio ágar-coco. Espécies do gênero Eurotium foram avaliadas quanto à síntese de esterigmatocistina, nos meios ágar-amendoim e trigo triturado. A porcentagem de grãos contaminados variou entre 0 e 100%, prevalecendo os gêneros Aspergillus, Penicillium e Fusarium. A espécie predominante foi a A. flavus (64%), seguida por E. amstelodami (19%), E. chevalieri (10%) e A. parasiticus (7%). A partir de 109 isolados de A. flavus, evidenciou-se que 73 isolados sintetizaram aflatoxinas B1 e B2, 20 sintetizaram B1, sete sintetizaram B1 e G1, três sintetizaram B1, B2 e G1 e em seis não foi detectada a síntese de aflatoxina. A síntese de esterigmatocistina pelas espécies E. amstelodami e E. chevalieri não foi detectada.

A nuclear factor I-like activity and a liver-specific repressor govern estrogen-regulated in vitro transcription from the Xenopus laevis vitellogenin B1 promoter.

A hormone-controlled in vitro transcription system derived from Xenopus liver nuclear extracts was exploited to identify novel cis-acting elements within the vitellogenin gene B1 promoter region. In addition to the already well-documented estrogen-responsive element (ERE), two elements were found within the 140 base pairs upstream of the transcription initiation site. One of them, a negative regulatory element, is responsible for the lack of promoter activity in the absence of the hormone and, as demonstrated by DNA-binding assays, interacts with a liver-specific transcription factor. The second is required in association with the estrogen-responsive element to mediate hormonal induction and is recognized by the Xenopus liver homolog of nuclear factor I.

Concentrações séricas de minerais e funções hepática e renal de frangos intoxicados com aflatoxina e tratados com montmorilonita sódica

O objetivo deste trabalho foi avaliar as concentrações séricas de minerais e as funções hepática e renal de frangos de corte intoxicados, experimentalmente, com 3 ppm de aflatoxina, e submetidos a diferentes concentrações de montmorilonita sódica na dieta. Foram utilizados 720 frangos, machos, da linhagem Cobb, divididos em seis tratamentos: T1, dieta normal; T2, dieta com aflatoxina (3 ppm); T3, dieta com montmorilonita sódica (0,25%); T4, dieta com aflatoxina (3 ppm) + montmorilonita sódica (0,25%); T5, dieta com montmorilonita sódica (0,5%); T6, dieta com aflatoxina (3 ppm) + montmorilonita sódica (0,5%), com seis repetições por tratamento. A dieta com 3 ppm de aflatoxina (T2) resultou em uma diminuição significativa na concentração sérica de proteínas totais, albumina, globulinas e aspartato amino transferase; a concentração sérica de ácido úrico diminuiu, significativamente, na dieta com aflatoxina (3 ppm) + momtmorilonita sódica a 0,25%; ocorreu diminuição significativa nas concentrações séricas de fósforo na dieta com aflatoxina + montmorilonita sódica a 0,5%. A aflatoxina na concentração de 3 ppm altera a função hepática de frangos de corte; o uso da montmorilonita sódica, na concentração de 0,5%, é eficaz na prevenção dos efeitos tóxicos da aflatoxina, mas diminui os níveis séricos de fósforo.

Vitellogenin genes A1 and B1 are linked in the Xenopus laevis genome.

Genomic clones containing the Xenopus laevis vitellogenin gene B1 have been isolated from DNA libraries and characterized by heteroduplex mapping in the electron microscope, restriction endonuclease analysis, and in vitro transcription in a HeLa whole-cell extract. Sequences from the 3'-flanking region of the previously isolated A1 vitellogenin gene were found in the 5'-flanking region of this B1 gene. Thus, the two genes are linked, with 15.5 kilobase pairs of DNA between them. Their length is about 22 kilobase pairs (A1 gene) and 16.5 kilobase pairs (B1 gene) and they have the following arrangement: 5'-A1 gene-spacer-B1 gene-3'. The analysis of heteroduplexes formed between the two genes revealed several regions of homology. Both genes are in the same orientation and, therefore, are transcribed from the same DNA strand. The possible events by which the vitellogenin gene family arose in Xenopus laevis are discussed.

Regulation of the vitellogenin gene B1 promoter after transfer into hepatocytes in primary cultures.

The estrogen-dependent and tissue-specific regulation of the Xenopus laevis vitellogenin gene B1 promoter has been studied by lipid-mediated DNA transfer into Xenopus hepatocytes in primary culture. Hepatocytes achieve an efficient hormonal control of this promoter through a functional interaction between the estrogen responsive elements and a promoter proximal region upstream of the TATA box, which is characterized by a high density of binding sites for the transcription factors CTF/NF-1, C/EBP and HNF3. DNA accessibility to restriction enzymes within the chromosomal copy of the vitellogenin gene B1 promoter shows that the estrogen responsive unit and the promoter proximal region are sensitive to digestion in uninduced and estrogen-induced hepatocytes but not in erythrocyte nuclei. Together, these findings support the notion that chromatin configuration as well as the interplay of promoter elements mediate proper hormone-dependent and tissue-specific expression of the B1 vitellogenin gene.

cis- and trans-acting elements of the estrogen-regulated vitellogenin gene B1 of Xenopus laevis.

Vitellogenin genes are expressed under strict estrogen control in the liver of female oviparous vertebrates. Gene transfer experiments using estrogen-responsive cells have shown that the 13 bp perfect palindromic element GGTCACTGTGACC found upstream of the Xenopus laevis vitellogenin gene A2 promoter mediates hormonal stimulation and thus, was called the estrogen-responsive element (ERE). In the Xenopus vitellogenin genes B1 and B2 there are two closely adjacent EREs with one or more base substitutions when compared to the consensus ERE GGTCANNNTGACC. On their own, these degenerated elements have only a low or no regulatory capacity at all but act together synergistically to form an estrogen-responsive unit (ERU) with the same strength as the perfect palindromic 13 bp element. Analysis of estrogen receptor binding to the gene B1 ERU revealed a cooperative interaction of receptor dimers to the two adjacent imperfect EREs which most likely explains the synergistic stimulation observed in vivo. Furthermore, a promoter activator element located between positions --113 and --42 of the gene B1 and functional in the human MCF-7 and the Xenopus B3.2 cells has been identified and shown to be involved in the high level of induced transcription activity when the ERE is placed at a distance from the promoter. Finally, a hormone-controlled in vitro transcription system derived from Xenopus liver nuclear extracts was exploited to characterize two additional novel cis-acting elements within the vitellogenin gene B1 promoter. One of them, a negative regulatory element (NRE), is responsible for repression of promoter activity in the absence of hormone. The second is related to the NF-I binding site and is required, together with the ERE, to mediate hormonal induction. Moreover, we detected three trans-acting activities in Xenopus liver nuclear extracts that interact with these regions and demonstrated that they participate in the regulation of the expression of the vitellogenin promoter in vitro.

Validação de métodos cromatográficos por clae para análise das vitaminas B1, B2, B6 e niacina naturalmente presentes em farinha de cereais

Complex B vitamins are present in some cereal foods and the ingestion of enriched products contributes to the recommended dietary intake of these micronutrients. To adapt the label of some products, it is necessary to develop and validate the analytical methods. These methods must be reliable and with enough sensitivity to analyze complex B vitamins naturally present in food at low concentration. The purpose of this work is to evaluate, with validated methods, the content of vitamins B1, B2, B6 and niacin in five cereal flours used in food industry (oat, rice, barley, corn and wheat).

Photometric procedure for quantitative analysis of Aflatoxin B1 in peanuts by thin-layer chromatography using charge coupled device detector

A photometric procedure was developed for determination of aflatoxin B1 in peanuts by TLC-CCD technique. The quantification and detection limit were 1.2 μg kg-1 and 0.4 ng per spot, respectively, with mean recovery of 98%. The CCD camera is sufficiently sensitive to detect small changes in spots fluorescence intensity and the results for performance confirmed the efficiency of the method. Another important property of CCD detector is its linearity for a wide range of luminous stimulus determined by analysis of five-point calibration curves using the intensity of AFB1 fluorescence versus AFB1 concentration (0.8 to 4.8 ng per spot). The method was applied to the analysis of thirty nine peanut samples and aflatoxin B1 levels ranged from 16 to 115 μg kg-1. The TLC-CCD and the photometric procedure developed in this study demonstrated to be a simple and efficient tool for quantitative analyses of AFB1 in peanut samples.

Qualidade sanitária e produção de fumonisina B1 em grãos de milho na fase de pré-colheita

Trinta e seis (36) cultivares de milho foram avaliadas em relação à incidência de grãos ardidos, mofados e produção de fumonisina B1. Amostras de 1,2 kg de grãos foram analisadas visualmente para a quantificação de grãos ardidos (Fusarium subglutinans), mofados (Penicillium oxalicum) e para a análise de fumonisina B1. Os grãos ardidos foram submetidos à análise de sanidade (papel de filtro com congelamento) visando identificar os fungos a eles associados. A cultivar Hatã 3052 apresentou 7,6% de grãos ardidos, ultrapassando o limite de tolerância que é de 6,0%. As cultivares AG 5011, HT 7105-3, Dina 1000 e C 701 apresentaram 16,8% , 3,4%, 3,2% e 3,1% de grãos mofados, respectivamente, acima do limite de tolerância que é de 3,0%. O fungo Fusarium subglutinans (Gibberella fujikuroi var. subglutinans) foi o causador de grãos ardidos, cuja detecção variou de 50,0 a 99,0%. A análise de variância mostrou diferenças significativas entre as cultivares com relação às incidências de grãos ardidos e de grãos mofados. Com relação à produção de fumonisina B1, as cultivares Hatã 3052, NB 6077 e 983 P produziram 7,0; 6,1 e 5,9 µg.g-1 de grãos, respectivamente, diferindo significativamente da cultivar P3071 (2,2 µg.g-1 de grãos). Conclui-se que há diferenças significativas entre as cultivares de milho em relação à produção de grãos ardidos e mofados, bem como acentuada interação entre as cultivares e o fungo toxigênico Fusarium subglutinans (Gibberella fujikuroi var. subglutinans) quanto à biossíntese de fumonisina B1 em grãos de milho.

Equine leukoencephalomalacia (ELEM) due to fumonisins B1 and B2 in Argentina

In August 2007 an outbreak of neurological disease and sudden death in Arabian horses occurred in a farm located in Coronel Rosales County, Buenos Aires Province, Argentina. The animals were on a pasture of native grasses and supplemented ad libitum with corn kernels and wheat bran. Three horses were observed having acute neurologic signs including blindness, four leg ataxia, hyperexcitability, aimless walking and circling, followed by death in two of them. Four other horses were found dead overnight without a history of neurologic signs. The morbidity, mortality and lethality rates were 11.6%, 10% and 85.7%, respectively. Grossly, the brain showed focal areas of hemorrhage, brown-yellow discoloration and softening of the sub-cortical white matter. The microscopic brain lesions consisted of extensive areas of malacia within the white matter of the cerebral hemispheres, brainstem and cerebellum, characterized by rarefaction of the white matter with cavitations filled with proteinaceous edema, multifocal hemorrhages and mild infiltration by neutrophils, and rare eosinophils. Swollen glial cells with abundant eosinophilic cytoplasm, distinct cell borders, intracytoplasmic deeply eosinophilic globules and eccentric, hyperchromatic, occasionally pyknotic nucleus were present throughout the areas of rarefaction hemorrhage, edema and necrosis. The feed supplements contained 12,490µg/kg of fumonisin B1 and 5,251µg/ kg of fumonisin B2. This is the first reported outbreak of ELEM associated with consumption of feed supplements containing high concentrations of fumonisins in Argentina.