A gene regulatory network for root epidermis cell differentiation in Arabidopsis.

The root epidermis of Arabidopsis provides an exceptional model for studying the molecular basis of cell fate and differentiation. To obtain a systems-level view of root epidermal cell differentiation, we used a genome-wide transcriptome approach to define and organize a large set of genes into a transcriptional regulatory network. Using cell fate mutants that produce only one of the two epidermal cell types, together with fluorescence-activated cell-sorting to preferentially analyze the root epidermis transcriptome, we identified 1,582 genes differentially expressed in the root-hair or non-hair cell types, including a set of 208 "core" root epidermal genes. The organization of the core genes into a network was accomplished by using 17 distinct root epidermis mutants and 2 hormone treatments to perturb the system and assess the effects on each gene's transcript accumulation. In addition, temporal gene expression information from a developmental time series dataset and predicted gene associations derived from a Bayesian modeling approach were used to aid the positioning of genes within the network. Further, a detailed functional analysis of likely bHLH regulatory genes within the network, including MYC1, bHLH54, bHLH66, and bHLH82, showed that three distinct subfamilies of bHLH proteins participate in root epidermis development in a stage-specific manner. The integration of genetic, genomic, and computational analyses provides a new view of the composition, architecture, and logic of the root epidermal transcriptional network, and it demonstrates the utility of a comprehensive systems approach for dissecting a complex regulatory network.

Performance of urinary NT-proBNP in ambulatory settings.

BACKGROUND: NT-proBNP (N-terminal prohormone of brain natriuretic peptide) has been established as a useful biomarker in plasma for children with congenital heart disease (CHD). Plasma values were shown to correlate well with urinary values. We designed a study to investigate the general utility of urinary NT-proBNP in children with and without CHD in an ambulatory setting. MATERIAL AND METHODS: 202 children (mean age 93 months (1-225 months)) were included in the analysis. We investigated the performance of urinary NT-proBNP values determined from spot urine as a diagnostic tool for different forms of congenital heart disease. RESULTS: Urinary NT-proBNP is a good diagnostic tool for children with congenital heart disease (ROC area under the curve 0.807). Combining these values with the Ross-classification further improves the diagnostic power (ROC area under the curve 0.831) Analysis also showed significant differences between Lg10 urinary NT-proBNP values of healthy controls and those of children after corrective surgery. Furthermore, children who have completed the stages of Fontan palliation showed higher values than age matched controls. CONCLUSIONS: Urinary NT-proBNP can be used in an ambulatory setting to discriminate between relevant and nonrelevant CHD and might be valuable as a follow up parameter for children after biventricular repair or univentricular palliation. Age dependant urinary NT-proBNP normal values for children could be an easy-to-use tool for general practitioners as well as specialised clinics.

Variables influencing the accuracy of 2-dimensional and real-time 3-dimensional echocardiography for assessment of small volumes, areas, and distances: an in vitro study using static tissue-mimicking phantoms.

OBJECTIVES: The aim of this study was to assess the validity, accuracy, and reproducibility of real-time 3-dimensional (3D) echocardiography for small distances, areas, and volumes. METHODS: Real-time 3D echocardiography using matrix technology was performed in small calibrated tissue-mimicking phantoms and compared with 2-dimensional (2D) echocardiography. In a systematic variation of variables on data acquisition and analysis including different 3D workstations (manual disk summation versus semiautomatic border detection), the relative contributions of sources of errors were determined. The clinical relevance of the in vitro findings was assessed in 5 neonates and infants. RESULTS: Distance calculation was valid (mean relative error ± SD, -0.15% ± 1.2%). Underestimation of areas and volumes was significant for both 2D and 3D echocardiography (area: 2D, -7.0% ± 2.9%; 3D, -6.0% ± 2.8%; volume: 2D, -13.1% ± 4.5%; 3D, -6.7% ± 2.5%; P < .05). Adjustment of compression and gain on data acquisition (difference of the means: 2D, 11.6%; 3D, 17.9%), gain on postprocessing (3D, 3.4%), and the border detection algorithm on analysis (2D, 4.8%; 3D, 16.6%) had a highly significant effect on volume and area calculations (P < .001). In vivo, compression and gain on acquisition (3D, 19.1%) and the 3D workstation on analysis (3D, 22.2%) had a highly significant impact on left ventricular volumetry (P < .001). CONCLUSIONS: Real-time 3D echocardiography is a reliable method for calculation of small distances, areas, and volumes comparable with the size of the neonatal and infant heart. Variables influencing boundary identification during image acquisition and analysis have a significant impact on 2D and 3D area and volume calculations. Standardized protocols are mandatory to avoid these sources of error in both clinical practice and research.

A novel hemoglobin, Bonn, causes falsely decreased oxygen saturation measurements in pulse oximetry.

BACKGROUND: A 4-year-old boy and his father exhibited low oxygen saturation measured transcutaneously by pulse oximetry, a finding that could not be confirmed by arterial blood gas analysis. Both patients exhibited slight hemolysis in their blood, and the boy had a microcytic anemia. There was no evidence of hypoxemia or methemoglobinemia. Despite the normal results from the arterial blood gas analysis, a right-to-left-shunt was assumed in the boy until a cardiology examination excluded this diagnosis. Sleep apnea syndrome was suspected in the father and treated with nocturnal positive pressure respiration based on the low oxygen saturation values obtained with pulse oximetry. Only after consultation with our laboratory was a hemoglobin variant suspected and investigated. METHODS: We performed hemoglobin protein analysis by HPLC, electrophoretic separation, and spectrophotometry and DNA sequence analysis of the alpha-globin gene. RESULTS: Both HPLC chromatographic separation and alkaline electrophoresis revealed a unique hemoglobin peak. In both patients, alpha-globin gene sequencing revealed a mutation resulting in a histidine-to-aspartatic acid substitution at position alpha87. The low oxygen saturation measurement by pulse oximetry was due to hemoglobin Bonn oxyhemoglobin having an absorption peak at 668 nm, near the 660 nm measured by pulse oximeters. CONCLUSION: Hemoglobin Bonn is a novel hemoglobin variant of the proximal alpha-globin that results in falsely low oxygen saturation measurements with pulse oximetry.

The bHLH genes GL3 and EGL3 participate in an intercellular regulatory circuit that controls cell patterning in the Arabidopsis root epidermis.

The specification of the hair and non-hair cells in the Arabidopsis root epidermis provides a useful model for the study of cell fate determination in plants. A network of putative transcriptional regulators, including the related bHLH proteins GLABRA3 (GL3) and ENHANCER OF GLABRA3 (EGL3), is known to influence the patterning of these cell types. Here, we analyze the expression and regulation of GL3 and EGL3 during root epidermis development. Although they are thought to act in both the hair and non-hair cell types, we surprisingly found that GL3 and EGL3 gene expression and RNA accumulation occurs preferentially in the developing hair cells. By analyzing the expression of GL3::GUS and EGL3::GUS reporter fusions in various mutant and overexpression lines, we discovered that the expression of both genes is negatively regulated by WER, GL3 and EGL3 in the developing non-hair cells, and positively regulated by the CPC and TRY proteins in the developing hair cells. Further, the analysis of a GL3-YFP translational fusion, expressed under the GL3 promoter, indicates that the GL3 protein moves from the hair cells to the non-hair cells. These results suggest that GL3/EGL3 accumulation in the N cells is dependent on specification of the hair cell fate, which itself is known to be influenced (via CPC-mediated lateral inhibition) by the non-hair cells. This bi-directional signaling mechanism defines a new regulatory circuit of intercellular communication to specify the epidermal cell types.

The bHLH genes GLABRA3 (GL3) and ENHANCER OF GLABRA3 (EGL3) specify epidermal cell fate in the Arabidopsis root.

The position-dependent specification of the hair and non-hair cell types in the Arabidopsis root epidermis provides a simple model for the study of cell fate determination in plants. Several putative transcriptional regulators are known to influence this cell fate decision. Indirect evidence from studies with the maize R gene has been used to suggest that a bHLH transcription factor also participates in this process. We show that two Arabidopsis genes encoding bHLH proteins, GLABRA3 (GL3) and ENHANCER OF GLABRA3 (EGL3), act in a partially redundant manner to specify root epidermal cell fates. Plants homozygous for mutations in both genes fail to specify the non-hair cell type, whereas plants overexpressing either gene produce ectopic non-hair cells. We also find that these genes are required for appropriate transcription of the non-hair specification gene GL2 and the hair cell specification gene CPC, showing that GL3 and EGL3 influence both epidermal cell fates. Furthermore, we show that these bHLH proteins require a functional WER MYB protein for their action, and they physically interact with WER and CPC in the yeast two-hybrid assay. These results suggest a model in which GL3 and EGL3 act together with WER in the N cell position to promote the non-hair cell fate, whereas they interact with the incomplete MYB protein CPC in the H position, which blocks the non-hair pathway and leads to the hair cell fate.